title: GLP-1 directed NMDA receptor antagonism for obesity treatment [II]

first: Lars

last: Ingerslev

overall_design: Comparative gene expression profiling analysis of RNA-seq data from hypothalamus which had been randomized to one of four treatment groups (Vehicle, GLP-1, MK-801/GLP-1, or conjugate 2). 5-6 replicates are sequenced for each tissue and treatment combination.

first: Jonas

last: Petersen

first: Mette

last: Ludwig

first: Vaida

last: Juozaityte

first: Pablo

last: Ranea-Robles

first: Charlotte

last: Svendsen

first: Eunsang

last: Hwang

first: Amalie

last: Kristensen

first: Nicole

last: Fadahunsi

first: Jens

last: Lund

first: Alberte

last: Breum

first: Cecilie

last: Mathiesen

first: Luisa

last: Sachs

first: Roger

last: Moreno-Justicia

first: Rebecca

last: Rohlfs

first: James

last: Ford

first: Jon

last: Douros

middle: E

middle: S


title: GLP-1 directed NMDA receptor antagonism for obesity treatment

status: Public on May 14 2024

city: Copenhagen

first: Lars

middle: Roed

last: Ingerslev

address: Blegdamsvej 3B

department: NNF Center for Basic Metabolic Research

country: Denmark

institute: Copenhagen University

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Exosomal miRNAs from serum sequencing

city: chengdu

first: tong

last: feng

state: -- None selected --

address: Acupuncture and Tuina College

country: China

institute: Chengdu University of TCM

type: Non-coding RNA profiling by high throughput sequencing

summary: Screening for dysregulated mirnas in serum exosomes in patients with osteoarthritis compared to healthy individuals.

overall_design: Serum was collected from 15 patients with osteoarthritis and 10 healthy individuals for serum exosome sequencing

first: Tong

last: Feng

first: Qi

last: Zhang


title: Genome wide identification of replication fork stalling/pausing sites and the interplay between RNA Pol II transcription and DNA replication progression

status: Public on May 13 2024

city: Birmingham

first: Marco

last: Saponaro

address: Vincent Drive

department: Institute of Cancer and Genomic Sciences

country: United Kingdom

institute: University of Birmingham


summary: DNA replication progression can be affected by the presence of physical barriers on the DNA, like RNA Polymerases, leading to replication stress and DNA damage. To characterize what happens at sites where DNA replication forks stall and pause, we establish a genome-wide approach to identify them. This approach uses multiple timepoints during S-phase, to identify replication fork/stalling hotspots throughout the genome. These sites are typically associated with increased DNA damage, overlap with fragile sites and with breakpoints of rearrangements identified in cancers, but do not overlap with replication origins. Overlaying these sites with a genome-wide analysis of RNA Polymerase II transcription, we found that replication fork stalling/pausing sites inside genes are directly related to transcription progression and activity. This would support data that indicate that transcription and replication can coexist over the same regions. We found instances where transcription activity by reducing histone density favors replication progression through genes, but also found that slowing down transcription elongation slows down directly replication progression through genes. Importantly, rearrangements found in cancers at transcription-replication collision sites can be detected in non-transformed cells and increased following treatment with ATM and ATR inhibitors. Altogether, our findings highlight how transcription and replication overlap during S-phase, with both positive and negative consequences for replication fork progression and genome stability.

overall_design: Genome-wide DNA replication profile of BJ-hTERT during S phase with or without indicated treatments generated by NGS

first: Patricia

last: Rojas

first: Jianming

last: Wang


title: Study of the tRNA-derived fragment expression profile in rat pancreatic islets of diabetes-susceptible low-protein (LP) exposed rats

city: Lausanne

first: Cecile

last: Jacovetti

address: Rue du Bugnon 9

department: Fundamental Neurosciences

country: Switzerland

institute: University of Lausanne

summary: In this study, we discovered cytosolic and mitochondrial fragments resulting from tRNA and mt-tRNA cleavage, which may act as new regulators of cellular and metabolic functions. The so-called LP rat model consists of 22-day-old rats whose mothers have been subjected to a low protein diet during gestation and lactation. These offspring have impaired pancreatic development with reduced ß-cell mass, glucose-stimulated insulin secretion defect and are more prone to develop diabetes in adulthood. We identified hundreds of these fragments as being changed in the pancreatic islets of LP rats and compared them to chow diet (CD) exposed rats.  In our analysis, we identified 6164 tRFs in the islets of LP rats, among which 256 exhibited significant changes (≥ 2 fold; adjusted p value ≤ 0.05) compared to controls. Of these, 113 tRFs showed increased levels, while 143 tRFs showed decreased levels in the LP rats. We focused our investigations on mitochondrial tRFs (mt-tRFs) that show significant alterations in both, adult pre-diabetic mice (db/db) and LP rats. We selected the mt-tRF-LeuTAA fragment  for further investigation, as its level is reduced in the islets of db/db mice and LP rats, while being abundant in ß-cells. mt-tRF-LeuTAA fragment is derived from the cleavage of tRNA-LeuTAA encoded by the mitochondrial genome. We demonstrated that mt-tRF-LeuTAA acts as a key regulator of mitochondrial OXPHOS functions, mitochondrial membrane potential, the insulin secretory capacity of ß-cells, and the insulin sensitivity of myotube muscle cells.

overall_design: In order to characterize the expression profile of tRFs (tRNA-derived fragments) under conditions that promote diabetes development, we isolated total RNA from pancreatic islets of four low-protein (LP) exposed rats and compared it to that of five chow diet (CD) exposed rats.

first: Chris

last: Donnelly

first: Véronique

last: Menoud

first: Mara

last: Suleiman

first: Cristina

last: Cosentino

first: Jonathan

last: Sobel

first: Keijing

last: Wu

first: Karim

last: Bouzakri

first: Piero

last: Marchetti

first: Claudiane

last: Guay

first: Bengt

last: Kayser

first: Romano

last: Regazzi


title: Effect of genomic and cellular environments on gene expression noise

status: Public on May 12 2024

city: St. Louis

first: SIQI

last: ZHAO

state: MO

address: Campus Box 8232 4515 McKinley Ave.

department: Genetics

country: USA

institute: Washington University in St. Louis

summary: Individual cells from isogenic populations often display large cell-to-cell differences in gene expression. This “noise” in expression derives from several sources, including the genomic and cellular environment in which a gene resides. Large-scale maps of genomic environments have revealed the effects of epigenetic modifications and transcription factor occupancy on mean expression levels, but leveraging such maps to explain expression noise will require new methods to assay how expression noise changes at locations across the genome.To address this gap, we present Single-cell Analysis of Reporter Gene Expression Noise and Transcriptome (SARGENT), a method that simultaneously measures the noisiness of reporter genes integrated throughout the genome and the global mRNA profiles of individual reporter-gene-containing cells. Using SARGENT, we perform the first comprehensive genome-wide survey of how genomic locations impact gene expression noise. We find that the mean and noise of expression correlate with different histone modifications. We quantify the intrinsic and extrinsic components of reporter gene noise and, using the associated mRNA profiles, assign the extrinsic component to differences between the CD24+ “stem-like” sub-state and the more “differentiated” sub-state. SARGENT also reveals the effects of transgene integrations on endogenous gene expression, which will help guide the search for “safe-harbor” loci.

overall_design: We developed a high-throughput method to test the effects of genomic environments on the mean and noise of gene expression. Our goal was to integrate a common transgene across the genome and then, for individual cells, measure both the transcripts produced from the transgene and the global mRNA profile. This allows us to compute the mean and noise of reporter gene expression at each location, and correlate reporter gene expression with the cellular mRNA state of each cell. Because every unique integration contains the same transgene, the measured differences in the mean and noise of reporter gene expression are directly attributable to the influence of genomic environments or cellular states.

first: Clarice

middle: K

last: Hong

first: Avinash

last: Ramu

first: Siqi

last: Zhao

first: Barak

middle: A

last: Cohen


title: Silencing of mt-tRF-LeuTAA fragment in pancreatic rat islets and investigation by RNA-sequencing of the downstream mechanisms activated by this fragment

first: Cecile

last: Jacovetti

summary: In this study, we discovered cytosolic and mitochondrial fragments resulting from tRNA and mt-tRNA cleavage, which may act as new regulators of cellular and metabolic functions. We selected the mt-tRF-LeuTAA fragment  for further investigation, as its level is altered in the islets diabetes-susceptible animal models, while being abundant in ß-cells. mt-tRF-LeuTAA fragment is derived from the cleavage of tRNA-LeuTAA encoded by the mitochondrial genome. We demonstrated that mt-tRF-LeuTAA acts as a key regulator of mitochondrial OXPHOS functions, mitochondrial membrane potential, the insulin secretory capacity of ß-cells, and the insulin sensitivity of myotube muscle cells. To gain a comprehensive understanding, we conducted transcriptomic and proteomic analyses on rat islets with silenced mt-tRF-LeuTAA for 72 hours. We transfected rat islet cells with an antisense oligonucleotides (antisense oligonucleotides to reduce mt-tRF-LeuTAA levels versus control oligos). These oligos specifically decreased the levels of the fragment (anti-tRF-LeuTAA) by more than 95% without affecting the host full-length tRNA mt-tRNA-LeuTAA. Inhibiting mt-tRF-LeuTAA led to significant differential expression of 4843 mRNA transcripts, cut-off adjusted P ≤ 0.05. To further investigate the cellular rearrangement associated with the inhibition of mt-tRF-LeuTAA, we conducted enrichment analysis using Gene Ontology Molecular Function terms on RNA-sequencing. At the transcriptomic level, we observed enrichment of pathways related to ion transport and various enzymatic activities, including mitochondrial oxidoreductase activity, when the mitochondrial fragment was repressed in rat islets.

overall_design: In order to characterize the gene expression profile of rat islet cells upon mt-tRF-LeuTAA inhibition, we transfected dispersed rat islet cells for 72hrs with antisense oligonucleotides to reduce mt-tRF-LeuTAA levels (anti-tRF-LeuTAAn n=6)  versus control oligos (anti-control, n=6). Afterwards we isolated total RNA and performed RNA-sequencing.


title: Immunomodulatory effects of glucocorticoid-induced leucine zipper (GILZ) expression in murine bone-marrow derived macrophages.

city: Saarbruecken

first: Thierry

middle: M.

last: Legroux

state: Saarland

address: Campus Saarbrücken

department: Pharmaceutical biology

country: Germany

institute: Saarland University

summary: Here, we provide evidence for the immunomodulatory effects of glucocorticoid-induced leucine zipper (GILZ) overexpression in macrophages, demonstrating enhanced antibacterial activity, protection from pyroptosis, and altered mitochondrial function. Additionally, our findings reveal a significant impact on MMP activity, suggesting a role for GILZ in influencing extracellular matrix remodeling and wound healing processes. The study contributes to explorations of GILZ in its significance in pro-resolving macrophage functions.

overall_design:
B6.129P2-Lyz2tm1(cre)Ifo/J mice with Cre recombinase expression under the endogenous Lyz2 promotor of the myeloid cell lineage were crossed with mice bearing loxP sites up and downstream of Gilz exon 6 resulting in a myeloid-specific knockout of GILZ (B6.129P2 Tsc22d3f/f Lyz2tm1(cre)Ifo/J, GILZ KO) (Bruscoli et al., 2012). In addition, mice bearing a Tsc22d3/Gilz‑1 cDNA knock-in under the control of the ROSA26 promoter preceded by a loxP-flanked stop cassette (Carceller et al., 2016) were crossed with B6.129P2-Lyz2tm1(cre)Ifo/J mice, resulting in myeloid-specific GILZ overexpression.
Employing RNA sequencing, we examined wild-type (WT), GILZ knockout (GILZ KO), and GILZ overexpressing (GILZ TG) bone-marrow derived macrophages (BMM) under untreated conditions and following 4 h lipopolysaccharide (LPS) treatment.
All analyses were conducted in the R programming language. DESeq2 analysis revealed differentially expressed genes (DEGs, p < 0.05) in the comparisons of GILZ KO vs. WT, GILZ TG vs. WT, and GILZ KO vs. GILZ TG under both untreated and LPS-treated conditions.

middle: M

first: Hanna

last: Schymik

first: Gilles

last: Gasparoni

first: Claude

last: Libert

first: Jörn

last: Walter

first: Britta

last: Diesel

first: Jessica

last: Hoppstädter

first: Alexandra

last: Kiemer


title: Vaccinia virus infection induces concurrent alterations in host chromatin architecture, accessibility, and gene expression [RNA-seq]

status: Public on May 11 2024

city: Los Alamos

first: Christina

middle: Rene

last: Steadman

state: New Mexico

address: Bikini Atoll Rd

department: Earth & Environmental Sciences Division

institute: Los Alamos National Laboratory

summary: Genomic DNA folds into complex configurations that produce particular local and globalstructures thought to profoundly impact genome function. To understand the dynamic nature ofthis relationship, we investigated the extent of host chromatin structural and functional changesin response to a viral agent. We performed comprehensive assessments of host architecture (Hi-C), accessibility (ATAC-seq), and gene expression (RNA-seq) in a paired manner in response toattenuated vaccinia (smallpox) virus. Over time, infection significantly increased long-rangeintra-chromosomal interactions and decreased chromatin accessibility. Fine-scale accessibilitychanges were independent of broad-scale chromatin compartment exchange, which increased (upto 12% of the genome) over time, underscoring potential independent mechanisms for global andlocal chromatin reorganization. The majority of differentially expressed genes, including thosedownregulated in immune responses, had concurrent alterations in local accessibility and loopdomain restructuring. Increased B compartmentalization, intra-chromosomal interactions, anddecreased inter-chromosomal interactions and chromatin accessibility together indicate thatinfection converts the host genome into a more condensed state with nearly equal bidirectionaldifferential gene expression. These changes in host chromatin features may have implications fordeveloping efficacious anti-viral countermeasures. Overall, our empirical data provides evidenceof orchestrated concurrent alterations in chromatin architecture, accessibility, and geneexpression in response to infection, further reinforcing the notion of coordinated structurefunctiondynamics of the genome.

overall_design: To investigate concurrent alterations in host chromatin accessibility and 3D organization as viralinfection progresses, we infected a susceptible cell line (Vero), derived from African greenmonkey, with Modified Vaccinia virus Ankara strain (MVA). We confirmedsuccessful infection and its gradual progression at 12-, 18-, and 24-hours post infection. At each time point, cells were collected from mock-infected (control) and MVAinfectedcell culture flasks for paired measurement of 3D genomic organization, chromatinaccessibility, and gene expression using Hi-C, ATAC, and RNA sequencing, respectively. We generated and sequenced 12 each Hi-C libraries, RNA-seq libraries, and ATAC libraries inlcuding two biological replcates per condition per timepoint.

first: Vrinda

last: Venu

first: Cullen

last: Roth

first: Samantha

middle: H

last: Adikari

first: Eric

last: Small

first: Shawn

middle: R

last: Starkenburg

first: Karissa

middle: Y

last: Sanbonmatsu


title: Construction of competing endogenous RNA networks in systemic lupus erythematosus by integrated analysis

status: Public on May 10 2024

city: Fuzhou city

first: He

last: Juanjuan

address: East Street, Gulou District,

institute: Fujian Provincial Hospital

summary: SLE is a disease characterised by immune inflammation and damage to multiple organs. Recent investigations have linked competing endogenous RNAs (ceRNAs) to lupus. However, the exact mechanism through which the ceRNAs network affects SLE is still unclear. This study aims to investigate the regulatory functions of the ceRNAs network, which are important pathways that control the pathophysiological processes of SLE. We constructed a novel circRNA-miRNA-mRNA ceRNA network (HSA circ 0000345- HSA miR-22-3-P-ESR1/SIRT1) that may have a major impact on SLE.

overall_design: CircRNA microarray for our tested assays were derived from bone marrow samples from three healthy individuals and three SLE patients in our hospital. The other sequencing data of circRNA, miRNA and mRNA were obtained from Gene Expression Omnibus (GEO) datasets. Using the limma package of R program, the differential expression of mRNA and miRNA in the GEO database was discovered. Then predicted miRNA-mRNA and circRNA-miRNA were established using miRMap, miRanda, miRDB, TargetScan and miTarBase. CircRNA-miRNA-mRNA ceRNA network was constructed using Cytoscape, and hub genes were screened using a protein-protein interaction network. Immune infiltration analysis of the hub gene was also performed by CIBERSORT and GSEA

first: Juanjuan

last: He

first: Yanfang

last: Wu


title: Study of the tRNA-derived fragment expression profile in mouse pancreatic islets of pre-diabetic obese db/db mice

first: Cecile

last: Jacovetti

summary: In this study, we discovered cytosolic and mitochondrial fragments resulting from tRNA and mt-tRNA cleavage, which may act as new regulators of cellular and metabolic functions. We analyzed hundreds of these fragments in the pancreatic islets of db/db mice and compared them to heterozygous control db/+ mice. At 16 weeks of age, db/db mice exhibit obesity, insulin resistance, and glucose intolerance. In our analysis, we identified 3858 tRFs in the islets of db/db mice, among which 342 exhibited significant changes (≥ 2 fold; adjusted p value ≤ 0.05) compared to controls. Of these, 199 tRFs showed increased levels, while 170 tRFs showed decreased levels in the pre-diabetic mice. Notably, a striking majority (147 out of 170) of the tRFs with reduced abundance in the islets of db/db mice were derived from the cleavage of tRNAs encoded by the mitochondrial genome. Our findings reveal a significant reshaping of mitochondrial tRFs in pre-diabetic conditions, coinciding with a well-established mitochondrial metabolic defect under these conditions. Specifically, we demonstrated that a fragment (named mt-tRF-LeuTAA) resulting from the cleavage of mt-tRNA-LeuTAA, encoded by the mitochondrial genome and found to be reduced in the islets of db/db mice, acts as a key regulator of mitochondrial OXPHOS functions, mitochondrial membrane potential, the insulin secretory capacity of ß-cells, and the insulin sensitivity of myotube muscle cells.

overall_design: In order to characterize the expression profile of tRFs (tRNA-derived fragments) in a pre-diabetic context, we isolated total RNA from pancreatic islets of three db/db mice and compared it to that of three heterozygous controls (db/+ mice).

first: Chris

last: Donnelly

first: Véronique

last: Menoud

first: Mara

last: Suleiman

first: Cristina

last: Cosentino

first: Jonathan

last: Sobel

first: Keijing

first: Karim

last: Bouzakri

first: Piero

last: Marchetti

first: Claudiane

last: Guay

first: Bengt

last: Kayser

first: Romano

last: Regazzi


title: Differential junction usage in paired RNA-seq of white and beige differentiated adipocytes from six human subjects

city: Oslo

first: Philippe

last: Collas

address: PO Box 1112 Blindern

department: Institute of Basic Medical Sciences

country: Norway

institute: University of Oslo

summary: Background: The beneficial effect of thermogenic adipocytes in maintaining body weight and protecting against metabolic disorders has raised interest in understanding the regulatory mechanisms defining white and beige adipocyte identity. Although alternative splicing has been shown to propagate adipose browning signals in mice, this has yet to be thoroughly investigated in human adipocytes.  Methods: We performed parallel white and beige adipogenic differentiation using primary adipose stem cells from 6 unrelated healthy subjects, and assessed differential gene and isoform expression in mature adipocytes by RNA sequencing. Results: We find 693 exon junctions with robust differential usage between white and beige adipocytes in all 6 subjects, mapping to 507 genes. Importantly, only 8% of these differentially spliced genes are also differentially expressed, indicating that alternative splicing constitutes an additional layer of gene expression regulation during beige adipocyte adipogenic differentiation. Functional classification of alternative isoforms point to a gain of function for key thermogenic regulators such as PPARG, CITED1 and PEMT. We find that a large majority of the splice variants arise from differential usage of transcription start sites (TSSs), with beige-specific TSSs being enriched for PPARγ and MED1 binding compared to white-specific TSSs. Finally, we validate beige specific isoform expression at the protein level for two thermogenic regulators, PPARγ and PEMT. Discussion: These results indicate that differential isoform expression through alternative TSS usage is an important regulatory mechanism for human adipocyte thermogenic specification.  Code for data processing and analysis are available at https://github.com/sarahhp/splicing_thermogenesis.

overall_design:
To study the differentiation of beige adipocytes in vitro, six human primary adipose stem cell (hASC) lines from female subjects were selected for similar white adipose differentiation capacity based on marker genes and proteins.
hASC cell lines from six subjects were differentiated in triplicate into both beige and white adipocytes in the presence or absence of rosiglitazone, respectively, and harvested for RNA-seq.
We investigated the difference between the white and beige adipocytes using differential gene and exon junction expression. Further, differential exon junctions were annotated with functionality scores from TRIFID.

first: Sarah

last: Hazell Pickering

first: Mohamed

last: Abdelhalim

first: Nolwenn

last: Briand


title: H2A.Z chaperones promote melanoma cell growth by facilitating transcription of E2F target genes through H2A.Z deposition and H4 acetylation

city: New York

first: Sina

last: Jostes

state: NY

address: 1470 Madison Avenue

institute: Icahn School of Medicine at Mt Sinai

type: Genome binding/occupancy profiling by high throughput sequencing

summary: High levels of the histone variant H2A.Z are characteristic for melanoma progression and correlate with poor prognosis of melanoma patients. Two chaperone complexes are reported to deposit H2A.Z isoforms into chromatin: SRCAP and P400-TIP60. YL1 is a common subunit of both complexes and directly binds to the H2A.Z-H2B dimer. Here, we showed that the knockdown of SRCAP (SRCAP-specific), P400 (P400-TIP60-specific) and YL1 (shared subunit) results in loss of H2A.Z deposition into chromatin in melanoma cells, confirming them as H2A.Z chaperone subunits in this system. Further, we demonstrated that H2A.Z, YL1 and the SRCAP-specific subunit ZNHIT1 co-localize at active promoters of cell cycle-related genes, such as the transcription factor E2F1. Knockdown of YL1, SRCAP and P400 downregulates E2F1 and its targets, resulting in a loss of proliferation and cell cycle arrest in melanoma cells as seen after knockdown of H2A.Z (Vardabasso et al. 2015). Strikingly, besides H2A.Z loss, we observed a dramatic loss of H4 acetylation in melanoma chromatin upon knockdown of H2A.Z chaperone subunits. The majority of genes whose promoters showed a reduction in H4 acetylation were previously bound by H2A.Z, YL1 and ZNHIT1. This is suggestive of a direct link between H2A.Z deposition and H4 acetylation to promote the expression of underlying genes. In agreement, the genes that lost H4 acetylation were enriched for E2F1 target genes. Interestingly, we additionally found that knockdown of YL1 did not only prohibit cell cycle progression, but also induces apoptosis, which is in contrast to H2A.Z knockdown affecting cell cycle only. In addition, YL1 (but not SRCAP or P400) was found to be overexpressed in melanoma and high YL1 levels were a predictor of poor melanoma patient outcome. These findings provide a rationale for targeting the H2A.Z-YL1 interaction as novel epigenetic strategy for melanoma treatment.

overall_design: Chromatin  inmunoprecipitation and sequencing of Parental, shScramble, and shYL1 Skmel147 of YL1, ZNHIT1 and H4ac

first: Chiara

last: Vardabasso

first: Joanna

last: Dong

first: Saul

last: Carcamo

first: Rajendra

last: Singh

first: Robert

last: Phelps

first: Austin

last: Meadows

first: Elena

last: Grossi

first: Dan

last: Hasson

first: Emily

last: Bernstein


title: Viral and host network analysis of the human cytomegalovirus transcriptome in latency

city: Tucson

first: Felicia

middle: Dare

last: Goodrum

state: AZ

address: 1657 E Helen St

department: Immunobiology

institute: University of Arizona

summary:
To investigate HCMV (and human host) gene expression profile during latency and reactivation, we infected THP-1 cells with the wildtype HCMV-TB40E or with recombinant viruses that cannot establish latency (UL138stop) or cannot reactivate (UL135stop).
After the establishment of viral latency, we treated cells with TPA to induce cellular differentiation and viral reactivation. We then performed gene expression profiling analysis using data obtained from RNA-seq over a timecourse of undifferentiated and differentiated cells
that are infected with each virus. Uninfected cells are included as a control for gene expression changes that are virally induced.

overall_design: Temporal gene expression profiling analysis of RNA-Seq data for THP1 cells infected with HCMV wildtype or recombinant viruses

first: Donna

last: Collins-McMillen

first: Diogo

last: de Oliveira Pessoa


title: MICROGLIAL CELL EXPRESSION OF THE TYPE 2 CANNABINOID RECEPTOR REGULATES IMMUNE-MEDIATED NEUROINFLAMMATION

city: Milwaukee

first: Anthony

middle: Elston

last: Zamora

state: Wisconsin

address: 8701 Watertown Plank Road

department: Medicine

institute: Medical College of Wisconsin

summary: Neuroinflammation is a recognized complication of immunotherapeutic approaches such as immune checkpoint inhibitor treatment, chimeric antigen receptor therapy, and graft versus host disease (GVHD) occurring after allogeneic hematopoietic stem cell transplantation.  While T cells and inflammatory cytokines play a role in this process, the precise interplay between the adaptive and innate arms of the immune system that propagates inflammation in the central nervous system remains incompletely understood.  Using a murine model of GVHD, we demonstrate that type 2 cannabinoid receptor (CB2R) signaling plays a critical role in the pathophysiology of neuroinflammation.  In these studies, we identify that CB2R expression on microglial cells induces an activated inflammatory phenotype which potentiates the accumulation of donor-derived proinflammatory T cells, regulates chemokine gene regulatory networks, and promotes neuronal cell death.  Pharmacological targeting of this receptor with a brain penetrant CB2R inverse agonist/antagonist selectively reduced neuroinflammation without deleteriously affecting systemic GVHD severity.  Thus, these findings delineate a therapeutically targetable neuroinflammatory pathway and has implications for the attenuation of neurotoxicity after GVHD and potentially other T cell-based immunotherapeutic approaches.

overall_design: Cells were isolated from the brains of mice that were transplanted with Bone marrow (BM) alone or BM plus adjunctive spleen cells (GVHD). To delineate the inflammatory environment in the brain, scRNAseq was performed on the immune cells isolated from brains of BM and GVHD mice. To delineate the effect of CB2R expression on microglia and determine whether these cells had a more inflammatory signature, scRNAseq was performed on sorted micrglial cells obtained from brains of WT versus CB2R-/- GVHD recipient mice.

first: Alison

last: Moe

first: Aditya

last: Rayasam

first: Garrett

last: Sauber

first: Ravi

last: Shah

first: Chengyin

last: Yuan

first: Bob

last: Moore

first: Marco

last: Colonna

first: Weiguo

last: Cui

first: Cecilia

middle: J

last: Hillard

first: William

last: Drobyski


title: New strategies for the identification of intronic variants related to splicing events in pancreas cancer

city: Goettingen

first: Gabriela

last: Salinas

state: Lower-Saxony

address: Kreuzbergring 57

department: Department of Pathology

institute: Universitaetsmedizin Goettingen


title: H2A.Z chaperones promote melanoma cell growth by  facilitating transcription of E2F target genes through H2A.Z deposition and H4 acetylation

first: Sina

last: Jostes

overall_design: To investigate the role of the H2A.Z Chaperone Subunits YL1, SRCAP and P400 in melanoma cells, we performed shRNA-mediated knockdown of each gene (VPS72, SRCAP, EP400) and performed RNA-seq analysis 3 or 6 days after infection. A scrambled shRNA (SCR) served as negative control.

first: Chiara

last: Vardabasso

first: Joanna

last: Dong

first: Rajendra

last: Singh

first: Robert

last: Phelps

first: Austin

last: Meadows

first: Saul

last: Carcamo

first: Elena

last: Grossi

first: Dan

last: Hasson


title: Induced pluripotent stem cell-derived fibroblasts efficiently engage senescence pathways but show increased sensitivity to stress inducers

city: montreal

first: Christian

last: Beausejour

state: Québec

address: 3175 Cote Ste-Catherine

department: Pharmacologie et physiologie

country: Canada

institute: Université de Montréal

summary: The risk of malignant transformation of induced pluripotent stem cells (iPSC)-derived cells in re-sponse to DNA damage is a concern knowing that the tumor suppressor genes TP53 and CDKN2A are transiently inactivated during reprogramming. Here, we evaluated the integrity of cellular senescence pathways and DNA double strand break (DSB) repair in iPSC-derived human fibro-blasts (i-HF) compared to their parental skin fibroblasts (HF). Using a variety of functional assays, we show that the capacity of i-HF to enter senescence and repair DSB is not compromised after damage induced by ionizing radiation (IR) or overexpression of H-RASV12. Still, i-HF lines are transcriptionally different from their parental lines, showing enhanced metabolic activity and higher expression of p53-related effector genes. As a result, i-HF lines generally exhibit increased sensitivity to various stresses, have an elevated senescence associated secretory phenotype (SASP) and cannot be immortalized unless p53 expression is knocked down. In conclusion, while our results suggest that i-HF are not at a greater risk of transformation, their overall hyperactivation of se-nescence pathways may impede their function as a cell therapy product.

overall_design:
To evaluate the tumorigenic potential of iPSC-derived cells,we used human fibroblasts (HF) isolated from skin biopsies of three different donors and measured their ability to enter senescence and to repair double strand DNA damage in comparison with their autologous iPSC-derived counterparts (i-HF).
HF were collected from three different donors (HF1: Adult male (41 years old), HF2: Adult female (39 years old), HF3: Foetal)
We also performed gene expression profiling analysis using three donors of parental and iPSC-derived fibroblasts at basal state (Ctl) and 8 hours post a 15 Gy dose of irradiation.

first: Marie-Lyn

last: Goyer

first: Cynthia

last: Desaulniers-Langevin

first: Anthony

last: Sonn

first: Georgio

last: Mansour Nehmo

first: Véronique

last: Lisi

first: Basma

last: Benabdallah

last: Beauséjour


title: Vaccinia virus infection induces concurrent alterations in host chromatin architecture, accessibility, and gene expression [Hi-C]

first: Christina

last: Steadman

first: Vrinda

last: Venu

first: Cullen

last: Roth

first: Samantha

last: Adikari

first: Eric

last: Small

first: Shawn

last: Starkenburg

first: Karissa

last: Sanbonmatsu


title: Vaccinia virus infection induces concurrent alterations in host chromatin architecture, accessibility, and gene expression

relation:
SuperSeries of: GSE248048
SuperSeries of: GSE248049
SuperSeries of: GSE248050


title: AML/T cell interactomics uncover correlates of patient outcomes and the key role of ICAM1 in T cell killing of AML

status: Public on May 09 2024

city: Palo Alto

first: Maria Grazia

last: Roncarolo

state: California

address: 265 Campus Drive

department: Pediatrics

institute: Stanford University

summary: T cells are important for the control of acute myeloid leukemia (AML), a common and often deadly malignancy. We observed that some AML patient samples are resistant to killing by human engineered cytotoxic CD4+ T cells. Single-cell RNA-seq of primary AML samples and CD4+ T cells before and after their interaction uncovered transcriptional programs that correlate with AML sensitivity or resistance to CD4+ T cell killing. Resistance-associated AML programs were enriched in AML patients with poor survival, and killing-resistant AML cells did not engage T cells in vitro. Killing-sensitive AML potently activated T cells before being killed, and upregulated ICAM1, a key component of the immune synapse with T cells. Without ICAM1, killing-sensitive AML became resistant to killing to primary ex vivo-isolated CD8+ T cells in vitro, and engineered CD4+ T cells in vitro and in vivo. Thus, ICAM1 on AML acts as an immune trigger, allowing T cell killing, and could affect AML patient survival in vivo.

overall_design:
To better Understand the determinants of AML sensitivity or resistance to T cell killing, we analyzed killing-sensitive (PARCEV and PARCHW) and -resistant (PATISD and PAPZCL) primary AML samples before and after co-culture with CD4^IL10 cells (LV10_072 and LV10_2441) using single-cell RNA sequencing.
Four Pediatric AML bone marrow aspirates were obtained from the Children's Oncology Group.  Primary AML samples and T cells were FACs-sorted for scRNA-seq at day 0.  Primary AML cells were co-cultured for 24 h with T cells at a 1:1 ratio and sorted for scRNA-seq.  This was repeated twice, so that each AML sample was co-cultured with T cells from two different donors.

first: Ece

middle: C

last: Sayitoglu

first: Bogdan

last: Luca

first: Allison

middle: P

last: Boss

first: Benjamin

last: Thomas

first: Robert

last: A Freeborn

first: Molly

last: Uyeda

first: Pauline

last: Chen

first: Yusuke

last: Nakauchi

first: Colin

last: Waichler

first: Norman

last: Lacayo

first: Rosa

last: Bacchetta

first: Ravindra

last: Majeti

first: Andrew

last: Gentles

first: Alma-Martina

last: Cepika

first: Maria

middle: G


title: Interplay of Salmonella enterica sv. Typhimurium and Bifidobacterium infantis during co-infection of gut epithelial cells

status: Public on May 08 2024

city: Davis

first: Bart

last: Weimer

state: CA

address: 1 Shields Ave

institute: UC Davis

summary: The deposited microarray data were generated in a study that integrated the gene expression profiles and metabolic responses of Caco2 cells incubated with Bifidobacterium infantis subsp. infantis and Salmonella enterica subsp. enterica sv. Typhimurium. The aim of this study was to investigate the interaction of B. infantis, S. Typhimurium, and host cells (Caco2) in the course of infection to understand the molecular mechanics of probiotic-pathogen-host interactions.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. LT2

overall_design: Differentiated Caco2 cells were incubated with S. Typhimurium, with B. infantis, or with a combination of S. Typhimurium and B. infantis. Gene expression measurments were analyzed at 30mins, 60mins, and 120mins after coculture and compared with the controls of the time-matched monocultures.

first: Claire

last: Shaw

first: Reed

last: Gann

first: Prerak

middle: T

last: Desai

first: Jigna

middle: D

last: Shah


title: MR1-restricted T cell clonotypes are associated with “resistance” to Mycobacterium tuberculosis infection

city: Seattle

first: Chetan

last: Seshadri

state: WA

address: 850 Republican St

institute: University of Washington

summary: T cells are required for protective immunity against Mycobacterium tuberculosis. We recently described a cohort of Ugandan household contacts of tuberculosis cases who appear to “resist” M. tuberculosis infection (resisters; RSTRs) and showed that these individuals harbor IFN-γ–independent T cell responses to M. tuberculosis–specific peptide antigens. However, T cells also recognize nonprotein antigens via antigen-presenting systems that are independent of genetic background, known as donor-unrestricted T cells (DURTs). We used tetramer staining and flow cytometry to characterize the association between DURTs and “resistance” to M. tuberculosis infection. Peripheral frequencies of most DURT subsets were comparable between RSTRs and latently infected controls (LTBIs). However, we observed a 1.65-fold increase in frequency of MR1-restricted T (MR1T) cells among RSTRs in comparison with LTBIs. Single-cell RNA sequencing of 18,251 MR1T cells sorted from 8 donors revealed 5,150 clonotypes that expressed a common transcriptional program, the majority of which were private. Sequencing of the T cell receptor α/T cell receptor δ (TCRα/δ) repertoire revealed several DURT clonotypes were expanded among RSTRs, including 2 MR1T clonotypes that recognized mycobacteria-infected cells in a TCR-dependent manner. Overall, our data reveal unexpected donor-specific diversity in the TCR repertoire of human MR1T cells as well as associations between mycobacteria-reactive MR1T clonotypes and resistance to M. tuberculosis infection.

overall_design: MR1-restricted T-cells (defined as live CD7+ MR1-5-OP-RU tetramer+ cells) were sorted from peripheral blood samples (n = 8). Surface protein expression, gene expression and paired T-cell receptor sequencing were performed on each sample. Samples from 4 ‘resistors’ (RSTR, defined as longitudinally negative for tuberculin skin test and IFNg release assay) and 4 latently infected donors (LTBI, defined as longitudinally positive for tuberculin skin test and IFNg release assay) have been included in this dataset. Samples were sequenced ex vivo without stimulation.


title: Elucidating the Onset of Cross-protective Immunity after Intranasal Vaccination with the Attenuated African Swine Fever Vaccine Candidate BA71ΔCD2

city: Bellaterra

first: Jordi

last: Argilaguet Marqués

state: Barcelona

address: CAMPUS UAB EDIF.CRESA

country: Spain

institute: IRTA-CReSA

summary: African swine fever (ASF) is a deadly disease of swine currently causing a worldwide pandemic leading to severe economic consequences for the porcine industry. The control of disease spread is hampered by the limitation of available effective vaccines. Live attenuated vaccines (LAVs) are currently the most advanced vaccine prototypes, providing strong protection against ASF. However, the significant advances achieved using LAVs must be complemented with further studies to analyse vaccine-induced immunity. Here we characterized the onset of cross-protective immunity triggered by the LAV candidate BA71ΔCD2. Intranasally vaccinated pigs were challenged with the virulent Georgia 2007/1 strain at days 3, 7 and 12 postvaccination. Only the animals vaccinated 12 days before challenge effectively controlled infection progression, showing low virus loads, minor clinical signs and lack of the unbalanced inflammatory response characteristic of severe disease. Contrarily, animals vaccinated 3 or 7 days before challenge just showed a minor delay of disease progression. Virus-specific antibody responses and whole blood transcriptome signatures demonstrated that control of infection is associated with the presence of virus-specific antibodies and a cytotoxic response before challenge. These results contribute to our understanding of protective immunity induced by LAV-based vaccines, encouraging their use in emergency responses in ASF affected areas.

overall_design: Twenty seven-week-old Landrace x Large white breed male pigs were divided into four groups of five pigs. Three groups were intranasally vaccinated with 10^6 pfu/animal of the attenuated strain BA71ΔCD2 at 3, 7 or 12-days pre-challenge. Each animal received 2 mL of the vaccine diluted in PBS via intranasal inoculation (1 mL/nostril). As a control group, the remaining 5 animals were inoculated with 2 mL of PBS. Each animal was intranasally inoculated with 2 mL containing 10^5 HAU50 of the highly virulent ASFV strain Georgia2007/1 (1 mL/nostril). Clinical signs were monitored daily throughout the length of the experiment. EDTA-blood, sera and nasal swabs were taken the days of vaccination and challenge, and at 3, 7 and 9 days post-challenge (dpc) to assess viral loads and host immune responses. Clinical signs were evaluated following standardized guidelines. Whole blood was collected and analysed at 0, 3 and 7 dpc by RNA-seq.

first: David

last: Marín-Moraleda

first: Jordana

last: Muñoz-Basagoiti

first: Aida

last: Tort-Miró

first: María

last: Navas

first: Marta

last: Muñoz

first: Enric

last: Vidal

first: Àlex

last: Cobos

first: Beatriz

last: Martín-Mur

first: Sochanwattey

last: Meas

first: Veronika

last: Motuzova

first: Chia-Yu

last: Chang

first: Francesc

last: Accensi

first: Sonia

last: Pina-Pedrero

first: Jose

middle: I

last: Núñez

first: Anna

last: Esteve-Codina

first: Boris

last: Gavrilov

first: Fernando

last: Rodriguez

first: Lihong

last: Liu

last: Argilaguet


title: The neuroprotective effect of TGF-β1 on mechanically injured cortical murine neurons by reducing trauma-induced autophagy and apoptosis

city: Shantou

first: Huixiong

last: Deng

address: Xinling 22 Rd

department: Guangdong Provincial Key Laboratory of Infectious Diseases and Molecular Immunopathology

institute: Shantou University Medical College

summary: Background Transforming growth factor β1 (TGF-β1) has a neuroprotective function in traumatic brain injury (TBI) through its anti-inflammatory and immunomodulatory properties. In our previous study, we found that TGF-β1 played a critical role in inhibiting apoptosis and increasing neuronal activity in murine cortical neurons following trauma. However, the precise mechanisms underlying the neuroprotective actions of TGF-β1 on the cortex require further investigation. Methods Thus, in this study, we were aimed to investigate the regulatory function of TGF-β1 on neuronal autophagy and apoptosis using an in vitro primary cortical neuron trauma-injury model. Results To establish the landscape of differentially expressed genes (DEGs) with or without TGF-β1 (10ng/ml) treatment for 24 hours, we performed RNA-sequencing. We observed significant enrichment of DEGs related to autophagy, apoptosis, and the lysosome pathway in trauma-injured cortical neurons. Additionally, transmission electron microscopy (TEM) confirmed the presence of autophagosomes as well as autophagolysosomes. Western blot analysis revealed upregulation of autophagy-related protein light chain 3 (LC3)-Ⅱ/LC3-Ⅰ, sequestosome 1 (SQSTM1)/p62, along with apoptosis-related protein Cleaved-caspase3 in trauma-injured primary cortical neurons. Furthermore, mechanically injured neurons showed an upregulation of lysosomal marker protein lysosomal marker protein (LAMP1) and lysosomal enzyme mature cathepsin D (mCTSD), but a decrease in the activity of CTSD enzyme. These results indicated that apoptosis was up-regulated in mechanically injured neurons at 24 hours, accompanied by lysosomal dysfunction and impaired autophagic flux. Notably, TGF-β1 significantly reversed these changes. Conclusions Therefore, our findings suggested that TGF-β1 exerted neuroprotective effects on mechanically injured neurons by reducing lysosomal dysfunction, decreasing the accumulation of autophagosomes and autophagolysosomes, and enhancing autophagic flux.

overall_design: We were aimed to investigate the regulatory function of TGF-β1 on neuronal autophagy and apoptosis using an in vitro primary cortical neuron trauma-injury model.To establish the landscape of differentially expressed genes (DEGs) with or without TGF-β1 (10ng/ml) treatment for 24 hours, we performed RNA-sequencing.

first: Yanlei

last: Li


title: Alveolar Macrophage Phenotypes in Schistosoma-Induced Pulmonary Hypertension

city: San Francisco

first: Rahul

last: Kumar

address: 1001 Potrero Ave, Rm.5K1

institute: University of California, San Francisco, USA

summary: Schistosomiasis, a prevalent cause of pulmonary hypertension (PH) globally, triggers type 2 inflammation. The role of alveolar macrophages (AMs) are not well defined. This study provides insights into the complex interplay of AMs subpopulations in Schistosoma-induced PH, shedding light on potential therapeutic targets for this global health concern.

overall_design: Cx3cr1+/GFP reporter mice were intraperitoneally sensitized and then intravenously challenged with S. mansoni eggs .Lungs alveolar macrophages (AMs) were sorted from unexposed and at day 1, 3 and 7 following Schistosoma IV challenge by flow cytometry. At each time point, IMs were sorted from of a pair of age- and sex-matched mice (1 male and 1 female) and were combined for subsequent analyses.  Single cells were captured using a Chromium Box (10X Genomics, Pleasanton, CA). Libraries were sequenced using the Illumina NovaSEQ 6000 sequencer (Illumina, Inc., San Diego, CA). Subsequently, RNA sequence reads were aligned to the Mus musculus GRCm39 reference genome using Cell Ranger v6 (10x Genomics). Downstream analysis was performed using the Seurat package in R.

first: Sushil

first: Claudia

last: Mickael

first: Dara

last: Fonseca Balladares

first: Kevin

last: Nolan

first: Michael

last: Lee

first: Linda

last: Sanders

first: Rubin

last: Tuder

first: Kurt

last: Stenmark

first: Brian

middle: B

last: Graham


title: Transcriptomic analysis of polysaccharide utilization loci reveals substrate preferences in two ruminal generalists Segatella bryantii TF1-3 and Xylanibacter ruminicola KHP1

city: Ljubljana

first: Urška

last: Murovec

address: Jamnikarjeva 101

country: Slovenia

institute: Biotechnical Faculty

summary: Bacteria of the genera Xylanibacter and Segatella  are one of the most dominant groups in the rumen microbiota. They are characterized by the ability to utilize different hemicelluloses and pectin of plant cell-wall as well as plant energy storage polysaccharides. The degradation is possible with the use of cell envelope bound multiprotein apparatuses coded in polysaccharide utilization loci (PULs), which have been shown to be substrate specific. The knowledge of PUL presence in rumen Xylanibacter and Segatella based on bioinformatic analyses is already established and transcriptomic and genetic approaches confirmed predicted PULs for a limited number of substrates. In this study, we transcriptomically identified additional different PULs in Xylanibacter ruminicola KHP1 and Segatella bryantii TF1-3. We also identified substrate preferences and found that specific growth rate and extent of growth impacted the choice of substrates preferentially used for degradation. These preferred substrates were used by both strains simultaneously as judged by their PUL upregulation. Lastly, β-glucan and xyloglucan were used by these strains in the absence of bioinformatically and transcriptomically identifiable PUL systems.

sample_organism:
Xylanibacter ruminicola
Segatella bryantii

overall_design: We were growing each bacteria in 11 different polysaccharides and glucose as reference. And performed RNAseq analysis for each polysaccharide (22 experiments) + 2 glucose. We also did the RNAseq in minimal medium with glucose and beta-glucan for S. bryantii TF1-3.

first: Tomaž

last: Accetto


title: Vaccinia virus infection induces concurrent alterations in host chromatin architecture, accessibility, and gene expression [ATAC-seq]

status: Public on May 07 2024

first: Christina

last: Steadman

first: Vrinda

last: Venu

first: Cullen

last: Roth

first: Samantha

last: Adikari

first: Eric

last: Small

first: Shawn

last: Starkenburg

first: Karissa

last: Sanbonmatsu


title: Interstitial Macrophage Phenotypes in Schistosoma-Induced Pulmonary Hypertension

first: Rahul

last: Kumar

summary: Schistosomiasis, a prevalent cause of pulmonary hypertension (PH) globally, triggers type 2 inflammation, with interstitial macrophages (IMs) derived from monocytes playing a crucial role. These IMs produce thrombospondin-1 (TSP-1), activating TGF-β and driving PH pathology. Two distinct IM subpopulations were identified: resident FOLR2+ IMs expressing monocyte recruitment factors, and recruited CCR2+ IMs expressing TSP-1. Upon exposure to Schistosoma, the CCR2+ subpopulation expanded. Flow cytometry and single-cell RNA sequencing confirmed these findings, revealing crosstalk between IM subpopulations. The resident FOLR2+ IMs increased expression of monocyte recruitment ligands, while the recruited CCR2+ IMs expressed elevated TSP-1, activating TGF-β and contributing to PH. This study provides insights into the complex interplay of IM subpopulations in Schistosoma-induced PH, shedding light on potential therapeutic targets for this global health concern.

overall_design: Cx3cr1+/GFP reporter mice were intraperitoneally sensitized and then intravenously challenged with S. mansoni eggs. Lung interstitial macrophages (IMs) were sorted from unexposed and at day 1, 3 and 7 following Schistosoma IV challenge by flow cytometry. At each time point, IMs were sorted from of a pair of age- and sex-matched mice (1 male and 1 female) and were combined for subsequent analyses.  Single cells were captured using a Chromium Box (10X Genomics, Pleasanton, CA). Libraries were sequenced using the Illumina NovaSEQ 6000 sequencer (Illumina, Inc., San Diego, CA). Subsequently, RNA sequence reads were aligned to the Mus musculus GRCm39 reference genome using Cell Ranger v6 (10x Genomics). Downstream analysis was performed using the Seurat package in R.

first: Sushil

first: Claudia

last: Mickael


title: Active mRNA transcripts obtained using ribosomal pulldown from mouse myocytes (myofibers)

city: Columbus

first: George

last: Kyriazis

state: Ohio

address: 460 W 12th Ave

institute: The Ohio State University

summary:
We have generated mice expressing a hemagglutinin tag in the ribosomes of myocytes (Myogenin-Cre x RiboTag-Rpl22fl/fl-HA) to be able to isolate active (polysome-bound) myocyte transcripts under various experimental factors (e.g. sex, age, exercise status)
RNA was obtained by immunoprecipitation of HA-tagged polysomes of left leg solei

overall_design: 5 wt control samples (sedentary) were analyzed to see the relative levels of myocyte expressed genes. This is a first release of a broader study on the effects of various experimental factors (e.g. sex, age, exercise status) on the active gene expression of myocytes

first: Joan

last: Serrano


title: Conserved epigenetic hallmarks of T-cell aging during immunity and malignancy

status: Public on May 06 2024

city: Memphis

first: Tian

last: Mi

state: TN

address: 262 Danny Thomas Pl MS351

department: Immunology

institute: St. Jude Children's Research Hospital

type: Methylation profiling by high throughput sequencing

summary: Chronological aging correlates with epigenetic modifications at specific loci, calibrated to species lifespan. Such ‘epigenetic clocks’ appear conserved among mammals, but whether they are cell-autonomous and restricted by maximal organismal lifespan remains unknown. We used a multi-lifetime murine model of repeat vaccination and memory T cell transplantation to test whether epigenetic aging tracks with cellular replication, and if such clocks continue ‘counting’ beyond species lifespan. We found that memory T cell epigenetic clocks tick independently of host age and continue through four lifetimes. Instead of recording chronological time, T cells recorded proliferative experience through modification of cell cycle regulatory genes. Applying this epigenetic profile across a range of human T cell contexts, we found that naïve T cells appeared ‘young’ regardless of organism age, while in pediatric patients, T-cell acute lymphoblastic leukemia (T-ALL) appeared to have epigenetically aged for up to 200 years. Thus, T cell epigenetic clocks measure replicative history and can continue to accumulate well-beyond organismal lifespan.

overall_design: 2 replicates of endogenous T cell samples, 3 replicates of half lifetime samples, 5 replicates of 2 lifetime samples, 3 replicates of 4 lifetime samples, 2 replicates of Naive T cells from aged mice, 2 replicates of TCM from aged mice, 2 replicates of TEM from aged mice

first: Andrew

last: Soerens

first: Caitlin

last: Zebley

first: David

last: Masopust

first: Ben

last: Youngblood


title: Altered chromatin topologies caused by balanced chromosomal translocation lead to central iris hypoplasia [Cut & Tag]

city: Guangzhou

first: Qingjiong

last: Zhang

state: Guangdong

address: Sun Yat-sen University

institute: Zhongshan Ophthalmic Center

summary: Despite the advent of genomic sequencing, molecular diagnosis remains unsolved in approximately half of patients with Mendelian disorders, largely due to unclarified functions of noncoding regions and the difficulty in identifying complex structural variations. In this study, we map a unique form of central iris hypoplasia in a large family to 6q15-q23.3 and 18p11.31-q12.1 using a genome-wide linkage scan. Long-read sequencing reveals a balanced translocation t(6;18)(q22.31;p11.22) with intergenic breakpoints. By performing Hi-C on induced pluripotent stem cells from a patient, we identify two chromatin topologically associating domains spanning across the breakpoints. These alterations lead the ectopic chromatin interactions between APCDD1 on chromosome 18 and enhancers on chromosome 6, resulting in upregulation of APCDD1. Notably, APCDD1 is specifically localized in the iris of human eyes. Our findings demonstrate that noncoding structural variations can lead to Mendelian diseases by disrupting the 3D genome structure and resulting in altered gene expression.

overall_design: Cleavage Under Targets and Tagmentation (CUT&Tag) for the histone modification H3K27ac in the induced pluripotent stem cells (iPSCs) from one patient and one normal control.


title: Altered chromatin topologies caused by balanced chromosomal translocation lead to central iris hypoplasia [RNA-Seq]

overall_design: RNAseq for gene expression changes in samples of the induced pluripotent stem cells (iPSCs) from one patient and one normal control.


title: Altered chromatin topologies caused by balanced chromosomal translocation lead to central iris hypoplasia [Hi-C]

overall_design: High-throughput chromosome conformation capture (Hi-C) for three dimensional genome structure in the induced pluripotent stem cells (iPSCs) from one patient and one normal control.


title: Deciphering bat influenza H18N11 infection dynamics in Jamaican fruit bats on a single cell level

city: Fort Collins

first: Tony

last: Schountz

address: 3185 Rampart Rd

institute: Colorado State University

summary: Jamaican fruit bats (Artibeus jamaicensis) naturally harbor a wide range of viruses of human relevance. These infections are typically mild in bats, suggesting unique features of their immune system. To better understand the immune response to viral infections in bats, we infected Jamaican fruit bats with the bat-derived influenza A virus H18N11. Using comparative single-cell RNA sequencing, we generated a single-cell atlas of the Jamaican fruit bat intestine and mesentery, the target organs of infection. Gene expression profiling showed that H18N11 infection resulted in a moderate induction of interferon-stimulated genes and transcriptional activation of immune cells. H18N11 infection was prominent in various leukocytes, including macrophages, B cells, and NK/T cells. Confirming these findings, human leukocytes, particularly macrophages, were also susceptible to H18N11, highlighting the zoonotic potential of this virus. Our study provides insight into the virus-host relationship and thus serves as a fundamental resource for further characterization of bat immunology.

overall_design: Jamaican fruit bats were infected with 5x10^5 TCID50 25 µL of either WT H18N11 (n=6) or the NS1-deficient variant delNS1 (n=6). At 3 and 9 dpi 3 animals each were euthanized and the mesentery and intestine of each bat was processed for scRNA-seq. Four uninfected control bats were included.

first: Geoffroy

last: Andrieux

first: Melanie

last: Boerries

first: Kevin

last: Ciminski

first: Susanne

last: Kessler


title: New strategies for the identification of intronic variants related to splicing events in pancreas cancer [iCELL8]

first: Gabriela

last: Salinas

summary: Most clinical diagnostic settings and genomic research focus almost exclusively on coding regions and essential splice sites, mostly ignoring non-coding variants. Indeed, the investigation of intronic mis-splicing variants interpreting mechanisms to disease-associated splicing events requires both genomic and transcriptomic data. Unfortunately, there are not many datasets where both are available, leading to the understanding of intronic variants in diseases full of gaps.      In this study, we present for the first time a full-length single nuclei RNA-sequencing (snRNA-Seq) approach improving the proper investigation of pathogenic mis-splicing intronic variants in pancreatic cancer showing its contribution to abnormal splicing changes and their transcriptional effects. Finally, we discuss the demands of further machine learning-based methods to process the growing number of single cell data and to enhance the precision and recall.

overall_design: Bulk RNA Seq from PDAC compared to single nuclei RNA Seq

first: Maren

last: Sitte

first: Emre

last: Taylan Duman

first: Elisabeth

last: Hessmann


title: Interfering with glycolysis potentiates the long-term effect of local tumor ablation via CD8+TCM cell differentiation [MWA or MWA+2DG group]

status: Public on May 04 2024

city: Nanjing

first: Xinyu

last: Tang

address: 300 Guangzhou Road

institute: The First Affiliated Hospital with Nanjing Medical University

summary: To evaluate the immune responses in tumor-draining lymph nodes induced by microwave ablation (MWA) or microwave ablation combined with 2DG (MWA+2DG), we then performed gene expression profiling analysis using data obtained from RNA-seq of 5 different lymph nodes from two group of mice.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for TDLNs on day 15 from mice in MWA or MWA+2DG groups.

first: Wenbin

last: Zhou


title: Interfering with glycolysis potentiates the long-term effect of local tumor ablation via CD8+TCM cell differentiation [MWA+PBS or MWA+2DG]

summary: To evaluate the genetic changes of systemic CD8+T cells induced by microwave ablation(MWA+PBS) or microwave ablation combined with 2DG(MWA+2DG),We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different spleens from two group of mice.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for CD8+T cells isolated from spleens of  mice on day 35 in MWA+PBS or MWA+2DG groups.


title: Interfering with glycolysis potentiates the long-term effect of local tumor ablation via CD8+TCM cell differentiation

status: Public on May 03 2024

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

overall_design: Refer to individual Series


title: Genomewide identification of replication fork stalling/pausing sites and the interplay between RNA Pol II transcription and DNA replication progression

city: Birmingham

first: Marco

last: Saponaro

address: Vincent Drive

institute: University of Birmingham

summary: DNA replication progression can be affected by the presence of physical barriers on the DNA, like the RNA Polymerases, leading to replication stress and DNA damage. Nonetheless, we do not know the overall influence of transcription on DNA replication progression. To characterize what happens at sites where DNA replication forks stall and pause, we establish a genome-wide approach to identify them. This approach uses multiple timepoints to identify replication fork/stalling hotspots as the replication progresses through the genome. These sites are typically associated with increased DNA damage, overlap with fragile sites and with breakpoints of rearrangements identified in cancers, but do not overlap with replication origins. Overlaying these sites with a genome-wide analysis of RNA Polymerase II transcription, we find that replication fork stalling/pausing sites inside genes are directly related to transcription progression and activity. We also find instances where transcription activity favors replication progression because reduces histone density, but also that slowing down transcription elongation slows down directly replication progression through genes. This would indicate that transcription and replication can coexist over the same regions. Importantly, rearrangements found in cancers overlapping transcription-replication collision sites are detected in non-transformed cells and increase following treatment with ATM and ATR inhibitors. Altogether, our findings highlight how transcription and replication overlap during S-phase, with both positive and negative consequences for replication fork progression and genome stability by the coexistence of these two processes.

overall_design:
Genome-wide profiles of RNAPII and PCNA in synchronous BJ-hTERT cells and generated by NEXT-seq in one biological repeats

Please note that the metagene profiles ('all_peaks.txt') are also peaks identified in the following BrdU-Seq samples (in GSE136288/PRJNA561836):

GEO Sample accns  SRA experiment accns  Biosample accns
GSM4044019   SRX6756306  SAMN12629357
GSM4044020   SRX6756307  SAMN12629356
GSM4044021   SRX6756308  SAMN12629355
GSM4044022   SRX6756309  SAMN12629354
GSM4044023   SRX6756310  SAMN12629353
GSM4044024   SRX6756311  SAMN12629352
GSM4044025   SRX6756312  SAMN12629351
GSM4044026   SRX6756313  SAMN12629350
GSM4044027   SRX6756314  SAMN12629349
GSM4044028   SRX6756315  SAMN12629348
GSM4044029   SRX6756316  SAMN12629347
GSM4044030   SRX6756317  SAMN12629346

BrdU peaks identified in each timepoint were all collected together in the file all_peals.txt
More specifically, BrdU peaks were called in:

GSM4044020(BrdU-Seq 16h 1) vs GSM4044019 (BrdU-Seq Input 1)
GSM4044021(BrdU-Seq 19h 1) vs GSM4044019 (BrdU-Seq Input 1)
GSM4044022(BrdU-Seq 22h 1) vs GSM4044019 (BrdU-Seq Input 1)
GSM4044023(BrdU-Seq 24h 1) vs GSM4044019 (BrdU-Seq Input 1)
GSM4044024(BrdU-Seq 28h 1) vs GSM4044019 (BrdU-Seq Input 1)

GSM4044026(BrdU-Seq 16h 2) vs GSM4044025 (BrdU-Seq Input 2)
GSM4044027(BrdU-Seq 19h 2) vs GSM4044025 (BrdU-Seq Input 2)
GSM4044028(BrdU-Seq 22h 2) vs GSM4044025 (BrdU-Seq Input 2)
GSM4044029(BrdU-Seq 24h 2) vs GSM4044025 (BrdU-Seq Input 2)
GSM4044030(BrdU-Seq 28h 2) vs GSM4044025 (BrdU-Seq Input 2)

first: Jianming

last: Wang

first: Patricia

last: Rojas


title: Single-cell RNA sequencing Reveals Obesity-Induced Inhibition of Testosterone Production and Damage of Blood-Testis Barrier Integrity

city: Xi'an

last: Cao

state: Shanxi

address: 76 Yanta West Road, Xi’an, Shaanxi 710061, China

institute: Xi'an Jiaotong University

summary: Obesity is accompanied by multiple known health risks and increased morbidity, and the prevalence of obesity continues to rise. Fertility in males of reproductive age is affected by the endocrine abnormalities and comorbidities associated with obesity, which has become a primary cause of male infertility. Single-cell RNA sequencing (scRNA-seq) of testes in obese individuals has not been done, so we chose to use this technique to study the mechanism of infertility in an obese mouse model.

overall_design: Male ICR mice were randomly divided into a conventional diet (CD) group and a high-fat diet (HFD) group at the age of 6 weeks. Testes of the two groups were collected and analyzed by scRNA-seq.

first: Pengxiang

last: Qu


title: New strategies for the identification of intronic variants related to splicing events in pancreas cancer [RNA-Seq]

summary:
Most clinical diagnostic settings and genomic research focus almost exclusively on coding regions and essential splice sites, mostly ignoring non-coding variants. Indeed, the investigation of intronic mis-splicing variants interpreting mechanisms to disease-associated splicing events requires both genomic and transcriptomic data. Unfortunately, there are not many datasets where both are available, leading to the understanding of intronic variants in diseases full of gaps.
In this study, we present for the first time a full-length single nuclei RNA-sequencing (snRNA-Seq) approach improving the proper investigation of pathogenic mis-splicing intronic variants in pancreatic cancer showing its contribution to abnormal splicing changes and their transcriptional effects. Finally, we discuss the demands of further machine learning-based methods to process the growing number of single cell data and to enhance the precision and recall.

last: Duman


title: Nucleotide depletion promotes cell fate transitions by inducing DNA replication stress

city: Cambridge

first: Brian

last: Do

state: MA

address: 500 Main St

department: Biology

institute: MIT

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: RNA-seq analysis of naïve and MASH liver tissue.

status: Public on May 02 2024

city: Hachioji

first: Kenichi

last: Asano

state: Tokyo

address: 1432-1 Horinouchi

department: Life Sciences

country: Japan

institute: Tokyo University of Pharmacy and Life Sciences

summary: This analysis was perfomed to explore microevironmental cues that may induce Egr2 expression in infiltrating monocytes.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for liver tissue from ND-fed mice and CDA-HFD-fed mice.

first: Ayaka

last: Iwata


title: A Cell Type-Specific Approach to Elucidate the Role of miR-96 in Inner Ear Hair Cells

city: Bethesda

first: Ronna

last: Hertzano

state: Maryland

address: 35 Convent Drive, Building 35A Room 1F127

department: Neurotology Branch

institute: NIDCD/NIH

summary: Mutations in miRNA-96, a microRNA expressed within the hair cells (HCs) of the inner ear, result in progressive hearing loss in both mouse models and humans. While previous studies have delved into miR-96 transcriptional cascades via whole organ of Corti microarray experiments of diminuendo (Mir96Dmdo) mice, they face limitations in pinpointing cell type-specific differentially expressed genes. This hinders the ability to conclusively determine if the effects of Mir96Dmdo are specifically within HCs and contribute to the observed abnormal Mir96Dmdo HC phenotype and determine the role of miR-96 in HCs. In this study, we generate the first HC-specific RNA-sequencing (RNA-seq) datasets from Mir96Dmdo; Atoh1/nGFP+ postnatal day 1 wildtype, heterozygous, and homozygous mutant mice. Our differential gene expression analysis between Mir96Dmdo homozygous mutant HCs compared to wildtype HCs identified 215 upregulated and 428 downregulated genes. Many significantly downregulated genes in Mir96Dmdo homozygous mutant HCs have established roles in HC development and/or known roles in deafness such as Myo15, Myo7a, Ush1c Gfi1, and Ptprq, some of which were not previously identified in other miR-96 datasets. In addition, active modules of   protein-protein interaction networks of significantly downregulated genes in Mir96Dmdo homozygous mutant HCs reveal enrichment in GO terms with biological functions such as sound perception and endocytosis. Genes significantly upregulated in Mir96Dmdo homozygous mutant HCs, which are more likely to be direct targets of miR-96, show higher expression in wildtype supporting cells compared to wildtype HCs, suggesting a role of miR-96 in suppressing non-HC genes during HC development. Finally, all generated HC-specific Mir96Dmdo RNA-seq datasets from this manuscript are now publicly available in the miR-96 specific gEAR profile (https://umgear.org/p?l=miR96).

overall_design: To investigate the role of miR-96 and further understand the transcriptional cascade of miR-96 in inner ear hair cells, we performed bulk RNA-sequencing on pooled cochlear and vestibular hair cells of  postnatal day (P) 1 wildtype, Mir96Dmdo heterozygous, and Mir96Dmdo homozgyous mutant mice (n=3 mice per genotype). Differential gene expression analysis was performed on Mir96Dmdo homozgyous mutant hair cells compared to wildtype controls.

first: Kathleen

last: Gwilliam

first: Michal

last: Sperber

first: Katherine

last: Perry

last: Rose

first: Laura

last: Morrison

first: Nikhil

last: Paladugu

first: Yang

last: Song

first: Beatrice

last: Milon

first: Ran

last: Elkon


title: beta-catenin in dendritic cells negatively regulates CD8 T cell im-mune responses through the immune checkpoint molecule Tim-3

city: Detroit

first: Qing-Sheng

last: Mi

state: Michigan

address: One Ford Place

institute: Henry Ford Health

summary: Recent studies have demonstrated that b-catenin in dendritic cells (DCs) serves as a key mediator in promoting both CD4 and CD8 T cell tolerance, although the mechanisms underlying how b-catenin exerts its functions remains incompletely understood. Here we report that activation of b-catenin leads to the up-regulation of inhibitory molecule T-cell immunoglobulin and mucin domain 3 (Tim-3) in type 1 conventional DCs (cDC1s). Using a cDC1-targeted vaccine model with anti-DEC-205 engineered to express the melanoma antigen--human gp100 (anti-DEC-205-hgp100), we demonstrated that CD11c-b-cateninactive mice exhibited impaired cross-priming and memory re-sponses of gp100-specific CD8 T (Pmel-1) cells upon immunization with anti-DEC-205-hgp100. Sin-gle cell RNA sequencing (scRNA-seq) analysis revealed that b-catenin in DCs negatively regulated transcription programs for effector function and proliferation of primed Pmel-1 cells, correlating with suppressed CD8 T cell immunity in CD11c-b-cateninactive mice. Further experiments showed that treating CD11c-b-cateninactive mice with anti-Tim-3 antibody upon anti-DEC-205-hgp100 vac-cination led to restored cross-priming and memory responses of gp100-specific CD8 T cells, sug-gesting that anti-Tim-3 treatment likely synergizes with DC vaccines to improve their efficacy. In-deed, treating B16F10-bearing mice with DC vaccines using anti-DEC-205-hgp100 in combination with anti-Tim-3 treatment resulted in significantly reduced tumor growth, compared to treatment with DC vaccine alone. Taken together, we have identified the b-catenin/Tim-3 axis as a novel mech-anism to inhibit anti-tumor CD8 T cell immunity, and that combination immunotherapy of a DC-targeted vaccine with anti-Tim-3 treatment leads to improved anti-tumor efficacy.

overall_design: We conducted scRNA-seq on transferred PMel-1 cells (from spleen) primed in WT or CD11c-b-catenin active mice.

first: Chunmei

last: Fu

first: Jie

last: Wang

first: Tianle

last: Ma

first: Congcong

last: Yin

first: Li

last: Zhou

first: Björn

middle: E

last: Clausen

first: Aimin

last: Jiang


title: Xbp1 overexpression in the gut and fat body extend Drosophila lifespan

city: LONDON

first: MengJia

last: Li

state: Please Select

address: flat38 midhopse house, midhope street

institute: University College London

summary: Manipulating the activity of transcription factor (TF) to regulate animal’s longevity is achieved in different species. However, deciphering the pro-longevity transcriptional programme triggered by the TFs activity is challenging. One obstacle is the multifunctional feature of TFs. The physiological functions of single TF could be diverse and tissues-dependent. The other is the gene regulatory network among the TFs, where multiple TFs can have synergetic or antagonistic impact on the same target genes. Here, we show Xbp1s overexpression in gut and fat body can extend lifespan in Drosophila. Importantly, Xbp1s activity triggers distinct transcriptional programmes in the two tissues, and Xbp1s induction in both tissues are beneficial for longevity. Furthermore, we reveal a pro-longevity gene regulatory network in the fat body, where Xbp1s and dFOXO impinge on the same target genes and induce identical transcriptional outcomes. dfoxo overexpression requires Xbp1 to extend lifespan. Lastly,  inducing Xbp1u, the precursor of Xbp1s, can also extend lifespan without triggering massive transcriptome change.

overall_design: S106>Xbp1s RU+/-  S106>Xbp1u RU+/- S106>wdah RU+/- all conditions with gut and fat body samples

first: Mengjia

first: Nazif

last: Alic


title: EGR2 is upregulated in monocyte-derived macrophages in MASH liver

first: Kenichi

last: Asano

summary: This analysis was perfomed to explore the transcription factors that were enhanced in MASH liver macrophages.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for liver mactophages from ND-fed mice and CDA-HFD-fed mice.

first: Ayaka

last: Iwata

first: Akira

last: Nishiyama


title: EGR2 is required for the acquisition of hLAM phenotype by monocyte-derived macrophages in MASH.

summary: This analysis was performed to understand the difference between WT and EGR2 KO macrophages in MASH liver.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for MASH liver mactophages from WT mice and EGR2-KO mice.

first: kenichi


title: Releasing the mitochondrial respiration brake MCJ/DnaJC15 enhances CD8 CAR-T cell therapy efficacy

city: Aurora

first: Tonya

last: Brunetti

state: CO

address: 12800 East 19th Avenue

department: Immunology and Microbiology

institute: University of Colorado Anschutz Medical Campus

summary: Metabolism of chimeric antigen receptor (CAR) T cells is emerging as an important area to improve CAR-T cell therapy in cancer treatment. Mitochondrial respiration is essential for survival and function of CAR-T cells, but developing strategies to specifically enhance mitochondrial respiration has been challenging. Here we identify MCJ/DnaJC15, an endogenous negative regulator of mitochondrial Complex I, as a metabolic target to enhance mitochondrial respiration in CD8 CAR-T cells. Loss of MCJ in CD8 CAR-T cells increases their in vitro and in vivo efficacy against mouse B cell leukemias. MCJ deficiency in TCR- specific CD8 cells also increases their efficacy against solid tumors in vivo. Furthermore, we reveal that human CD8 cells express MCJ and that silencing MCJ expression increases mitochondrial metabolism and anti-tumor activity of human CAR-T cells. Thus, we demonstrate the unique therapeutic potential of targeting MCJ to enhance the metabolism and efficacy of adoptive T cell therapies.

overall_design: MCJ/DnaJC15 is an endogenous negative regulator of Complex I and mitochondrial respiration. The goal of these RNAseq experiments is to characterize the phenotype of mouse WT CD8 CAR-T cells and MCJ KO CD8 CAR-T cells after three expansions with IL-2

first: Meng-Han

last: Wu

first: Felipe

last: Valenca-Pereira

first: Francesca

last: Cendali

first: Emily

middle: L

last: Giddings

first: Catherine

last: Pham-Danis

first: Michael

last: Yarnell

first: Amanda

last: Novak

first: Jorge

last: Henao-Mejia

first: Richard

middle: A

last: Flavell

first: Angelo

last: D’Alessandro

first: M

last: Kohler

first: Mercedes

last: Rincon

first: Scott

last: Thompson


title: Gene expression profiles at the single-nucleus level of the antennae of wild-type and orco mutant Harpegnathos saltator

city: New York

first: Bogdan

last: Sieriebriennikov

state: NY

address: 100 Washington Sq E

institute: New York University

summary: H. saltator orco mutant has a reduced number of cells in the antennae. We performed single-nucleus sequencing of the antennae and compared the mutant to the wild type to identify any differentially abundant cell types.

overall_design: Nuclei were isolated from dissociated antennae of wild-type and orco mutant H. saltator ants by staining with Hoechst and FACS sorting stained particles. Nuclei were analyzed using snRNA-seq.

first: Kayli

last: Sieber

first: Olena

last: Kolumba

first: Jakub

last: Mlejnek

first: Shadi

last: Jafari

first: Hua

last: Yan


title: CD11c and Mincle discriminates hLAMs from moKCs in MASH liver

status: Public on May 01 2024

first: Kenichi

last: Asano

summary: This analysis was performed to understand the difference between subpopulations of monocyte-derived macrophages.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for liver macrohpages.

first: Ayaka

last: Iwata

first: Akira

last: Nishiyama

first: Kenchi


title: Dissecting the Mechanisms of Transcriptional Synergy in Response to Proinflammatory Cytokines in Endothelial Cells

city: Nashville

first: Lindsey

middle: "Joey"

last: Stolze

address: 1211 Medical Center Dr

institute: Vanderbilt University Medical Center

summary: Combinatorial cytokine signaling is implicated in the pathogenesis of diseases of the cardiovascular system ranging from acute combinatorial cytokine signaling observed in complications from SARS-COVID19 infection to chronic combinatorial cytokine signaling observed in atherosclerosis. There remains a large gap of our understanding how combinatorial cytokine signaling interface to drive proinflammatory programs. We hypothesize that stimulation of HAECs with IFNg and TNFa promotes synergistic gene transcription through the coordinated interplay between p65, STAT1, p300/CBP, and BRD4. Our work demonstrates through the generation of novel statistical framework that IFNg and TNFa dual stimulation for one hour results in the synergistic induction of a small subset of proinflammatory genes. Prolonged dual cytokine stimulation results in synergistic levels of protein expression and cell death. Treatment with A-485 and/or JQ1 reduces levels of synergistic gene and protein expression along with improvements in cell viability. Defining synergistic responses and identifying their unique transcriptional dependencies opens the door to understanding how disparate cytokine signaling drives proinflammatory responses and how these responses can be targeted.

overall_design:
teloHAECs were treated for one hour with either vehicle, IFNg, TNFa, or both IFNg and TNFa before cells were harvested for RNA. We then performed compartive gene expression profile analysis between all four conditions.

teloHAECs were treated for one hour with either vehicle, IFNg, TNFa, or both IFNg and TNFa before cells were harvested for to perform ATAC-seq. We then performed comparative anlysis to determine differerntial peaks that were specific to the dual cytokine condition.

Inhibitor treated ATAC: teloHAECs were treated for one hour with vehicle or both IFNg and TNFa (SYN) then either DMSO or an inhibitor (A485 or JQ1) before cells were harvested for to perform ATAC-seq. We then performed comparative anlysis to determine differential peaks. CUTandRUN: teloHAECs were treated for one hour with vehicle or both IFNg and TNFa (SYN) then either DMSO or an inhibitor (A485 or JQ1) before cells were harvested for to perform ATAC-seq. We then performed comparative anlysis to determine differential peaks. CUT&RUN: TELO HAECs were treated for one hour with vehicle or both IFNg and TNFa (SYN) in the presence or absence of DMSO or inhibitors (A485 or JQ1). Cells were fixed in 1% formaldhyde for 1 min before being quenched with glycine. Cells were bound to Concavalin MyOne beads and incubated with antibody for 24 hours. Samples were washed and then pAG-MNase bound for 1 hour at 4C, after which CaCL2 was added and pAG-MNase cleavage went for 30 minutes with samples submerged in ice. DNA was purified via a phenol-chloroform isolation after which the purified DNA was utilized to generate sequencing libraries. DNA libraries were generated using the NEBNext Ultra II DNA Library Prep Kit with the following PCR cycling adjustmentsfor the PCR Enrichment of adaptor ligated DNA  a. 45 sec at 98ºC  b. 15 sec at 98ºC  c. 10 sec at 60ºC  d. Repeat Steps b-c for a total of 14 cycles e. 1 min at 72ºC  final extension. We then performed compartative analysis to determine differential peaks.

first: Ronan

last: Bracken

first: Joey

first: Quanhu

last: Sheng

first: Jonathan

last: Brown


title: Bulk gene expression profiles of adult and pupal antennae in wild-type and orco mutant Harpegnathos saltator


title: Cyclic compressive loading induces a mature meniscal cell phenotype in mesenchymal stem cells with an atelocollagen-based scaffold

city: Suita

first: Shohei

last: Oyama

state: Osaka

address: 2-2 Yamada-oka

department: Department of Musculoskeletal Regenerative Medicine

institute: Osaka University Graduate School of Medicine

summary: we investigated how cyclic compressive loading (CCL) could impact MSCs using three-dimensional cultures in atelocollagen-based meniscal substitute (ACMS). We extracted MSCs from the meniscus, synovium, and articular cartilage, cultured them in three-dimensional cultures, and submitted to CCL for seven days. We then compared the transcriptomes of MSCs treated with and without CCL. Our RNA-seq analysis revealed that CCL induced significant transcriptome changes, significantly affecting chondrocyte-related genes, including SOX9, TGFB1, and PRG4 upregulation. CCL-induced transcriptional differentiation from meniscus progenitors toward mature meniscal cells.

overall_design: We obtained mesenchymal stem cells (MSCs) from meniscus, synovium, and cartilage tissues derived from patients with osteoarthritis in order to compare the characteristics of MSCs among these tissues. Subsequently, to investigate the changes in gene expression induced by cyclic compression loading, the MSCs cultured in three dimensions were subjected to one hour of repeated compressive loading per day for seven days.


title: A culture method with berbamine, a plant alkaloid, enhances CAR-T cell efficacy through modulating cellular metabolism

city: Fujisawa-shi

first: Shin-ichiro

last: Takayanagi

state: Kanagawa

address: 2-26-1, Muraoka-Higashi

department: KIRIN Central Research Institute

institute: Kirin Holdings

summary: Memory T cells demonstrate superior in vivo persistence and antitumor efficacy. However, methods for manufacturing less differentiated T cells are not yet well-established. Here, we show that producing chimeric antigen receptor (CAR)-T cells using berbamine (BBM), a natural compound found in the Chinese herbal medicine Berberis amurensis, enhances the antitumor efficacy of CAR-T cells. BBM is identified through cell-based screening of chemical compounds using induced pluripotent stem cell-derived T cells, leading to improved viability with a memory T cell phenotype. Transcriptomics and metabolomics using stem cell memory T cells reveal that BBM broadly enhances lipid metabolism. Furthermore, the addition of BBM downregulates the phosphorylation of p38 mitogen-activated protein kinase and enhanced mitochondrial respiration. CD19-CAR-T cells cultured with BBM also extend the survival of leukaemia mouse models due to their superior in vivo persistence. This technology offers a straightforward approach to enhancing the antitumor efficacy of CAR-T cells.

overall_design: CD8β+CD45RA+CCR7+CD45RO-CD95+ cells were purified from healthy donor-derived peripheral blood mononuclear cells. Purified cells were cultured in the presence of berbamine (BBM) or water for 12 days.  Then, the cells were prepared for this RNA-sequencing.

first: Sayaka

last: Chuganji

first: Ken

last: Fukumoto

first: Shin

last: Kaneko


title: Gene regulatory landscape of cerebral cortex folding

city: Belfast

first: Aditi

last: Singh

state: Please Select State

address: 97 Lisburn Road, Belfast

department: The Wellcome-Wolfson Institute for Experimental Medicine, SMDBS

institute: Queen's University Belfast

summary: Our brains accommodate a largely folded cerebral cortex that associates to our advanced brain functions. Several theories have been proposed to explain the cortical folding process, reasoning the mechanical forces as neuronal tension in underlying layers, cellular expansion and glial progenitor’s diversity in the OSVZ; but the mechanistic insights and underlying genomics changes causing the appearance of cortical folds is still illusive. Importantly, no studies have attempted to comprehensively characterize the gene regulatory networks underlying cortical folding during development. Here, by using ferret as a model system, we have compared the transcriptomes of germinal layers between sulci and gyri, of different cortical areas and across two developmental stages (E30 & E34), to achieve a comprehensive understanding of the spatio-temporal dynamics of gene expression of cortical progenitors. Furthermore, we have also characterized the epigenetic landscape of germinal layers at E30 and E34, a critical period for their development, to correlate changes in chromatin at promoter and enhancer regions with the observed changes in gene expression. Our preliminary results indicate towards a clear transformational axis of gene regulation between germinal layers. By performing motif analysis of differentially expressed gene (DEGs), we reveal transcription factors which might have a critical role in determining cortical folding. The genes targeted by these factors belong to important pathways implicated in proliferation and neurogenesis. We highlight potential candidates in cortical folding through functional validation studies and acetylation (H3K27ac) levels for these genes correlate with their expression state. Importantly, these are critical for cell adhesion, migration and proliferation processes in-line with previous studies stating that proliferative divisions cause neocortical expansions. Our findings will have strong impact on the clinical interventions for neurological disorders relating to cortical malformation, while at the same time enhancing our understanding of molecular circuitry underlying gyrencephalic brain development and folding.

overall_design: 1. RNA-seq was performed at embryonic day 30 (E30), 34 (E34) in microdissected ferret cortex for gyrus and sulcus regions;   2. H3K27ac (abcam, ab4729) ChIP-seq was performed at embryonic day 30 and 34 in microdissected ferret cortex for gyrus and sulcus regions; 3. E37 ferret embryos previously electroporated at E34 were obtained for scRNAseq library preparation from microdissected ferret cortex.

first: Lucia

last: Del Valle Anton

first: Camino

last: De juan Romero

first: Victor

last: Borrell

first: Vijay

middle: K

last: Tiwari


title: Bulk gene expression profiles of adult antennae in wild-type and orco mutant Harpegnathos saltator

first: Bogdan

last: Sieriebriennikov

summary: H. saltator orco mutant has a reduced number of cells in the antennae. We performed RNA-seq of the antennae and compared the mutant to the wild type to identify any differentially expressed odorant receptor genes.

overall_design: RNA was extracted from the antennae of adult wild-type and orco mutant H. saltator.

first: Kayli

middle: R

last: Sieber

first: Olena

last: Kolumba

first: Jakub

last: Mlejnek

first: Shadi

last: Jafari

first: Hua

last: Yan


title: SNP genotyping array of human naive pluripotent stem cells II

city: San Francisco

first: Akira

last: Kunitomi

state: CA

address: 1650 Owens Street

institute: The J. David Gladstone Institutes

type:
SNP genotyping by SNP array
Genome variation profiling by SNP array

summary: Human pluripotent stem cells have two major pluripotent states, primed and naive, and the heterogeneity among cell lines in each pluripotent state remains a major unresolved problem. We showed that the overexpression of H1FOO-DD, which has a short expression period by fusing the destabilized domain to the maternal-specific linker histone H1FOO, together with OCT4, SOX2, KLF4 and LMYC in human somatic cells improves the quality of reprogramming to primed and naive pluripotency.

overall_design: Genomic DNA from the 30 samples were hybridized onto the Infinium OmniExpress24 v1.4 DNA Analysis Kit (Illumina), and intensities were scanned by iScan (Illumina) following the manufacturer’s protocol.

first: Masaki

last: Nomura


title: Well-ST-seq: cost-effective and cellular level spatial transcriptomics with high RNA capture efficiency

city: Changchun

first: Zhengyang

last: Jin

state: Jilin

address: No. 1163 Xinmin Street

institute: Jilin University

summary: Spatial transcriptomic technologies are promising tools to reveal fine anatomical profiles of tissues. In the case of methodologies utilizing barcoded probe arrays, achieving a balance among probe barcoding complexity, cost, gene capture sensitivity, and spatial resolution is crucial for accelerating the spreading speed of spatial transcriptomic in basic science and clinical work. Here, we developed spatially cellular-level RNA-capture probe arrays using miniaturized microfluidic and microarray technologies. By leveraging the predetermined and cost-effective probe fixation characteristics of this methodology, we significantly reduced the consumable cost of the probe array to $0.31/mm2 and fabrication time to approximately 2 hours. Furthermore, the modification of the RNA-capture probe on sequencing slides by microfluidic chip does not rely on large imaging or printing instruments. Notably, the efficiency of the transcript captured by the probe array is even comparable to conventional single-cell RNA sequencing. Based on this technology, the stacked three-dimensional transcriptome atlas and the spatial cell heterogeneity of mouse brains were successfully visualized. Taken together, we present an experimental and analytical framework for the spatial investigation of mouse brain structures and cell phenotypes.

overall_design: Applying new approach – Well-ST-seq for spatial transcriptomes sequencing to mouse tissues in different resolution (10 μm, 20 μm and 30μm).

first: Nianzuo

last: Yu

first: Shoujun

last: Zhu

first: Zhiying

last: Hu

first: Xiaoduo

last: Tang

first: Jingwen

last: Fang

first: Chongyang

last: Liang

first: Junhu

last: Zhang

first: Bai

last: Yang


title: EGR2 biases the differentiation of liver infiltrating monocytes into hLAMs.

first: Kenichi

last: Asano

summary: This analysis was performed to understand the heterogeneity and differentiation pathway of monocyte-derived macrophages.

overall_design: A single cell RNA-seq analysis of CD45.2+Lin (CD90.2, B220, NK1.1, SiglecF)-CD11b+ liver innate immune cells from one ND-fed WT mouse, one ND-fed EGR2-KO mouse, one CDA-HFD-fed WT mouse, and one CDA-HFD-fed EGR2-KO mouse.

first: Ayala

last: Iwata

first: Shigeyuki

last: Shichino

first: Takao

last: Seki


title: Direct transposition of native DNA for sensitive multimodal single-molecule sequencing IV

first: Arjun

middle: S

last: Nanda

state: California

address: 1700 Owens St

department: Gladstone Institute for Data Science & Biotechnology

institute: Gladstone Institutes / UCSF

summary: Concurrent readout of sequence and base modifications from long unamplified DNA templates by PacBio single-molecule sequencing requires large amounts of input material. Here we adapt Tn5 transposition to introduce hairpin oligonucleotides and fragment (tagment) limiting quantities of DNA for generating PacBio-compatible circular molecules. We developed two methods that implement tagmentation and use 90–99% less input than current protocols: (1) single-molecule real-time sequencing by tagmentation (SMRT-Tag), which allows detection of genetic variation and CpG methylation; and (2) single-molecule adenine-methylated oligonucleosome sequencing assay by tagmentation (SAMOSA-Tag), which uses exogenous adenine methylation to add a third channel for probing chromatin accessibility. SMRT-Tag of 40 ng or more human DNA (approximately 7,000 cell equivalents) yielded data comparable to gold standard whole-genome and bisulfite sequencing. SAMOSA-Tag of 30,000–50,000 nuclei resolved single-fiber chromatin structure, CTCF binding and DNA methylation in patient-derived prostate cancer xenografts and uncovered metastasis-associated global epigenome disorganization. Tagmentation thus promises to enable sensitive, scalable and multimodal single-molecule genomics for diverse basic and clinical applications.

overall_design: Chromatin profiling with m6dA methyltransferase footprinting of mESCs, followed by transposase-mediated PCR-free library construction ex situ (SAMOSA-Tag ex situ) and single molecule sequencing.

first: Ke

last: Wu

first: Sivakanthan

last: Kasinathan

first: Megan

last: Ostrowski

first: Ansuman

last: Satpathy

first: E

last: Sweet-Cordero

first: Hani

last: Goodarzi

first: Vijay

last: Ramani


title: MPRA in primary human macrophages

city: London

first: James

last: Lee

address: The Francis Crick Institute, 1 Midland Rd

institute: The Francis Crick Institute

summary: We investigated an intergenic haplotype on chr21q22, linked to five different inflammatory diseases. We used a functional approach (massively-parallel reporter assay; MPRA) to first identify active enhancers at the locus in primary human macrophages, and then determine if candidate variants within these regulatory regions might alter enhancer activity. In doing so, we discovered a mechanism that orchestrates macrophage responses during chronic inflammation and delineated how the risk haplotype increases expression of the causal gene, ETS2.

sample_organism:
Homo sapiens
synthetic construct

platform_organism:
Homo sapiens
synthetic construct

overall_design: The study consists of a massively parallel reporter assay, in which the minimal promoter in the vector was replaced by the RSV promoter. This was necessary in order for the assay to work in primary macrophages. An oligo library was designed to test the chr21q22 candidate SNPs within the 99% credible set (based on SuSiE fine-mapping of IBD GWAS data) and to tile the region containing these candidate SNPs. Allelic constructs were tiled across each SNP in triplicate, and each was tagged by 30 unique 11nt barcodes. Each tiling construct was tagged by 6 unique barcodes. Transfections were performed into inflammatory (TPP) macrophages from 8 healthy individuals. Raw data are provided as Illumina reads of the 11nt barcode from mRNA extracted 24 hours after transfection, or from 4 replicates of the input MPRA vector library. We also provide tab-delimited files containing the raw barcode counts for each sample and the quantile normalized element counts (sum of barcodes tagging a single genomic sequence).