title: Aneuploidy induces premature aging in yeast due to defects in Ribosome Quality Control [MoBYSeqData]

city: Madison

first: Audrey

last: Gasch

state: WI

address: 425G Henry mall

institute: University of Wisconsin

summary: Premature aging is a hallmark of Down syndrome, caused by trisomy of human chromosome 21;  but the reason is unclear and difficult to study in humans.  We used an aneuploid model in wild yeast to show that chromosome amplification disrupts nutrient-induced cell-cycle arrest, quiescence entry, and healthy aging, across genetic backgrounds and amplified chromosomes.  We discovered that these defects are due in part to aneuploidy-induced dysfunction in Ribosome Quality Control (RQC).  Aneuploids entering quiescence display aberrant ribosome profiles, accumulate RQC intermediates, and harbor an increased load of protein aggregates. Although they have normal proteasome capacity, aneuploids show signs of ubiquitin dysregulation, which impacts cyclin abundance to disrupt arrest.  Remarkably, inducing ribosome stalling in euploids produces similar aberrations, while up-regulating limiting RQC subunits or proteins in ubiquitin metabolism alleviates many of the aneuploid defects.  Our results raise major implications for other aneuploidy disorders including Down syndrome.

overall_design: Haploid euploid and aneuploid (carrying one additional copy of chromosome XII) yeast strains were transformed with a barcoded CEN plasmid library. For each strain, we took samples at the start of the experiment (Time 0), 24 hours after start (Day 1), and 28 days after start (Day 28) to quantify the effects of gene duplication on chronological aging. Each strain was measured in biological triplicate (3 replicates from 2 independent transformations of the plasmid library).

middle: P

first: Leah

middle: E

last: Escalante


title: GTestimate: Improving relative gene expression estimation in scRNA-seq using the Good-Turing estimator

status: Public on Jul 31 2024

city: Vienna

first: Martin

last: Fahrenberger

state: Vienna

address: Dr.-Bohr-Gasse 9

department: Structural and Computational Biology

country: Austria

institute: Max Perutz Labs

type: Expression profiling by high throughput sequencing

summary: Single-cell RNA-seq suffers from unwanted technical variation between cells, caused by its complex experiments and shallow sequencing depths. We present GTestimate a new normalization method based on the Good-Turing estimator, which improves upon conventional methods by accounting for unobserved genes. We validate GTestimate using new ultra-deep sequencing data, generated via a novel cell targeted PCR-amplification approach, and show substantial improvements in cell-cell distance estimation and downstream results.

overall_design:
A human cerebral-organoid scRNA-seq library was split into two aliquotes and sequenced; once as a ‘typical’ run and once after cell targeted PCR-amplification (cta-seq). This second run enabled a ‘ultra-deep’ sequencing depth for a small set of targeted cells.

***************************************************************
Submitter statement concerning raw data: Raw sequencing data could not be included in this submission due to patient privacy concerns. Raw-data will be made available through controlled access at the European Genome-Phenome Archive at a later date.
***************************************************************

first: Christopher

last: Esk

first: Arndt

last: von Haeseler


title: RNA-seq after chmA knockdown in PhiKZ-infected  PAO1 cells

city: Würzburg

first: Jörg

last: Vogel

address: Josef-Schneider-Straße 2

department: Institute for Molecular Infection Biology

country: Germany

institute: Würzburg University

summary: ChmA forms the phage nucleus in PhiKZ-infected PAO1 cells, we aimed to illuminate the effects on the transcriptome upon knockdown (KD) of ChmA translation through anti-sense-based technology. We identified phage transcripts that are affected in level upon KD of ChmA and clustering allowed to group these together that likely are expressed at a similar stage and dependency in the phage replication cycle.

sample_organism:
Pseudomonas aeruginosa
Pseudomonas phage phiKZ

overall_design: PAO1 cells were pretreated with anti-sense-based molecules against the start codon and ribosome binding site of the chmA transcript. Cells were infected with PhiKZ and at defined time-points post-infection, cells were harvested. RNA was extracted from cells and sequenced.

first: Milan

last: Gerovac


title: Aneuploidy induces premature aging in yeast due to defects in Ribosome Quality Control [RNASeqData]

department: Genetics

country: USA

overall_design: Euploid and select aneuploid Saccharomyces cerevisiae strains (haploids with one additional copy of chromosome IV, XII, XIV, or XV). We cultured each strain in rich media, and allowed cultures to exhaust glucose and enter quiescence. Samples for RNAseq were harvested at log-phase (proliferative growth) and at days 1, 3, and 7 after start of experiment to quantify differential gene expression as euploid and aneuploid cultures enter quiescence. Each strain and timepoint was measured in biological duplicate. An equivalent amount of Schizosaccharomyces pombe was doped into each S. cer sample for downstream normalization.


title: Molecular mechanisms of Target organ damage in youth with primary hypertension [bulk RNA-Seq 1]

city: Cincinnati

first: Kwangmin

last: Choi

state: Ohio

address: 333 Burnet Avenue

department: Experimental Hematology & Cancer Biology

institute: Cincinnati Children's Hospital Medical Center

summary: High blood pressure or Primary Hypertension is associated with target organ damage (TOD) in both adults and youth, where TOD predicts the adverse cardiovscular (CV) events. The mechanisms involved in blood pressure mediated TOD are limitedly understood in onset of  TOD. To understand this,  first using RNA sequencing, we identified candidate differentially expressed genes  in circulating PBMCs of study participants with and without hypertension and elevated  left ventrivlar mass index . Next, through miRNA and whole genome bisulphite sequencing of circulating PBMCs we determined the epigenetic regulators of TOD. Also, using LCMS, serum proteomics, the key circulatory protein involved in advancing CV- TOD events was identified. In conclusion,  multiomics approach was utilized to define the circulatory regulators involved in BP mediated CV-TOD.

overall_design: The design and methods of the SHIP AHOY (Study of High Blood Pressure in Pediatrics, Adult Hypertension Onset in Youth) project have been discussed elsewhere  (Mendizabal B, et al., 2018, Hypertension . 2018 Sep;72(3):625-631.). Briefly, in the current cross-sectional study, youth aged 11 to less than 19 years with BP levels (N=397) across the BP distribution, were enrolled at five clinical sites (Cincinnati, Houston, Philadelphia, Rochester, and Seattle) from 2015 to 2017 from pediatric specialty clinics (four nephrology clinics, and one joint nephrology and cardiology clinic). Efforts were made to achieve body mass index (BMI) balance across BP groups by periodically checking differences in BMI percentile among groups and actively recruiting normotensive obese controls. Out of 397 subjects, 132 adolescents of both sexes and all races with normal (Cut off: SBP <80th%), and high- systolic blood pressure (Cut off: SBP >90th%), and then stratified by with or without TOD, like Low LVMI (Left ventricular mass index) (Cut off: <38.6 gm/m2.7), and High LVMI (Cut off: >38.6 gm/m2.7), were selected in the present study cohort. Urine and Blood samples (for PBNCs and serum) were collected to investigate genetic, epigenetic, and proteomics changes that influence the development of hypertensive TOD in youth.  We excluded youth with symptomatic severe hypertension, on antihypertensive or lipid-lowering medication in the past 6 months, with diabetes mellitus (type 1 or 2), kidney disease, or other chronic medical conditions. All study participants and their parents provided written informed consent and assent according to local investigational review board requirements.

first: Tammy

middle: M

last: Brady

first: Kalyani

last: Ananthamohan

first: Mohammed

last: Arif

first: Stephen

last: Daniels

first: Bonita

last: Falkner

first: Michael

last: Ferguson

first: Joseph

middle: T

last: Flynn

first: Coral

last: Hanevold

last: Hooper

first: Julie

last: Inglefinger

first: Marc

last: Lande

first: Lisa

middle: J

last: Martin

first: Meyer

last: SK

first: Mark

last: Mitsnefes

first: Bernard

last: Rosner

first: Joshua

middle: A

last: Samuels

first: Gina

last: Kuffel

last: Zilliox

first: Richard

middle: C

last: Becker

first: Elaine

last: Urbina

first: Sakthivel

last: Sadayappan


title: Premature aging in aneuploid yeast is influenced by aneuploidy-induced defects in Ribosome Quality Control [RiboSeq]

first: Audrey

last: Gasch

overall_design: Euploid and select aneuploid Saccharomyces cerevisiae strains (haploids with one additional copy of chromosome XII) were cultured in rich media and allowed to exhaust glucose and enter stationary phase. Samples for ribosome profiling were harvested at log-phase and 4 days after start of experiment. Each strain and timepoint was measured in biological duplicate.

first: Leah

last: Escalante


title: Ad-SGE-DKK3 Gene Therapy Against Resistance to Immune Checkpoint Blockade in Malignant Mesothelioma

status: Public on Jul 30 2024

city: Houston

first: Sung Wook

last: Kang

state: TX

address: one plaza

department: Surgery

institute: Baylor college of medicine

overall_design: This phase 2 study evaluates the combination of Ad-SGE-Dickkopf-3 (DKK3) gene therapy and nivolumab in patients with chemotherapy-refractory epithelioid malignant pleural mesothelioma (MPM). This clinical study was based on our preclinical study to overcome primary immune resistance by combining Ad-SGE-DKK3 gene therapy with anti-PD-1 therapy through DKK3-driven immune modulation in mesothelioma. The study enrolled 12 patients between 2019 and 2022, who were refractory to pemetrexed-platin-based chemotherapy. Treatment involved CT-guided intratumoral injections of Ad-SGE-DKK3 and systemic nivolumab administration. The primary goal was to assess the objective response rate (ORR), with secondary and exploratory objectives focusing on safety, durable clinical benefit (DCB), survival, and biomarker changes. Of the enrolled patients, 75% completed the Ad-SGE-DKK3 treatment protocol. The study observed a 16.6% ORR with 41.7% maintaining stable disease, which reached 58.3% DCB rate. The median overall survival was 14.5 months, and the median progression-free survival was 4.5 months. Adverse events of grade 3 were noted in 41.7% of patients. Serial analysis of tumor biopsies and serum biomarkers indicated that patients with DCB had increased tumor-infiltrating CD8 T cells, decreased circulating memory CD8 T cells, and sustained lower concentrations of soluble mesothelin and M-CSF than progressors. The combination therapy demonstrated a favorable safety profile and promising efficacy in this patient population. ClinicalTrials.gov identifier: NCT04013334.

first: Hyun-Sung

last: Lee

first: Bryan

last: Burt

first: Sung


title: Molecular mechanisms of Target organ damage in youth with primary hypertension

first: Kwangmin

last: Choi

type: Methylation profiling by high throughput sequencing

type: Non-coding RNA profiling by high throughput sequencing

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Molecular mechanisms of Target organ damage in youth with primary hypertension [bisulfite-Seq]

overall_design: The design and methods of the SHIP AHOY (Study of High Blood Pressure in Pediatrics, Adult Hypertension Onset in Youth) project have been discussed elsewhere.1 Briefly, in the current cross-sectional study, youth aged 11 to less than 19 years with BP levels (N=397) across the BP distribution, were enrolled at five clinical sites (Cincinnati, Houston, Philadelphia, Rochester, and Seattle) from 2015 to 2017 from pediatric specialty clinics (four nephrology clinics, and one joint nephrology and cardiology clinic). Efforts were made to achieve body mass index (BMI) balance across BP groups by periodically checking differences in BMI percentile among groups and actively recruiting normotensive obese controls. Out of 397 subjects, 132 adolescents of both sexes and all races with normal (Cut off: SBP <80th%), and high- systolic blood pressure (Cut off: SBP >90th%), and then stratified by with or without TOD, like Low LVMI (Left ventricular mass index) (Cut off: <38.6 gm/m2.7), and High LVMI (Cut off: >38.6 gm/m2.7), were selected in the present study cohort. Urine and Blood samples (for PBNCs and serum) were collected to investigate genetic, epigenetic, and proteomics changes that influence the development of hypertensive TOD in youth.  We excluded youth with symptomatic severe hypertension, on antihypertensive or lipid-lowering medication in the past 6 months, with diabetes mellitus (type 1 or 2), kidney disease, or other chronic medical conditions. All study participants and their parents provided written informed consent and assent according to local investigational review board requirements.

first: Tammy

last: Brady

first: Kalyani

last: Ananthamohan

first: Mohammed

last: Arif

first: Stephen

last: Daniels

first: Bonita

last: Falkner

first: Michael

last: Ferguson

first: Joseph

last: Flynn

first: Coral

last: Hanevold

last: Hooper

first: Julie

last: Inglefinger

first: Marc

last: Lande


title: Molecular mechanisms of Target organ damage in youth with primary hypertension [smallRNA-Seq 1]

overall_design: The design and methods of the SHIP AHOY (Study of High Blood Pressure in Pediatrics, Adult Hypertension Onset in Youth) project have been discussed elsewhere (Mendizabal B, et al., 2018, Hypertension . 2018 Sep;72(3):625-631.).  Briefly, in the current cross-sectional study, youth aged 11 to less than 19 years with BP levels (N=397) across the BP distribution, were enrolled at five clinical sites (Cincinnati, Houston, Philadelphia, Rochester, and Seattle) from 2015 to 2017 from pediatric specialty clinics (four nephrology clinics, and one joint nephrology and cardiology clinic). Efforts were made to achieve body mass index (BMI) balance across BP groups by periodically checking differences in BMI percentile among groups and actively recruiting normotensive obese controls. Out of 397 subjects, 132 adolescents of both sexes and all races with normal (Cut off: SBP <80th%), and high- systolic blood pressure (Cut off: SBP >90th%), and then stratified by with or without TOD, like Low LVMI (Left ventricular mass index) (Cut off: <38.6 gm/m2.7), and High LVMI (Cut off: >38.6 gm/m2.7), were selected in the present study cohort. Urine and Blood samples (for PBNCs and serum) were collected to investigate genetic, epigenetic, and proteomics changes that influence the development of hypertensive TOD in youth.  We excluded youth with symptomatic severe hypertension, on antihypertensive or lipid-lowering medication in the past 6 months, with diabetes mellitus (type 1 or 2), kidney disease, or other chronic medical conditions. All study participants and their parents provided written informed consent and assent according to local investigational review board requirements.

first: Lisa

last: Martin

first: Meyer

last: SK

first: Mark

last: Mitsnefes

first: Bernard

last: Rosner

first: Joshua

last: Samuels

first: Gina

last: Kuffel

last: Zilliox

first: Richard

last: Becker

first: Elaine

last: Urbina

first: Sakthivel

last: Sadayappan


title: AP-3-84 effects on gene expression in thioglycollate-induced, ID8 ascites or bone-marrow derived macrophages

status: Public on Jul 26 2024

city: Philadelphia

first: Priyankara

last: Wickramasinghe

state: PA

address: 3601 Spruce Street

department: Bioinformatics

institute: The Wistar Institute

summary: Here we studied the effects of AP-3-84 treatment on gene expression in different types of macrophages: thioglycollate-induced macrophages, ascites macrophages from ID8 ovarian cancer-bearing mice, bone marrow-derived macrophages. We also studied whether AP-3-84 effects are mediated by p53 using bone marrow-derived macrophages from p53 KO mice. We found the dramatic changes in gene expression upon AP-3-84 treatment in all types of macrophages with the induction of major stress response programs and apoptosis signaling.

overall_design: We isolated thioglycollate-induced macrophages, ascites macrophages from ID8 ovarian cancer-bearing mice. We also generated M0, M1 or M2-polarized macrophages using hematopoietic bone marrow stem cells from wild-type B6 or p53 KO B6 mice. We then treated these different types of macrophages with 10 uM AP-3-84 for 4 hours and analyzed gene expression changes by bulk RNA Sequencing.

first: Evgenii

middle: N

last: Tcyganov

first: Taekyoung

last: Kwak

first: Luis

last: Montaner


title: Parvalbumin expression does not account for discrete electrophysiological profiles of glutamatergic ventral pallidal subpopulations.

status: Public on Jul 29 2024

city: Saint Louis

first: Meaghan

last: Creed

state: MO

address: 4370 Duncan Ave

department: Anesthesiology

institute: Washington University

summary: The ventral pallidum (VP) has emerged as a critical node in the mesolimbic reward system. Modulating the VP can impact the subjective valuation of rewards, reward motivation, and reward seeking under conflict, making it an attractive target for clinical neuromodulation therapies that manage substance use disorders. To understand how to rationally modulate the VP, we need a better understanding of the electrophysiological properties of VP neurons and the molecular and biophysical determinants of these properties. Here, we used patch-clamp electrophysiology to characterize the intrinsic properties of glutamatergic VP (VPGlu) neurons and observed two distinct electrophysiological profiles: VPGlu neurons that undergo depolarization block in response to progressively increasing current injection amplitudes and those that were resistant to depolarization block. To explore the mechanisms that could contribute to these distinct profiles, we used targeted ribosome affinity purification to identify ion channel subunits and regulatory proteins by isolating actively transcribed mRNA selectively from VPGlu neurons. We then used this transcriptomic information to implement a Markov Chain Monte Carlo method to parameterize a large population of biophysically distinct multicompartment models of VPGlu neurons conforming to either subpopulation. Based on prior literature suggesting parvalbumin (PV) is expressed in a subset of VPGlu neurons, and that PV expression governs the firing properties of those neurons, we tested the hypothesis that PV expression accounted for differences in subgroups, by increasing the maximal firing frequency and conferring resistance to depolarization block. In contrast, our model determined that PV expression at physiological levels had no effect on maximum firing rate. However, supraphysiological expression levels of PV appeared to induce a depolarization block in previously depolarization block-resistant neuron models, suggesting that other intracellular calcium-binding proteins could play a role in determining the firing phenotype of VPGlu neurons. We corroborated this result with single-cell patch-clamp RT-PCR, which confirmed that PV expression did not distinguish the two electrophysiologically distinct subpopulations. Together, these findings establish that VPGlu neurons are composed of biophysically distinct subpopulations that have not been appreciated in prior studies interrogating the function of this population. With the advent of novel tools for cell-type specific pharmacology and targeted neurostimulation, this understanding will be critical for developing strategies to rationally modulate VPGlu cells to treat disorders characterized by maladaptive reward seeking.

overall_design: We used translating ribosome affinity purification (TRAP) to isolate RNA bound to ribosomes in glutamatergic neurons in the ventral pallidum and performed bulk RNA sequencing and differential gene expression analysis to compare this TRAP fraction to the input fraction.

last: Fang

first: Robert

last: Graham


title: Cancer-associated snaR-A noncoding RNA interacts with core splicing machinery and disrupts processing of mRNA subpopulations

status: Public on Jul 24 2024

city: Urbana

first: Sihang

last: Zhou

state: IL

address: 601 S Goodwin Ave

institute: University of Illinois Urbana-Champaign

summary: RNA polymerase III (Pol III) activity in cancer is linked to the production of small noncoding (nc)RNAs that are otherwise silent in most tissues. snaR-A (small NF90-associated RNA isoform A) - a hominid-specific ncRNA shown to enhance cell proliferation, migration, and invasion - is a cancer-emergent Pol III product that remains largely uncharacterized despite promoting growth phenotypes. Here, we applied a combination of genomic and biochemical approaches to predict the biogenesis and subsequent protein interactions of snaR-A to better understand its role as a putative driver of cancer progression. By profiling the chromatin landscapes across a multitude of primary tumor types, we show that predicted snaR-A upregulation is broadly linked with unfavorable outcomes among cancer patients. At the molecular level, we unexpectedly discover widespread interactions between snaR-A and mRNA splicing factors, including SF3B2 - a core component of the U2 small nuclear ribonucleoprotein (snRNP). We find that SF3B2 levels are sensitive to high snaR-A abundance, and that depletion of snaR-A alone is sufficient to decrease intron retention levels across subpopulations of mRNA enriched for U2 snRNP occupancy. snaR-A sensitive genes are characterized by high GC content, close spatial proximity to nuclear bodies concentrated in pre-mRNA splicing factors, and functionally enriched for proteins involved in deacetylation and autophagy. We highlight examples of splicing resolution and increased protein levels following snaR-A depletion for a wide-ranging set of factors, suggesting snaR-A-driven splicing defects may have far-reaching effects that re-shape the cellular proteome. These findings help clarify the molecular activities and consequences of snaR-A express- ion in cancer, and altogether establish a novel mechanism through which Pol III overactivity may promote tumorigenesis.

overall_design: To investigate the effect of snaR-A on splicing of mRNAs, we performed snaR-A knockdown using three different siRNAs targeting snaR-A in HEK293T cells and deep sequenced the samples. The experiment included three biological replicates for both the scramble knockdown and the snaR-A knockdown. Raw mRNA signal counts were extracted.

first: Kevin

last: Van Bortle


title: Molecular mechanisms of Target organ damage in youth with primary hypertension [bulk RNA-Seq 2]

first: Kwangmin

last: Choi

first: Tammy

last: Brady

first: Kalyani

last: Ananthamohan

first: Mohammed

last: Arif

first: Stephen

last: Daniels

first: Bonita

last: Falkner

first: Michael

last: Ferguson

first: Joseph

last: Flynn

first: Coral

last: Hanevold

last: Hooper

first: Julie

last: Inglefinger

first: Marc

last: Lande

first: Lisa

last: Martin

last: SK

last: Mitsnefes


title: Neutralization of truncated procalcitonin by antibody treatment protects vascular barrier integrity during systemic inflammation

status: Public on Jul 22 2024

city: Münster

first: Klaus

last: Schughart

address: Von-Esmarch-Str 56

department: Virology

institute: University Münster

summary: Differentially expressed genes were identified in lung endothelial cells of CLP-treated mice compared to sham-treated controls and after treatment with an antibody against truncated murine procalcitonin.

overall_design: Sepsis was induced in mice by CLP (Cecal Ligation Puncture). Some mice were treated with an antibody against murine truncated procalcitonin. Endothelial cells from the lungs of CLP-treated, CLP plus antibody treatment, sham-treated and untreated mice were isolated and subjected to RNAseq analysis.

first: Nana-Maria

last: Wagner

first: Laura

last: Brabenec

first: Katharina

last: Hellenthal


title: Uniform dynamics of cohesin-mediated loop extrusion in living human cells

status: Public on Jul 28 2024

city: PARIS

first: Christophe

last: Zimmer

address: 25-28 rue du docteur Roux

department: Computational Biology

country: France

institute: Institut Pasteur

summary: Most animal genomes are partitioned into Topologically Associating Domains (TADs), created by cohesin-mediated loop extrusion and defined by convergently oriented CCCTC-binding factor (CTCF) sites. The dynamics of loop extrusion and its regulation remain poorly characterized in vivo. Here, we tracked the motion of TAD anchors in living human cells to visualize and quantify cohesin-dependent loop extrusion across multiple endogenous genomic regions. We show that TADs are dynamic structures whose anchors are brought in proximity about once per hour and for 6-19 min (~16% of the time). Moreover, TADs are continuously subjected to extrusion by multiple cohesin complexes. Remarkably, despite strong differences of Hi-C patterns between the chromatin regions, their dynamics is consistent with the same density, residence time and speed of cohesin. Our results suggest that TAD dynamics is governed primarily by the location and affinity of CTCF binding sites, which allows genome-wide predictive models of cohesin-dependent chromatin interactions.

overall_design: We performed Capture Micro-C in HCT116 cells treated or not with 500 𝞵M auxin (leading to RAD21 depletion) for 3 hours. We used cell lines with TetO and CuO repeats inserted in the genome at the L1, L2, T1 and No Loop genomic regions.

first: Thomas

last: Sabaté

first: Benoît

last: Lelandais

first: Marie-Cécile

last: Robert

last: Szalay

first: Jean-Yves

last: Tinevez

first: Edouard

last: Bertrand


title: Molecular mechanisms of Target organ damage in youth with primary hypertension [smallRNA-Seq 2]

first: Kwangmin

last: Choi

first: Tammy

last: Brady

first: Kalyani

last: Ananthamohan

first: Mohammed

last: Arif

first: Stephen

last: Daniels

first: Bonita

last: Falkner

last: Ferguson

last: Flynn

last: Hanevold

last: Hooper

first: Meyer

first: Mark

first: Bernard

last: Rosner

first: Joshua

last: Samuels

first: Gina

last: Kuffel

last: Zilliox

first: Richard

last: Becker

first: Elaine

last: Urbina

first: Sakthivel

last: Sadayappan


title: ARID1A-BAF coordinates ZIC2 genomic occupancy for epithelial to mesenchymal transition in cranial neural crest lineage commitment.

status: Public on Jul 23 2024

city: London

first: Marco

last: Trizzino

state: UK

address: Imperial College Road

department: Life Sciences

country: United Kingdom

institute: Imperial College London

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The BAF chromatin remodeler regulates lineage commitment including cranial neural crest cell (CNCC) specification. BAF subunit mutations cause Coffin-Siris Syndrome (CSS), a congenital disorder characterized by distinct craniofacial features and intellectual disability. Approximately 50% of CSS patients carry mutations in one of the mutually exclusive BAF subunits, ARID1A/ARID1B. While Arid1a deletion in mouse neural crest causes severe craniofacial phenotypes, little is known about the role of ARID1A in CNCC specification. Using CSS patient-derived ARID1A+/- iPSCs to model CNCC specification, we discovered that ARID1A-haploinsufficency impairs epithelial to mesenchymal transition (EMT), a process necessary for CNCC delamination and migration from the neural tube. Furthermore, wild-type ARID1A-BAF regulates enhancers associated with EMT genes. ARID1A-BAF binding at these enhancers is impaired in heterozygotes, while binding at promoters is unaffected. At the sequence level, these EMT enhancers contain binding motifs for ZIC2, and ZIC2 binding at these sites is ARID1A-dependent. When excluded from EMT enhancers, ZIC2 relocates to neuronal enhancers, triggering aberrant neuronal gene activation. In mice, deletion of Zic2 impairs NCC delamination, while ZIC2 overexpression in chick embryos at pre-migratory neural crest stages elicits abnormal cell delamination from the neural tube. These findings reveal a novel ARID1A-ZIC2 axis essential for EMT and CNCC delamination.

overall_design:
RNA-seq of two Coffin Siris Syndrome patient-derived ARID1A-haploinsufficient iPSC lines and two control iPSC lines  (3 replicates per line)
RNA-seq of two Coffin Siris Syndrome patient-derived ARID1A-haploinsufficient cell lines and two control cell lines at day 10 of in vitro cranial neural crest cell specification (3 replicates per line)
RNA-seq of two Coffin Siris Syndrome patient-derived ARID1A-haploinsufficient cell lines and two control cell lines at day 14 of in vitro cranial neural crest cell specification in early maintenance media (3 replicates per line)
RNA-seq of two Coffin Siris Syndrome patient-derived ARID1A-haploinsufficient cell lines and two control cell lines at day 14 of in vitro cranial neural crest cell specification in late maintenance media (at least 2 replicates per line)
ChIP-seq of ZIC2 in two Coffin Siris Syndrome patient-derived ARID1A-haploinsufficient cell lines and two control cell lines at day 10 of in vitro cranial neural crest cell specification
ChIP-seq of H3K27ac in two Coffin Siris Syndrome patient-derived ARID1A-haploinsufficient cell lines and two control cell lines at day 10 of in vitro cranial neural crest cell specification
ChIP-seq of ARID1A in two Coffin Siris Syndrome patient-derived ARID1A-haploinsufficient cell lines and two control cell lines at day 10 of in vitro cranial neural crest cell specification
ATAC-seq of two Coffin Siris Syndrome patient-derived ARID1A-haploinsufficient cell lines and two control cell lines at day 10 of in vitro cranial neural crest cell specification (2 replicates per line)

first: Samantha

last: Barnada

first: Aida

last: Giner de Gracia

first: Cruz

last: Morenilla-Palao

first: María

last: López-Cascales

first: Chiara

last: Scopa

first: Francis

last: Waltrich

first: Maria

last: Cicardi

first: Jonathan

last: Karlin

first: Davide

last: Trotti

first: Kevin

last: Peterson

last: Brugmann

first: Gijs

last: Santen

first: Steven

middle: B

last: McMahon

first: Eloísa

last: Herrera


title: Comparison of transcriptome landscape between adenine-induced and nephrectomy-induced uremic cardiomyopathy

city: Guangzhou

first: Jing-Fu

last: Bao

address: 1838 North Guangzhou Ave

country: China

institute: Southern Medical University

summary: Adenine and nephrectomy can be used to establish models of uremic cardiomyopathy in rodents. Although these two approaches cause similar phenotypes of heart, adenine may directly act on cardiomyocytes and lead to different molecular changes from nephrectomy-induced uremic cardiomyopathy. Transcriptome analysis on left ventricles showed 789 upregulated and 345 downregulated genes in mice with nephrectomy-induced uremic cardiomyopathy compared to control mice, while 432 upregulated and 155 downregulated genes in mice with adenine diet-induced uremic cardiomyopathy compared to control mice. Functional analysis revealed that these DEGs were involved in pathways related to immune inflammation, metabolic responses, and synaptic transmission. Further analysis demonstrated that nephrectomy-induced uremic cardiomyopathy has enhanced cell cycle process/extracellular matrix remodeling and suppressed fatty acid metabolism, whereas adenine diet-induced uremic cardiomyopathy only exhibits increased inflammatory response/extracellular matrix remodeling. Compared with nephrectomy-induced uremic cardiomyopathy, adenine diet-induced uremic cardiomyopathy showed improved oxidative phosphorylation, aerobic electron transport chain, and tricarboxylic acid cycle, and suppressed cell cycle-related process, indicating that adenine and its derivates may affect the process of chronic kidney induced cardiac changes.

overall_design: To measure transcriptomic changes genome-wide, we performed RNA sequencing using left ventricular apexes of mice with nephrectomy-induced and adenine diet -induced uremic cardiomyopathy


title: Peyer’s patches are required for Bacteroides fragilis to induce protective systemic IgA

status: Public on Jul 25 2024

city: Syracuse

first: Joel

last: Wilmore

state: NY

address: 766 Irving Ave

department: Microbiology and Immunology

institute: SUNY Upstate Medical University

summary: Gut-educated IgA-secreting plasma cells that disseminate beyond the mucosa and into systemic tissues can help prevent disease in several contexts. Here we show, the commensal bacteria Bacteroides fragilis (Bf), is an efficient inducer of systemic IgA responses. The generation of bone marrow IgA plasma cells and high levels of serum IgA specific to Bf requires robust intestinal colonization. Bf-specific IgA responses were severely diminished in mice lacking Peyer’s patches, but not mice lacking a cecal patch. Colonization resulted in few changes in the host transcriptional profile in the gut, suggesting a commensal relationship. High levels of Bf-specific serum IgA, but not IgG, provided protection from peritoneal abscess formation in a bowel perforation model of Bf dissemination. These findings demonstrate a critical role for bacterial colonization and Peyer’s patches in the induction of robust systemic IgA responses that confer protection from bacterial dissemination originating from the gut.

overall_design: Mice were treated with B. fragilis by oral gavage and given either 1 or 6 doses and then euthanized 2 or 4 weeks after treatment. A 2cm piece of colon or small intestine were washed to remove feces and then RNA was isolated for RNAseq analysis.

middle: R

first: Joshua

last: Harris


title: Molecular mechanisms of Target organ damage in youth with primary hypertension [bulk RNA-Seq 3]

first: Kwangmin

last: Choi

first: Tammy

last: Brady

first: Kalyani

last: Ananthamohan

first: Mohammed

last: Arif

first: Stephen

last: Daniels

first: Bonita

last: Falkner

first: Michael

last: Ferguson

first: Joseph

last: Flynn

first: Coral

last: Hanevold

last: Hooper

first: Julie

last: Inglefinger

first: Marc

last: Lande

first: Lisa

last: Martin

first: Meyer

last: SK

first: Mark

last: Mitsnefes

first: Bernard

last: Rosner

last: Samuels

first: Gina

last: Kuffel

last: Zilliox

first: Richard

last: Becker

first: Elaine

last: Urbina

first: Sakthivel

last: Sadayappan


title: Identifications of vernalization regulatory elements by epigenomic approaches in winter wheat (ATAC-Seq)

city: Beijing

first: Zefu

last: Lu

state: Haidian

address: No.12 Zhongguancun South St.,Haidian District, Beijing, P.R.China

institute: Institute of Crop Sciences

summary:
Epigenetic modification plays a key role in the vernalization process and maintains gene expression states after low temperature. In this study, the Chinese main cultivar Aikang58 was used to construct the chromatin accessibility maps, expression profile and histone modification (including H3K27me3, H3K27ac, H3K36me3 and H3K4me3) maps of winter wheat before and after vernalization.  We provided a crucial data resource for deep understanding of epigenome in vernalization. What’s more, important core regulatory elements and corresponding transcription factors were discovered, and the complex regulatory network of wheat vernalization was analyzed.
ATAC-seq of leaf, axillary bud and shoot apex before and after vernalization


title: Satellite glial contact enhances differentiation and maturation of human induced sensory neurons

city: Boston

first: Chelsey

last: LeBlang

state: Massachusetts

address: 360 Longwood Ave

department: Cancer Biology

institute: Dana Farber Cancer Institute

summary: Induced sensory neurons (iSNs) generated from pluripotent stem cells are used to model human peripheral neuropathies, however current differentiation protocols produce sensory neurons with an embryonic phenotype. Peripheral glial cells contact sensory neurons early in development and contribute to formation of the canonical pseudounipolar morphology, but these signals are not encompassed in current iSN differentiation protocols. Here, we show that terminal differentiation of iSNs in co-culture with rodent Dorsal Root Ganglion satellite glia (rSG) advances their differentiation and maturation. Co-cultured iSNs develop pseudounipolar morphology through contact with rSGs. This transition depends on semaphorin-plexin guidance cues and on glial gap junction signaling. In addition to morphological changes, iSNs terminally differentiated in co-culture exhibit enhanced spontaneous action potential firing, more mature gene expression, and increased susceptibility to paclitaxel induced axonal degeneration. Thus, iSNs differentiated in coculture with rSGs provide a better model for understanding human peripheral neuropathies.

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design: To determine the impact of iSN-rDRG glial coculture on iSN development we compared human mRNA  from iSN- rDRG glial cocultures (Glia_iSN_Co) with iSNs cultured alone (iSN only). We also explored the impact of maturation media on glial cell populations comparing rat mRNA from rDRG glial cultures maintained in baseline media (Glia Only),rDRG glial cultures treated with maturation media (Glia_NCATS), and rDRG glial cultures that were cocultured with iSNs (Glia_iSN_co).

first: Rosalind

last: Segal


title: Identifications of vernalization regulatory elements by epigenomic approaches in winter wheat (RNA-Seq)

summary: Epigenetic modification plays a key role in the vernalization process and maintains gene expression states after low temperature. In this study, the Chinese main cultivar Aikang58 was used to construct the chromatin accessibility maps, expression profile and histone modification (including H3K27me3, H3K27ac, H3K36me3 and H3K4me3) maps of winter wheat before and after vernalization.  We provided a crucial data resource for deep understanding of epigenome in vernalization. What’s more, important core regulatory elements and corresponding transcription factors were discovered, and the complex regulatory network of wheat vernalization was analyzed.

overall_design: RNA-seq of leaf, axillary bud and shoot apex before and after vernalization


title: Profiling the rat cortical single-cell transcriptome reveals the potential mechanism of specific mode electroacupuncture stimulation opening the blood-brain barrier

city: Hangzhou

first: Congcong

last: Ma

state: Zhejiang

address: No. 548, binwen Road

institute: Zhejiang Chinese Medical University

summary: The existence of the blood-brain barrier (BBB) dramatically limits the drug treatment of central nervous system diseases such as stroke, neurodegenerative diseases, and brain tumors. Due to the complexity of the structure and function of the blood-brain barrier, it has been challenging to develop central nervous system drug delivery strategies and reveal critical biological mechanisms. Our previous studies have confirmed that specific mode electroacupuncture stimulation (EA) can effectively open the BBB in rats. Here, we utilized single-cell RNA sequencing (scRNA-seq) to map the cell population of rat cerebral cortex comprehensively. We identified 23 cell subsets and 8 types of cells in the brain obtained by cell annotation, and explored the potential mechanism of EA to open the blood-brain barrier by bioinformatics analysis of specific cell subsets and cell types, as well as cell communications. In summary, scRNA-seq uncovers the transcriptional changes of the cerebral cortex under EA at the single cell level; our results provide rich insights into the molecular and cellular architecture changes of the brain after EA intervention and provide a new reference for BBB material transport and its drug delivery research in health and disease, providing new potential strategies into the mechanism of BBB opening and provide essential information for designing rational treatment regimens and optimizing drug delivery.

overall_design: Rats were randomly divided into electroacupuncture group and control group ( n = 5 ). Rats in the EA group received electroacupuncture intervention, and rats in the CON group received only the same restraint treatment. The frontal lobe tissues of bilateral cerebral cortex of rats were taken for single cell sequencing.

first: Zhaoxing

last: Jia

first: Lin

last: Gan

first: Xianming

last: Lin


title: Identifications of vernalization regulatory elements by epigenomic approaches in winter wheat


title: Cadherins modulate the self-organizing potential of gastruloids

status: Public on Jul 18 2024

city: LAUSANNE

first: ALEXANDRE

last: MAYRAN

state: Waadt

address: Station 19

country: Switzerland

institute: EPFL


title: Identifying regulatory variants using multiome single-cell sequencing in prostate cell lines

city: TAMPA

first: Liang

last: Wang

state: FL

address: 14059 Riveredge Drove

institute: Moffitt Cancer Center

summary: Despite significant progresses, the genetic roles of regulatory elements in gene expression still remain largely unknown in prostate cells. Recent development in single cell sequencing has made it possible to combine ATAC-seq and RNA-seq to determine genome-wide linkages between chromatin accessibilities and gene expression. To test the feasibility of using single cell multiome sequencing in dissecting regulatory linkages between chromatin accessibilities and gene expressions, we applied 10X Multiome ATAC + Gene Expression platform to simultaneously encapsulate Tn5 transposase tagged nuclei from multiple prostate cell lines. Based on these multiomic data, we developed subsampling linkage analysis to identify linkage associations between open chromatin and gene expressions. Moreover, we implemented an innovative analytical method to investigate RNA expression alterations related to germline variant locus accessibilities at single cell levels, i.e., expression quantitate accessible loci (eQAL) analysis. Our data and methodology will complement traditional eQTL analysis and foresee more genetic applications aiming to clarify regulatory elements by single cell sequencing.

overall_design: Multiomic profilings in human prostate cell lines were captured using 10x Multiome ATAC + Gene Expression assay, and targeted hybridization enrichment was further used to collect signals on eQTL locus.


title: FOXQ1 Regulates Mitochondrial Cristae Formation and Energy Metabolism in Mouse Brain Endothelial Cells [RNA-seq]

status: Public on Jul 16 2024

first: Jianwei

last: Jiao

state: Beijing

address: 1 Beichen West Road

department: State Key Laboratory of Reproductive Biology

institute: Institute of Zoology, Chinese Academy of Sciences

summary:
Purpose: To gain  futher insight into how FOXQ1 regulates the brain endothelial cells ,RNA-seq was used to analyze the genome-wide changes resulting from the cerebral cortical endothelial cells  of E18 FOXQ1 conditional knock out mice and littermate wild-type.
Methods: Total RNA from E18 Flow sorting of endothelial cells of wild-type(WT) and FOXQ1fl/fl;Tie2-Cre mice was extracted. Specifically, Agilent 2100 Bioanalyze was used to quality controlled and quantified. then, total RNA was converted to cDNA and bound the library. RNA-sequencing analysis was used by the Illumina HiSeq 6000 platform in Annoroad Genomics
Approximately  one thousand transcripts showed differential expression between the wild-type(WT) and FOXQ1fl/fl;Tie2-Cre mice cortical endothelial cells, with a fold change ≥1.5 and p value <0.05. These results reflected FOXQ1 plays a role in brain endothelial cells.
Conclusions:  RNA-seq based transcriptome characterization would provide a overall understanding how FOXQ1 works in endothelial cells.

overall_design: Total mRNA profiles of E18 wild-type(WT) and FOXQ1fl/fl;Tie2-Cre mice were generated by deep sequencing, using Illumina HiSeq 6000.

first: Wenzheng

last: Zou


title: Identifications of vernalization regulatory elements by epigenomic approaches in winter wheat (ChIP-Seq)

overall_design: ChIP-seq of leaf and vernalized leaf, including H3K27me3, H3K27ac, H3K36me3 and H3K4me3


title: Cadherins modulate the self-organizing potential of gastruloids (ATAC-seq)

summary: Gastruloids have recently emerged as an efficient four-dimensional model for studying some aspects of post-implantation embryonic patterning. Their ability to undergo gastrulation-like processes leading to the self-organization into highly reproducible biological objects, has opened new avenues to investigate early embryonic patterning. Here, we sought to uncover the molecular and cellular mechanism underlying this remarkable property. We report that self-organization competence is associated with a cell-specific coordination of a Cadherin switch. We find that N-Cadherin hinders gastruloids morphogenetic competence, for its inactivation leads to the formation of trunk-like structures without relying on the otherwise requested extra-cellular matrix analogues such as Matrigel. In contrast, the repression of E-Cadherin by Snai1 is critical for self-organization: Snai1 establishes a cell-specific repressive pace by triggering the repression of a pluripotency-associated transcription program and its chromatin landscape, thus allowing a proper transition from E- to N-Cadherin to occur. Altogether, this work establishes a molecular mechanism that integrates the exit from pluripotency and the pace of cell differentiation, leading to the observed self-organizing potential of gastruloids.

overall_design: ATAC-seq on gastruloids from WT and mutant conditions

first: Alexandre

last: Mayran

first: Rita

last: Amândio

first: Lucille

last: Lopez-Delisle

first: Denis

last: Duboule


title: Single-cell transcriptomic profiling of mouse enteric neurons and enteric mesenchymal cells in enteric neurosphere cultures.

first: Rhian

last: Stavely

state: MA

address: 185 Cambridge Street, CPZN, 6100

department: Pediatric Surgery

institute: Massachusetts General Hospital

summary: This study was undertaken to define the molecular subtypes of cells within enteric neurospheres. Methods: We performed single-cell RNA sequencing on 3-dimensional neurosphere cultures derived from the small intestines of Plp1::GFP mice. These data show profiling results from the PLP1::GFP negative cell fraction ancillary to the positive fraction in series GSE184981. Results: We identify populations of enteric neurons and fibroblasts in the PLP1::GFP negative cell fraction of enteric neurospheres.

overall_design: Use of 10X Genomics technology to analyze single-cell transcriptomics in 3-dimensional neurosphere cultures derived from the small intestine after performing FACS to purify the PLP1::GFP negative cell fraction.

first: Richard

last: Guyer

first: Allan

last: Goldstein


title: Glial Guidance Drives Postnatal Intestinal Reinnervation by Enteric Neurons

summary: This study investigates the phenomenon of postnatal plasticity within the enteric nervous system (ENS), specifically investigating the reinnervation potential of post-mitotic enteric neurons. Employing BAF53b-Cre for selective tracing, the reinnervation capabilities of postnatal enteric neurons in multiple model systems are shown. Denervated enteric neurons exhibit the ability to regenerate neurites in vitro, with neurite complexity and direction notably influenced by contact with enteric glial cells (EGCs). In vivo nerve fibers from transplanted enteric neurons exclusively interface with EGCs. Resident EGCs are sustained after Cre dependent ablation of enteric neurons and govern the architecture of the ENS by reinnervating enteric neurons. Transplantation experiments underscore the swift reintegration and reinnervation potential of post-mitotic neurons, leading to restored muscle function within two weeks. Optogenetic investigations further delineate time-dependent functional recovery via transplantation of isolated enteric ganglia. These revelations demonstrate the structural and functional reinnervation capacity of post-mitotic enteric neurons, underscored by EGC guidance.

overall_design: To ablate enteric neurons expressing inducible diphtheria toxin receptor (iDTR), BAF53b::iDTR mice were generated. Diphtheria toxin (Sigma Aldrich, St. Louise, MO, USA, #D0564) was prepared by dilution in PBS and India ink to a final dose of 0.33ng/μL of DT and 10% India ink at final volume.  Mice were anesthetized by isoflurane to perform laparotomy and delivery of 4μL of the DT solution by microinjection to the muscularis propria with the India ink used to confirm the injection site. Injection of the solution DT was performed in littermate mice lacking iDTR expression as experimental controls.

first: Ahmed

last: Rahman

first: Christopher

middle: Y

last: Han

first: Abigail

last: Leavitt

first: Leah

last: Ott

first: Ryo

last: Hotta


title: FOXQ1 Regulates Mitochondrial Cristae Formation and Energy Metabolism in Mouse Brain Endothelial Cells [CUT&TAG]

summary: To explore FOXQ1’s transcriptional role, we utilized an HA-tagged FOXQ1 plasmid for CUTTag enabling high-resolution mapping of FOXQ1 binding sites on the genome.

overall_design: Examination of gene which FOXQ1 binding within primary brain vessel endothelial cells.


title: Cortical stimulation-based transcriptome shifts on Parkinson's disease animal model

status: Public on Jul 11 2024

city: Incheon

first: Mi-Ryung

address: 265, Harmony-ro, Yeonsu-gu, Incheon, Republic of Korea

country: South Korea

institute: Incheon National University

summary: Parkinson's disease is the second most prevalent neurodegenerative disorder, characterized by the degeneration of dopaminergic neurons. Significant improvements in gait balance, particularly step length and velocity, were revealed by less-invasive wireless cortical stimulation. Transcriptome sequencing was performed to demonstrate the cellular mechanism, specifically targeting the primary motor cortex where the stimulation was applied. Our findings indicated that the differentially expressed genes (DEGs), initially down-regulated following Parkinson's disease induction, were subsequently restored to normal levels after cortical stimulation. We propose these DEGs as a potential target for motor disorder treatment in Parkinson's disease. These genes are implicated in crucial processes such as astrocyte-mediated blood vessel development and microglia-mediated phagocytosis of damaged motor neurons, suggesting their significant roles in improvement of behavior disorder. Moreover, these biomarkers not only facilitate rapid and accurate diagnosis of Parkinson's disease but also assist precision medicine approaches.

overall_design: This study evaluated the efficacy of cortical stimulation using a cutting-edge graphene-based electrode array and wireless technology to mitigate the symptoms in a rat model of PD induced by 6-OHDA. Specifically, the graphene array was surgically placed in the primary motor cortex of the rats, followed by daily wireless stimulation at 130 Hz for one hour, over two weeks. The participants were divided into three groups: control (Ctrl), PD, and Stimulation (Stim). This study aimed to explore the biological mechanisms that facilitate the observed behavioral improvements in patients with PD following stimulation through comprehensive transcriptome analysis using NGS techniques.

first: Johyeon

last: Nam

first: Hongseong

last: Shin

first: Chaeyeon

last: You

first: Eunha

last: Baeg

first: Jae

middle: G

last: Kim

first: Sunggu

last: Yang


title: Cadherins modulate the self-organizing potential of gastruloids (RNA-seq)

first: ALEXANDRE

last: MAYRAN

overall_design: Bulk RNA-seq on gastruloids from WT and mutant conditions

first: Alexandre

last: Mayran

first: Anne-Catherine

last: Cossy

first: Lucille


title: Hox gene activity directs physical forces to differentially shape chick small and large intestinal epithelia

status: Public on Jul 06 2024

first: Hasreet

last: Gill

address: 77 Avenue Louis Pasteur, NRB 360

institute: Harvard Medical School

summary: Hox genes are highly conserved, master regulators of spatial patterning in the embryo, but how these factors trigger regional morphogenesis has largely remained a mystery. In the developing gut, Hox genes help demarcate identities of the small and large intestines early in embryogenesis, which ultimately leads to their specialization in both form and function. While the midgut forms villi, the hindgut develops flat, brain-like sulci that resolve into heterogeneous outgrowths. Combining mechanical measurements of the embryonic chick intestine and mathematical modeling, we demonstrate that the posterior Hox gene HOXD13 regulates biophysical phenomena that shape the hindgut lumen. We further show that HOXD13 acts through the TGFβ pathway to thicken, stiffen, and promote isotropic growth of the subepithelial mesenchyme; together, these features lead to hindgut surface buckling. TGFβ, in turn, promotes collagen deposition to affect mesenchymal geometry and growth. We thus identify a cascade of events downstream of positional genetic identity that direct posterior intestinal morphogenesis.

overall_design: To more specifically identify which genes are commonly differentially expressed within the subepithelial mesenchyme of the midgut with Hoxd13 misexpression and the hindgut, relative to the midgut, a spatial sequencing approach called Light-seq  was employed (Kishi et al., 2022). In situ reverse transcription was performed in midgut, hindgut, and RCAS-Hoxd13 tissue sections on slides. Light-directed crosslinking was used to iteratively barcode manually defined regions--the subepithelial mesenchyme, inner longitudinal muscle layer, and inner superficial side of the circumferential muscle layer. Barcoded cDNA was then extracted from slides and used for downstream qPCR amplification and library preparation. Each sequenced library contained reads from all three compartments. Downstream barcode parsing then enabled assignment of each read to the tissue layer of interest. Though three layers were barcoded and sequenced, only the inner mesenchyme layer was used for differential gene expression analysis comparing conditions (midgut, hindgut, midgut + Hoxd13).

middle: K

first: Sifan

last: Yin

first: Nandan

middle: L

last: Nerurkar

first: John

last: Lawlor

first: ChangHee

last: Lee

first: Tyler

last: Huycke

first: L

last: Mahadevan

first: Clifford

last: Tabin


title: Cadherins modulate the self-organizing potential of gastruloids (scRNA-seq)

overall_design: scRNA-seq on gastruloids from WT and mutant conditions

last: Lopez-Delisle

first: Dominique

last: Kolly

first: Denis

last: Duboule


title: Computational design of sequence-specific DNA-binding proteins

status: Public on Jul 05 2024

city: Durham

first: Raluca

last: Gordan

state: NC

address: 101 Science Dr, CIEMAS 2179

department: Center for Genomic and Computational Biology

institute: Duke University

summary: Sequence-specific DNA-binding proteins (DBPs) play critical roles in biology and biotechnology, and there has been considerable interest in the engineering of DBPs with new or altered specificities for genome editing and other applications. While there has been some success in reprogramming naturally occurring DBPs using selection methods, the computational design of new DBPs that recognize arbitrary target sites remains an outstanding challenge. We describe a computational method for the design of small DBPs that recognize specific target sequences through interactions with bases in the major groove.

overall_design: Universal protein-binding microarray (PBM) experiments were performed for small, recombinant, DNA-binding protein designs 1, 3, 5, 6, 9, 35, & 48. Briefly, universal PBMs involved binding of his-tagged proteins to double-stranded 15K Agilent microarrays containing a DNA library designed to cover all possible 9-bp sequences, with every 7-mer occurring in at least 16 different spots on the array. This design allows comprehensive and unbiased characterization of the binding specificity of transcription factors for all possible 7-bp sequences.

first: Olivier

last: Boivin


title: Persistent alterations in glucose metabolism, gut microbiome, and adipose tissue transcriptome after two-week sleep recovery in chronically sleep-fragmented female mice

status: Public on Jun 30 2024

first: Ling

last: Zhong

address: Wangfujing Street

department: Department of Endocrinology

institute: Peking Union Medical College Hospital

summary: Introduction: Chronic sleep fragmentation (SF) is prevalent in contemporary human society, highlighting detrimental effects on glucose metabolism and adipose tissue morphology, which is closely linked to gut microbiota composition. However, it remains unclear whether sleep recovery (SR) after prolonged SF can ameliorate glucose metabolism, influence the transcriptome of inguinal white adipose tissue (iWAT), and whether these effects align with alterations in the gut microbiota.  Methods: Mice were subjected to 8 weeks of SF and subsequently allowed 2 weeks of SR. We assessed glucose tolerance through intraperitoneal glucose tolerance tests (ipGTT), analyzed gut microbiota via 16s rDNA amplicon sequencing, and examined transcriptomic alterations in iWAT using RNA sequencing.  Results: Despite the two-week SR following chronic SF, significant glucose intolerance persisted, accompanied by subtle shifts in the gut microbiota and alterations in gene expression within iWAT. The top hub genes Ncapg, Cenpe, and Tik were identified from the protein-protein interaction network.  Conclusion: Even followed by a brief period of SR, prolonged SF still led to ongoing glucose intolerance and alterations in the adipose tissue transcriptome in mice. These changes were intertwined with modifications in the gut microbiome. The shifts in gut microbiota may play a pivotal role in understanding the sustained negative effects of SF.

overall_design: RNA sequencing was conducted on 15-week-old female C57BL/6N mice who were subjected to  8 weeks of sleep fragmentation and subsequently allowed 2 weeks of sleep recovery.


title: Determine the role of ALK5 signaling in murine brain pericytes at embryonic day E13.5

city: New Haven

first: Jui

last: Dave

state: CT

address: 300 George St

department: Cardiovascular Research Center

institute: Yale University

summary: Pericytes (PCs) are microvascular mural cells which constituent the embryonic blood brain barrier (BBB) along with endothelial cells (ECs). During brain development, germinal matrix (GM) - a highly vascularized region rich in neuronal-glial precursors, is selectively vulnerable to hemorrhage in premature infants. The transforming growth factor β (TGFβ) pathway plays a crucial role in barrier development by regulating cellular cross talk between PCs and ECs. Indeed, murine embryos lacking TGFβ receptor activin receptor-like kinase 5 (Alk5) in brain PCs (mutants) develop gross germinal matrix hemorrhage-intraventricular hemorrhage (GMH-IVH) and culminate in perinatal lethality. Mutant GM vessels display reduced PC and collagen coverage, abnormal vessel dilation and EC hyperproliferation. However, the mechanistic link between PC-specific deletion of ALK5 and aberrant EC behavior remains elusive. Herein, using bulk RNA sequencing from human brain PCs lacking ALK5 (siALK5) as well as murine vascular cells [PCs and ECs] isolated from embryonic brain at embryonic days E11.5 and  E13.5 we establish that angiopoietin 2 (ANGPT2), a secreted angiogenic growth factor, is robustly repressed by the TGFβ pathway in PCs. Conversely, mutants lacking PC-ALK5 secrete higher levels of ANGPT2 resulting in overactivation of tyrosine protein kinase receptor (TIE2) and culminating in EC hyperproliferation, vessel dilation, BBB breakdown and GMH. PC-specific Angpt2 deletion or pharmacological inhibition in mutants improves GM vessel morphology, reduces EC proliferation and attenuates GMH pathogenesis. Taken together, we demonstrate that loss of TGFβ-mediated ANGPT2 repression in PCs is detrimental for BBB integrity and identify ANGPT2 as an important pathological target for GMH-IVH.

overall_design: To evaluate ALK5-mediated differential gene expression in percityes during cerebrovascular development at E13.5, perciytes were isolated from murine embryonic brains of Pdgfrb-Cre, Alk5(flox/+) [control] and Pdgfrb-Cre, Alk5(flox/flox) [mutant] mice also bearing ZS green by flow cytometry. Total RNA was isolated and subjected to bulk RNA sequencing.

first: Daniel

last: Greif


title: Transcriptional Remodeling in Human Cardiac Fibroblasts and Myocytes Induced by Oxygen-Dependent Crosstalk

city: Los Angeles

first: Megan

last: McCain

state: CA

address: 1024 Downey Way

institute: USC

summary: Myocardial infarctions are caused by coronary occlusions that deprive downstream myocardial tissue of oxygenated blood, leading to localized necrosis of cardiac myocytes. Hypoxic injury drives a remodeling process in which cardiac fibroblasts activate and synthesize matrix proteins to form a scar. During this process, cardiac cells are exposed to an oxygen gradient at the infarct border zone that transitions from 0% oxygen at the site of injury to 10% oxygen in healthy myocardium. However, the impact of any crosstalk between hypoxic cardiac fibroblasts and neighboring, normoxic (10% oxygen) cardiac cells is unexplored because conventional research tools cannot re-create spatially heterogeneous oxygen landscapes. Here, we used a microfluidic device to co-culture hypoxic cardiac fibroblasts with (1) normoxic cardiac fibroblasts or (2) normoxic human induced pluripotent stem cell (hiPSC)-derived cardiac myocytes. We then compared their transcriptome to the same cells cultured in uniform hypoxia or normoxia. These data contribute to more advanced understanding of how oxygen-dependent crosstalk between cardiac cells impacts the wound healing process post-myocardial infarction.

overall_design: Primary human adult cardiac fibroblasts or human induced pluripotent stem cell (hiPSC)-derived cardiac myocytes were cultured on fibronectin-coated PDMS microfluidic devices designed to expose cells to 0% oxygen, 10% oxygen, or adjacent co-culture regions of 0% and 10% oxygen. After 24h of oxygen modulation, cells from each half of the co-culture region were immediately separated with a razor blade and all conditions (0% control, 0% in co-culture, 10% in co-culture, and 10% control) were lysed. RNA was isolated and sequenced, and gene expression was analyzed using Partek Flow Genomics Software.

first: Natalie

last: Khalil

first: Divya

last: Gupta

first: Sarah

last: Parker


title: Oncogenic KRAS-dependent stromal interleukin-33 directs the pancreatic microenvironment to promote tumor growth [scRNA-seq]

status: Public on Jun 26 2024

city: Ann Arbor

first: Marina

last: Pasca di Magliano

state: MI

address: 1500 E Medical Center Dr

institute: University of Michigan Medical School

summary: Here, we sought to understand the function of stromal interleukin-33 (IL-33) in PanIN and PDA. We collected pancreatic tissues from various mouse models of PanIN and PDA for single cell RNA sequencing to assess IL-33 expression across the microenviornment. We also implanted syngeneic orthotopic pancreatic tumors into mice harboring a conditional stromal IL-33 knockout and sequenced the resulting tumors.

overall_design: Pancreata from various mouse models of pancreatic tumorigenesis (spontaneous or orthotopic injection) collected and subjected to single cell RNA sequencing.

first: Katelyn

last: Donahue

first: Yaqing

last: Zhang

first: Emily

last: Lasse-Opsahl


title: Chromatin accessibility of genomic regions in human mesenchymal stem cells cultured in sliding hydrogels (SGs) and chemical hydrogels (CGs) undergoing chondrogenesis.

status: Public on Jun 29 2024

city: Palo Alto

first: Fan

last: Yang

state: California

address: Orthopedic Surgery, 240 Pasteur Drive, Biomedical Innovation Building 1200

institute: Stanford University

summary: Cells can deform their local microenvironment by exerting forces through pushing and pulling. Such interactions result in behaviors such as cell spreading, cell migration, or cell volume expansion, which occur on timescales of hours to days. However, whether cells can deform local niches at shorter timescales and how such interactions impact long-term cell fates remain largely unknown. Here we report “cell dancing”- a previously unknown cell behavior characterized by seconds-to-minutes scale deformation of the pericellular niche and enhanced cytoskeletal and nuclear dynamics. Using hydrogels that allow cells to deform their local 3D niche and mesenchymal stem cell chondrogenesis as a model system, we identify early-stage cell dancing essential for enhancing stem cell differentiation in 3D. This is mediated through nuclear mechanotransduction pathways that repress global chromatin accessibility, which is necessary for cell dancing-enhanced differentiation. We further demonstrate that cell dancing induces nuclear deformations and phospholipase A2 (PLA2) signaling, and small molecules targeting this pathway can further boost stem cell chondrogenesis. Finally, we validate that cell dancing can enhance stem cell differentiation towards multiple lineages and occurs in other commonly used hydrogel systems. These results establish cell dancing as a novel cell behavior in 3D that may be broadly applied to promote desirable cell fates and tissue outcomes.

overall_design: ATAC-seq analysis was conducted for two conditions (SG and CG) at two time points (16 hours and 48 hours)

first: Manish

last: Ayushman

first: Eunice

last: Lopez-Fuentes

first: Alejandro

last: Sweet-Cordero


title: Effect of TWIST1 dosage changes on chromatin accessibility in hESC-derived cranial neural crest cells

city: Stanford

first: Sahin

last: Naqvi

state: CA California

address: 265 Campus Drive, Lokey Stem Cell Research Building/SIM1

summary: To assess the effects of quantitative TWIST1 dosage changes on chromatin accessibility in hESC-derived cranial neural crest cells, we performed genome editing of the H9 hESC lines to tag TWIST1 with FKBPV36 and V5, Upon differentiating edited hESC to cranial neural crest cells using an established protocol, addition of differing concentrations of the degrader molecule dTAGV-1 results in different TWIST1 concentrations, as measured by V5 intracellular staining and flow cytometry. We then profiled chromatin accessibility in each of these TWIST1 concentrations

overall_design: Genome-wide chromatin accessibility profiling (ATAC-Seq) of TWIST1-tagged H9 hESC-dervied cranial neural crest cells (CNCCs)


title: Determine the role of ALK5 signaling in human brain pericytes treated with TGFβ.

overall_design: To evaluate ALK5-mediated differential gene expression in percityes, cultured human brain perciytes were trasnfecetd with Scr (control siRNA)or siALK5 (siRNA targeting ALK5) and then treated with or without TGFβ. Total RNA was isolated and subjected to bulk RNA sequencing.


city: BOSTON

first: CHANGHEE

last: LEE

address: 77 AVENUE LOUIS PASTEUR

overall_design: Single-cell RNA sequencing of the chick embryonic hindgut at E15, and embryonic midgut at E15 with and without ectopic, sparse, inducible HOXD13 misexpression in the mesoderm; Single-cell RNA sequencing of the chick embryonic E14 midgut and hindguts. All samples with biological multiplicates.

first: Hasreet

last: Gill

first: Sifan

last: Yin

first: Nandan

last: Nerurkar

first: John

last: Lawlor

first: ChangHee

last: Lee

first: Tyler

last: Huycke

first: L

last: Mahadevan

first: Clifford

last: Tabin


title: Hepatic insulin resistance promotes fructose-enriched diet-induced fatty liver diseases.

status: Public on Jun 23 2024

first: rongya

last: tao

state: Ma

address: 300 longwood ave

institute: Boston children's hospital

summary: Understanding mechanisms causing MAFLD (Metabolic Associated Fatty Liver Disease) and its progression to MASH (metabolic dysfunction-associated steatohepatitis) is clinically important and scientifically challenging. Hepatic insulin resistance is a common component in the progression of MAFLD in patients and experimental animals; however, hepatic steatosis caused by the HFD45% (high-fat diet) decreases during chronic hepatic IR generated by inactivation of Irs1/2 (LDKO), AKT1/2, or InsR 1-3—which is inconsistent with the expected relationship between IR and MAFLD in humans4. Here we found that complete hepatic insulin resistance promotes the fructose-enriched GAN diet-induced MAFLD, including acute inflammation and MASH in LDKO mice. Unexpectedly, fructose phosphorylation catalyzed by hepatic Khk (ketohexokinase) was not required as acute MAFLD progressed strongly in LDKOKhkL/L mice fed the GAN diet. FoxO1 activated during hepatic IR induces Fst (Follistatin) expression and secretion from the liver of LDKO mice. Inactivation of hepatic FoxO1 in LTKO mice (LDKO•FoxO1L/L) or Fst in LDKOFstKO mice prevented acute MAFLD during the GAN diet. Consistently, overexpression of hepatic Fst promoted GAN diet-induced MAFLD/MASH and hepatic carcinoma. Mechanistically, circulating Fst promoted adipose tissue IR and lipolysis, which can deliver FFA (free fatty acid) to the liver for esterification with excess Gro3P (glycerol 3-phosphate) generated by fructose metabolism, although hepatic DNL (de novo lipogenesis) decreased strongly in LDKO mice while. Since circulating FST correlates positively with both T2D and MAFLD in humans, our results suggests that hepatic FST induced by progressive hepatic IR might promote MAFLD/MASH during the consumption of sugar-sweetened food and beverages consumed frequently by people and animals with T2D.

overall_design: We generated the LDKO (Irs1L/L:Irs2L/L:CreAlb)  as previously described.  Male Cntr and LDKO mice were fed HFD, GAN, or HFruD between 5-15 wks (n=2). Gene expression was determined by RNAseq. Moreover,  C57BL/6 mice were fed chow, HFD or GAN diet  and infected at 33 weeks without or with Fst315AAV-TBG (Fst315 mice) or GFPAAV-TBG (GFP mice.Liver samples were collected after 5 hours fasting

first: Rongya

last: Tao

first: Morris

last: White

first: Kyle

last: Copps

first: Oliver

last: Stöhr


title: Single cell RNA sequencing of adult regenerating mouse digit tips

status: Public on Jul 03 2024

first: Jessica

last: Lehoczky

address: 60 Fenwood Rd.

department: Orthopedic Surgery

institute: Brigham and Women's Hospital

summary: Innate regeneration following digit tip amputation is one of the few examples of epimorphic regeneration in mammals. Digit tip regeneration is mediated by the blastema, the same structure invoked during limb regeneration in some lower vertebrates. By genetic lineage analyses in mice, the digit tip blastema has been defined as a population of heterogeneous, lineage restructed progenitor cells. These previous studies, however, do not comprehensively evaluate blastema heterogeneity or address lineage restruction of closely related cell types. Here we report one additional single cell RNA sequencing sample (28 days post-amputation) to add to our already published 38,000 cells from mouse digit tip blastemas and unamputated control digit tips used to generate an atlas of the cell types participating in digit tip regeneration.

overall_design: Adult wildtype FVB/NJ mice used to generate 28 days post-amputation digit tips for single cell RNA sequencing by 10X genomics. GSM4276219, GSM4276220, GSM4276221, GSM4276222, and GSM4276223 from GSE143888 were re-used for the current study


title: UBE2D3 is an Intracellular Checkpoint in Pancreatic Ductal Carcinoma that Restricts CD8+ T-Cell Antitumor Immunity II

first: Shiqun

last: Wang

state: Zhejiang Province

address: No. 150, Fucheng Road, Hangzhou Economic and Technological Development Zone

institute: Institute of Cancer and Basic Medicine, Chinese Academy of Sciences (CAS)

summary: Ubiquitylation is pivotal in regulating cellular responses, with its aberration implicated in tumor immune evasion. However, the impact of ubiquitin-conjugating enzymes (E2s) on this evasion remains unclear. Here, we employ a systematic approach to demonstrate that in pancreatic ductal carcinoma (PDAC), the inflammatory microenvironment induces overexpression of the E2 enzyme UBE2D3, contributing to tumor progression through non-oncogene codependent disorders. Through gene expression analyses and functional investigations, we elucidate a mechanism wherein cancer cells evade T cell immune responses by UBE2D3 binding to the ubiquitin ligase KLHL13 to co-ubiquitinate TAP2. This K63-linkage ubiquitination at the lysine 245 site of TAP2 impedes antigenic peptide transport by the TAP1/TAP2 complex, hindering p-MHC assembly and presentation in cancer cells. We demonstrate that genetic inhibition of UBE2D3 enhances tumor-specific CD8+ T cell proliferation and extends effector-memory-like phenotypes. Building on this, we develop a small-molecule inhibitor, QX-6, targeting the active site of UBE2D3 to disrupt its function. Pharmacologic inhibition of UBE2D3 blocks the ubiquitylation of antigen presentation-related substrates, leading to increased p-MHC presentation by cancer cells and reduced T cell exhaustion. Using immunocompetent and humanized models, we elucidate the therapeutic efficacy of targeting UBE2D3. Furthermore, we evaluate the synergistic anti-tumor effects of QX-6 in combination with KRAS-targeted TCR-T cell immunotherapy. In summary, our study reveals a post-translational modification mechanism wherein the intracellular checkpoint UBE2D3 regulates the TAP2 switch to control cancer cell evasion of CTLs, presenting a potential immunotherapeutic strategy to improve PDAC treatment outcomes.

overall_design: For single cell sequencing, C57/BL6 mice bearing KPC-shScramble or KPC-shUBE2D3-1 tumors for 30 d were euthanized. Three tumors from the same group were pooled to prepare single-cell suspension for isolating enough tumor-infiltrating CD8+ T cells. These cells were isolated by FCM, selecting live, single cells expressing CD45, CD3, and CD8.

first: Wenyan

last: Yang

first: Jiangjiang

last: Qin


title: Transcriptomic analysis of human naive B cells stimulated by CD40L, CpG, anti-IgM IgG, IL-4 or combinations

first: Rui

last: Guo

address: 150 Harrison Ave.

department: School of Medicine

institute: Tufts University

summary: B-lymphocytes play major adaptive immune roles, producing antibody and driving cell mediated responses. However, how B-cells acutely differentiate in response to receptor signaling codes, including T-cell dependent versus independent cues, remains incompletely understood. To gain insights, we used multi-omic profiling to characterize ex vivo primary human B-cell transcriptomic, proteomic and metabolomic remodeling by B-cell receptor (BCR), Toll-like receptor 9 (TLR9), CD40-ligand (CD40L), interleukin-4 (IL4) or combinations thereof, highlighting  key stimulus-specific phenotypes.

overall_design:
Primary human B-cells were treated in culture in the absence of stimulation or stimulated by, CD40L only, CpG only, IL4 only, aIgM only, CD40L+CpG, CD40L+IL4, CpG+ aIgM, CD40L+ aIgM, and CD40L+ aIgM+IL4. Cells were treated for 24 hours. For each experiment, cells from 3 donors were isolated and treated separately.
Primary human B-cells knocked out for BCAT1.

first: Benjamin

last: Gewurz


title: Platelet Reactivity ExpreSsion Score Predicts Cardiovascular Risk

city: New York City

first: Matthew

last: Muller

address: 550 First Avenue

department: Department of Medicine

institute: New York University

summary: Platelets are key mediators of atherothrombosis; yet, limited tools exist to identify individuals with a hyperreactive platelet phenotype. Here we derive and validate a Platelet Reactivity ExpreSsion Score (PRESS) integrating platelet aggregation responses and RNA sequencing in multiple cohorts. PRESS performs well in identifying a hyperreactive phenotype in patients with cardiovascular disease (AUC [cross-validation] 0.81, 95% CI 0.68 to 0.94), with similar discrimination in an independent cohort of healthy participants (AUC [validation] 0.77, 95% CI 0.75 to 0.79); accuracy 80%, sensitivity 71%, and specificity 86%. Next, we validate PRESS in multiple cohorts of patients with platelet RNASeq available. Following multivariable adjustment, PRESS was significantly higher in MI (β=1.8, p=0.02), and individuals with a score above the median more frequently develop a future cardiovascular event (adjusted HR 1.90, CI 1.07 to 3.36), p=0.027). Altogether, we provide strong evidence that PRESS is a robust prognostic marker, which could have an essential role in facilitating a personalized approach to cardiovascular risk management.

overall_design: RNA-seq of purified platelets from patients with cardiovascular disease.

first: Jeffrey

middle: S

last: Berger

first: MacIntosh

last: Cornwell

first: Yuhe

last: Xia

first: Nathaniel

last: Smilowitz

first: Jonathan

middle: D

last: Newman

first: Caron

last: Rockman

first: Kelly

middle: V

last: Ruggles

first: Deepak

last: Voora

first: Judith

last: Hochman

first: Tessa

last: Barrett


title: Patchseq Data of T1D Donor Islets vs ND Matched Controls

status: Public on Jun 25 2024

city: Edmonton

first: Patrick

last: MacDonald

state: Alberta

address: 6-126 Li Ka Shing Centre

country: Canada

institute: University of Alberta

summary: Will be updated closer to manuscript finalization. Patchseq allows for combined electrical and transcriptomic data to be obtained at single cell resolution. Here we performed patchseq on islet cells from human donors

overall_design: Isolated human islets were dispersed to single cells, and whole cell patch clamp electrophysiology was performed. Following, the cell was sent for scRNA-seq (SmartSeq). XXH12 wells were left empty and A3A11 and V2A11 failed to sequence. Since they were empty fastq files, they have not been uploaded based on GEO's recommendations.

first: Theodore

last: dos Santos

first: Cara

last: Ellis

first: Jessica

last: Ewald

first: Robert

last: Jones

first: Joan

last: Camunas-Soler

first: Stephen

last: Quake


title: Effect of FL-Fc-DM1 drug on gene expression of THP-1 cells pretreated with cytarabine.

city: Shenzhen

first: Dengyang

last: Zhang

state: Guangdong

address: Zhenmei road, guangming district

department: Scientific research center

institute: Seventh affiliated hospital, Sun Yat-sen University

summary:
Cytarabine is one of the first line chemo drugs to treat acute myeloid leukemia (AML).
However, cytarabine can induce AML cell cell cycle arrest at G0/G1 phase. These cell-cycle-arrested AML cells resist cytarabine, show a senescence-like phenotype or stem-cell like phenotype.
To eliminate these cell-cycle arrested cells, we developed a FL-Fc-DM1 drug by conjugating FLT3 liangd Fc fusion protein and mertansine. We used the THP-1 cells as a AML model.
The total mRNA of THP-1 cells treated with different drugs (cytarabine, FL-Fc-DM1, FL-Fc) were sequenced.

overall_design:
THP-1 cells were cultured in RPMI 1640 medium supplemented with 10% FBS.
In the control group, THP-1 cells were cultured for 4 days. In the cytarabine group, THP-1 cells were treated with 20 µM cytarabine for 4 days. In the FL-Fc-DM1 treatment group, THP-1 cells were treated by cytarabine for 3 days and then treated with FL-Fc-DM1 (5 nM) for another 24 hours in the absence of cytarabine. In the FL-Fc treatment group, THP-1 cells were treated by cytarabine for 3 days and then treated with FL-Fc (5 nM) for another 24 hours in the absence of cytarabine.
We than analyze the transcriptome differences between different groups.

first: Yun

last: Chen


title: Senescent cells cluster CTCF on nuclear speckles to sustain their splicing program [Chipmentation]

status: Public on Jun 17 2024

city: Göttingen

first: Argyris

last: Papantonis

address: Robert-Koch-Str. 40

department: Institute of Pathology

institute: University Medical Center Göttingen

summary: Senescence —the end-point of the replicative life span of normal cells— is characterized by a complex sequence of molecular and biochemical events. One of these is the dramatic reorganization of CTCF into large senescence-induced CTCF clusters (SICCs). However, the molecular determinants, genomic consequences, and functional purpose of SICCs remained unknown. Here, we combine super-resolution imaging, 3D genomics, and functional assays with computational modelling to address all aspects of SICC emergence. First, we used modelling and data from the overexpression of nuclear factors lost upon senescence entry to explain how changes in the nuclear environment allow for CTCF clustering. Second, we discovered that cells entering senescence repurpose SRRM2 —a key component of nuclear speckles— and BANF1 —a protein known to act as ‘molecular glue’ for chromosomes during mitosis— to cluster CTCF on the surface of speckles, while rewiring higher-order genome architecture. Last, we could show that SICCs are required for the faithful execution of the senescence splicing program as their disruption by SRRM2 or BANF1 depletion reverts splicing choices. Together, the senescence-specific clustering of CTCF on nuclear speckles is a new paradigm of a change in the biochemical environment of the nucleus being translated into its architectural reorganization to allow for the deployment of a splicing program driving homeostatic deregulation.

overall_design: We performed ChIPmentation of the Nuclear Speckle component SON and the transcription factor HMGB2, in order to indentify the interaction with CTCF as well as their contribution towards genome architecture and gene expression.

first: Shyam

last: Ramasamy

first: Vassiliki

last: Varamogianni Mamatsi


title: UBE2D3 is an Intracellular Checkpoint in Pancreatic Ductal Carcinoma that Restricts CD8+ T-Cell Antitumor Immunity

status: Public on Jun 21 2024

first: Shiqun

last: Wang

overall_design: For bulk RNA-seq, 1 x 106 KPC-shScramble and KPC-shUBE2D3-1 cells were inoculated in 6-well plates. After 24 h, cells were harvested. Total RNA was isolated and purified using TRIzol (ThermoFisher, 15596018) following the manufacturer’s protocol. Comparative gene expression profiling analysis of RNA-seq data for KPC-shScramble and KPC-shUBE2D3 cell line in vitro

first: Wenyan

last: Yang

first: Jiangjiang

last: Qin


title: In vivo CRISPR base editing for treatment of Huntington’s disease

status: Public on Jul 12 2024

city: Urbana

first: Pablo

last: Perez-Pinera

state: il

address: 1406 West Green Street

institute: University of Illinois Urbana-Champaign

summary: Huntington's disease (HD) is an inherited and ultimately fatal neurodegenerative disorder caused by an expanded polyglutamine-encoding CAG repeat within exon 1 of the huntingtin (HTT) gene, which produces a mutant protein that destroys striatal and cortical neurons.Importantly, a critical event in the pathogenesis of HD is the proteolytic cleavage of the mutant HTT protein by caspase-6, which generates fragments of the N-terminal domain of the protein that form highly toxic aggregates. Given the role that proteolysis of the mutant HTT protein plays in HD, strategies forpreventing this process hold potential for treating the disorder.By screening 141 CRISPR base editor variants targeting splice elements in the HTT gene, we identified platforms capable of producing HTT protein isoforms resistant to caspase-6-mediated proteolysis via editing of the splice acceptor sequence for exon 13. When delivered to the striatum of a rodent HD model, these base editors induced efficient exon skipping and decreased the formation of the N-terminal fragments, which in turn reduced HTT protein aggregation and attenuatedstriatal and cortical atrophy.Collectively, these results illustrate the potential for CRISPR base editing to decrease the toxicity of the mutant HTT protein for HD.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Mammalian cell transfections with CRISPR base editors to modify DNA bases in HTT exon and influence RNA splicing to investigate its impact on caspase-6 induced proteolysis. Stereotaxic injections using AAV delivery into mice brains to study DNA editing and exon skipping using base editing tools

first: Shraddha

last: Shirguppe

first: Michael

last: Gapinske

first: Devyani

last: Swami

first: Nicholas

last: Gosstola

first: Pankaj

last: Acharya

first: Angelo

last: Miskalis

first: Dana

last: Joulani

first: Maddie

last: Szkwarek

first: Abhishek

last: Bhattacharjee

first: Gianna

last: Elias

first: Michelle

last: Stilger

first: Jackson

last: Winter

first: Wendy

last: Woods

first: Daphine

last: Anand

first: Colin

last: Lim

first: Thomas

last: Gaj


title: Adiponectin regulated by IAA in Paneth cells controls renewal and differentiation of gut stem cells to maintain homeostasis of epithelial cells (small intestinal epithelium)

city: Tianjin

first: Yun Huan

last: Gao

address: 94 Weijin Road, Nankai District, Tianjin

institute: Nankai University

summary: Crypt cells (mainly stem and transit amplifying (TA) cells) in Adipfl/flpVilli-CreT (gut epithelial cell adiponectin conditioned knockout) mice exhibited stronger ability of renewal and differentiation. ScRNA-seq found that there had much more enrichment intestinal stem and TA cells in Adipfl/flpVilli-CreT mice. The renewal and development of gut epithelial cells in irradiated or DSS (dextran sulfate sodium) treated Adipfl/flpVilli-CreT mice was faster than control Adipfl/flp mice. In vitro culture gut organoid of Adipfl/flpVilli-CreT mice exhibited faster growth than those from Adipfl/flp mice.

overall_design: To further determine the effects of adiponectin from Paneth cells on the gut stem and TA cells, we performed scRNAseq on crypt-enriched whole small intestinal epithelium from control Adipfl/fl mice and Adipfl/flpVilli-CreT mice.

first: Xiaomin

last: Su

first: Juanjuan

last: Wang

first: Hang

last: Liu

first: Rongcun

last: Yang


title: Genomic heterogeneity drives mechanical adaptation in human tumor cells [RNA-seq]

city: College Station

first: Ting-Ching

address: 101 Bizzell St

institute: Texas A&M University

summary: The progression of many solid tumors is accompanied by temporal and spatial changes in the stiffness of the extracellular matrix (ECM). Cancer cells adapt to soft and stiff ECM through mechanisms that are not fully understood. In particular, it is well known that there is significant genetic heterogeneity from cell to cell in tumors, but how ECM stiffness as a parameter might interact with that genetic variation is not known. Here, we used the method of experimental evolution to study response of genetically variable and clonal tumor cell populations to ECM stiffness. Cell fitness increased on soft ECM over a period of several weeks in genetically variable but not clonal populations. DNA barcode-enabled clonal tracking revealed that sustained culture on soft ECM selects for a few genetic variants. These data provide the first evidence that ECM stiffness exerts natural selection on genetically variable tumor populations. Genome-wide analysis including RNA-seq, ATAC-seq and DNA methylation profiling reveal substantial differences in gene expression between selected populations and ancestral cells which are partially explained by epigenetic modifications. Soft-selected cells are highly migratory with enriched oncogenic signatures and exhibit highly unusual behaviors like spreading and traction force generation on ECMs as soft as 1 kPa. Cell spreading is the directly selected trait but not levels of integrins or adhesion proteins like talin. Overall, these data show that ECM stiffness in solid tumors may drive malignant behaviors through evolution by natural selection.

overall_design: To explore the mechanism underlying cell evolution on soft ECM, we performed whole genome transcriptome sequencing (RNA-seq).

first: Suchitaa

last: Sawhney

first: Daylin

last: Morgan

first: Richard

last: Bennett

first: Richa

last: Rashmi

first: Marcos

last: Estecio

first: Amy

last: Brock

first: Irtisha

last: Singh

first: Charles

middle: F

last: Baer

first: Jonathan

last: Licht

first: Tanmay

middle: P

last: Lele


title: RNA sequencing for wild type and shENO1 Daudi cells.

status: Public on Jun 18 2024

city: Dalian

first: Jinsong

last: Yan

state: Liaoning

address: 467 Zhongshan Road, Dalian 116027, Liaoning, P. R. China

department: Department of Hematology

institute: the Second Hospital of Dalian Medical University

summary: We systematically evaluated the expression pattern and function of ENO1 in BL and clarified that ENO1, regulated by cMyc, can promote BL cell proliferation and metastasis by activating TGFβ1 signaling pathways in a plasmin-mediated manner. Through structure-based virtual screening, we successfully discovered Ciwujianoside E (L-06) as a novel ENO1 inhibitor which can limit BL cell proliferation and dissemination in vitro and in vivo.

overall_design: Examination of drug sensitivity for ENO1 knockdown in Burkitt’s lymphoma cell line Daudi


title: Multiplexed single-cell characterization of alternative polyadenylation regulators (MPRA)

status: Public on Jun 15 2024

city: New York

first: Madeline

last: Kowalski

address: 101 6th Ave

institute: New York Genome Center

summary: Most mammalian genes have multiple polyA sites, representing a substantial source of transcript diversity regulated by the cleavage and polyadenylation (CPA) machinery. To better understand how these proteins govern polyA site choice we introduce CPA-Perturb-seq, a multiplexed perturbation screen dataset of 42 CPA regulators with a 3’ scRNA-seq readout that enables transcriptome-wide inference of polyA site usage. We develop a framework to detect perturbation-dependent changes in polyadenylation and characterize modules of co-regulated polyA sites. We find groups of intronic polyA sites regulated by distinct components of the nuclear RNA life cycle including elongation, splicing, termination, and surveillance. We train and validate a multi-task deep neural network (APARENT-Perturb) for tandem polyA site usage, delineating a cis-regulatory code that predicts perturbation response and reveals interactions between regulatory complexes. Our work highlights the potential for multiplexed single-cell perturbation screens to further our understanding of post-transcriptional regulation.

overall_design: We conducted a pooled single-cell CRISPR screen (in both HEK293FT and K562 cell lines) targeting 42 genes previously implicated in regulating cleavage and polyadenylation.  We inferred polyA site usage at single-cell resolution from a scRNA-seq readout, and determine how each of these perturbations effects polyA site usage. We validate our polyA site quantifications by performing 3'RACE followed by Illumina sequencing at 7 loci in NT and NUDT21-perturbed cells. We also built a sequence-based deep learning model (APARENT-Perturb) to describe how sequence affects perturbation, and design an MPRA to validate these sequence-based predictions on polyA site usage.

middle: H

first: Hans-Hermann

last: Wessels

first: Johannes

last: Linder

first: Carol

last: Dalgarno

first: Isabella

last: Mascio

first: Saket

last: Choudhary

first: Austin

last: Hartman

first: Yuhan

last: Hao

first: Anshul

last: Kundaje

first: Rahul

last: Satija


title: in vitro construction of an embryonic caudal organizer

status: Public on Jul 08 2024

city: Hang Zhou

first: Tao

last: cheng

state: Zhe Jiang

address: 886 Yuhangtang Road

department: Institute of genetics

institute: Zhejiang University

summary: Purpose: Characterization of cell types in bmp4 explants and BMP4 induced human embryoids. Methods:  Single cell transcriptomes of bmp4 explants were harvested at 6 hpf, 8 hpf,10 hpf, 12 hpf, 18 hpf and 24 hpf. Single cell transcriptomes of BMP4 induced human embryoids were harvested at Day 5 and Day 6. Libraries were prepared using Single Cell 3′Library & gel Bead kit v3.1 (10x Genomics, Cat# PN-1000121) according to the manufacturer’s protocol for 10000 cell recovery. Results: paraxial mesoderm, lateral plate mesoderm, and their derivatives were detected in bmp4 explants, and NMP, somite and neural cell fates were detected in bmp4 induced human embryoids. Conclusions: A well-organized caudal-like structure was induced in bmp4 explants and caudal cell fates were specified in BMP4 induced human embryoids.

sample_organism:
Danio rerio
Homo sapiens

platform_organism:
Danio rerio
Homo sapiens

overall_design: bmp4 injected zebrafish explants or BMP4 induced human embryoids  were harvested  for scRNA-seq.

last: Cheng

first: Yan-Yi

last: Xing

first: Ying

last: Huang


title: Multiplexed single-cell characterization of alternative polyadenylation regulators (scRNA-Seq)


title: Multiplexed single-cell characterization of alternative polyadenylation regulators

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Adiponectin regulated by IAA in Paneth cells controls renewal and differentiation of gut stem cells to maintain homeostasis of epithelial cells

status: Public on Jun 14 2024

first: Yun Huan

last: Gao

summary: We here demonstrate that adiponectin in Paneth cells can suppress renewal and differentiation of gut stem cells through adiponectin receptor 1 (AdipR1). Crypt cells (mainly stem and transit amplifying (TA) cells) in Adipfl/flpVilli-CreT (gut epithelial cell adiponectin conditioned knockout) mice exhibited stronger ability of renewal and differentiation. ScRNA-seq found that there had much more enrichment intestinal stem and TA cells in Adipfl/flpVilli-CreT mice. The renewal and development of gut epithelial cells in irradiated or DSS (dextran sulfate sodium) treated Adipfl/flpVilli-CreT mice was faster than control Adipfl/flp mice. In vitro culture gut organoid of Adipfl/flpVilli-CreT mice exhibited faster growth than those from Adipfl/flp mice. The gut epithelial cells in AdipR1 knockout (KO) mice had much longer crypts than control mice. Interestingly, indole-3-acetic acid (IAA) from gut microbiota could control the expression of adiponectin to promote the renewal and proliferation of gut stem cells. In vitro culture organoid also showed faster development after exposing to IAA. Thus, adiponectin, which is regulated by gut microbiota derived IAA controls the renewal and differentiation of gut stem cells to maintain the homeostasis of epithelial cells.

overall_design: To further illustrate the function of adiponectin from gut Paneth cells, we again generated in vitro gut organoids from the intestines of Adipfl/flpVilli-CreT mice (Adip KO) and from control Adipfl/fl mice(WT).

first: Xiaomin

last: Su

first: Juanjuan

last: Wang

first: Rongcun

last: Yang


title: A system for functional studies of the major virulence factor of malaria parasites

city: Hamburg

last: Allweier

address: Bernhard-Nocht-Straße 74

institute: Bernhard Nocht Institute for Tropical Medicine

summary: The major virulence factor of Plasmodium falciparum parasites, PfEMP1 is expressed by a multigene family, termed var genes. Here selection linked integration (SLI) was utilized to modify var genes in P. falciparum parasites to select for parasite populations expressing a single var gene. Bulk RNA was isolated from ring stage parasites of these SLI parasite populations and analyzed with next generation sequencing. The proportion of exon 2 transcripts of var genes normalized to transcripts per million was determined per cell line to confirm the predominant expression of the desired var gene.

overall_design: RNA-seq samples of Plasmodium falciparum infected human erythorcyes

first: Jakob

last: Cronshagen

first: Paolo

last: Mesén-Ramírez

first: Jan

last: Stäcker

first: Anna

last: Viktoria Vaaben

first: Gala

last: Ramón-Zamorano

first: Isabel

last: Naranjo

first: Susann

last: Ofori

first: Pascal

middle: W

last: Jansen

first: Joëlle

last: Hornebeck

first: Florian

last: Kieferle

first: Elicia

last: Martin

first: Agnes

last: Murk

first: Carolina

last: Castro-Peña

first: Richárd

last: Bártfai

first: Thomas

last: Lavstsen

first: Iris

last: Bruchhaus

first: Tobias

last: Spielmann


title: Multiplexed single-cell characterization of alternative polyadenylation regulators (3PRACE)

first: Madeline

last: Kowalski

first: Hans-Hermann

last: Wessels

first: Johannes

last: Linder

first: Carol

last: Dalgarno

first: Isabella

last: Mascio

first: Saket

last: Choudhary

first: Austin

last: Hartman

first: Yuhan

last: Hao

first: Anshul

last: Kundaje

first: Rahul

last: Satija


title: Estrogen-induced chromatin looping changes identify a subset of functional regulatory elements

status: Public on Jun 12 2024

city: Salt Lake City

first: Jason

last: Gertz

state: UT

address: 2000 circle of hope drive

department: Oncological Sciences

institute: University of Utah

summary: Transcriptional enhancers can regulate individual or multiple genes through long-range three-dimensional (3D) genome interactions, and these interactions are commonly altered in cancer. Yet, the functional relationship between changes in 3D interactions associated with regulatory regions and differential gene expression appears context-dependent. In this study, we used HiChiP to capture changes in 3D genome interactions between active regulatory regions of endometrial cancer cells in response to estrogen treatment and uncovered significant differential long-range interactions that are strongly enriched for estrogen receptor  (ER) bound sites (ERBS). The ERBS anchoring differential loops with either a gene’s promoter or distal regions were correlated with larger transcriptional responses to estrogen compared to ERBS not involved in differential interactions. To functionally test this observation, CRISPR-based Enhancer-i was used to deactivate specific ERBS, which revealed a wide range of effects on the transcriptional response to estrogen. However, these effects are only subtly and not significantly stronger for ERBS in differential loops. In addition, we observed an enrichment of 3D interactions between the promoters of estrogen up-regulated genes and found that looped promoters can work together cooperatively. Overall, our work suggests that changes in 3D genome structure upon estrogen treatment identify some functionally important regulatory regions; however, these changes aren’t required for a transcriptional response to E2 in endometrial cancer cells.

overall_design: Endometrial cancer cell lines treated with estrogen or vehicle control and profiled using H3K27ac Hi-ChIP

first: Hosiana

last: Abewe

first: Jeffery

last: Vahrenkamp