title: Hyper-IgE Syndrome due to an Elusive Novel Intronic Homozygous Variant in DOCK8

city: Darlinghurst

first: Cindy

middle: S.

last: Ma

state: New South Wales

address: 384 Victoria St

institute: Garvan Institute of Medical Research

summary: Rare, biallelic loss-of-function mutations in DOCK8 result in a combined immune deficiency characterized by severe and recurrent cutaneous infections, eczema, allergies, and susceptibility to malignancy, as well as impaired humoral and cellular immunity and hyper-IgE. The advent of next-generation sequencing technologies has enabled the rapid molecular diagnosis of rare monogenic diseases, including inborn errors of immunity. These advances have resulted in the implementation of gene-guided treatments, such as hematopoietic stem cell transplant for DOCK8 deficiency. However, putative disease- causing variants revealed by next-generation sequencing need rigorous validation to demonstrate pathogenicity. Here, we report the eventual diagnosis of DOCK8 deficiency in a consanguineous family due to a novel homozygous intronic deletion variant that caused aberrant exon splicing and subsequent loss of expression of DOCK8 protein. Remarkably, the causative variant was not initially detected by clinical whole-genome sequencing but was subsequently identified and validated by combining advanced genomic analysis, RNA-seq, and flow cytometry. This case highlights the need to adopt multipronged confirmatory approaches to definitively solve complex genetic cases that result from variants outside protein-coding exons and conventional splice sites.

overall_design:
RNA-seq on Illumina NextSeq 550, 150bp paired end reads

Processed data were not produced for these samples.

middle: S

first: Stuart

middle: G

last: Tangye

first: Paul

last: Gray

first: Bethany

last: Pillay

first: Jin

last: Yap

first: William

last: Figgett

first: John

last: Reeves

first: Sarah

last: Kummerfeld

first: Jennifer

last: Stoddard

first: Gulbu

last: Uzel

first: Hule

last: Jing

first: Helen

last: Su

first: Dianne

last: Campbell

first: Anna

last: Sullivan

first: Leslie

last: Burnett

first: Jane

last: Peake


title: Identification of cell-types in the mouse stomach

status: Public on May 19 2023

city: Wako

first: Itoshi

last: NIKAIDO

state: Saitama

address: 2-1 Hirosawa

department: Center for Biosystems Dynamics Research

country: Japan

institute: RIKEN

type: Expression profiling by high throughput sequencing

summary: We determined the cell types present in the mouse stomach by single-cell RNA-seq with Quartz-Seq2.

overall_design: We dissociated mouse adult stomach to single-cell by enzymatic treatment. Single cell was sorted into lysis buffer in each well of a 384-well PCR plate using MoFlo Astrios EQ cell sorter. Sequence library DNA was prepared according to Quartz-Seq2 method. We analyzed the sequence library DNA using NextSeq500. Digital expression matrix was calculated with fastq files.

first: Hitomi

last: Takada

first: Yohei

last: Sasagawa

first: Mika

last: Yoshimura

first: Kaori

last: Tanaka

first: Yoshimi

last: Iwayama

first: Tetsutaro

last: Hayashi

first: Ayako

last: Isomura-Matoba

last: Nikaido

first: Akira

last: Kurisaki


title: Neoantigen-Specific CD8 T Cells with High Structural Avidity Preferentially Reside in and Eliminate Tumors (VDJ)

status: Public on May 18 2023

city: Lausanne

first: David

last: Barras

state: Vaud

address: AGORA, Bugnon 25A

department: Oncology

country: Switzerland

institute: Centre hospitalier universitaire vaudois

summary: The success of cancer immunotherapy depends in part on the strength of antigen recognition by T cells. We characterized the T cell receptor (TCR) functional (antigen sensitivity) and structural (monomeric pMHC-TCR off-rates) avidities of 371 CD8 T cell clones specific for neoantigens, tumor-associated antigens (TAAs) or viral antigens isolated from tumors or blood of patients and healthy subjects. T cells from tumors exhibit stronger functional and structural avidity than their blood counterparts. Relative to TAA, neoantigen-specific T cells are of higher structural avidity and, consistently, are preferentially detected in tumors. Effective tumor infiltration in mice models is associated with high structural avidity and CXCR3 expression. Based on TCRs biophysicochemical properties, we derived and applied an in silico model predicting TCR structural avidity and validated the enrichment in high avidity T cells in patients’ tumors. These observations indicate a direct relationship between neoantigen recognition, T cell functionality and tumor infiltration. These results delineate a rational approach to identify potent T cells for personalized cancer immunotherapy.

overall_design: Profiling of expanded TILs from four melanoma patients by live cell-sorted single cell RNAseq

first: Julien

last: Schmidt

first: Johanna

last: Chiffelle

first: Marta

last: Perez

first: Morgane

last: Magnin

first: Vincent

last: Zoete

first: George

last: Coukos

first: Alexandre

last: Harari


title: PHDs-seq: A Large-scale Phenotypic Screening Method for Drug Discovery through Parallel Multi-readout Quantification [Targeted RNA-seq: KF-b3_0723]

city: 海淀区

first: Jun

last: Li

state: 北京市

address: 北京市海淀区颐和园路5号北京大学王克桢楼207

country: China

institute: Peking University

summary: High-throughput phenotypic screening is a cornerstone of drug development and the main technical approach for stem cell research. However, simultaneous detection of activated core factors responsible for cell fate determination and accurate assessment of directional cell transition are difficult using conventional screening methods that focus on changes in only a few biomarkers. The PHDs-seq (Probe Hybridization based Drug screening by sequencing) platform was developed to evaluate compound function based on their transcriptional effects in a wide range of signature biomarkers. In this proof-of-concept demonstration, several sets of markers related to cell fate determination were profiled in adipocyte reprogramming from dermal fibroblasts. After validating the accuracy, sensitivity and reproducibility of PHDs-seq data in molecular and cellular assays, a panel of 128 signalling-related compounds was screened for the ability to induce reprogramming of keloid fibroblasts (KF) into adipocytes. Notably, the potent ATP-competitive VEGFR/PDGFR inhibitor compound, ABT869, was found to promote the transition from fibroblasts to adipocytes. This study highlights the power and accuracy of the PHDs-seq platform for high-throughput drug screening in stem cell research, and supports its use in basic explorations of the molecular mechanisms underlying disease development.

overall_design: Targeted gene expression profiling analysis of PHDs-seq data for human skin cells (KF) treated with (DMEM/high glucose, 1% ITS, 0.5mM isobutylmethylxanthine, 1 μM dexamethasone) plus 128 compounds, respectively.

last: Chi

first: Yang

last: Yang

first: Zhongya

last: Song

last: Yong

first: Xin

last: Zhou

last: Liu

last: Zhao


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [HU117_CBE_PEM-seq]

city: Beijing

first: Jinchun

last: Wu

address: No. 5, Yiheyuan street

summary: Cohesin loss-of-function mutations are frequently observed in tumors, but the mechanism is unclear. Here, we found that depletion of RAD21, a core subunit of cohesin, leads to massive genome-wide DNA breaks, up to five-fold, and 147 translocation hotspot genes that are co-mutated with cohesin in multiple cancers. Increased DNA damages are independent of RAD21-loss-induced transcription alteration and loop anchor elimination. However, damage-induced chromosomal translocations coincide with the asymmetrically distributed Okazaki fragments of DNA replication, suggesting that RAD21 depletion causes replication stresses evidenced by the slower replication speed and increased stalled forks. Mechanistically, approximately 30% of the human genome exhibits an earlier replication timing after RAD21 depletion, caused by the early initiation of >900 extra dormant origins. Correspondingly, most translocation hotspot genes lie in timing-altered regions. Therefore, we conclude that cohesin dysfunction causes replication stresses induced by excessive DNA replication initiation, resulting in gross DNA damages that may promote tumorigenesis.

overall_design: We employed  primer-extension-mediated sequencing (PEM-seq) to study the level of genome instability under replication stresses.

first: Zhengrong

last: Zhangding

first: Xuhao

first: Jiazhi

last: Hu


title: Atoh1b-dependent genes in early zebrafish inner ear development

city: Dresden

first: Stefan

last: Hans

address: Fetscherstraße 105

department: CRTD - Center for Regenerative Therapies Dresden

country: Germany

institute: Technische Universität Dresden

summary: Identification of genes regulated by the distal-less homeodomain transcription factor Atoh1b. To this aim, we performed gene expression profiling analysis using data obtained from RNA-seq of 3 samples each from wt control as well as atoh1b knockdown.

overall_design: Generation of a transcriptome dataset of control and Atoh1b-depleted zebrafish embryos using the pax8:DsRed transgene, which labels the developing OEPD and kidney anlagen. At OEPD stages, we performed fluorescence-activated cell sorting (FACS) and the isolated transcripts from sorted cells of control and Atoh1b-depleted embryos were used for transcriptome profiling. Comparison of control versus Atoh1b-depleted samples reveals numerous differentially expressed genes.

first: Diana

last: Ezhkova

first: Simone

last: Schwarzer

first: Sandra

last: Spieß

first: Michaela

last: Geffarth

first: Anja

last: Machate

first: Daniela

last: Zöller

last: Stucke

first: Dimitra

last: Alexopoulou

first: Mathias

last: Lesche

first: Andreas

last: Dahl


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [MYC-repli-stress]

first: Yang


title: Transcriptome-based chemical screens identify CDK8 as a common barrier in multiple cell reprogramming systems [ATAC-seq]

first: Jun

last: Li

address: 北京市海淀区颐和园路5号北京大学王克桢楼207

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Fibroblasts can be chemically induced to pluripotent stem cells (CiPSCs) through an extraembryonic endoderm (XEN)-like state or directly converted into other differentiated cell lineages. However, the mechanisms underlying chemically-induced cell fate reprogramming remain unclear. Here, a transcriptome-based screen of biologically active compounds uncovered that CDK8 inhibition was essential to enable chemically-induced reprogramming from fibroblasts into XEN-like cells, then CiPSCs. RNA-seq analysis showed that a CDK8 inhibitor, MSC2530818, inhibited pro-inflammatory pathways that suppress chemical reprogramming, and facilitated the induction of a multi-lineage priming state, indicating the establishment of plasticity in fibroblasts. CDK8 inhibition also resulted in chromatin accessibility profile and Pol II occupation profile similar to that under initial chemical reprogramming. Moreover, CDK8 inhibition greatly promoted transgene-mediated reprogramming of mouse fibroblasts into hepatocyte-like cells, and chemical reprogramming of human fibroblasts into adipocytes. These collective findings thus define CDK8 as a general molecular barrier in multiple cell reprogramming processes, and as a common target for inducing plasticity and cell fate conversion.

overall_design: Comparative chromation states of ATAC-seq data in MEFs with MEF medium, CFAE+8, CFAE+8+MSC, CFAE+6 (C, CHIR99021; F, Forskolin; A, AM580; E, EPZ04777; 6, E-616452; 8, A-83-01; MSC, MSC2530818) induction for 4 days, respectively.

first: Yunfei

last: Bai

first: Zhongya

last: Song

last: Yong


title: CRAC of yeast RNA polymerase II in Topoisomerase 1 and 2 depleted cells

city: Montpellier

first: Domenico

last: Libri

address: 1919 route de Mende

country: France

institute: IGMM - CNRS

summary: We obtained  RNA polymerase II profiles across the genome of S.cerevisiae cells harbouring Top1-AID and Top2-AID tagged proteins. Cells were treated with anauxin analog (IAA) to induce degradation of Top1-AID and Top2-AID.  This allowed us to compare RNAPII in absence of Top1 and Top2 activity to the control condition (absence of IAA treatment).

overall_design: The nascent RNA was crosslinked to Rpb1 (RNAPII) and sequenced after purification of the target protein under stringent and denaturing conditions. This reveals the position of the target protein at the nucleotide resolution level.

first: Umberto

last: Aiello


title: ATF3 promotes endothelial cell response to acute lung injury

city: Philadelphia

first: Michael

middle: Patrick

last: Morley

state: PA

address: 3400 Civic Center Blvd, Bldg 421

department: Penn Cardiovascular Institute

country: USA

institute: Perelman School of Medicine at the University of Pennsylvania

summary: Following acute injury, the capillary vascular bed in the lung must be repaired to reestablish gas exchange between pulmonary endothelial cells (ECs) lining these vessels and the alveolar epithelium. However, the factors that control EC stress response and drive regeneration of pulmonary capillaries remain incompletely understood. Viral infections such as influenza and COVID-19 may indirectly damage lung vasculature through loss of epithelial gas exchange partners or through signaling from infiltrating immune cells. To prevent excessive tissue damage and to renew the endothelium, ECs must both withstand cellular stress and proliferate after injury. Here, we show that the transcription factor and immediate early gene Atf3 is essential for both responses in the mouse lung after influenza infection. Atf3 expression defines a subpopulation of capillary ECs enriched in genes involved in cellular response to stress, angiogenesis, and vascular development. Endothelial loss of ATF3 results in defective alveolar regeneration: in the absence of ATF3, ECs exhibit increased apoptosis and decreased proliferation, resulting in an emphysema-like phenotype with enlarged alveolar airspaces lined with regions of lost vasculature. These data implicate ATF3 as an essential component of the vascular response to acute lung injury that is required for successful lung regeneration.

overall_design: This experiment isolated tdTomato+ endothelial cells from Cdh5-CreERT2; ROSA26-tdTomato mice (control or "Atf3-WT" endothelial cells) and Cdh5-CreERT2; ROSA26-tdTomato; Atf3 lox/lox mice (experimental or "Atf3-KO" ECs). Single-cell suspensions were prepared from whole mouse lungs, and tdTomato+ cells were isolated by FACS. Cells were pelleted and lysed for RNA extraction, cDNA preparation, and RNA sequencing library preparation using the NEB Protocol for Low Input RNA kit (NEB#E6420). Sequencing was performed by Genewiz (now Azenta Life Sciences).

first: Terren

middle: K

last: Niethamer

middle: P

first: Edward

middle: E

last: Morrisey


title: RNAseq analysis of dorsal root ganglia from global Nav1.7 knockout mice

city: South San Francisco

first: James

last: Maksymetz

state: CA

address: 1 DNA Way

department: Neuroscience

institute: Genentech, Inc

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

overall_design: Refer to individual Series


title: Dlx3b/4b-dependent genes in early zebrafish inner ear development

first: Stefan

last: Hans

summary: Identification of genes regulated by the distal-less homeodomain transcription factors Dlx3b and Dlx4b (Dlx3b/4b). To this aim, we performed gene expression profiling analysis using data obtained from RNA-seq of 3 samples each from wt control as well as dlx3b/4b knockdown.

overall_design: Generation of a transcriptome dataset of control and Dlx3b/4b-depleted zebrafish embryos using the pax8:DsRed transgene, which labels the developing OEPD (otic-epibranchial progenitor domain) and kidney anlagen. At OEPD stages, we performed fluorescence-activated cell sorting (FACS) and the isolated transcripts from sorted cells of control and Dlx3b/4b-depleted embryos were used for transcriptome profiling. Comparison of control versus Dlx3b/4b-depleted samples reveals numerous differentially expressed genes.

first: Diana

last: Ezhkova

first: Simone

last: Schwarzer

first: Sandra

last: Spieß

last: Geffarth

first: Johanna


title: Mechanistic impacts of bacterial diet on dopaminergic neurodegeneration in a Caenorhabditis elegans α-synuclein model of Parkinson's disease

city: Tuscaloosa

first: Guy

last: Caldwell

state: AL

address: 300 Hackberry Lane

department: Biological Sciences

institute: The University of Alabama

summary:
Using distinct mutant strains deficient in key aspects of gene silencing machinery, we provide evidence that neuroprotection appears to stem from dsRNAs that originate from the bacteria themselves, and Systemic RNA Interference Defective (SID) mutants deficient in dsRNA-mediated gene silencing do not allow for neuroprotection from the bacteria.
Transcriptomic analysis between worms reared on OP50 and HB101 E. coli for multiple generations in an a-syn background revealed that the primary cellular processes involved in neuroprotection of this nature included but were not limited to endopeptidase activity, transcriptional regulation, calcium ion binding activity, and water channel activity.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for C. elegans in an a-synuclein background reared on OP50 or HB101 E. coli, 3 biological replicates.

first: Anthony

middle: L

last: Gaeta

first: Luke

last: McKay

first: Candice

middle: M

last: Keogh

first: Karolina

last: Willicott

first: Tyler

last: Altman

middle: A

first: Kim


title: Stuxnet fine-tunes Notch dose during development using a functional Polycomb response element

first: Tao

last: He

address: Summer Palace Road No. 5

institute: Peking university

summary:
Evolutionarily conserved Notch signaling is highly sensitive to changes
in Notch receptor dose caused by intrinsic and environmental
fluctuations. It is well known that epigenetic regulation responds
dynamically to genetic, cellular and environmental stresses. However,
it is unclear whether the Notch receptor dose is directly regulated at the
epigenetic level. Here, by studying the role of the upstream epigenetic
regulator Stuxnet (Stx) in Drosophila developmental signaling, we find
that Stx promotes Notch receptor mRNA expression by counteracting
the activity of Polycomb repressive complex 1 (PRC1). In addition, we
provide evidence that Notch is a direct PRC1 target by identifying and
validating in vivo the only bona fide Polycomb response element (PRE)
among the seven Polycomb group (PcG)-binding sites revealed by
DamID-seq and ChIP-seq analysis. Importantly, in situ deletion of this
PRE results in increased Notch expression and phenotypes resembling
Notch hyperactivation in cell fate specification. These results not only
underscore the importance of epigenetic regulation in fine-tuning the
Notch activity dose, but also the need to assess the physiological
significance of omics-based PcG binding in development.

overall_design: Three samples: Dam, Dam-Pc, and Dam-Pc and Stx drove by nub-Gal4.

first: Mengyuan

last: Yi

first: Min

last: Liu

first: Alan

middle: J

last: Zhu


title: Ageing restructures the basal transcriptome of the core hypothalamic osmoregulatory control centre and alters the response to dehydration

city: Bristol

first: Ghadir

last: ElSamad

address: Whitson St.

country: United Kingdom

institute: University of Bristol

summary:
To investigate  the polyadenylated transcriptomes of the Wistar Han rat SON in EH from 3-month old euhydrated and dehydrated rats and from 18-month old euhydrated and dehydrated rats.
We then performed gene expression profiling analysis using data obtained from adult and aged rat in EH and Dh conditions

overall_design: Comparative gene expression profiling analysis of RNA-seq data for the Control and stimulated SON in adult and aged rats.

first: Audrys

last: Pauza

first: Andre

last: Mecawi

first: Michael

last: Greenwood

first: David

last: Murphy


title: Sox9 Deletion Causes Severe Intervertebral Disc Degeneration Characterized by Apoptosis, Matrix Remodeling, and Compartment-Specific Transcriptomic Changes

first: Makarand

last: Risbud

address: 1015 Walnut Street

institute: Thomas Jefferson University

summary:
Transcriptomic analysis of nucleus pulposus (NP) and annulus fibrosus (AF) tissues from intervertebral discs of 3-month-old mice with the conditional postnatal deletion of Sox9.
The transcription factor Sox9 is essential for the maintenance and health of the intervertebral disc of mice. We examined the transcriptomic profiles of nucleus pulposus and annulus fibrosus cells in control and Sox9 conditional knock-out mice using microarrays.

overall_design: Tissues from the nucleus pulposus (NP) and annulus fibrosus (AF) were harvested from the intervertebral discs of four Sox9 control (Sox9CTR) (Sox9fl/fl) and four Sox9 conditional knock-out mice (Sox9cKO) (AcanCreERT2Sox9fl/fl) between 3 and 4 months of age. NP and AF tissues were separately pooled from lumbar and caudal discs in each animal for RNA extraction and hybridization using Affymetrix microarrays.

first: Maria

last: Tsingas

first: Olivia

last: Ottone

middle: V


title: Senohemostatic role for DPP4 in atherosclerosis

city: Baltimore

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

institute: NIA-IRP, NIH

summary: We performed scRNA sequencing to understand the composition and differential gene expression changes in aortic cells from LDLR-/- mice fed a normal diet (ND), high-fat diet (HFD), or HFD + DPP4i. We combined this technique with CITE-sequencing to measure the cell surface protein expression (DPP4) and transcriptomic changes associated with our experimental conditions as well as changes associated with DPP4 cell surface expression in all aortic cells.

overall_design: LDL receptor knock out (Ldlr−/−) mice on the C57BL/6 background purchased from Jackson labs (Catalog #002207) were bred to obtain homozygous Ldlr−/− knock out mice.  Mice of both sexes were entered in the study at 8 weeks of age when normal chow was replaced with an atherogenic diet (42% Fat, 0.2% cholesterol, Harlan atherogenic diet TD.88137).  For the treatment groups, mice were gavaged with 10 mg/kg/day of Vildagliptin from SelleckChem or an equal volume of DMSO, 5 days per week for 16 weeks. Mice were anesthetized and blood was extracted using cardiac puncture.  The whole aorta was collected after left ventricular perfusion with 10 mL of PBS and quickly transferred to cold PBS. To prepare a single-cell suspension, perivascular adipose tissue was removed and whole aortas from three mice were cut into ~1-mm pieces and digested with an enzyme solution consisting of 10 mg/ml Collagenase II (Sigma) and 1 mg/ml elastase (Worthington) for 15 min at 37 °C. The cell suspension was strained through a 40-μm filter and centrifuged at 500 x g for 5 min. Cells were resuspended in 100 µl Cell Staining Buffer with 15% FBS and 5 µl of Human TruStain FcX™ Fc Blocking reagent (or 0.5 µl of TruStain FcX™ PLUS (anti-mouse CD 16/32) blocking reagent.  Cells were incubated for 10 minutes at 4°C, followed by the addition of 1 µg of DPP4 (CD26)-APC antibody and incubated for 30 minutes at 4°C.  Cells were incubated with TotalSeq™ APC secondary antibody for 30 minutes at 4C. Next, cells were washed with 1 ml of Cell Staining Buffer, spun at 400g for 5 minutes, and resuspended in 1 mL of PBS + 1X EDTA (0.5M = 100x).  Cells were stained with viability dye as described in the Flow Cytometry method and single, viable cells were collected for 10X library preparation.


title: Neoantigen-Specific CD8 T Cells with High Structural Avidity Preferentially Reside in and Eliminate Tumors (GEX)

last: Barras

first: Julien

last: Schmidt

first: Johanna

last: Chiffelle

first: Marta

last: Perez

first: Morgane

last: Magnin

first: Vincent

last: Zoete

first: George

last: Coukos

first: Alexandre

last: Harari


title: ENO1 promotes liver carcinogenesis through YAP1-dependent arachidonic acid metabolism

city: Guangzhou

first: Linchong

last: Sun

state: Guangdong

address: B2 Building, South China University of Technology, No. 382, Waihuandong Road, Guangzhou Higher Education Mega Center, Panyu District

institute: South China University of China

summary: Enolase 1 (ENO1) is a glycolytic enzyme that plays essential roles in various pathological activities including cancer development. However, the mechanisms underlying ENO1-contributed tumorigenesis are not well explained. Here, we uncover that ENO1, as an RNA-binding protein, binds to the cytosine-uracil-guanine-rich elements of YAP1 messenger RNA to promote its translation. ENO1 and YAP1 positively regulate alternative arachidonic acid (AA) metabolism by inverse regulation of PLCB1 and HPGD (15-hydroxyprostaglandin dehydrogenase). The YAP1/PLCB1/HPGD axis-mediated activation of AA metabolism and subsequent accumulation of prostaglandin E2 (PGE2) are responsible for ENO1-mediated cancer progression, which can be retarded by aspirin. Finally, aberrant activation of ENO1/YAP1/PLCB1 and decreased HPGD expression in clinical hepatocellular carcinoma samples indicate a potential correlation between ENO1-regulated AA metabolism and cancer development. These findings underline a new function of ENO1 in regulating AA metabolism and tumorigenesis, suggesting a therapeutic potential for aspirin in patients with liver cancer with aberrant expression of ENO1 or YAP1.

overall_design: RNA immunoprecipitation sequencing (RIP-seq) analysis in HEK293T cells overexpressing ENO1, primary antibodies against ENO1 (11204-1-AP, Proteintech) were used in the RIP-seq assay. PLC cells with ENO1 or YAP1 knockdown were cultured with DMEM, then total RNA was extracted using Trizol Reagent followed by RNA-seq to identify the co-regulated genes by ENO1 and YAP1.

first: Shengqi

last: Shen

first: Huafeng

last: Zhang

first: Ping

last: Gao


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [q3C-seq]

first: Jinchun

last: Wu

overall_design: We employed  quantitative 3C-seq (q3C) to quantify the elimination of loop after RAD21 depletion.

first: Yang

last: Liu

first: Zhengrong

last: Zhangding

first: Xuhao

first: Jiazhi

last: Hu


title: Neoantigen-Specific CD8 T Cells with High Structural Avidity Preferentially Reside in and Eliminate Tumors

first: David


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [mES_Repli_Timing]

overall_design: We applied NAIL-seq to identify the replication timing in WT and RAD21-depleted mES cells.


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [NAIL-seq, EdUHU]

summary: Cohesin mutations are frequently observed in tumors, but the mechanism is unclear. Here, we found that depletion of RAD21, a core subunit of cohesin, leads to gross DNA breaks and consequently elevates genome-wide chromosomal translocation levels up to five-fold in mammalian cells. These damages are independent of transcription alteration and loop anchor robustness. However, identified translocations occur asymmetrically on the chromatin Watson and Crick strands, coincident with Okazaki fragments of DNA replication. Moreover, early DNA replication initiates at about one thousand extra dormant origins after RAD21 depletion, which is independent of CTCF, resulting in earlier replicating timing at approximately 30% or 13% of the human or mouse genomic regions, respectively. And the translocation hotspot genes arising in RAD21-depleted cells are accompanied by dysregulated replication timing. Therefore, we conclude that cohesin dysfunction leads to perturbed DNA replication timing, which further results in gross DNA damages that may be involved in tumorigenesis.

overall_design: We applied NAIL-seq to identify the early replication initiation zones in RAD21-depleted cells.


title: Perturbation on DNA replication by cohesin dysfunction leads to gross genomic damages [PEM-seq-mutant]

overall_design: We employed  primer-extension-mediated sequencing (PEM-seq) to study the level of genome instability after RAD21-depletion.


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [PEM-seq: RAD21-shRNA]

overall_design: We employed  primer-extension-mediated sequencing (PEM-seq) to study the level of genome instability afterr RAD21 knockdown.


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [HU120-CBE_NAIL-seq_END-seq]

overall_design: We employed  NAIL-seq and END-seq to study the replication initiation and DNA breaks after depletion of CBE at c-MYC locus.


title: Differences in the neural basis and transcriptomic patterns in acute and persistent pain-related anxiety-like behaviors

first: Lianyan

last: Huang

address: 74 Zhongshaner Rd

institute: Sun Yat-sen University

summary: To investigate the different mechanism of Formalin induced acute pain and CFA induced persistent pain in mPFC and BNST, mice were treated by Formalin exposure or CFA exposure or without treatment and their mPFC and BNST were dissected to perform RNA seq.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for mPFC and BNST under different treatment (no treatment as  control, Formalin exposure and CFA exposure).

first: Shunchang

last: Fang

first: Yuxin

last: Qin


title: Effect of QKI7 on global RNA stability

city: Monrovia

first: Jianjun

last: Chen

department: Systems Biology

institute: Beckman Research Institute of City of Hope

summary: N7-methylguanosine (m7G) modification, routinely occurring at the 5’ cap of mRNA or within tRNA and rRNA, also exists internally in mRNA. Although essential for mRNA translation as well as stress response, the “reader” protein for mRNA internal m7G modification is still unrevealed. Here, we reported that Quaking protein (QKI), especially QKI7, can selectively recognize the internal mRNA m7G decoration in the cytosol of various cell types. We identified over 1000 confident m7G-modified and QKI binding RNA targets with a conserved motif, “GANGAN (N=A/U/G)”. More strikingly, internal m7G reader QKI7 directly interacts with the SG core protein G3BP1 and can shuttle a subset of m7G-modified transcripts into SG mRNA pool under oxidative stress condition. Additionally, by sequestering mRNA within SGs, QKI7 modulates the translation efficiency of selected transcripts. Moreover, in line with the observation that doxorubicin triggers the assembly of SGs, QKI7 mediates the sensitivity of cancer cells to chemotherapy drug treatment.

overall_design: To study the influence of QKI on mRNA stability, we performed mRNA stability proﬁling in U2OS cells (sgNS, sgQKI, sgQKI+QKI7-WT, and sgQKI+QKI7-△KH). Briefly, U2OS cells were treated  with 5 µg/mL actinomycin D and 200 µM sodium arsenite. The total RNA was extracted at indicated time point.

first: Zhicong

last: Zhao

first: Lei

last: Dong

first: Rui

last: Su


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [PRO-seq]

overall_design: We applied PRO-seq to compare the expression difference of WT and RAD21-depleted cells.


title: Perturbation on DNA replication by cohesin dysfunction leads to gross genomic damages [NAIL-seq mES-EdUHU]

overall_design: We applied NAIL-seq to identify the early replication initiation zones in mES RAD21-depleted cells.


title: Perturbation on DNA replication by cohesin dysfunction leads to gross genomic damages [END-seq]

summary: Cohesin loss-of-function mutations are frequently observed in tumors, but the mechanism is unclear. Here, we found that depletion of RAD21, a core subunit of cohesin, leads to massive DNA breaks, up to five-fold, and 147 translocation hotspot genes that are co-mutated with cohesin in multiple cancers. DNA damages are independent of RAD21-loss-induced transcription alteration and loop anchor elimination. However, translocation junctions coincide with the asymmetrically distributed Okazaki fragments of DNA replication, suggesting that RAD21 depletion causes replication stresses evidenced by the slower replication speed and increased stalled forks. Mechanistically, approximately 30% of the human genome exhibits an earlier replication timing after RAD21 depletion, caused by the early initiation of >900 extra dormant origins. Correspondingly, most translocation hotspot genes lie in timing-altered regions. Therefore, we conclude that cohesin dysfunction causes severe replication stresses induced by extra DNA replication, resulting in gross DNA damages that may induce tumorigenesis.

overall_design: We applied END-seq to detect the genome-wide DNA breaks in WT and RAD21-depleted K562 cells.


title: Single-cell chromatin accessibility of developing murine pancreas identifies cell state-specific gene regulatory programs

city: San Francisco

first: Julie

middle: B

last: Sneddon

state: California

address: 500 Parnassus Ave.

institute: UCSF

summary: A number of studies have reported cell heterogeneity within the developing mouse pancreas, as well as the transcriptional profiles corresponding to various cell states. The upstream mechanisms that initiate and maintain gene expression programs across cell states, however, remain largely unknown. Here, we applied single-nucleus ATAC-Seq to developing mouse pancreas to generate an atlas of chromatin accessibility, at single-cell resolution. Our goals were first, to generate an atlas of chromatin accessibility of embryonic mouse pancreas, at single-cell resolution, that can serve as a resource for the field. We aimed to provide such a resource not only for epithelial cells within the pancreas, but for non-epithelial (e.g., mesenchymal) as well. Our second goal was to identify gene regulatory networks governing cell fate transitions through integration of single-cell chromatin accessibility and gene expression data.

overall_design: To investigate chromatin accessibility of the developing pancreas, we performed snATAC-Seq on E14.5 and E17.5 murine pancreas from eFev-YFP mice using the 10x Genomics scATAC Seq platform. We profiled both whole pancreas, as well as eFev-YFP+ sorted cells, the latter designed to enriched for endocrine progenitor cells.

first: Sean

last: de la O

first: Zhe

first: Xinaki

last: Yao


title: K562_WT_Replication_Timing

first: Jinchun

summary: Cohesin participates in loop formation by extruding DNA fibers from its ring-shaped structure. Cohesin dysfunction eliminates chromatin loops but only causes modest transcription perturbation, which cannot fully explain the frequently observed mutations of cohesin in various cancers. Here, we found that DNA replication initiates at more than one thousand extra dormant origins after acute depletion of RAD21, a core subunit of cohesin, resulting in earlier replicating timing at approximately 30% of the human genomic regions. In contrast, CTCF is dispensable for suppressing the early firing of dormant origins that are distributed away from the loop boundaries. Furthermore, greatly elevated levels of gross DNA breaks and genome-wide chromosomal translocations arise in RAD21-depleted cells, accompanied by dysregulated replication timing at dozens of hotspot genes. Thus, we conclude that cohesin coordinates DNA replication initiation to ensure proper replication timing and safeguards genome integrity.

overall_design: We applied NAIL-seq to identify the replication timing in WT cells.


title: K562_RAD21_mAC_Replication_Timing

overall_design: We applied NAIL-seq to identify the replication timing in RAD21-depleted cells.

first: Yang


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [PEM-seq]

summary: Cohesin loss-of-function mutations are frequently observed in tumors, but the mechanism is unclear. Here, we found that depletion of RAD21, a core subunit of cohesin, leads to massive genome-wide DNA breaks, up to five-fold, and 147 translocation hotspot genes that are co-mutated with cohesin in multiple cancers. Increased DNA damages are independent of RAD21-loss-induced transcription alteration and loop anchor elimination. However, damage-induced chromosomal translocations coincide with the asymmetrically distributed Okazaki fragments of DNA replication, suggesting that RAD21 depletion causes replication stresses evidenced by the slower replication speed and increased stalled forks. Mechanistically, approximately 30% of the human genome exhibits an earlier replication timing after RAD21 depletion, caused by the early initiation of >900 extra dormant origins. Correspondingly, most translocation hotspot genes lie in timing-altered regions. Therefore, we conclude that cohesin dysfunction causes replication stresses induced by excessive DNA replication initiation, resulting in gross DNA damages that may promote tumorigenesis.

first: Zhengrong


title: Single-cell transcriptomics of human-skin-equivalent organoids

city: Irvine

first: Scott

last: Atwood

address: 4340 McGaugh Hall, Ring Rd

institute: University of California, Irvine

summary: Several methods for generating human-skin-equivalent (HSE) organoid cultures are in use to study skin biology; however, few studies thoroughly characterize these systems. To fill this gap, we use single-cell transcriptomics to compare in vitro HSEs, xenograft HSEs, and in vivo epidermis. By combining differential gene expression, pseudotime analyses, and spatial localization, we reconstruct HSE keratinocyte differentiation trajectories that recapitulate known in vivo epidermal differentiation pathways and show that HSEs contain major in vivo cellular states. However, HSEs also develop unique keratinocyte states, an expanded basal stem cell program, and disrupted terminal differentiation. Cell-cell communication modeling shows aberrant epithelial-to-mesenchymal transition (EMT)-associated signaling pathways that alter upon epidermal growth factor (EGF) supplementation. Last, xenograft HSEs at early time points post transplantation significantly rescue many in vitro deficits while undergoing a hypoxic response that drives an alternative differentiation lineage. This study highlights the strengths and limitations of organoid cultures and identifies areas for potential innovation.

overall_design: We report transcriptomes of human skin equivalents.

first: Adam

middle: R

last: Stabell

middle: X


title: Integrative analysis of hepatic transcriptional profiles reveals genetic regulation of atherosclerosis in hyperlipidemic Diversity Outbred-F1 mice

city: Davis

first: Brian

last: Bennett

address: 430 West Health Sciences Drive

institute: USDA ARS-WHNRC

summary:
Purpose: To investigate the sex-dependence of liver transcriptome in Diversity Outbred (DO)-F1 mice
Methods: Total RNA was extracted from snap-frozen liver using miRVana total RNA isolation kit (Thermo Fisher Scientific, Waltham, MA, USA) according to the manufacturer’s protocol. The quality and amount of liver RNA were evaluated using a Bioanalyzer (Agilent, Inc., Santa Clara, CA). The average RNA-integrity score for 162 DO-F1 liver samples was 9.01 ± 0.4. RNA samples from 85 females and 77 males were submitted to the UC Davis DNA Technologies Core at the Genome Center. The RNA-seq libraries were constructed from 1 µg total RNA after poly-A library preparation. To minimize technical variability, all samples were assigned to each lane and the pooled libraries were sequenced on two lanes of the Illumina NovaSeq 6000 sequencing (Illumina Inc., San Diego, CA, USA) to achieve paired-end reads of at least 25 million 150 bp. Only R1 was used in the analysis and only R1 was submitted.
Results: Our results demonstrate the tremendous effects of sex on hepatic gene expression. In support of this, genetic loci associated with the transcripts frequently showed sex specificity. We revealed sex-specific candidate genes that were mapped to the quantitative trait loci for aortic lesion area and whose expression was regulated locally regulated via global liver transcriptome.
Conclusions: Our study provide a valuable data resource to the research community and show that liver transcriptomic analysis identified diet- or strain-specific pathways to pathogenesis of metabolic syndrome.

overall_design: Liver mRNA profiles of 24-week old Diversity Outbred-F1 mice

first: Excel

last: Que

first: Myungsuk

last: Kim


title: Unique molecular signatures sustained in circulating monocytes and regulatory T cells inconvalescent COVID-19 patients

status: Public on May 17 2023

city: Chicago

first: Huiping

last: Liu

state: IL

address: 303 E Superior St Room 5-250

department: Pharmacology

institute: Northwestern University

summary: We used single-cell RNA sequencing to analyze the immune response of two distinct cell types to COVID-19. We drew blood from convalescent patients and examined monocytes and regulatory T cells.

overall_design: Monocytes and Tregs were sorted using fluorescence activated sorting (FACS) and sequenced using a 10X genomics library preparation kit and an Illumina HiSeq sequencer.

first: Andrew

middle: D

last: Hoffmann

first: Sam

last: Weinberg

first: Suchitra

last: Swaminathan

first: Shuvam

last: Chadhuri

first: Hannah

middle: F

last: Almubarak

first: Matthew

last: Schipma

first: Chengsheng

last: Cmao

first: Xinkun

last: Wang

first: Lamiaa

last: El-Shennawy

first: Nurmaa

last: Dashzeveg

first: Juncheng

last: Wei

first: Paul

last: Mehl

first: Laura

last: Shihadah

first: Ching

last: Wai

first: Carolina

last: Ostiguin

first: Yuzhi

last: Jia

first: Paolo

last: D'Amico

first: Neale

first: Yuan

last: Luo

first: Alexis

last: Demonbreun

first: Michael

last: Ison

first: Deyu


title: Rubella virus tropism and single cell responses in human primary tissue and microglia-containing organoids

first: Chang

last: Kim

address: 505 Parnassus Ave

summary: scRNA-seq of rubella virus infected cortical organoids

overall_design: Two organoid lines with and without microglia that were treated with RV or uninfected and performed scRNA-seq on conditions pooled using Multi-seq

first: Kim

last: Chang


title: Adoptive transfer of type 2 diabetes by hematopoietic stem cells reprogrammed in utero by vitamin D deficiency

first: Ruteja

last: Barve

state: MO

address: 4515 McKinley Ave

department: Genetics

institute: Washington University

summary: This study demonstrates that transplantation of fetal liver hematopoietic stem cells (FL-HSCs) from fetuses subjected to vitamin D (VD) deficiency in utero induced persistent insulin resistance to VD sufficient recipient mice.  Transcriptional changes associated with intrauterine vitamin D deficiency where identified through microarray analysis of mouse BMMs from vitamin D deficient and sufficient conditions.

overall_design: Gene expression profiles of Vitamin D sufficent and deficient BMMs were comapared and pathway analysis performed.

first: Jisu

last: Oh

first: Carlos

last: Bernal-Mizrachi


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [RAD21_ChIPseq]

first: Jinchun

last: Wu

address: No. 5, Yiheyuan street

institute: Peking University

overall_design: We applied ChIP-seq to check the amount of chromatin-bound RAD21 or CTCF.

first: Yang

last: Liu

first: Zhengrong

last: Zhangding

first: Xuhao


title: PP2Ac Deficiency Enhances Tumor Immunogenicity by Activating STING-Type I Interferon Signaling in Glioblastoma [RNA]

city: Memphis

first: Beisi

last: Xu

state: Tennessee

address: 262 Danny Thomas Pl

department: Center for Applied Bioinformatics

institute: St Jude Children's Research Hosipital

summary: Glioblastoma (GBM) is an incurable cancer despite aggressive treatment paradigms. Current immunotherapies, such as CTLA-4 and/or PD-1 blockade, have yet to demonstrate benefits for GBM. A key barrier is the immunologically “cold” nature of GBM defined by insufficient tumor antigen presentation and lack of T cells infiltration. We previously demonstrated that pharmacologic inhibition of Protein Phosphatase-2A (PP2A), using a small molecule inhibitor, synergized with PD1 blockade in multiple preclinical tumor models including GBM. However, PP2A is ubiquitously expressed in many cell types and regulates many cellular pathways. The cell type or molecular mechanism responsible for PP2A-modulated tumor immunity is poorly understood. Here, we demonstrate that genetic ablation of PP2A in glioma cells sufficiently promotes tumor immunogenicity by enhancing 1) dsDNA production and thereby cGAS-Type I interferon (IFN) signaling, 2) MHC-I expression and 3) tumor mutational burden. PP2A deficient glioma enhances DC activation and cross presentation to promote clonal expansion of tumor antigen specific CD8 cells. PP2A deficiency also markedly sensitize tumors to immune checkpoint blockade and radiotherapy. Single cell analysis of murine glioma demonstrates that PP2A deficient tumors have 1) increased infiltration of CD8+ T cell, NK cell and cDC1 cells, 2) reduced infiltration of immunosuppressive tumor associated macrophages, 3) promotion of IFN signaling in both myeloid and tumor cells, and 4) reduced glioma-associated gene signature that predicts poor prognosis in glioma patients. This is the first study to establish a role for PP2A in regulating dsDNA-cGAS-STING signaling and collectively, our results suggest that PP2A is a promising novel target to enhance anti-tumor immunity for glioma.

overall_design: We implanted subcutanous or intracranial SB28 murine glioma in WT and LysMcrePP2Acfl/fl mice. 18 days after implantation, tumor tissue was harvested, and single cell suspensions were obtained and sent for sequencing.

first: Das

last: Oishika

first: Mondal

last: Isha

first: Xu

last: Beisi

first: Lu

last: Rongze

first: Ho

last: Winson


title: Identify QKI-bound transcripts via RIP-seq

first: Jianjun

last: Chen

overall_design: To identify the directly bound transcripts of QKI (QKI5, QKI6, and QKI7), RNA immunoprecipitation sequencing (RIP-seq) was conducted in HepG2 cells stable expressing QKI proteins. Briefly, HepG2 cells were infected with lentivirus, pmiRNA1-3 x Flag-QKI. Only the GFP-positive cells were used for study and expanded in DMEM medium. RIP assay was performed with Flag antibody (F3165, Sigma).

first: Zhicong

last: Zhao

first: Lei

last: Dong

first: Rui

last: Su


title: Gene expression changes in lung adenocarcinoma cells from overexpression of p53-regulated tumor-suppressive lncRNA (PTSL)

first: Ramkrishna

last: Mitra

state: Pennsylvania

address: 233 S 10th street

summary: RNA-sequencing was performed to evaluate gene expression changes following ectopic expression of PTSL or vector control in lung adenocarcinoma cells. Genes and pathways that induce proliferation were reduced with elevated levels of PTSL.

overall_design: From RNA-sequencing gene expression profiles were generated in PTSL-expressing lung adenocarcinoma cells and control lung adenocarcinoma cells.

first: Christine

last: Eischen

first: Clare

last: Adams


title: PP2Ac Deficiency Enhances Tumor Immunogenicity by Activating STING-Type I Interferon Signaling in Glioblastoma [scRNA]


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [MCM5_ChIPseq]

first: Jinchun

overall_design: We applied ChIP-seq to depict the distribution of chromatin-bound MCM5 with and without RAD21 during G1 phase.

first: Jiazhi

last: Hu


title: Impact of Major Facilitator Superfamily Domain containing 2a (Mfsd2a) mediated lipid transport on oligodendrocyte lineage development at single cell level.

city: Singapore

first: Vetrivel

last: Sengottuvel

address: 8 College Road

department: Cardiovascular and Metabolic Disorders

country: Singapore

institute: Duke NUS Medical School

summary: Mfsd2a is involved in transport of essential fatty acids such as docosahexaenoic acid (DHA) from the periphery across blood brain barrier (BBB) into the brain parenchyma. Loss of Mfsd2a in humans leads to microcephaly and hypomyelination. The significance of lipid species transported by Mfsd2a on oligodendrocyte lineage development is not known. Using OPC specific deletion of Mfsd2a (2aOKO) in mice we found that OPC cell state, differentiation and oligodendrocytes maturation is disrupted resulting in hypomyelination compared to 2afl/fl controls. Single cell RNAsequencing analysis was performed on total oligodendrocyte population from postnatal mouse brain using Ribo-TRAP mouse lines that labels complete oligodendrocyte lineage with GFP fluorescence to understand the influence of Mfsd2a on oligodendrocyte development.

overall_design: Olig2-EGFP/Rpl10a (OL-Ribo-TRAP) BAC-transgenic mouse line from Jax, Stock # 030265 crossed to 2afl/fl and 2aOKO was used for the study. Total oligodendrocyte (OL) lineage population from postnatal day 8 mouse brain of 2afl/fl-OL-Ribo-TRAP (control) and 2aOKO-OL-Ribo-TRAP (OPC specific Mfsd2a knockout) were isolated using GFP fluorescence based flow cytometry. GFP positive cells from both these genotypes were analysed using single cell RNAsequencing.

first: Monalisa

last: Hota

first: Sujoy

last: Ghosh

first: David

last: Silver


title: Recognition of internal mRNA N7-methylguanosine by QKI shuttles transcripts into stress granules and regulates translation

summary: Recent studies reported that N7-methylguanosine (m7G) modification exists in internal mRNAs; however, the “reader” protein for mRNA internal m7G modification is still unrevealed. Here, by performing m7G MeRIP-seq and RIP-seq, we identified quaking protein (QKI) as a novel internal m7G modification reader. Internal mRNA m7G modification acts as a key player in mRNA translation under stress condition. To explore the mechanism underlying the function of QKI in translation regulation, we employed RNA-seq and Ribo-seq, and demonstrated that QKI7 regulates the translation efficiency of a subset of internal m7G-modified transcripts under stress condition.

overall_design: Refer to individual Series


title: Effect of METTL1 KO on the binding ability of QKI7

overall_design: To study whether METTL1 mediates the binding ability between QKI7 and its target transcripts, RNA immunoprecipitation sequencing (RIP-seq) was conducted in HepG2-cas9 cells stably expressing QKI7 protein with or without METTL1 KO. Briefly, HepG2-cas9 cells were infected with lentivirus, pmiRNA1-3 x Flag-QKI7. After that, METTL1 was depleted using sgRNA. Only the GFP-positive cells were used for study and expanded in DMEM medium. RIP assay was performed with Flag antibody (F3165, Sigma).


title: Identify QKI7-bound transcripts via RIP-seq

overall_design: To identify the directly bound transcripts of QKI7, RNA immunoprecipitation sequencing (RIP-seq) was conducted in U2OS cells stable expressing QKI proteins. Briefly, U2OS cells were infected with lentivirus, pmiRNA1-3 x Flag-QKI7. Only the GFP-positive cells were used for study and expanded in DMEM medium. RIP assay was performed with Flag antibody (F3165, Sigma).


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes [NAIL-seq]

last: Wu

overall_design: We applied NAIL-seq to identify the early replication initiation zones in RAD21-depleted cells.

first: Yang

last: Liu

first: Zhengrong

last: Zhangding

first: Xuhao


title: Transcriptional changes in oligodendrocyte precursor cells (OPC) and immature oligodendrocytes (O4) of postnatal mouse brain due to Major Facilitator Superfamily Domain containing 2a (Mfsd2a) mediated lipid transport.

summary: Mfsd2a plays an important role in accretion of essential fatty acids such as docosahexaenoic acid (DHA) from the periphery into the brain. Loss of Mfsd2a in humans leads to microcephaly and hypomyelination. The precise impact of lipid species transported by Mfsd2a on myelination is not known. Using OPC specific deletion of Mfsd2a (2aOKO) in mice we found that OPC cell state, differentiation and oligodendrocytes maturation is disrupted resulting in hypomyelination. RNAsequencing and differential gene expression analysis was performed on OPC and O4 cells from postnatal mouse brain to understand the influence of Mfsd2a on oligodendrocyte development.

overall_design: OPC and O4 cells from postnatal day 8 mouse brain of 2afl/fl and 2aOKO were isolated using magnetic cell sorting (MACS Miltenyi microbeads kit). Total RNA was extracted from both these cell types for RNAsequencing. 3-4 mice per genotype was used.


title: Transcriptome Mapping of Internal N7-methylguanosine modification via m7G MeRIP-seq [m7g.2]

summary: Previous study found the presence of internal N7-methylguanosine (m7G) within mammalian mRNA. We performed antibody-based m7G MeRIP-seq for transcriptome-wide mapping the internal m7G sites in U2OS cells.

overall_design: The U2OS cells were collected and total RNA were isolated from the whole cells via Qiazol. The poly(A) RNAs were further enriched from the total RNA and then subjected to m7G IP.


title: Postnatal expression of CD38 in astrocytes regulates synapse formation and adult social memory

city: Kanazawa City

first: Tsuyoshi

last: Hattori

state: Ishikawa

address: 13-1 Takara-Machi

department: Department of Neuroanatomy

institute: Kanazawa University

summary: To investigate effect of CD38 knockout on gene expression in astrocytes, we performed RNA-seq of WT and CD38 KO primary astrocytes.

overall_design: Comparative gene expression profiling analysis of RNA-seq data from WT and CD38 KO primary astrocytes

first: Takayuki

last: Kannon

first: Kazuyoshi

last: Hosomichi

first: Atsuhi

last: Tajima

first: Osamu

last: Hori


title: Determine the effects of QKI7 on translation efficiency via ribosome profiling (Ribo-seq)

first: Jianjun

last: Chen

summary: N7-methylguanosine (m7G) modification, routinely occurring at the 5’ cap of mRNA or within tRNA and rRNA, also exists internally in mRNA. Although essential for mRNA translation as well as stress response, the “reader” protein for mRNA internal m7G modification is still unrevealed. Here, we reported that Quaking protein (QKI), especially QKI7, can selectively recognize the internal mRNA m7G decoration in the cytosol of various cell types. We identified over 1000 confident m7G-modified and QKI binding RNA targets with a conserved motif, “GANGAN (N=A/U/G)”. More strikingly, internal m7G reader QKI7 directly interacts with the SG core protein G3BP1 and can shuttle a subset of m7G-modified transcripts into SG mRNA pool under oxidative stress condition. Additionally, by sequestering mRNA within SGs, QKI7 modulates the translation efficiency of selected transcripts. Moreover, in line with the observation that doxorubicin triggers the assembly of SGs, QKI7 mediates the sensitivity of cancer cells to chemotherapy drug treatment.

overall_design: Two 15-cm dishes empty vector or QKI7 overexpression U2OS cells were treated with 0.5mM NaAsO2 for 1 hour before harvesting the cells. The samples were subjected to ribosome profiling and both the input RNA and footprint RNAs were used for sequencing.

first: Zhicong

last: Zhao

first: Lei

last: Dong

first: Rui

last: Su


title: PP2Ac Deficiency Enhances Tumor Immunogenicity by Activating STING-Type I Interferon Signaling in Glioblastoma

first: Beisi

last: Xu

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


title: Transcriptome Mapping of Internal N7-methylguanosine modification via m7G MeRIP-seq [m7g.1]

summary: Previous study found the presence of internal N7-methylguanosine (m7G) within mammalian mRNA. We performed antibody-based m7G MeRIP-seq for transcriptome-wide mapping the internal m7G sites in HepG2 cells.

overall_design: The HepG2 cells were collected and total RNA were isolated from the whole cells via Qiazol. The poly(A) RNAs were further enriched from the total RNA and then subjected to m7G IP.


title: PIE-Seq: Identifying RNA-binding Protein Targets by Dual RNA-deaminase Editing and Sequencing

first: Xiangbin

last: Ruan

address: 6054 S. DREXEL AVE.

institute: University of Chicago

summary: RNA-binding proteins (RBPs) are essential for posttranscriptional gene regulation, but it remains a challenge to identify RNA targets of RBPs from specific cell types. Here we present PIE-Seq to investigate Protein-RNA Interaction with dual-deaminase Editing and Sequencing by conjugating C-to-U and A-to-I RNA deaminases to RBPs. We benchmark the method and demonstrate its sensitivity in single cells, its application in the developing brain, and its scalability with 25 human RBPs involved in diverse biological functions. Bulk PIE-Seq reveals canonical binding features and target genes for RBPs such as PUM2 and NOVA1, and nominates new target genes for most tested RBPs such as SRSF1 and TDP-43/TARDBP. Integrative analyses show that homologous RBPs frequently bind to comparable sequences and target gene sets while different RBP families bind to distinct targets, indicating that PIE-Seq is robust to uncovering RBP targets. Single-cell PIE-PUM2 uncovers comparable targets to bulk samples and applying PIE-PUM2 to the developing mouse neocortex identifies neural-progenitor- and neuron-specific target genes. In summary, the dual-deaminase PIE-Seq joins the editable nucleotide space for single editors and provides an orthogonal approach and resource to uncover RBP targets in mice and human cells.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: HEK293FT cells or developing dorsal brain cells were transfected with different plasmids for 48 or 24 hours, transfected cells were sorted by FACS, and then total RNA was extracted from each group for RNA-Sequencing.

first: Xiaochang

last: Zhang


title: Increased G3BP2-Tau interaction in Tauopathies is a natural defense against Tau aggregation

city: Basel

first: Marco

last: Terrigno

address: Grenzacherstrasse 124

institute: F. Hoffmann-La Roche AG

summary: Many RNA-binding proteins (RBPs), particularly those associated with RNA granules, promote pathological protein aggregation in neurodegenerative diseases. Here, we demonstrate that G3BP2, a core component of stress granules, directly interacts with Tau and inhibits Tau aggregation. In the human brain, the interaction of G3BP2 and Tau is dramatically increased in multiple Tauopathies and precedes neurofibrillary tangle (NFT) formation in Alzheimer’s disease (AD). Surprisingly, Tau pathology is significantly elevated upon loss of G3BP2 in human neurons and brain organoids. Moreover, we find that G3BP2 masks the microtubule-binding region (MTBR) of Tau, thereby inhibiting Tau aggregation. Our study defines a novel role for RBPs as a line of defense against Tau aggregation in Tauopathies.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for hiPSC-derived glutamatergic neurons  (FujiFilm Cellular Dynamics; Cat.R1034) after treatment with G3BP2 siRNA (Horizon, Cat. E-015329-00) or NT siRNA (Horizon, Cat. D-001910-10).

first: Congwei

last: Wang

first: Juan

last: Li

first: Tania

last: Distler

first: Nikhil

last: Pandya

first: Martin

last: Ebeling

first: Stefka

last: Tyanova

first: Jeroen

last: Hoozemans

first: Anke

last: Dijkstra

first: Luisa

last: Fuchs

first: Shengqi

last: Xiang

first: Azad

last: Bonni

first: Fiona

last: Grüninger

first: Ravi

last: Jagasia


title: Sex-Specific and Opposed Effects of FKBP51 in Glutamatergic and GABAergic Neurons: Implications for Stress Susceptibility and Resilience

status: Public on May 16 2023

city: Munich

first: Mathias

middle: V.

last: Schmidt

state: Bayern

address: Kraepelinstrasse 2-10

institute: Max Planck Institute of Psychiatry

summary: Mental health disorders often arise as a combination of environmental and genetic factors. The FKBP5 gene, encoding the GR co-chaperone FKBP51, has been uncovered as a key genetic risk factor for stress related illness. However, the exact cell type and region-specific mechanisms by which FKBP51 contributes to stress resilience or susceptibility processes remain to be unravelled. FKBP51 functionality is known to interact with the environmental risk factors age and sex, but so far data on behavioral, structural and molecular consequences of these interactions are still largely unknown. Here we report the cell type- and sex-specific contribution of FKBP51 to stress susceptibility and resilience mechanisms under the high-risk environmental conditions of an older age, by using two conditional knockout models within glutamatergic (Fkbp5Nex) and GABAergic (Fkbp5Dlx) neurons of the forebrain. Specific manipulation of Fkbp5 in these two cell types led to opposing effects on behavior, brain structure and gene expression profiles in a highly sex-dependent fashion. The results emphasize the role of FKBP51 as a key player in stress-related illness and the need for more targeted and sex-specific treatment strategies.

overall_design: 5-6 mice per condition (2 mouselines; 2 brain regions; 2 sexes; 2 genotypes (wildtype or knockout))

first: Lotte

last: van Doeselaar

first: Tibor

last: Stark

first: Shiladitya

last: Mitra

first: Huanqing

last: Yang

first: Joeri

last: Bordes

first: Linda

last: Stolwijk

first: Clara

last: Engelhardt

first: Veronika

last: Kovarova

first: Sowmya

last: Narayan

first: Lea

last: Brix

first: Margherita

last: Springer

first: Jan

last: Deussing

last: Lopez

first: Michael

last: Czisch


title: Twist1 regulates stem cell properties in mesenchymal stem cells

city: Osaka-Sayama

first: Toshiyuki

last: Takehara

state: Osaka

address: 377-2, Ohno-Higashi

department: Faculty of Medicine

institute: Kindai Universty Hospital

summary: To investigate the function TWIST1 in the regulation of stemness in mesenchymal stem cell, we established human mesenchymal stem cell in TWIST1-overexpression.

overall_design: Comparative gene expression profile analysis of microarray data for human mesenchymal stem cell and its TWIST1 overexpression cell line.

first: Kensuke

last: Toriumi

first: Tatsufumi

last: Mori

first: Joe

last: Hasei

first: Takeshi

last: Teramura


title: Isw1 contributes to cohesin association with centromeres and pericentromeric regions

city: Wrocław

first: Ireneusz

middle: Bernard

last: Litwin

address: Kanonia 6/8

department: Academic Excellence Incubator - Research Centre for DNA Repair and Replication

country: Poland

institute: University of Wroclaw

summary: Cohesin is a highly conserved, multiprotein complex whose canonical function is to hold sister chromatids together to ensure accurate chromosome segregation. Cohesin association with chromatin relies on the separate Scc2-Scc4 complex that enables cohesin ring opening and topological entrapment of sister DNAs. Cohesin loading at centromeres also requires the CTF19 kinetochore subcomplex that recruits Scc2-Scc4, ensuring efficient and targeted cohesin binding. After loading at centromeres, cohesin rapidly translocates to neighboring pericentromeric regions. In an attempt to better understand how sister chromatid cohesion is regulated, we performed a proteomic screen in budding yeast that identified the Isw1 chromatin remodeler as a cohesin binding partner. We found that while cohesin can associate with both Isw1-containing complexes, ISW1a and ISW1b, however only ISW1a localizes to centromeres. There, ISW1a utilizes its chromatin remodeling activity to allow efficient translocation of cohesin loaded through CTF19 at centromeres to pericentromeres. Consistently, lack of Isw1 results in cohesin and cohesin loader retainment at CENs and periCENs. Interestingly, lack of Isw1 resulted in changes in the nucleosomal landscape at the CENs and pericentromeric regions, suggesting that chromatin organization may be a crucial determinant of cohesin loading. Finally, we provide evidence that the role of ISW1a at the CENs is to restrict the activity of the RSC complex. Taken together, our results support the notion of a key role of chromatin environment for cohesin functions.

sample_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: Calibrated Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for Scc1-9Pk and Rad21-6HA in Saccharomyces cerevisiae and Schizosaccharomyces pombe cells, respectively.

first: Małgorzata

last: Nowicka

first: Ewa

last: Maciaszczyk-Dziubińska

first: Paulina

last: Tomaszewska

first: Katarzyna

last: Markowska

first: Robert

last: Wysocki

first: Karol

last: Kramarz


title: Chimeric TIM-4 receptor-modified T cells targeting phosphatidylserine mediates both cytotoxic anti-tumor responses and phagocytic uptake of tumor-associated antigen for T cell cross presentation

city: South San Francisco

first: Brandon

last: Cieniewicz

address: 201 Haskins Way, Suite 230

institute: CERo Therapeutics

summary: To investigate the gene expression induced in CER-1236 CER T cells after binding to phosphatidylserine

overall_design: Comparative gene expression profiling analysis of RNA-seq data for CER-1236 and control CER-1251 T cells stimulated on plate-bound phosphatidylserine

first: Daniel

last: Corey


title: Transcriptomic Profiling of human granulosa cells between women with advanced maternal age with different ovarian reserves

city: jinan

first: zhicheng

last: jia

state: shandong

address: Wenhua East Road, Lixia District, Jinan City

department: The First Clinical College

institute: Shandong University of Traditional Chinese Medicine

summary: Age-related diminished ovarian reserve (DOR) is not absolute. Some advanced maternal age (AMA) still have normal ovarian reserve (NOR) and often show better pregnancy outcomes. Exploring the transcriptomic profile of AMA could lead to new ideas for mitigating age-related diminished ovarian reserve.

overall_design: the transcriptomic profile of AMA with different ovarian reserves and provide a new perspective for the age-related diminished ovarian reserve.

first: Zhi-cheng

last: Jia

first: Ying

last: Guo

first: Yong-qian

last: Li

first: zhengao

last: sun


title: Genetic inactivation of essential HSF1 reveals an isolated transcriptional stress response selectively induced by protein misfolding

city: Stockholm

first: Michela

last: Ciccarelli

state: Sweden

address: MBW, Hus F Plan 4 Frescativägen 8 Stockholm, Stockholms län [SE-01]

department: Department of Molecular Biosciences, The Wenner-Gren Institute

country: Sweden

institute: University of Stockholm

summary:
Heat Shock Factor 1 (Hsf1) in yeast drives the basal transcription of key proteostasis factors and its activity is induced as part of the core heat shock response. In this study we stringently test the role of Hsf1 under basal and stress conditions using a newly constructed hsf1∆ strain.
To assess how cells mount transcriptional stress responses when Hsf1 is inactivated, we performed mRNA-sequencing (mRNA-seq) upon hear shock (HS) or treatment with azetidine-2-carboxylic acid (AzC).

overall_design: Comparative gene expression profiling analysis of mRNA-seq data for the hsf1∆, WT, HSF1 and msn2/4∆strains. The experimet has been performed in triplicate. The WT strain was used as a control

first: Anna

last: Masser

first: Jayasankar

last: Mohanakrishnan Kaimal

first: Jordi

last: Planells

first: Claes

last: Andréasson


title: Cohesin maintains replication timing to suppress DNA damage on cancer genes

city: Beijing

first: Jinchun

last: Wu

overall_design: Refer to individual Series


title: Enhancing human iPSC-derived retinal organoids by incorporating microglia-like cells

city: Newcastle Upon Tyne

first: Majlinda

last: Lako

address: International Centre for Life, Central Parkway

department: Institute of Genetic Medicine and Institute for Ageing

institute: Newcastle University

summary: Microglia are the primary resident immune cells in the retina. They regulate neuronal survival and synaptic pruning, which makes them essential for normal development. Following injury, they mediate adaptive responses and under pathological conditions can trigger neurodegeneration exacerbating the effect of the disease. Retinal organoids derived from human induced pluripotent stem cells (hiPSCs) are increasingly used for a range of applications, including disease modelling, development of new therapies and in the study of retinogenesis. Despite their close resemblance to the in vivo retina, they lack some key physiological features including immune cells. To enhance the retinal organoid model, we engineered an hiPSC co-culture system containing the hiPSC-derived retinal organoids and hiPSC-derived microglia-like (iMG) cells and tested their retinal invasion capacity and function. We incorporated iMG into retinal organoids at 13 weeks and tested their effect on function and development at 15 and 21.5 weeks of differentiation. Our key findings showed that iMG cells were able to respond to endotoxin challenge in monocultures and when co-cultured with the organoids. Single cell RNA-Seq transcriptomic analyses, protein expression, electron microscopy imaging and electrophysiological recordings showed that retinal organoids developed normally, expressed key markers, and retained their ability to generate spiking activity in response to light. Thus, this new co-culture immunocompetent in vitro retinal model provides a platform with greater relevance to the in vivo human retina.

overall_design: We engineered an hiPSC co-culture system containing the hiPSC-derived retinal organoids and hiPSC-derived microglia-like (iMG) cells and tested their retinal invasion capacity and function.

first: Valeria

last: Chichagova

first: Maria

last: Georgiou

first: Madeleine

last: Carter

first: Birthe

last: Dorgau

first: Gerrit

last: Hilgen

first: Joseph

last: Collin

first: Rachel

last: Queen

first: Git

last: Chung

first: Jila

last: Ajeian

first: Marina

last: Moya-Molina

first: Philip

last: Hewitt

first: Michael

last: Schmitt

first: Evelyn

last: Sernagor

first: Lyle

last: Armstrong

first: Francois

last: Poignon

first: Stephan

last: Kustermann


title: HDAC2-SP1 orchestrated M2-like macrophage phenotype drives lung cancer growth – novel intervention strategy

city: Bad Nauheim

first: Carsten

last: Kuenne

address: Ludwigstrasse 43

department: Bioinformatics

institute: Max Planck Institute for Heart and Lung Research

summary: Tumor-associated macrophages (TAMs), including anti-tumor M1-like TAMs and pro-tumor M2-like TAMs, are transcriptionally dynamic innate immune cells with diverse roles in lung cancer development. Epigenetic regulators are key controllers of macrophage fate in the spatially heterogeneous tumor microenvironment. Here, we demonstrate that the spatial proximity of histone deacetylase 2 (HDAC2) – overexpressing M2-like macrophages to tumor cells significantly correlates with poor overall survival of lung cancer patients. Suppression of HDAC2 in TAMs altered macrophage polarization, migration, and associated signaling pathways related to interleukin, chemokine, cytokine, and T cell activation. In various co-culture systems of TAMs and cancer cells, suppressing HDAC2 in TAMs results in reduced proliferation and migration and increased apoptosis of cancer cell lines and primary lung cancer cells, as well as attenuated endothelial cell tube length. HDAC2 regulates the pro-tumor macrophage phenotype via acetylation of histone H3 and transcription factor SP1. Myeloidcell–specificc deletion of Hdac2 (Hdac2f/fLysmCre) and pharmacological inhibition of class I HDACs in four different murine lung cancer models (intravenous, intratracheal, tumor relapse, and KrasLA2-driven) induced M2-like to M1-like macrophage phenotypic switching, altered infiltration of CD4 and CD8 T cells and retarded tumor growth and tumor microenvironmental angiogenesis. ThuTAM-specific HDAC2 expression may provide a novel strategy for lung cancer stratification and therapy.

overall_design: To understand effect of HDAC2 inactivation in macrophages from tumor-bearing mice, LLC1 cells were intravenously injected in Hdac2fl/fl and Hdac2fl/fLysMCre mice, followed by harvesting of bone marrow at day 20 to generate bone marrow derived macrophages (BMDMs)

first: Sarode

last: Kumar

first: Stefan

last: Günther


title: USP28 controls SREBP2 and the mevalonate pathway to drive tumour growth in squamous cancer

city: Heidelberg

first: Felix

middle: Christian Eduard

last: Vogel

address: Im Neuenheimer Feld 280

department: Division of Tumor Metabolism and Microenvironment

institute: German Cancer Research Center

summary: SREBP2 controls the expression of enzymes involved in the mevalonate pathway (MVP), a biosynthetic process that drives the synthesis of dolichol, heme A, ubiquinone and cholesterol but also provides substrates for protein prenylation, and that is frequently deregulated in cancer. We show here that SREBP2 is a novel substrate for the deubiquitinating enzyme USP28 and that USP28 regulates SREBP2 stability independent of FBXW7. Inhibition of USP28 reduces MVP activity and renders cancer cells highly sensitive to MVP inhibition by statins. Moreover, statin sensitivity of USP28 depleted cells was rescued by the addition of geranyl-geranyl-pyrophosphate, a substrate for protein prenylation. We also provide evidence that SREBP2 participates in the regulation of gene expression signature associated with squamous cell carcinoma (SCC) and that SREBP2 and enzymes of the MVP are overexpressed in SCC from different tissue origins. Finally, deletion of SREBP2 attenuated tumour growth in a mouse model of lung cancer. Our findings suggest that SREBP2 is a novel substrate for USP28 and that combinatorial targeting of the MVP together with USP28 inhibition could be a therapeutic strategy for the treatment of squamous cell carcinomas.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for A431 cells and its KD derivatives (shUSP28_1, shUSP28_2, shSREBP2_1, shSREBP2_2, and shREN [control]).

first: Carina

last: Maier

middle: C

first: Almut

last: Schulze


title: Gene expression profile at single cell level of stromal cells from fresh and cryopreserved tonsils and lymph node

city: Philadelphia

first: Joshua

middle: Daniel

last: Brandstadter

address: BRB II/III, Room 446, 421 Curie Blvd

institute: University of Pennsylvania

summary: Non-hematopoietic lymph node stromal cells (LNSCs) regulate lymphocyte trafficking, survival, and function for key roles in host defense, autoimmunity, alloimmunity, and lymphoproliferative disorders. However, study of LNSCs in human diseases is complicated by a dependence on viable lymphoid tissues, which are most often excised prior to establishment of a specific diagnosis. Here, we demonstrate that cryopreservation can be used to bank lymphoid tissue for the study of LNSCs in human disease. Using human tonsils, lymphoid tissue fragments were cryopreserved for subsequent enzymatic digestion and recovery of viable non-hematopoietic cells. Flow cytometry and single-cell transcriptomics identified comparable proportions of LNSC cell types in fresh and cryopreserved tissue. Moreover, cryopreservation had little effect on transcriptional profiles, which showed significant overlap between tonsils and lymph nodes. The presence and spatial distribution of transcriptionally defined cell types was confirmed by in situ analyses. Our broadly applicable approach promises to greatly enable research into the roles of LNSC in human disease.

overall_design: Tonsils and lymph nodes were processed fresh or cryopreserved as whole tissue before thawing for enzymatic digestion and fluorescence-activated cell sorting for CD45-EpCAM- cells and analyzed using scRNAseq.

first: Angelina

last: De Martin

first: Mechthild

last: Lϋtge

first: Antonio

last: Ferreira

first: Brain

middle: T

last: Gaudette

first: Yves

last: Stanossek

first: Shumei

last: Wang

last: Gonzalez

first: Edward

last: Camiolo

first: Gerald

last: Wertheim

first: Bridget

last: Austin

first: David

last: Allman

first: Adam

last: Bagg

first: Megan

middle: S

last: Lim

last: Fajgenbaum

first: Jon

last: Aster

first: Burkhard

last: Ludewig

first: Ivan

last: Maillard


title: A novel TetR-like transcriptional regulator in Stenotrophomonas maltophilia is involved in fatty acid metabolism, including quorum sensing signal turnover

city: Bellaterra

first: Isidre

last: Gibert

state: Barcelona

address: Carrer de la Vinya, s/n

department: Institut de Biotecnologia i de Biomedicina (IBB)

country: Spain

institute: Universitat Autònoma de Barcelona (UAB)

summary: Stenotrophomonas maltophilia is an emerging opportunistic multidrug-resistant pathogen frequently co-isolated with other relevant nosocomial pathogens in respiratory tract infections. S. maltophilia uses the endogenous DSF quorum sensing (QS) system to regulate virulence processes but can also respond to exogenous AHL signals produced by neighboring bacteria. A whole-transcriptome sequencing analysis was performed for S. maltophilia strain K279a in the exponential and stationary phases and in exponential cultures after a treatment with exogenous DSF or AHLs. Among the common top upregulated genes, the putative TetR-like regulator Smlt2053 was selected for functional characterization. This regulator was found to sense long-chain fatty acids, including the QS signal DSF, and activate a β-oxidation catabolic pathway.

overall_design: In total six samples were analyzed (2 different conditions with three independent samples), and all cultures were done in LB at 37C and samples collected at an OD550 of 1.5. S. maltophilia K279aΔsmlt2053 mutant was compared to its wild type variant.

first: Xavier

last: Coves

first: Pol

last: Huedo

first: Marc

last: Bravo Bravo

first: Òscar

last: Conchillo-Solé

first: Andromeda-Celeste

last: Gomez

first: Anna

last: Esteve-Codina

last: Dabad

last: Daura

first: Daniel

last: Yero


title: PD-1 blockade potentiates neoadjuvant chemotherapy in NSCLC via increasing CD127+ and KLRG1+ CD8 T cells

city: Tianjin

first: Xiubao

last: Ren

address: West Huan-Hu Rd, Ti Yuan Bei, Hexi District

institute: Tianjin Medical University Cancer Institute & Hospital,

summary: The combination of PD-1 blockade with neoadjuvant chemotherapy (NAC) has achieved unprecedented clinical success in non-small-cell lung cancer (NSCLC) compared to NAC alone, but the underlying mechanisms by which PD-1 blockade augments the effects of chemotherapy remain incompletely elucidated. Single-cell RNA sequencing was performed on CD45+ immune cells isolated from surgically resected fresh tumor tissues of seven NSCLC patients receiving NAC or neoadjuvant pembrolizumab and chemotherapy (NAPC). Multiplex fluorescent immunohistochemistry was performed on paired FFPE tissues before and after NAC or NAPC from 65 resectable NSCLC patients, and results were validated with GEO dataset. NAC resulted in an increase only of CD20+ B cells, whereas NAPC increased the infiltration of CD20+ B cells, CD4+ T cells, CD4+CD127+ T cells, CD8+ T cells, CD8+CD127+ and CD8+KLRG1+ T cells. Synergistic increase in B and T cells promotes favorable therapeutic response after NAPC. Spatial distribution analysis discovered that CD8+ T cells and their CD127+ and KLRG1+ subsets were in closer proximity to CD4+ T/CD20+ B cells in NAPC versus NAC. GEO dataset validated that B-cell, CD4, memory and effector CD8 signatures correlated with therapeutic responses and clinical outcomes. The adding of PD-1 blockade to NAC promoted anti-tumor immunity through T and B cells recruitment in the tumor microenvironment and induced tumor-infiltrating CD8+ T cells skewed toward CD127+ and KLRG1+ phenotypes，which may be assisted by CD4+ T cells and B cells. Our comprehensive study identified key immune cell subsets exerting anti-tumor responses during PD-1 blockade therapy and that may be therapeutically targeted to improve upon existing immunotherapies for NSCLC.

overall_design: Xiubao,Ren

first: Zhenzhen

last: Hui

first: Yulin

first: Dong

last: Zhang

first: Yulong

last: Chen

first: Wenwen

last: Yu

first: Jie

last: Cao

first: Liang

last: Liu

first: Tao

last: Wang

first: Shanshan

last: Xiao

first: Liuqing

last: Zheng

first: Yue

last: Pu

first: Feng

last: Wei

first: Jian

last: You


title: RNA-sequencing for dynamic epi-transcriptomic landscape mapping with disease progression in patient primary breast tumours.

city: Dublin

first: Leonie

last: Young

state: Dublin

address: 31A York Street

department: Surgery

country: Ireland

institute: RCSI

summary: Advanced breast cancer is characterised by enhanced tumour adaptability to therapeutic pressure and the metastatic microenvironment. Transcriptome differences in 14 primary breast tumour samples (n = 14 samples) are uncovered through this comparative RNA-seq analysis of patients that responded well to therapy (n = 7) and patients who had disease recurrence on endocrine treatment (n = 7). RNA sequencing data is deposited as log2 transformed median-of-ratios (DESeq2) normalised counts gene expression values (44934 genes IDs; n = 14 tumour samples).

overall_design:
RNA sequencing was carried out on patient primary breast tumour samples for patients that responded well to therapy (n = 7) and patients who had disease recurrence on endocrine treatment (n = 7).
>>> Raw RNA sequencing data for the paired primary and metastatic samples are not published openly in order to protect participant identities, but will be made available upon request and under regulatory compliance via a data usage agreement (DUA). Please contact the corresponding author with data access requests that will be granted once the DUA is signed. <<<

first: Mihaela

last: Ola

first: Sinéad

last: Cocchiglia

first: Stephen

last: Keelan

first: Sara

last: Charmsaz

first: Daniela

last: Ottaviani

first: Seán

last: Hickey


title: Deregulated long non-coding transcripts in M1 and M2 macrophages

first: Carsten

last: Kuenne

type: Non-coding RNA profiling by high throughput sequencing

summary: Non-coding transcripts make up around 98 % of RNAs in the transcriptome of cells, while only around 2 % are coding for proteins. Long non-coding RNAs (> 200bp) are known to have high regulatory functions in various cellular processes. Here, we aimed to characterize the distinct macrophage phenotypes regarding their expression of long non-coding RNAs, which might be involved in the activation process.

overall_design: RNA-sequencing of PBMC derived unstimulated (M0) or in vitro polarized (M1 or M2) macrophages

first: Rajkumar

last: Savai

first: Stefan

last: Guenther

first: Annika

last: Karger


title: CLARITY study: Expression data from RRMS patients treated with Oral Cladribine versus Placebo at week 96

city: Billerica

first: Irina

last: Kalatskaya

state: MA

address: 45 Middlesex Turnpike

department: Clinical Measurement Sciences

institute: EMD Serono Research and Development Institute

summary:
The CLARITY trial (NCT00213135) was designed as a double-blind, placebo-controlled study to allow the best comparison of absolute efficacy and safety of oral cladribine in RRMS subjects. 1326 patients with relapsing MS were randomized (1:1:1) to receive cladribine tablets 3.5 mg/kg or 5.25 mg/kg bodyweight or placebo. Gene expression data in whole blood samples at 96 weeks were prepared according to standard Affymetrix protocols
Gene expression data in whole blood samples at 96 weeks were available from patients randomized to placebo (n=57), cladribine tablets 3.5 mg/kg (n=62), and cladribine tablets 5.25 mg/kg (n=70).

overall_design: Transcriptome of the whole blood from RRMS patients either treated with high (5.25 mg/kg) or low dose (3.5 mg/kg) of cladribine or from placebo arm at 96 weeks after start day.

first: Ursula

last: Boschert


title: RNA-sequencing for dynamic epi-transcriptomic landscape mapping with disease progression in ER-positive breast cancer

summary: Advanced breast cancer is characterised by enhanced tumour adaptability to therapeutic pressure and the metastatic microenvironment. Transcriptome differences in three ER positive (ER+) cell models are uncovered through this RNA-seq analysis of MCF7 (endocrine sensitive), LY2 (endocrine resistant) and T347 (derived from an ER-positive, treatment resistant brain metastatic patient tumour) cells.

overall_design: We report the RNA-sequencing of three ER positive (ER+) cell models: MCF7, LY2 and T347 cells, and examine the gene expression differences.

first: Sinead


title: Circulating small RNA profiling of patients with alveolar and cystic echinococcosis

status: Public on May 15 2023

city: Buenos Aires

first: Marcela

middle: Alejandra

last: Cucher

address: Paraguay 2155, Piso 13

country: Argentina

institute: IMPaM, UBA-CONICET

summary: Alveolar (AE) and cystic (CE) echinococcosis are two parasitic diseases caused by the tapeworms Echinococcus multilocularis and E. granulosus sensu lato (s. l.), respectively. Currently, AE and CE are mainly diagnosed by means of imaging techniques, supported by serology and clinical and epidemiological data. However, no viability markers that indicate parasite state during infection are available. Extracellular small RNAs (sRNAs) are short non-coding RNAs that can be secreted by cells through association with extracellular vesicles, proteins, or lipoproteins. Circulating sRNAs can show altered expression in pathological states; hence, they are intensively studied as biomarkers of several diseases. Here, we profiled the sRNA transcriptomes in the context of AE and CE to identify novel biomarkers to aid in medical decisions. For this, endogenous and parasitic sRNAs were analysed by sRNA sequencing in sera from disease negative, positive, and treated patients and patients harbouring a non-parasitic lesion. Consequently, 20 differentially expressed sRNAs associated with AE, CE and/or non-parasitic liver lesion were identified. Our results represent an in-depth characterisation of the effect E. multilocularis and E. granulosus s.l. exert on the extracellular sRNA landscape in human infections and provide a set of novel candidate biomarkers for both AE and CE detection.

overall_design: small RNA sequencing from serum samples of AE and CE patients was performed. Patients with hepatic non-parasitic lesions (NPL) were also included. The AE set is composed of  Negative,  Positive and Treated patients. The CE set is composed of CE negative patients with no cysts (Negative), CE negative patients with non-parasitic lesion (NPL), CE positive with active cyst (CE1-2), CE positive with transitional cyst (CE3a and CE3b), CE positive with inactive cyst (CE4, CE5) ) that reached inactivation spontaneously and CE positive with albendazole treatment (Treated).

first: Mara

last: Mariconti

first: Tommaso

last: Manciulli

first: Ambra

last: Vola

last: Rosenzvit

first: Klaus

last: Brehm

first: Laura

last: Kamenetzky

first: Enrico

last: Brunetti


title: Lin28 is essential for the maintenance of chicken primordial germ cells

city: Fukuoka

first: Seung June

last: Kwon

state: Fukuoka

address: 744 Motooka, Nishi-ku

department: Graduate School of Systems Life Sciences

institute: Kyushu University

summary:
To investigate the function of Lin28A and B in primordial germ cell (PGC) , we generated chicken PGCs in which Lin28A or B has been knocked out by CRISPR/Cas9.
We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different cell types.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for (1) control, (2) Lin28A knockout, (3) Lin28B knockout chicken PGCs.

first: Seung

first: Katsuya

last: Suzuki

first: Yuji

last: Atsuta


title: Maladaptive TGF-β Signals to the Alveolar Epithelium Drive Fibrosis after COVID-19 Infection

city: los angeles

first: Changfu

last: Yao

state: Ca

address: 8700 Beverly Blvd. AHSP Room A9401

department: Lung and Regenerative Medicine Institutes

institute: cedars-sinai medical center

summary: We have performed single cell RNA sequencing on explant tissue of post-acute sequelae SARS-CoV-2 infection  (PASC) patients. The study focused on epithelial cells (Epcam+) and immune cells (CD45+) cells. We further compared our PASC sample with Donor and IPF samples from GSE146981 and GSE135893.

overall_design: We isolated single cells from PASC explant samples, further FACS sorted CD45+ cells and EpCAM+ (CD326) cells for single cell RNA sequencing using 10x Genomic 3 prime Next Gem kit.

first: Peter

last: Chen

first: Barry

last: Stripp

first: Cory

last: Hogaboam

first: Milena

last: Espindola


title: Lymph-Node-Based CD3+ CD20+ Cells Emerge from Membrane Exchange between T Follicular Helper Cells and B Cells and Increase Their Frequency following Simian Immunodeficiency Virus Infection

city: Atlanta

first: Gregory

middle: K

last: Tharp

state: GA

address: 954 Gatewood Dr

department: Developmental and Cognitive Neuroscience

institute: Yerkes National Primate Research Center

summary: CD4+ T follicular helper (TFH) cells are key targets for human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) replication and contribute to the virus reservoir under antiretroviral therapy (ART). Here, we describe a novel CD3+ CD20+ double-positive (DP) lymphocyte subset, resident in secondary lymphoid organs of humans and rhesus macaques (RMs), that appear predominantly after membrane exchange between TFH and B cells. DP lymphocytes are enriched in cells displaying a TFH phenotype (CD4+ PD1hi CXCR5hi), function (interleukin 21 positive [IL-21+]), and gene expression profile. Importantly, expression of CD40L upon brief in vitro mitogen stimulation identifies, by specific gene- expression signatures, DP cells of TFH-cell origin versus those of B-cell origin. Analysis of 56 RMs showed that DP cells (i) significantly increase following SIV infection, (ii) are reduced after 12 months of ART in comparison to pre-ART levels, and (iii) expand to a significantly higher frequency following ART interruption. Quantification of total SIV-gag DNA on sorted DP cells from chronically infected RMs showed that these cells are susceptible to SIV infection. These data reinforce earlier observations that CD20+ T cells are infected and expanded by HIV infection, while suggesting that these cells phenotypically overlap activated CD4+ TFH cells that acquire CD20 expression via trogocytosis and can be targeted as part of therapeutic strategies aimed at HIV remission.

overall_design: LN cells from 9 Rhesus Macaques were FACS-sorted and gene expresson profiles were examined.

first: Sadia

last: Samer

first: Ankita

last: Chowdhury

first: Tomas

last: Wiche Salinas

first: Perla

middle: M

last: Del Rio Estrada

first: Morgan

last: Reuter

first: Steven

last: Bosinger

first: Barbara

last: Cervasi

first: James

last: Auger

first: Kiran

last: Gill

first: Yuria

last: Ablanedo-Terrazas

first: Gustavo

last: Reyes-Teran

first: Jacob

last: Estes

first: Michael

last: Betts

first: Guido

last: Silvestri

first: Mirko

last: Paiardini


title: Transcriptional changes in 6 month post-fertilization zebrafish bbs2 -/- mutant retinas versus wild-type siblings during ongoing photoreceptor degeneration.

city: Cleveland

first: Brian

last: Perkins

address: 9500 Euclid Ave

institute: Cleveland Clinic

summary: Bulk RNA-seq transcriptional profiling was performed on 6 month post-fertilization bbs2 -/- mutant zebrafish retinas and bbs2 +/+ wild-type siblings to investigate the transcriptional changes that occur during progressive photoreceptor degeneration.

overall_design: Total RNA from the retinas of 4 bbs2 -/- homozygote mutant and 4 bbs2 +/+ wild-type siblings was used to prepare libraries for RNA-seq and pairwise comparisons of gene expression.

first: Sarah

middle: E

last: Grabinski

first: Adrian

last: Reich


title: Dynamic epi-transcriptomic landscape mapping with disease progression in ER-positive breast cancer

first: Leonie

last: Young

type: Methylation profiling by high throughput sequencing

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


title: MeRIP-sequencing for dynamic epi-transcriptomic landscape mapping with disease progression in ER-positive breast cancer

summary: Advanced breast cancer is characterised by enhanced tumour adaptability to therapeutic pressure and the metastatic microenvironment. Targeting epi-transcriptomic modulators may reverse cellular adaptability, offering new therapeutic strategies to treat metastatic disease. This study looks into the dynamic adaptations that occur in cancer cells in response to therapeutic pressure and metastatic evolution by profiling mRNA epi-transcriptomic modifications in models of disease progression. ER positive (ER+) cell models were used to represent progressive breast cancer, MCF7 (endocrine sensitive), LY2 (endocrine resistant) and T347 (derived from an ER-positive, treatment resistant brain metastatic patient tumour) cells. MeRIP sequencing was undertaken to determine genome-wide RNA methylated regions.

overall_design: We report the MeRIP-sequencing of three ER positive (ER+) cell models: MCF7, LY2 and T347 cells, and examine the epi-transcriptomic differences.

first: Stephen

last: Keelan

first: Mihaela

last: Ola

first: Sara

last: Charmsaz

first: Sinead

last: Cocchiglia


title: Lipid nanoparticle delivery promotes cell-type-specific distribution of antisense oligonucleotides in the brain

city: South San Francisco

first: Benjamin

middle: I

last: Laufer

address: 1 DNA Way

institute: Genentech

summary: Antisense oligonucleotides (ASOs) are being actively investigated as potential therapeutics for a broad range of neurodegenerative diseases. While these small oligonucleotides have been effective in the clinic, many basic questions regarding ASO internalization, trafficking, and modes of enhancing delivery remain. To address these questions, we investigated how lipid nanoparticle (LNP) delivery affects ASO uptake and distribution in the brain. We show that ASOs are internalized and active in central nervous system (CNS) cell types both in vivo and in vitro. While differential cellular activity and polarization states do not affect ASO potency, encapsulating ASOs in LNPs increases ASO activity up to 100-fold in cultured CNS cells. This dramatic increase in efficacy is facilitated by the intracellular trafficking of ASOs away from lysosomes or enhanced ASO uptake, which is cell-type-specific. We assessed the translatability of these results by screening ASO-LNPs in vitro and intracerebroventricularly injecting top performing formulations in mice. ASO-LNP delivery induced a strong ASO-dependent microgliosis response in the brain revealing that LNP encapsulation cannot mask ASO-mediated toxicity. However, LNP-delivered ASOs did not downregulate mRNA levels in bulk tissue due to changes in ASO distribution. While ASOs distribute widely across the brain after bulk injection, ASO-LNPs are preferentially internalized by cells lining the blood vessels and ventricles. Consistent with our findings in cells, ASOs localize to the endolysosomal system following both ASO and ASO-LNP delivery in the brain as determined using immunoelectron microscopy. These data provide valuable insights into how LNPs regulate ASO uptake and distribution in the brain, and support further development of ASO-LNPs for treating CNS disorders.

overall_design: RNA from mouse brain after a single intracerebroventricular (ICV) bolus injection of ASO, LNP-encapsulated ASO, LNP, or Saline.

first: Amy

last: Byrnes

first: Brad

middle: A

last: Friedman

first: Casper

last: Hoogenraad


title: Transcriptomic analyses of cutaneous immune network in patiens with atopic dermatitis challenged with house dust mite (HDM) patch test

city: Southampton

first: Marta

middle: Ewa

last: Polak

state: Hampshire

address: Southampton General Hospital, Mailpoint 813, LE59,

department: Clinical and Experimental Sciences

institute: University of Southampton

summary: Our study investigates the activation and transcriptional programming of primary skin cells (FACS-purified Langerhans cells and single cell analysis of the whole biopsy)  from patients with atopic dermatitis (n=22, nr = rensponding, nnr=nonresponding), exposed to a control or HDM patch test. 6 mm biopsies were taken from control and HDM patch test site 48h post application.

overall_design: Patients with moderate to severe atopic dermatitis eczema severity scores (EASI) median = 17.70, IQR:10.20 – 30.90, max = 51.40) under the care of a dermatologist in a tertiary referral centre were recruited to the study. 48h post application of a patch test to buttock skin, 11 out of the 28 patients were HDM-reactive, while In 12 patients, HDM did not induce a visible response, creating “HDM-non-reactive” group. 4 patients reacted to both HDM and control patch test "irritant", and one showed redness which was not patch test localised "undefined". 6 mm biopsies were taken from all patch test sites, and processed for flow cytometry or single cell RNA-seq analysis. CD207, CD1a, HLA-DR+ Langerhans cells (LCs)were FACS-sorted into RLT buffer and processed for bulk RNA-seq. Non-LCs cells were sorted and cryopreserved for further analysis with Constellation 10X. Freshly dissociated whole skin biopsies from 6 biopsies were suspended in RNAse-out buffer and processed on ice to the co-encapsulation of single cells with genetically-encoded beads (Drop-seq).

first: Sofia

last: Sirvent-Bernal

first: Andres

last: Vallejo


title: Emx2 KO RNAseq

city: Salt Lake City

first: Shihai

last: Jia

state: ut

address: 20 South 2030 East, Room 320 BPRB

department: Department of Neurobiology

institute: University of Utah

summary: Emx2Cre/Cre vs. Emx2WT/WT by RNAseq

overall_design: 10 utricles were dissected and pooled for each technical replicate and stored in RNAlater at 4C until genotypes were verified.  RNA extraction was completed via Qiagen RNeasy micro Kit with DNAse I column digest for 4 technical replicates of each genotype. Concentrations were validated with Qubit and quality scores using Agilent High Sensitivity RNA ScreenTape Assay prior to RNA analysis. RNA library prep was completed using Illumina TruSeq Stranded Total RNA Library Prep Ribo-Zero Gold kit. Sequencing was completed with NovaSeq2 x 150bp Sequencing 200M read-pairs per sample. Subsequent alignments identified human RNA transcript read contamination in a single replicate, which was removed prior to DESeq2 analysis to identify upregulated and downregulated transcripts.

first: Evan

last: Ratzan

first: Litty

last: Paul

first: Chris

last: Stubbens

last: Dalley