title: Transcription factor FOXM1 specifies chromatin DNA to extracellular vesicles

city: Changsha

first: Yunsheng

state: Hunan province

address: Tianma Road

institute: Hunan University

summary: Extracellular vesicle DNAs (evDNAs) hold significant diagnostic value for various diseases and facilitate transcellular transfer of genetic material. Our study identifies transcription factor FOXM1 as a mediator for directing chromatin genes or DNA fragments (termed FOXM1-chDNAs) to extracellular vesicles (EVs). FOXM1 binds to MAP1LC3/LC3 in the nucleus, and FOXM1-chDNAs, such as the DUX4 gene and telomere DNA, are designated by FOXM1 binding and translocated to the cytoplasm before being released to EVs through the secretory autophagy during lysosome inhibition (SALI) process involving LC3. Disrupting FOXM1 expression or the SALI process impairs FOXM1-chDNAs incorporation into EVs. FOXM1-chDNAs can be transmitted to recipient cells via EVs and expressed in recipient cells when they carry functional genes. This finding provides an example of how chromatin DNA fragments are specified to EVs by transcription factor FOXM1, revealing its contribution to the formation of evDNAs from nuclear chromatin. It provides a basis for further exploration of the roles of evDNAs in biological processes, such as horizontal gene transfer.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for FOXM1 and LC3 in A549 cells; Extracellular vesicle DNA-sequencing (evDNA-seq) for extracellular vesicles (EVs) from A549 cells

first: Ping

last: Fu

first: Yousong

last: Peng

first: Yongjun

last: Tan


title: Retinoic acid drives intestine-specific adaptation of effector ILC2s originating from distant sites [scRNA-Seq]

status: Public on Sep 15 2023

city: Hamburg

first: Nikhat

last: Shaikh

address: Martinistraße 52

country: Germany

institute: University Medical Center Hamburg-Eppendorf

type: Expression profiling by high throughput sequencing

summary: Adaptation of immune cells to tissue-specific microenvironments is a crucial process in homeostasis and inflammation. Here, we show that murine effector type 2 innate lymphoid cells (ILC2s) from various organs are equally effective in repopulating ILC2 niches in other anatomical locations where they adapt tissue-specific phenotypes of target organs. Single-cell transcriptomics of ILC2 populations revealed upregulation of retinoic acid (RA) signaling in ILC2s during adaptation to the small intestinal microenvironment, and RA signaling mediated reprogramming of kidney effector ILC2s towards the small intestinal phenotype in vitro and in vivo. Inhibition of intestinal ILC2 adaptation by blocking RA signaling impaired worm expulsion during Strongyloides ratti infection, indicating functional importance of ILC2 tissue imprinting. In conclusion, this study highlights that effector ILC2s retain the ability to adapt to changing tissue-specific microenvironments, enabling them to exert tissue-specific functions, such as promoting control of intestinal helminth infections.

overall_design: To investigate transcriptional programs sepcific to kidney and SILP ILC2s and those flexible during ILC2 adaptation to the small-intestinal microenvironment, we performed single-cell RNA sequencing of kidney and SILP ILC2s from donor mice and from those of recipient mice. ILC2s from kidney were transferred into alymphoid mice (recipient mice) and analysed after 2 and 8 weeks of transfer

first: Alex

last: Waterhölter

first: Ann-Christin

last: Gnirck

first: Martina

last: Becker

first: Virginia

last: Adamiak

first: Lena

last: Henneken

first: Malte

last: Wunderlich

first: Wiebke

last: Hartmann

first: Lara

last: Linnemann

first: Tobias

middle: B

last: Huber

first: Christian

middle: F

last: Krebs

first: Ulf

last: Panzer

first: Richard

middle: M

last: Locksley

first: Christoph

last: Wilhelm

first: Minka

last: Breloer

first: Jan-Eric

last: Turner


title: Stimulation of immunity-linked genes by membrane disruption is linked to Golgi function and the ARF-1 GTPase

city: Worcester

first: Amy

last: Walker

state: MA

address: 373 Plantation Street

country: USA

institute: UMass Medical School

summary: Immunity-linked genes (ILGs) are activated by pathogens but also may respond to imbalances in lipids. Why pathogen attack and metabolic changes both impact ILG activation is unclear. We find that ILGs are activated when membrane phosphatidylcholine ratios change in secretory organelles in C. elegans. RNAi targeting of the ADP-ribosylation factor ARF-1, which disrupts the Golgi, also activates ILG expression, suggesting that activation of this membrane stress response could occur outside the ER. Our data argue that ILG upregulation is a coordinated response to changes in trafficking resulting from intrinsic cues (changes in membrane lipids) or extrinsic stimulation (increased secretion during immune response). Indeed, a focused RNAi screen of ILGs uncovered defects in secretion of two GFP reporters as well as accumulation of a pathogen-responsive CUB-domain fusion protein. These results also suggests that genes shared between the classical pathogen responses and lipid stress may act to counteract stress on secretory function.

overall_design:
Lysis of young adult C. elegans was performed in 0.5% SDS, 5% β-ME, 10 mM EDTA, 10 mM Tris-HCl ph7.4, 0.5 mg/ml Proteinase K, before purification of RNA by TRI-Reagent (Sigma). cDNA was produced with Transcriptor First-strand cDNA kits (Roche), and RT-PCR was performed using Kappa SYBR Green 2X Mastermix.
RNA for sequencing was purified using RNAeasy columns (Qiagen). RNA sequencing (including library construction was performed by BGI (Hong Kong). Reads were analyzed through the Dolphin analysis platform (https://dolphin.umassmed.edu/), using DeSeq to identify differentially expressed genes. Gene set enrichment was performed using WormCat (www.wormcat.com).

first: Matthew

middle: J

last: Fanelli

first: Christofer

last: Welsh

first: Dominique

middle: S

last: Lui

first: Lorissa

last: Smulan

middle: K


title: Circular extrachromosomal DNA promotes tumor heterogeneity in high-risk medulloblastomas [scRNA+ATAC-seq]

city: La Jolla

first: Lukas

last: Chavez

state: CA

address: 10901 North Torrey Pines Road

department: Cancer Genome and Epigenetics Program, NCI-Designated Cancer Center

institute: Sanford Burnham Prebys Medical Discovery Institute

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Circular extrachromosomal DNA (ecDNA) in patient tumor genomes is an important driver of oncogenic gene expression, evolution of drug resistance, and poor patient outcomes. Applying computational methods for detection and reconstruction of ecDNA across a retrospective cohort of 481 medulloblastoma (MB) tumors from 465 patients, we identify circular ecDNA in 82 patients (18%). Patients with ecDNA+ MB were more than twice as likely to relapse and three times as likely to die within 5 years of diagnosis. Individual tumors harbor multiple ecDNA lineages, each containing distinct amplified oncogenes. Multimodal sequencing, imaging, and CRISPR inhibition experiments in MB models reveal intratumoral heterogeneity of ecDNA copy number per cell and frequent putative "enhancer rewiring" events on ecDNA. This study reveals the frequency and diversity of ecDNA in a subset of highly aggressive MB tumors, and suggests copy number heterogeneity and enhancer rewiring as clinically relevant features of ecDNA in MB.

overall_design: 10X Multiome ATAC and Gene Expression single-nucleus sequencing of primary SHH MB tumor RCMB56-ht.

first: Owen

last: Chapman

first: Jill

middle: P

last: Mesirov


title: Comprehensive comparison of female germ cell development in vitro and in vivo identifies epigenetic gene regulation crucial for oocyte development and embryonic competence [scRNA-seq]

city: Basel

first: Antoine

last: Peters

address: Maulbeerstrasse 66

country: Switzerland

institute: Friedrich Miescher Institute

summary: Single cell transcriptomics and functional assessment in a stage-specific manner provides a comprehensive comparison of entire female germ cell development in vitro and in vivo.

overall_design: To identify possible causes underlying the overall low oogenic and embryonic competence of in vitro developed oocytes, we performed extensive comparative transcriptome analysis between in vivo and in vitro generated germ cells, at different stages of their development.

first: Eishi

last: Aizawa

first: Evgeniy

middle: A

last: Ozonov

first: Yumiko

last: Kawamura

middle: H

first: Anton

last: Wutz


title: MAF Amplification licenses Estrogen Receptor α to Drive Breast Cancer Metastasis [ER ChIP-seq]

city: Barcelona

first: Adria

last: Caballe

address: C/ Baldiri Reixac 10

department: biostatistics

country: Spain

institute: IRB

type: Genome binding/occupancy profiling by SNP array

summary: We performed genome-wide mapping of estrogen receptor (ER) binding sites in control and MAF-overexpressing MCF7 cells to assess the consequences of estrogen (E2) stimulation upon metastasic MAF expression. To this end, we cultured MCF7 cells in hormone-deprived (HD) medium for 72 h and then E2 or vehicle was added for 1h prior to chromatin immunoprecipitation (ChIP). Samples were generated in triplicate. We report that E2 induces extensive ER recruitment to chromatin and that ER-binding is gained and expanded upon MAF overexpression.

overall_design: Examination of ER binding sites in control and MAF-overexpressing MCF7 cells before and after E2 stimulation.

first: Alicia

last: Llorente

first: Cecilia

last: Ballaré

first: Enrique

last: Blanco

first: Adrià

last: Caballé

first: Roger

last: Gomis

first: Luciano

last: Di Croce


title: Circular extrachromosomal DNA promotes tumor heterogeneity in high-risk medulloblastomas [pooled CRISPRi]

overall_design: Pooled CRISPRi proliferation screen of 32,530 sgRNAs targeting 645 accessible loci on the D458 ecDNA. Cells were lentivirally transformed with CRISPR-dCas9-KRAB and relative sgRNA abundances were counted before and after 21 days.


title: MAF Amplification licenses Estrogen Receptor α to Drive Breast Cancer Metastasis.[ChIP-Seq]

summary: We performed genome-wide mapping of H3k27ac and H3K4me3 sites in control and MAF-overexpressing MCF7 cells to assess the consequences of estrogen (E2) stimulation and MAF overexpression recruitment to chromatin. To this end, we cultured MCF7 cells in hormone-deprived (HD) medium for 72 h and then E2 or vehicle was added for 1h prior to chromatin immunoprecipitation (ChIP). Samples were generated in triplicate.

overall_design: Examination of H3K27ac and H3K4me3 sites in control and MAF-overexpressing MCF7 cells before and after E2 stimulation.

first: Teresa

last: Blasco


title: MAF Amplification licenses Estrogen Receptor α to Drive Breast Cancer Metastasis.

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

overall_design: Refer to individual Series


title: MAF Amplification licenses Estrogen Receptor α to Drive Breast Cancer Metastasis [dataset 1]

summary: We performed mRNA profiling of control and MAF-overexpressing MCF7 cells to assess the transcriptional consequences of estrogen receptor (ER) activation upon metastatic MAF expression in the presence or absence or ER (using a protac for ER degradation). To this end, we cultured MCF7 cells in hormone-deprived (HD) medium for 72 h. At this point, we treated the cells with vehice or ER-protac for 24h and then estrogen (E2) or vehicle was added for 6h prior to RNA extraction. Samples were generated in triplicate. We report that MAF supports the expression of genes involved in metastatic functions and that E2 treatment and ER expression correlates with E2 early and late gene responses and cell cycle regulation. Additionally, a set of MAF and ER-dependent gene responses was identified. Our results show that MAF expression causes an expansion of ER-mediated transcriptional events in MCF7 cells exposed to E2.

overall_design: To assess gene expression changes governed by MAF and estrogen receptor (ER), control or MAF-overexpressing MCF7 cells were maintained in hormone-deprived medium for 72 h. Then, ER-protac or vehicle was added for  24h. The day after E2 or vehicle was added for 6h prior to RNA extraction and mRNA profiling.

first: Maria

middle: T

middle: R


title: MAF Amplification licenses Estrogen Receptor α to Drive Breast Cancer Metastasis [dataset 2]

summary: We mapped chromatin accessibility on control and MAF-overexpressing MCF7 cells to assess the consequences of estrogen (E2, estrogen cepetor (ER) and metastatic MAF expression on the chromatin landscape. To this end, we cultured MCF7 cells in hormone-deprived (HD) medium for 72 h and a specific ER-protac or vehicle was added for 24h for ER degradation.  The day after, estrogen (E2) or vehicle was added for 1h prior to DNA purification. Samples were generated in duplicate. We report changes in chromaitn accessibility depending on both MAF expression and E2 stimulation.

overall_design: In order to assess changes in chromatin accessibility governed by MAF, ER and estrogen (E2), control or MAF-overexpressing MCF7 cells were maintained in hormone-deprived medium for 72 h. Then, an ER-protac or vehicle was added for 24h and cells were stimulated with E2 or vehicle for 1h. DNA was purified for ATAC-seq analysis


title: DUX4-r neomorphic activity depending on GTF2I in acute lymphoblastic leukemia [RNA-seq]

city: Milan

first: Davide

last: Gabellini

address: Via Olgettina 60

department: Division of Genetics and Cell Biology

country: Italy

institute: SAN RAFFAELE SCIENTIFIC INSTITUTE

summary: Translocations producing rearranged versions of the transcription factor DUX4 (DUX4-r) are one of the most frequent causes of B-cell acute lymphoblastic leukemia (B-ALL). DUX4-r retains the DNA binding domain of wt DUX4, but is truncated on the C-terminal transcription activation domain. The precise mechanism through which DUX4-r causes leukemia is unknown and no targeted therapy is currently available. We found that the rearrangement leads to both a loss and a gain of function in DUX4-r. Loss of CBP/EP300 transcriptional co-activators interaction and inability to bind and activate repressed chromatin. Gain of interaction with the transcription factor GTF2I, which redirects DUX4-r toward leukemogenic targets. Importantly, this neomorphic activity exposes an Achilles' heel whereby DUX4-r positive leukemia cells are exquisitely sensitive to GTF2I targeting, which inhibits DUX4-r leukemogenic activity. Our work elucidates the molecular mechanism through which DUX4-r causes leukemia and suggest a possible therapeutic avenue tailored to this B-ALL subtype.

overall_design: Examination of transcriptomic differences between REH cells expressing WT DUX4, DUX4- DUX4-IGH and DUX4-del50

first: Daniele

last: Campolungo

first: Anna

last: Tascini


title: Comprehensive comparison of female germ cell development in vitro and in vivo identifies epigenetic gene regulation crucial for oocyte development and embryonic competence [bulk RNA-seq]

first: Antoine

summary: NOT PROVIDED; REQUESTED

overall_design: We performed bulk RNA-Seq analysis of BVSC-iPSC-derived PGCLCs at day 8 of in vitro culture (termed d6PGCLC, representing PGCLCs 6 days after initiating in vitro differentiation from EpiLCs) and PGCs isolated from E12.5 female gonads by FACS for both SSEA1 and integrin β3 expression.


title: Effect of omipalisib on transcriptome and signaling pathways in leukemic cell line OCI-AML3

city: Taipei

first: Yu-Hsuan

last: Fu

address: No.1, Chang-Te St

department: Department of Clinical Laboratory Sciences and Medical Biotechnology

country: Taiwan

institute: National Taiwan University

summary: To identify molecular effects of the dual PI3K/mTOR inhibitor omipalisib (GSK2126458), we applied gene expression profiling in human leukemic cell lines OCI-AML3 using Whole Transcriptome Shotgun Sequencing (RNA-seq). After OCI-AML3 cell were treated with 50 nM omipalisib for 24 h, differentially expressed genes (DEGs) of 1245 genes upregulated and 811 were downregulated were found compared with vehicle group. Down-regulated genes enriched in cell cycle, DNA metabolic process, PI3K/AKT/mTOR signaling, MYC pathway, E2F pathway, Gly, Ser and Thr metabolism, Cys and Met metabolism, carbon metabolism, oxidative phosphorylation, mitochondrial biogenesis and respiratory electron transporter chain.

overall_design: Gene expression in OCI-AML3 was measured after exposure for 24 h to 50 nM omipalisib (GSK2126458) or to the vehicle control (DMSO).

first: Chi-Yang

last: Tseng

first: Liang-In

last: Lin


title: MAF Amplification licenses Estrogen Receptor α to Drive Breast Cancer Metastasis [RNA-seq]

summary: We performed mRNA profiling of control and MAF-overexpressing MCF7 cells to assess the transcriptional consequences of estrogen receptor (ER) activation upon metastatic MAF expression. To this end, we cultured MCF7 cells in hormone-deprived (HD) medium for 72 h and then estrogen (E2) or vehicle was added for 6h prior to RNA extraction. Samples were generated in triplicate. We report that MAF supports the expression of genes involved in metastatic functions and that E2 treatment correlates with E2 early and late gene responses and cell cycle regulation. Additionally, a set of MAF and E2-dependent gene responses was identified. Our results show that MAF expression causes an expansion of ER-mediated transcriptional events in MCF7 cells exposed to E2.  Additionally, we silenced KDM1A expression in MAF-overexpressing MCF7 cells to test whether this histone demethylase mediates the transcriptional consequences of MAF overexpression. Samples with KDM1A knockdown were generated in duplicates. We report that KDM1A partially regulates MAF/E2-dependent gene responses.

overall_design: To assess gene expression changes governed by MAF and estrogen (E2), control or MAF-overexpressing MCF7 cells were maintained in hormone-deprived medium for 72 h. Then, E2 or vehicle was added for 6h prior to RNA extraction and mRNA profiling.  To evaluate the effect of KDM1A expression on the regulation of MAF/E2-dependent gene responses, expression of the histone demethylase was silenced in MAF-overexpressing cells using shRNA followed by selection with 2µg/mL puromycin for 72 h. Then cells were cultured in hormone-deprived medium before E2 stimulation as previously explained.


title: MAF Amplification licenses Estrogen Receptor α to Drive Breast Cancer Metastasis


title: Modular Pooled Discovery of Synthetic Knockin Sequences to Program Durable Cell Therapies [scRNA-seq]

status: Public on Sep 14 2023

city: San Francisco

first: Franziska

last: Blaeschke

address: 1650 Owens Street

institute: Gladstone Institutes

summary: Chronic stimulation can cause T cell dysfunction and limit efficacy of cellular immunotherapies. CRISPR screens have nominated gene targets for engineered T cells, but improved methods are required to compare large numbers of synthetic knockin sequences to reprogram cell functions. Here, we developed Modular Pooled Knockin Screening (ModPoKI), an adaptable platform for modular construction of DNA knockin libraries using barcoded multicistronic adaptors. We built two ModPoKI libraries of 100 transcription factors (TFs) and 129 natural and synthetic surface receptors. Over 30 ModPoKI screens across human TCR and CAR T cells in diverse conditions identified a transcription factor AP4 (TFAP4) construct that enhanced fitness of chronically-stimulated CAR T cells and anti-cancer function in vitro and in vivo. ModPoKI’s modularity allowed us to generate a ~10,000-member library of TF combinations. Non-viral knockin of a combined BATF-TFAP4 polycistronic construct further enhanced function. ModPoKI facilitates discovery of complex gene constructs to program cellular functions.

overall_design: ModPoKI-Seq (scRNA-Seq with barcode assignment) of T cells after knockin of NY-ESO-1 TCR and TF library

first: Yan

middle: Y

last: Chen

first: Ryan

last: Apathy

first: Bence

last: Daniel

first: Andy

first: Peixin

first: Katalin

last: Sandor

first: Wenxi

last: Zhang

first: Zhongmei

last: Li

first: Cody

last: Mowery

first: Tori

middle: N

last: Yamamoto

first: William

last: Nyberg

first: Angela

last: To

first: Ruby

last: Yu

first: Raymund

last: Bueno

first: Min

middle: C

last: Kim

first: Ralf

last: Schmidt

first: Daniel

last: Goodman

first: Tobias

last: Feuchtinger

first: Justin

last: Eyquem

first: Chun

last: Ye

first: Julia

last: Carnevale

first: Ansuman

last: Satpathy

first: Eric

last: Shifrut

first: Theodore

middle: L

last: Roth

first: Alexander

last: Marson


title: MAF Amplification licenses Estrogen Receptor α to Drive Breast Cancer Metastasis [MAF ChIP-seq]

first: Adria

last: Caballe

summary: We performed genome-wide mapping of MAF binding sites in control and MAF-overexpressing MCF7 cells to assess the consequences of estrogen (E2) stimulation on MAF recruitment to chromatin. To this end, we cultured MCF7 cells in hormone-deprived (HD) medium for 72 h and then E2 or vehicle was added for 1h prior to chromatin immunoprecipitation (ChIP). Samples were generated in triplicate. We report that MAF binding is largely independent of E2

overall_design: Examination of MAF binding sites in control and MAF-overexpressing MCF7 cells before and after E2 stimulation.

first: Alicia

last: Llorente

first: Cecilia

last: Ballaré

first: Enrique

last: Blanco

first: Adrià

last: Caballé

first: Roger

last: Gomis

first: Luciano

last: Di Croce


title: MAF Amplification licenses Estrogen Receptor α to Drive Breast Cancer Metastasis [ATAC-seq]

summary: We mapped chromatin accessibility on control and MAF-overexpressing MCF7 cells to assess the consequences of estrogen (E2) and metastatic MAF expression on the chromatin landscape. To this end, we cultured MCF7 cells in hormone-deprived (HD) medium for 72 h and estrogen (E2) or vehicle was added for 1h prior to DNA purification. Samples were generated in quadruplicate. We report changes in chromaitn accessibility depending on both MAF expression and E2 stimulation.

overall_design: In order to assess changes in chromatin accessibility governed by MAF and estrogen (E2), control or MAF-overexpressing MCF7 cells were maintained in hormone-deprived medium for 72 h. Then, E2 or vehicle was added for 1h and DNA was purified for ATAC-seq analysis


title: Gene expression profiling and the effect of HDAC9 in cisplatin resistance of esophageal squamous cell carcinoma

city: Shantou

first: Yang

last: Chen

address: No.69, Dongxia North Road, Guangxia New Town, Shantou, Guangdong,

country: China

institute: Second Affiliated Hospital, Shantou University Medical College,

summary: Esophageal cancer is one of the most serious health problems globally because of its high incidence and mortality rate. Cisplatin (CDDP) is the primary treatment but limited by the drug resistance. However, the mechanism of cisplatin resistance remains unclear. Herein, the CDDP-resistant cell line TE-1/CDDP was established to compared with TE-1, the expression profiles of circRNAs   mRNA  and lncRNA were compared between TE-1/CDDP and TE-1 using microarray analysis, and used this as a basis for exploring possible mechanisms of drug resistance in genes such as HDAC9.

overall_design: We establish CDDP-resistant strains of TE-1, the samples were hybridized using Hybridization Ove for 17 hours at 65°C.T he chips were then scanned with an Sure Scan Dx scanner.


title: Circular extrachromosomal DNA promotes tumor heterogeneity in high-risk medulloblastomas [Hi-C]

first: Lukas

last: Chavez

overall_design: Bulk Hi-C of frozen primary medulloblastomas (MB***). Hi-C of fresh patient-derived xenograft (PDX) mouse model (RCMB56). Hi-C of cell line D458.

first: Owen

last: Chapman

first: Jill

last: Mesirov


title: Single cell insights into epithelial morphogenesis in the neonatal mouse uterus

city: Pullman

first: Kimberly

last: Davenport

state: WA

address: 100 Dairy Rd

department: Animal Science

institute: Washington State University

summary: The uterus is an essential organ for reproduction in mammals, however the cellular and molecular mechanisms that direct its development remain insufficiently understood. To fill this knowledge gap, the epithelium of the neonatal mouse uterus was isolated and subjected to single cell transcriptome (scRNA-seq) analysis. Substantial differences in transcriptional profiles and cell types were evident as the epithelium developed from postnatal days 1 to 15. Both the undifferentiated epithelium and determined luminal epithelium were heterogeneous and contained several different cell clusters. Novel glandular epithelial-specific genes (Gas6 and Cited4) were validated by in situ hybridization during uterine development. Trajectory analyses provided a framework for understanding epithelium maturation and differentiation of the glandular epithelium. A candidate set of transcription factors and gene regulatory networks were identified that regulate epithelium lineage specification and morphogenesis. This atlas provides foundational insights important for future mechanistic studies focused on uterine epithelium morphogenesis during the critical window of postnatal development.

overall_design: Nuclei were isolated and libraries prepared following the 10X Genomics platform protocols

first: Thomas

middle: E

last: Spencer

first: Makenzie

last: Lowke

first: Pramod

last: Dhakal

first: Andrew

last: Kelleher


title: Comparative gene expression profiling analysis of RNA-seq data for bovine macrophages infected with two Myocbacterium bovis strains of variable virulence and co-cultured with autologous lymphocytes.

city: CABA

first: Maria

middle: Mercedes

last: Bigi

address: Paraguay 2155 piso 5 M1

country: Argentina

institute: INSTITUTO DE INVESTIGACIONES BIOMEDICAS

summary: In this study we compared the transcriptome of bovine macrophages infected with two Mycobacterium bovis strains of variable virulence, co-cultured with autologous lymphocytes. We used the highly virulent M. bovis strain called Mb04-303 and the attenuated Mb534 strain. We first observed that only the infection of bovine macrophages with the virulent strain, Mb04-303, induced in peripheral bovine mononuclear cells a powerful innate immune response capable of controlling the intracellular mycobacterial replication.By RNAseq analysis we found that infections with Mb04-303 downregulated the KEAP1-NFE2L2 pathway that encodes a transcriptional factor involved in antioxidant genes and inflammasome activation and upregulated the type 1 interferon signalling pathway, compared to the infections with the attenuated Mb534 strain. T

overall_design:
In this study macrophages infected with Mb04-303 showed reduced replication only when autologous lymphocyte were added to the infected macfophages, while macrophages infected with the attenuated M. bovis strain, Mb534, showed low bacterial loads in both conditions, with and without autologous lymphocytes.
We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different co-culturres after 24 hs post infection.

first: Federico

first: María


title: Modular Pooled Discovery of Synthetic Knockin Sequences to Program Durable Cell Therapies [ChIP-Seq]

first: Franziska

last: Blaeschke

overall_design: ChIP-Sequencing with BATF and TFAP4 antibodies using primary human GD2 CAR T cells with BATF, TFAP4, BATF-TFAP4 or control (tNGFR) knockin

first: Yan

first: Ryan

last: Apathy

first: Bence

last: Daniel

first: Andy

first: Peixin

first: Katalin

last: Sandor

first: Wenxi

last: Zhang

first: Zhongmei

last: Li

first: Cody

last: Mowery

first: Tori

last: Yamamoto

first: William

last: Nyberg

first: Angela

last: To

first: Ruby

last: Yu

first: Raymund

last: Bueno

first: Min

last: Kim

first: Ralf

last: Schmidt

first: Daniel

middle: B

last: Goodman

first: Tobias

last: Feuchtinger

first: Justin

last: Eyquem

first: Chun

last: Ye

first: Julia

last: Carnevale

first: Ansuman

last: Satpathy

first: Eric

last: Shifrut

first: Theodore

last: Roth

first: Alexander

last: Marson


title: Temporal chromatin accessibility changes define transcriptional states essential for osteosarcoma metastasis

city: Cleveland

first: Peter

last: Scacheri

state: OH

address: Biomedical Research, 2109 Adelbert Rd Building, 6th FL, Room 647

department: Genetics and Genome Sciences

institute: Case Western Reserve University School of Medicine


title: Circular extrachromosomal DNA promotes tumor heterogeneity in high-risk medulloblastomas

first: Lukas

last: Chavez


title: In vivo-like nearest neighbor parameters accurately predict fractional RNA base-pairing in cells

city: University Park

first: Elizabeth

last: Jolley

state: PA

address: 239 Chemistry Bldg, Pennsylvania State University

institute: Pennsylvania State University

summary: In this study, we conducted a thermodynamic analysis of RNA helix stability in the Eco80 artificial cytoplasm, which mimics in vivo conditions. Eco80 contains 80% of Escherichia coli metabolites, with biological concentrations of metal ions including 2 mM free Mg2+ and 29 mM metabolite-chelated Mg2+. We determined a set of Watson-Crick free energy nearest-neighbor parameters in Eco80 and found that helices are less stable by ∆∆Go37 ~ 1 kcal/mol in comparison to the traditional 1 M NaCl condition. Analysis indicates that Eco80 reduces the stability of three nearest-neighbor pairs. We applied this information to update the nearest-neighbor model using the RNAstructure package. We determined that our in vivo-like corrections have minimal effects on the prediction of RNA secondary structures determined in vitro and in silico. In contrast, in vivo-like corrections markedly improve prediction of fractional RNA base pairing in E. coli, as benchmarked with in vivo RNA chemical probing data using both DMS and EDC. In summary, our thermodynamic and chemical probing analyses of RNA helices in Eco80 indicate that RNA secondary structures are less stable in cells than in artificially stable in vitro buffer conditions such as 1 M NaCl.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: Structure-seq2 libraries constructed from RNA extracted from Escherichia coli grown to mid-exponential phase at 37 °C in LB media. Control samples, DMS-treated samples, and EDC-treated samples were prepared as biological replicates. Libraries were sequenced on an Illumina NextSeq platform in single-end mode.

first: Jacob

last: Sieg

first: Melanie

last: Hout

first: Paul

last: Babitzke

first: Philip

last: Bevilacqua


title: ACD15, ACD21 and SLN regulate accumulation and mobility of MBD6 to silence genes and transposable elements

city: Los Angeles

first: Lucia

last: Ichino

address: 610 Charles E Young Dr East

department: MCDB

institute: University of California, Los Angeles


title: ACD15, ACD21 and SLN regulate accumulation and mobility of MBD6 to silence genes and transposable elements [RNA-seq]

summary: DNA methylation mediates silencing of transposable elements and genes in part via recruitment of the Arabidopsis MBD5/6 complex, which contains the methyl-CpG-binding domain (MBD) proteins MBD5 and MBD6, and the J-domain containing protein SILENZIO (SLN). Here we characterize two additional complex members: α-crystalline domain containing proteins ACD15 and ACD21. We show that they are necessary for gene silencing, bridge SLN to the complex, and promote higher order multimerization of MBD5/6 complexes within heterochromatin. These complexes are also highly dynamic, with the mobility of complex components regulated by the activity of SLN. Using a dCas9 system, we demonstrate that tethering the ACDs to an ectopic site outside of heterochromatin can drive massive accumulation of MBD5/6 complexes into large nuclear bodies. These results demonstrate that ACD15 and ACD21 are critical components of gene silencing complexes that act to drive the formation of higher order, dynamic assemblies.

overall_design: This submission includes 35 RNA-seq libraries.

first: Brandon

last: Boone

first: Shuya

last: Wang

first: Jason

last: Gardiner

first: Jaewon

last: Yun

first: Yasaman

last: Jami-Alahmadi

first: Jihui

last: Sha

first: Cristy

last: Mendoza

first: Bailey

last: Steelman

first: Aliya

last: van Aardenne

first: Sophia

last: Kira-Lucas

first: Isabelle

last: Trentchev

first: James

last: Wohlschlegel

first: Steven

last: Jacobsen


title: ACD15, ACD21 and SLN regulate accumulation and mobility of MBD6 to silence genes and transposable elements [ChIP-seq]

overall_design: This submission includes 11 ChIP-seq libraries.


title: Designing a single cell ATAC-Seq (scATAC-Seq) dataset to validate long read RNA-Seq isoforms and benchmark scATAC-Seq analysis methods II

city: Parkville

first: Matthew

last: Ritchie

state: Victoria

address: 1G Royal Parade

country: Australia

institute: The Walter and Eliza Hall Institute of Medical Research

summary: Single cell sequencing technology has been widely used for understanding the heterogeneity of complex tissue and for identifying novel cell types or cell states. Previous efforts of single cell profiling are mostly performed by measuring transcriptomes using single cell RNA sequencing (scRNA-seq). scRNA-seq is relatively well developed and around 500 analysis tools are currently available for performing different tasks. In the past five years, assays for profiling the single cell chromatin accessibility landscape have emerged and provide extra information about gene regulation at the epigenetic level. Due to its simplicity and sensitivity, single cell Assays for Transposase-Accessible Chromatin using sequencing (scATAC-seq) is widely used to obtain chromatin accessibility. This data will be used to comprehensively evaluate scATAC-seq data analysis tools and gaps in analysis workflows together with publicly available bulk ATAC-Seq and scATAC-seq data using optimised universal evaluation metrics. Furthermore, this data will be used to validate novel isoforms identified from long-read scRNA-Seq study.

overall_design: Our experiment utilized the 5 human lung adenocarcinoma cell lines H2228, H1975, A549, H838 and HCC827. For the single cell designs, the five cell lines were mixed equally and processed by 10X Chromium.

first: Haoyu

last: Yang

first: Shanika

last: Amarasinghe

first: Jafar

last: Jabbari

first: Daniel

last: Brown

first: Mei

last: Du


title: Modular Pooled Discovery of Synthetic Knockin Sequences to Program Durable Cell Therapies [RNA-2]

first: Franziska

last: Blaeschke

overall_design: RNA-Sequencing using primary human GD2 CAR T cells with various knockin conditions on day 6, 7, 14 or 15 after electroporation. Day 15 condition got stimulated with target cells.

first: Yan

last: Chen

first: Ryan

last: Apathy

first: Bence

last: Daniel

first: Andy

first: Peixin

first: Katalin

last: Sandor

first: Wenxi

last: Zhang

first: Zhongmei

last: Li

first: Cody

last: Mowery

first: Tori

last: Yamamoto

first: William

last: Nyberg

first: Angela

last: To

first: Ruby

last: Yu

first: Raymund

last: Bueno

first: Min

last: Kim

first: Ralf

last: Schmidt

last: Goodman

first: Tobias

last: Feuchtinger

first: Justin

last: Eyquem

first: Chun

last: Ye

first: Julia

last: Carnevale

first: Ansuman

last: Satpathy

first: Eric

last: Shifrut

first: Theodore

last: Roth

first: Alexander

last: Marson


title: Real-time quantitative PCR analysis of rat's brain mRNAs

status: Public on Sep 13 2023

city: Moscow

first: Diana

last: Salikhova

address: Miklukho-Maklaya, 8

department: Institute of molecular and cellular medecine

country: Russia

institute: RUDN University

summary: We investigated the therapeutic effect of the extracellular vesicles secreted by glial progenitor cells (GPC-EV) derived from human induced pluripotent stem cell in a traumatic brain injury model. The injury was modeled using the dosed concussion to the open brain method. The male Wistar rats were performed traumatic brain injury (TBI), and after 24 hours animals were randomly divided into two groups: group 1  — rats with intranasal administration of PBS (control), and group 2  — rats with similar administration of extracellular visicles (EV) derived from glial progenitor cells. PCR analysis of brain tissues ( cortex, striatum, hippocampus) were performed on postoperative day 7 and 14 to investigate genes associated with apoptosis, and inflammation.

overall_design: qPCR gene expression profiling.  The brain tissue was collected at 7 and 14 days post-TBI, as indicated in the summary.

middle: I


title: STAT4 promotes Bhlhe40 induction to drive protective IFN-g from natural killer cells during viral infection [STAT4 CUT&RUN]

city: New York

first: Hyunu

state: NY

address: 417 E 68th St

department: Immunology

institute: Memorial Sloan Kettering Cancer Center

summary: NK cells represent a cellular component of the mammalian innate immune system, and mount rapid responses against viral infection, including the secretion of the potent anti-viral effector cytokine IFN-g. Following mouse cytomegalovirus (MCMV) infection, Bhlhe40 was the most highly induced transcription factor in NK cells among the basic helix-loop-helix family. Bhlhe40 upregulation in NK cells depended upon IL-12 and IL-18 signals, with the promoter of Bhlhe40 enriched for STAT4 and the permissive histone H3K4me3, and STAT4-deficient NK cells showing an impairment of Bhlhe40 induction and diminished H3K4me3. Transcriptomic and protein analysis of Bhlhe40-deficient NK cells revealed a defect in IFN-g production during MCMV infection, resulting in diminished protective immunity following viral challenge. Finally, we provide evidence that Bhlhe40 directly promotes IFN-g by binding throughout the Ifng loci in activated NK cells. Thus, our study reveals how STAT4-mediated control of Bhlhe40 drives protective IFN-g secretion by NK cells during viral infection.

overall_design:
To investigate the binding sites of STAT4 in NK cells during homeostasis and 3 hour IL-12 + IL-18 cytokine stimulation, we performed STAT4 CUT&RUN using WT and STAT4-/-(STAT4KO) mice.
All cells for each conditions were sorted and cultured in complete IMDM + IL-15 media for 5 days prior. These cells were then washed and incubated in complete IMDM media without IL-15 for 1 hour prior to addition of appropriate stimulus, unstimulated or 3 hour IL-12 + IL-18. After stimulation, these CD49b+ NK cells were FACS sorted and CUT&RUN was performed, where replicates are biologically paired across stimulation conditions.

first: Joseph

last: Sun


title: Modular Pooled Discovery of Synthetic Knockin Sequences to Program Durable Cell Therapies

first: Franziska

last: Blaeschke

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

overall_design: Refer to individual Series


title: Modular Pooled Discovery of Synthetic Knockin Sequences to Program Durable Cell Therapies [RNA-1]

overall_design: RNA-Sequencing using primary human GD2 CAR T cells vs CD19 CAR T cells on day 6 vs day 15 after electroporation (tNGFR control knockin)

first: Yan

last: Chen

first: Ryan

last: Apathy

first: Bence

last: Daniel

first: Andy

first: Peixin

first: Katalin

last: Sandor

first: Wenxi

last: Zhang

first: Zhongmei

last: Li

first: Cody

last: Mowery

first: Tori

last: Yamamoto

first: William

last: Nyberg

first: Angela

last: To

first: Daniel


title: STAT4 promotes Bhlhe40 induction to drive protective IFN-g from natural killer cells during viral infection [RNA-Seq]

last: Kim

overall_design:
To investigate the function of Bhlhe40 in NK cells during homeostasis and acute MCMV infection, we generated mixed bone marrow chimera from WT and Bhlhe40-/- mice into irradiated hosts.
RNA-seq was performed on FACS sorted Ly49H+ NK cells at d0 and d2 in vivo MCMV infection, where WT and Bhlhe40-/- NK cells are paired per replicates.

first: Lau

middle: M

last: Colleen

first: Kim

last: Hyunu

first: Sun

last: Joseph


title: Single Cell Analysis of a Mouse Model of Benign Prostate Hyperplasia Links Oxidative Stress to Progenitor Cell Properties

city: Paris

first: Mickael

middle: Mathieu

last: Menager

address: 24 Boulevard du Montparnasse

country: France

institute: Institut Imagine

summary: scRNAseq of prostate cells from 6 and 8 month old Pb-PRL whole prostate mice with enlarged prostate after 28 days of treatment with antioxidant molecule; Anethole tritione (ATT) or coconut oil (vehicle).

overall_design: Two Pb-PRL transgenic mice (6 and 8 months old) expressing prolactin under the control of the probasin promoter were treated by daily gavage for 30 days with a dose of 60mg/kg/day of Anethole tritione. Two other Pb-PRL transgenic mice (6 and 8 months old) were gavaged with coconut oil, the vehicle used to dissolve ATT. The prostates were then dissected , digested with collagenase and FACS sorted to harvest live cells, excluding Ter119+ red blood cells.

first: Leïla

last: DOS SANTOS

first: Francesco

last: CARBONE

first: Emeline

last: PACREAU

first: Marine

last: LUKA

first: Mickaël

last: MENAGER

first: Jacques-Emmanuel

last: GUIDOTTI

first: Vincent

last: GOFFIN


title: Whole genome microarray gene expression profiling of peripheral blood mononuclear cells (PBMC) isolated from old and middle-aged, voluntary human research participants

city: Zurich

first: Ursula

last: Quitterer

address: Winterthurerstrasse 190

department: Molecular Pharmacology

institute: ETH Zurich

summary: With increasing age of the global population, age-associated diseases such as cardiovascular diseases, cancer or diabetes are on the rise in many countries. To identify cellular processes and markers of aging in humans, this study performed comparative whole genome microarray gene expression profiling of peripheral blood mononuclear cells (PBMC) isolated from two age groups, i.e., an old age group (age: 75-89 years) and a younger, middle-aged group (age: 35-50 years). Total RNA was isolated from the PBMC and subjected to whole genome microarray gene expression profiling. The whole genome gene expression data of human PBMC reveal age-associated changes in transcript levels and contribute to a better understanding of biological processes underlying aging and the pathogenesis of age-associated diseases.

overall_design: The study was performed with total RNA isolated from peripheral blood mononuclear cells (PBMC) of two age groups of voluntary human research participants. The first age group encompasses old human research participants (age: 75-89 years; n=5), and the second group are middle-aged human research participants aged 35-50 years (n=4).

first: Joshua

last: Abd Alla


title: Comparative multi-omics analyses of cardiac mitochondrial stress in three mouse models of frataxin deficiency

first: Nicole

last: Sayles

address: 407 East 61st Street

institute: Weill Cornell

summary: The goal of this study was to analyze the molecular changes that occur in the hearts of three mouse models of frataxin deficiency.

overall_design: Libraries were sequenced with single-end 86 bps on an Illumina NextSeq500 sequencer

first: Giovanni

last: Manfredi


title: Modular Pooled Discovery of Synthetic Knockin Sequences to Program Durable Cell Therapies [RNA-3]

first: Franziska

last: Blaeschke

overall_design: RNA-Sequencing using primary human GD2 CAR T cells with TFAP4, tNGFR, BATF-TFAP4, BATF-RFP, RFP-TFAP4 or RFP-tNGFR knockin. This dataset was combined with ATAC- and ChIP-Seq.

first: Yan

last: Chen

first: Ryan

last: Apathy

first: Bence

last: Daniel

first: Andy

first: Peixin

first: Katalin

last: Sandor

first: Wenxi

last: Zhang

first: Zhongmei

last: Li

first: Cody

last: Mowery

first: Tori

last: Yamamoto

first: William

last: Nyberg

first: Angela

last: To

first: Ruby

last: Yu

first: Raymund

last: Bueno

first: Min

first: Ralf

last: Schmidt

last: Goodman

first: Tobias

last: Feuchtinger

first: Justin

last: Eyquem

first: Chun

middle: J

last: Ye

first: Julia

last: Carnevale

first: Ansuman

last: Satpathy

first: Eric

last: Shifrut

first: Theodore

last: Roth

first: Alexander

last: Marson


title: Optimization of a universal allogeneic CAR-T cells combining CRISPR and transposon-based technologies for treatment of Acute Myeloid Leukemia

city: PAMPLONA

first: Juan R

last: Rodriguez-Madoz

address: AV PIO XII 53

institute: CIMA

summary: Despite the potential of CAR-T therapies for hematological malignancies, their efficacy in patients with relapse and refractory Acute Myeloid Leukemia has been limited. The aim of our study has been to develop and manufacture a CAR-T cell product that addresses some of the current limitations. We initially compared the phenotype of T cells from AML patients and healthy young and elderly controls. This analysis showed that T cells from AML patients displayed a predominantly effector phenotype, with increased expression of activation (CD69 and HLA-DR) and exhaustion markers (PD1 and LAG3), in contrast to the enriched memory phenotype observed in healthy donors. This differentiated and more exhausted phenotype was also observed, and corroborated by transcriptomic analyses, in CAR-T cells from AML patients engineered with an optimized CAR construct targeting CD33, resulting in a decreased in vivo antitumoral efficacy evaluated in xenograft AML models. To overcome some of these limitations we have combined CRISPR-based genome editing technologies with virus-free gene-transfer strategies using Sleeping Beauty transposons, to generate CAR-T cells depleted of HLA-I and TCR complexes for allogeneic approaches. Our optimized protocol allows one-step generation of edited CAR-T cells that show a similar phenotypic profile than non-edited CAR-T cells, with equivalent in vitro and in vivo antitumoral efficacy. Moreover, genomic analysis of edited CAR-T cells revealed a safe integration profile of the vector, with no preferences for specific genomic regions, with a highly specific editing of the HLA-I and TCR, without significant off-target sites. Finally, production of edited CAR-T cells at a larger scale allowed the generation and selection of enough HLA-IKO/TCRKO CAR-T cells that would be compatible with clinical applications. In summary, our results demonstrate that CAR-T cells from AML patients, although functional, present phenotypic and functional features that could compromise their antitumoral efficacy, compared to CAR-T cells from healthy donors. The combination of CRISPR technologies with transposon-based delivery strategies allow the generation of HLA-IKO/TCRKO CAR-T cells, compatible with allogeneic approaches, that would represent a promising option for AML treatment.

overall_design: As mentioned in the paper, the my96(45) CAR, containing the 45aa hinge from CD8a, was selected for the rest of the studies. Using our 4-1BB aforementioned second-generation CAR construct, we sequenced the generated CAR-T cells (CD33-CAR-T) from 4 AML patients that were compared to CD33-CAR-T cells from 3 young (adult) and  3 elderly (senior) healthy donors. RNA-seq libraries from these CAR-T cells were prepared following MARS-seq protocol adapted for bulk RNA-seq (23,24) with minor modifications. RNA-seq libraries quantification was done with Qubit 3.0 Fluorometer (Life Technologies) and size profiles were examined using Agilent’s 4200 TapeStation System. Libraries were sequenced in an Illumina NextSeq500 at a sequence depth of 10 million reads per sample. Samples were aligned to the Human genome (GRCh38) with STAR (v2.6.1). Gene expression was quantified with quant3p (github.com/ctlab/quant3p). Downstream analyses were performed in R (v3.6.2).

first: Cristina

last: Calviño

first: Maria

last: Calleja-Cervantes

first: Mikel

last: Hernaez

first: Felipe

last: Prosper

first: Juan


title: STAT4 promotes Bhlhe40 induction to drive protective IFN-g from natural killer cells during viral infection

first: Hyunu

last: Kim


title: Gene expression profile (RNAseq) of peritoneal mouse macrophages in Endotoxic shock and Tolerance

first: Antonio

last: Postigo

state: BARCELONA

address: Casanova 143. Cellex 1B

institute: IDIBAPS/ICREA

summary:
Purpose: Characterize the gene expression profile of of peritoneal mouse macrophages in Endotoxic shock and Tolerance through RNA sequencing
Methods: RNA sequencing of RNA from peritoneal macrophages in Endotoxic shock and Tolerance isolated by peritoneal lavage and FACS sorting (F4/80+ CD11b+)
Results:  Endotoxic shock and Tolerance peritoneal mouse macrophages display differential gene expression.
Conclusions: Endotoxic shock and Tolerance peritoneal mouse macrophages display differential gene expression.

overall_design: RNA sequencing low Imput  of RNA from peritoneal macrophages isolated by peritoneal lavage and FACS sorting (F4/80+ CD11b+)  (2-3 mice pooled for each sample)

first: Marlies

last: Cortes

first: Anna

last: Esteve-Codina


title: STAT4 promotes Bhlhe40 induction to drive protective IFN-g from natural killer cells during viral infection [BHLHE40 CUT&RUN]

overall_design:
To investigate the binding sites of Bhlhe40 in NK cells during homeostasis and 3 hour IL-12 + IL-18 cytokine stimulation, we performed Bhlhe40 CUT&RUN using WT mice.
CUT&RUN was performed on FACS sorted mouse CD49b+ NK cells, where replicates are biologically paired across conditions.

first: Kim

last: Hyunu

first: Sun

last: Joseph


title: SIKs regulate HDAC7 stabilization and cytokine recall in late-stage T cell effector differentiation

status: Public on Sep 12 2023

city: Boston

first: Alberto

middle: Marin

last: González

address: 3 Blackfan St

department: PCMM

institute: Boston Children's Hospital

summary: Understanding the mechanisms underlying the acquisition and maintenance of effector function during T cell differentiation is important to unraveling how these processes can be dysregulated in the context of disease and manipulated for therapeutic intervention. Herein, we report the identification of a novel regulator of murine T cell differentiation through the evaluation of a previously unreported activity of the kinase inhibitor, BioE-1197. Specifically, we demonstrate liver kinase B1 (LKB1) mediated activation of salt-inducible kinases (SIKs) epigenetically regulates cytokine recall potential in effector CD8+ and Th1 cells. Evaluation of this phenotype revealed SIK mediated phosphorylation dependent stabilization of histone deacetylase 7 (HDAC7) occurred during late-stage effector differentiation. HDAC7 stabilization increased nuclear HDAC7 levels which correlated with both global and cytokine loci specific reductions in the activating transcription mark, histone 3 lysine 27 acetylation (H3K27Ac). Accordingly, HDAC7 stabilization diminished transcriptional induction of cytokine upon re-stimulation. Inhibition of this pathway during differentiation produced effector T cells epigenetically poised for enhanced cytokine recall. This work identifies a novel target for enhancing effector T cell functionality.

overall_design: Murine CD8+ T cells were isolated from the spleen and lymph nodes of OT-1 mice and activated with plate bound anti-CD3 and soluble anti-CD28. Cells were treated at the time of activation and throughout differentiation with BioE-1197 or DMSO (Control). On day six after activation and expansion in IL-2, remaining live CD8+ T cells were harvested, fixed, and subjected to ChIP-Seq against H3K27Ac histone mark. Experiments were done in biological triplicates.

first: Rachel

middle: S

last: Helms

last: Marin-Gonzalez

first: Chirag

last: Patel

first: Im-Hong

last: Sun

first: Jiayu

last: Wen

first: Robert

middle: D

last: Leone

first: Bridget

last: Duvall

first: Run-Duo

last: Gao

first: Taekjip

last: Ha

first: Takashi

last: Tsukamoto

first: Barbara

last: Slusher

first: Joel

last: Pomerantz

first: Jonathan

last: Powell


title: Potential role of Bcl2 in lipid metabolism and synaptic dysfunction of age-related hearing loss

city: Shenzhen

first: Yue

last: Liu

address: No. 3004, Longgang Section, Longgang Avenue, Longgang District

institute: Longgang E.N.T Hospital & Shenzhen Key Laboratory of E.N.T, Institute of E.N.T

summary: Age-related hearing loss (ARHL) is a prevalent disease that affects millions of people worldwide. BCL-2 gene plays a tremendous role in cellular senescence, and it was confirmed that the BCL-2 family is closely associated with neurological disorders and ARHL. In addition to modulating apoptosis and oxidative stress, as well as other effects, this study we report here that BCL-2 plays a crucial position in regulating phospholipid and lipid metabolism in  the developing cochlear tissues of rats and a sensory epithelial cell line(OC1), including the cholesterol synthesis pathway that is differentially expressed by genes such as LSS,NSDHL, and CYP51, which further demonstrates the mechanism of the action of BCL-2 in hair cells. In conclusion our data provide a new molecular framework for the regulatory mechanism of BCL-2 down-regulation in senescent cells. These findings provide valuable insights into the molecular mechanisms of ARHL and may set the stage for new therapeutic approaches for this prevalent age-related disease.

overall_design: To investigate the potential mechanism of BCL-2 action in ARHL, we established OC1 cells with knocked down BCL-2 targeting sequences by siRNA.                                                                                We performed gene expression profiling using RNA-seq data obtained from cells that knocked down the target gene in different targeting sequences. A comparison of the gene expression profiling analysis of RNA-seq data from OC1 cells was performed.

first: Cong

last: Fan

first: Huasong

last: Zhang


title: Modular Pooled Discovery of Synthetic Knockin Sequences to Program Durable Cell Therapies [ATAC-Seq]

first: Franziska

last: Blaeschke

address: 1650 Owens Street

institute: Gladstone Institutes

summary: Chronic stimulation can cause T cell dysfunction and limit efficacy of cellular immunotherapies. CRISPR screens have nominated gene targets for engineered T cells, but improved methods are required to compare large numbers of synthetic knockin sequences to reprogram cell functions. Here, we developed Modular Pooled Knockin Screening (ModPoKI), an adaptable platform for modular construction of DNA knockin libraries using barcoded multicistronic adaptors. We built two ModPoKI libraries of 100 transcription factors (TFs) and 129 natural and synthetic surface receptors. Over 30 ModPoKI screens across human TCR and CAR T cells in diverse conditions identified a transcription factor AP4 (TFAP4) construct that enhanced fitness of chronically-stimulated CAR T cells and anti-cancer function in vitro and in vivo. ModPoKI’s modularity allowed us to generate a ~10,000-member library of TF combinations. Non-viral knockin of a combined BATF-TFAP4 polycistronic construct further enhanced function. ModPoKI facilitates discovery of complex gene constructs to program cellular functions.

overall_design: ATAC-Sequencing using primary human GD2 CAR T cells with TFAP4, tNGFR, BATF-TFAP4, BATF-RFP, RFP-TFAP4 or RFP-tNGFR knockin

first: Yan

last: Chen

first: Ryan

last: Apathy

first: Bence

last: Daniel

first: Andy

first: Peixin

middle: A

first: Katalin

last: Sandor

first: Wenxi

first: Zhongmei

last: Li

first: Cody

last: Mowery

first: Tori

last: Yamamoto

first: William

last: Nyberg

first: Angela

last: To

first: Ruby

last: Yu

first: Raymund

last: Bueno

first: Min

last: Kim

first: Ralf

last: Schmidt

first: Daniel

last: Goodman

first: Tobias

last: Feuchtinger

first: Justin

last: Eyquem

first: Chun

last: Ye

first: Julia

last: Carnevale

first: Ansuman

last: Satpathy

first: Eric

last: Shifrut

first: Theodore

last: Roth

first: Alexander

last: Marson


title: Calorie Restriction Outperforms Bariatric Surgery in Reversing Obesity-Driven Tumor Progression in a Mouse Model of Triple Negative Breast Cancer-RNAseq

city: chapel hill

first: michael

middle: francis

last: coleman

state: North Carolina

address: 235 dauer drive

institute: UNC

summary: lean control, obese, and formerly obese C57BL6N mice which underwent weight loss via low-fat diet or verticle sleeve gastrectomy were injected with E0771 cells and tumor growth was monitored

overall_design: DNA was extracted from contralateral normal mammary tissue and subject to RNAseq

first: Michael

middle: F

last: Coleman

first: Stephen

last: Hursting


title: Abnormal accumulation of extracellular vesicles in hippocampal dystrophic axons and regulation by the primary cilia gene intraflagellar transport homolog 88 in Alzheimer's disease

city: Daegu

first: Jaemyung

last: Jang

address: 61, Cheomdan-ro, Dong-gu

country: South Korea

institute: Korea Brain Research Institute

summary: This study identified the specific types of hippocampal neurons whose axons are vulnerable to amyloid-β accumulation. We used the inDropsTM platform to sequence 8 libraries: 5 libraries from the hippocampus of Alzheimer’s disease mice at 12,20, and, 24 weeks of age, and 3 libraries from the hippocampus of normal age-matched mice for the purpose of finding the selectively vulnerable hippocampal neuron type. 22,233 cells with 72,146 transcriptomes were collected after quality control was completed. When the 5,468 neurons that were marked by the expression of Snap25, Syt1, Ndrg4, and Meg3 were examined, we found that three types of neurons, such as somatostatin, chromogranin B, and calbindin-expressing neurons in the hippocampus, were selectively vulnerable to amyloid-β overexpression.

first: Seungeun

last: Yeo

first: HyunJin

last: Jung

first: MiSuk

last: Lee

first: Soonbong

last: Ba

first: Youngshik

last: Choe


title: Calorie Restriction Outperforms Bariatric Surgery in Reversing Obesity-Driven Tumor Progression in a Mouse Model of Triple Negative Breast Cancer-RRBS

type: Methylation profiling by high throughput sequencing

overall_design: DNA was extracted from contralateral normal mammary tissue and subject to RRBS


title: Gene expression profile at single cell level of newborn and adult mouse dorsal skin organoids

city: Taichung

first: Yung-Chih

last: Lai

address: 7F., No. 6, Xueshi Road, North District

department: Integrative Stem Cell Center

institute: China Medical University Hospital

summary: The formation of tissue patterns is critical for organ functions in development and regeneration. To advance the use of organoids for organ regeneration, we learn principles that govern the skin organoids to regenerate hairs. We show the formation of epidermal-dermal coupled cysts function as competent morphogenetic units (CMU) which harbor the ability of skin organoids to regenerate. ScRNA-sequencing shows the emergence of cell types and new cell interactions during CMU formation. In newborn skin organoids, epidermal cells undergo apical-basal polarization via the IFNr-PKR-PKC signaling module. Dermal-Tgfb regulates Gsk3 to establish basement membranes between the epidermal cyst and dermal cells. Meanwhile, VEGF signaling mediates dermal cell attachment to the cyst. Adult cells cannot form organoids but can be induced to generate CMUs and regenerate hairs by adding IFNr or VEGF. We compare the similar principles and different paths used to establish morphogenetic competency in developing skin, wound-induced hair neogenesis, and organoids.

overall_design: Cells were derived from the dorsal skin of neonatal mice within 24 hours of birth. The skin was floated in 0.25% trypsin solution at 4°C to separate into the dermis and epidermis. Epidermal cells were cut with scissors, pipetted, and filtered through a 70um cell strainer followed by centrifugation. The dermis was digested in 0.35% collagenase for 20 minutes, then filtered through a 70um cell strainer followed by centrifugation. The dissociated epidermal cells and dermal cells were mixed at a ratio of 1:9 and were dropped onto the upper chamber of a transwell culture insert, and the lower chamber was filled with 700ul DMEM/F12 (Corning) culture medium containing 10% FBS (Gibco). For adult cell culture, the growth factors were added from D0 to D4, followed by adding MMP14 recombinant protein from D4 to D7. PKC inhibitors were added throughout the cultivation period. The cells were cultured in a 5% CO2 at 37°C incubator, with the culture medium being changed every other day.

first: Mingxing

last: Lei

first: Cheng-Ming

last: Chuong


title: Delineating the Interplay Between Oncogenic Pathways and Immunity in Anaplastic Wilms Tumors: Implications for Prognosis and Therapeutic Vulnerability

city: Houston

first: Xiaoping

last: Su

state: Texas

address: 7300 Brompton St, Apt 5633

institute: MD Anderson Cancer Center

summary: Through multi-omics profiling, we identified and validated a desert-like DAWT subtype that is closely associated with poor outcomes. This subtype is further characterized by TP53 alterations, cGAS-STING pathway downregulation, low CD8 infiltration, a unique signature of human endogenous retroviruses, and activation of oncogenic pathways involving HDAC and DNA repair.

overall_design: Paired-end RNAseq of Anaplastic Wilms Tumors

first: Gabriel

last: Malouf


title: Circadian ribosome profiling reveals a role for the Period2 upstream opening reading frame in sleep.

city: Suita

first: Arthur

last: Millius

state: Osaka

address: iFReC 6F

department: iFReC

country: Japan

institute: Osaka University

summary: Many mammalian proteins have circadian cycles of production and degradation, and many of these rhythms are altered post-transcriptionally. We used ribosome profiling to examine post-transcriptional control of circadian rhythms by quantifying RNA translation in the liver over a 24-h period from circadian-entrained mice transferred to constant darkness conditions and by comparing ribosome binding levels to protein levels for 16 circadian proteins. We observed large differences in ribosome binding levels compared to protein levels, and we observed delays between peak ribosome binding and peak protein abundance. We found extensive binding of ribosomes to upstream open reading frames (uORFs) in circadian mRNAs, including the core clock gene Period2 (Per2). An increase in the number of uORFs in the 5'UTR was associated with a decrease in ribosome binding in the main coding sequence and a reduction in expression of synthetic reporter constructs. Mutation of the Per2 uORF increased luciferase and fluorescence reporter expression in 3T3 cells without altering the phase or period and increased luciferase expression in PER2:LUC MEF cells. Mutation of the Per2 uORF in mice increased PER2 protein levels and reduced total sleep time compared to that in wild-type mice. These results suggest that post-transcriptional processes shape translation of mRNA transcripts, which can impact physiological rhythms and sleep.

overall_design: Ribosome profiling and RNA sequencing of three C57BL/6N mouse livers from wild-type and Per2uORF mutant mice

first: Dimitri

last: Perrin

first: Hiroki

middle: R

last: Ueda


title: Single-cell dissection of the multicellular ecosystem and molecular feature sunderlying microvascular invasion in hepatocellular carcinoma

city: Wenzhou

first: Meng

last: Zhou

address: 270 Xueyuan Xi Road

institute: Wenzhou Medical University

summary: Microvascular invasion (MVI) is a significant pathological feature of hepatocellular carcinoma (HCC) associated with poor outcomes, early recurrence, and intrahepatic metastasis in post-surgical resection and transplant patients. However, the tumor microenvironment (TME) and transcriptional programs underlying MVI remain poorly understood. In this study, we conducted single-cell RNA sequencing of 46,789 individual cells from 10 samples of MVI+ (MVI present) and MVI- (MVI absent) HCC patients, providing a comprehensive single-cell atlas of MVI. Single-cell dissection of the multicellular ecosystem revealed the emergence or expansion of cycling T cells, LAMP3+ dendritic cells, TREM2+ macrophages, myofibroblasts, and tumor-promoting arterial i endothelial cells within the MVI+ TME. MVI+ malignant cells are characterized by high proliferation rates and the ability to evade immune detection through the upregulation of non-classical MHC class I molecules. In contrast, MVI-malignant cells exhibit an inflammatory milieu.  Furthermore, we identified the CCL16/CCR1 interaction between TREM2+ macrophages and malignant cells as a contributing factor in MVI formation and tumor progression. Notably, we identified a unique multicellular module that plays the dominant role in shaping the immunosuppressive microenvironment of MVI and correlates with poor prognosis. In summary, these findings enhance our understanding of the underlying cellular and molecular mechanisms driving MVI and provide valuable insights for improving clinical diagnosis and developing more effective treatment strategies.

overall_design: Ten independent surgical resected specimens were collected from five HCC patients (3 MVI present patients and 2 MVI absent patients), including 5 HCC tissues and 5 adjacent non-tumor tissues for single-cell RNA sequencing . Inclusion criteria for patients comprised the absence of apparent metastases, a singular tumor of dimensions between 3 and 5 cm, and no prior therapeutic intervention.

first: Ke

last: Li

first: Zhaoyang

last: Lu

first: Yifeng

last: Cui


title: Lymph node dendritic cells harbor inducible replication-competent HIV despite years of suppressive ART

city: Epalinges

first: Mathilde

last: Foglierini-Perez

address: Chemin de Boveresses 155

institute: CHUV

summary: The comprehensive characterization and quantification of the HIV tissue reservoirs is required to design appropriate therapeutic intervention(s) to achieve a cure. While pioneering studies demonstrated that HIV replication and spreading mainly occur in lymphoid tissues, the identification of specific cell subsets harboring replication competent virus in lymphoid tissues has long been neglected.  In this context, we and others have recently shown that gut memory CD4 T cells, lymph node T follicular helper (Tfh) cells and tissue macrophages represent major HIV/SIV tissue reservoirs. Notably, Tfh cell differentiation is a multi-stage process that requires long-lasting interactions with lymph node (LN) dendritic cells (DCs) and pre-germinal center B cells in a specific cytokine/chemokine microenvironment. Lymph node DCs are endowed with an exceptional T-cell stimulatory potential and can either migrate from the periphery to the draining lymph node (migratory DCs) or locate in the LN for their entire life span (resident DCs). On the basis of these unique properties, long-term persistence of LN DCs infected with replication competent virus may represent the initial trigger of viral rebound post ART interruption and may therefore represent a major obstacle to HIV cure. We demonstrate that LN migratory DCs are infected with replication competent virus and persist despite suppressive ART. In addition, we identified the mechanisms associated with i) LN DC susceptibility to HIV infection and ii) long term persistence of HIV-infected LN DCs i.e. proliferation of infected cells in tissue sanctuaries.

overall_design: 3 anti-retroviral therapy treated HIV-infected individuals

first: Riddhima

last: Banga

first: Matthieu

last: Perreau

first: Francesco

last: Procopio

first: Erica

last: Lana

first: Gregory

last: Takashi

first: Annamaria

last: Kauzlaric

first: Olivia

last: Munoz

first: Craig

last: Fenwick

last: Foglierini

first: Matthias

last: Cavassini

first: Jean-Marc

last: Corpataux

first: Mauro

last: Delorenzi

first: Mathias

last: Lichterfeld

first: Giuseppe

last: Pantaleo


title: Spatiotemporal insight into early pregnancy governed by immune-featured stromal cells [RNA-seq]

city: Beijing

first: Min

last: Yang

state: Beijing

address: No. 5, yiheyuan road, haidian district,

institute: Peking University

summary: Endometrial decidualization connecting embryo implantation and placentation is transient, but essential for successful pregnancy, which however is not systematically investigated. Here, by using a novel single-cell spatial transcriptomic profiling technology (scStereo-seq), we were able to visualize and define key and dynamic functional decidual hubs assembled by distinct immune, endothelial, trophoblast and decidual stromal cells (DSCs) during early pregnancy in mice. We classified DSC subclusters and described their transdifferentiation trajectory controlled by transcription factor cascades. Interestingly, we discovered a novel type of DSCs expressing immune cell-specific genes, termed immune DSCs (iDSCs). Whereas immature DSCs attracted immune cells and induced decidual angiogenesis at the mesenchymal-to-epithelial transition (MET) hub during decidualization initiation, iDSCs enabled immune cell recruitment, angiogenesis facilitation and cytotoxic effect induction to form immune cell assembling and angiogenesis hubs, which eventually allow decidual homeostasis maintenance and decidualization-to-placentation transition. We spatiotemporally decode mouse early pregnancy events controlled by immune DSCs.

overall_design: In vitro decidualization was induced from primary endometrialum stromal cells by using progesterone (P4) and cAMP-stimulation. Then, one primary endometrial stromal cells (control) and samples were collected at different time points (Day1-Day5) and subjected to bulk RNA sequencing.

first: Jennie

last: Ong


title: Spatiotemporal insight into early pregnancy governed by immune-featured stromal cells [scRNA-seq]

overall_design: Embryo-derived tissues (the yolk sac, placenta), and maternal-derived decidua tissue, were dissected and dissociated. The single cells were sorted using fluorescence-activated cell sorting (FACS) and subjected to single-cell RNA sequencing using the 10x Genomics Single Cell 3’ Library kit.


title: Glucocorticoid receptors drive breast cancer cell migration and metabolic reprograming via pSer134-GR-induced expression of PDK4

city: Minneapolis

first: Carol

last: Lange

state: MN

address: 2231 6th ST SE

department: Masonic Cancer Center

institute: University of Minnesota

summary: Corticosteroids act on the glucocorticoid receptor (GR; NR3C1) to resolve inflammation and are routinely prescribed to breast cancer patients undergoing chemotherapy treatment to alleviate side effects. Triple negative breast cancers (TNBCs) account for 15-20% of diagnoses and lack expression of estrogen and progesterone receptors as well as amplified HER2, but often express high GR levels. GR is a mediator of TNBC progression to advanced metastatic disease, however the mechanisms underpinning this transition to more aggressive behavior remain elusive. We previously showed that tissue/cellular stress (hypoxia, chemotherapies) as well as factors in the tumor microenvironment (TGFβ, HGF) activate p38 MAPK, which phosphorylates GR on Ser134. In the absence of ligand, p-Ser134-GR further upregulates genes important for responses to cellular stress, including key components of the p38 MAPK pathway. Herein, we show that GR Ser134 is required for TNBC metastatic colonization to the lung. To understand the mechanisms of p-Ser134-GR action in the presence of GR agonists, we examined glucocorticoid-driven transcriptomes in CRISPR (knock-in) models of TNBC cells expressing wild-type or phospho-mutant (S134A) GR. We identified dexamethasone- and p-Ser134-GR-dependent regulation of specific gene sets controlling TNBC migration (NEDD9, CSF1, RUNX3) and metabolic adaptation (PDK4, PGK1, PFKFB4). TNBC cells harboring S134A GR displayed metabolic reprogramming that was phenocopied by PDK4 knockdown. PDK4 knockdown or chemical inhibition also blocked cancer cell migration. Our results reveal a convergence of GR agonists (i.e., host stress) with cellular stress signaling whereby pSer134-GR critically regulates TNBC metabolism, an exploitable target for the treatment of this deadly disease.

overall_design: Examination of the effect of Dexamethasone in TNBC cell line MDA-MB-231 with mutation in the glucocorticoid receptor on Ser134.

first: Amy

last: Dwyer

first: Carlos

last: Perez Kerkvliet


title: Differential voluntary feed intake and whole transcriptome profiling in the hypothalamus of young sheep offered crude protein and phosphorus deficient diets

city: Armidale

first: David

last: Innes

state: New South Wales

address: University of New England

department: School of Environmental and Rural Science

institute: University of New England

summary: A reduction in voluntary feed intake is observed in ruminants consuming nutrient deficient diets, such as those with a low CP or P content, and has been attributed to active metabolic regulation, rather than a physical constraint. The hypothalamus is the key integrator of feed intake regulation in mammals. The objectives of this experiment were to 1) establish a model of metabolic feed intake regulation in ruminants consuming diets of variable CP and P content, and 2) determine key biochemical pathways and influential points of regulation within the hypothalamus. Merino wethers [n = 40; 23.7 ± 1.4 kg liveweight (mean ± SD)] were fed one of five dietary treatments (n = 8/treatment) for 63 days in individual pens. The treatments included targeted combinations of high (H) and low (L) CP (110 and 55 g/kg DM) and high and low P (2.5 and 0.7 g/kg DM) with 9 MJ metabolisable energy (ME) per kg DM which were fed ad libitum (UMEI; unrestricted ME intake) resulting in four experimental diets (HCP-HP-UMEI, LCP-HP-UMEI, HCP-LP-UMEI and LCP-LP-UMEI). An additional nutritional treatment (HCP-HP-RMEI) restricted intake of the HCP-HP diet to an equivalent ME intake of wethers consuming the LCP-LP-UMEI treatment. Wethers offered the LCP-HP-UMEI, HCP-LP-UMEI and LCP-LP-UMEI treatments consumed 42, 32 and 49% less total DM (P ≤ 0.05), respectively than the HCP-HP-UMEI treatment, and this was not attributable to any physical limitation of the rumen. Plasma concentrations of urea nitrogen and inorganic phosphate indicated that these nutrient deficiencies were successfully established. To assess potential mechanisms, RNA-seq was conducted on samples from the arcuate nucleus (ARC), ventromedial hypothalamus and lateral hypothalamus of the wethers, yielding a total of 301, 8 and 148 differentially expressed genes across all pairwise comparisons, respectively. The expression of NPY, AGRP and CARTPT, known for their regulatory role in mammalian feed intake regulation, had a similar transcriptional response in the ARC of wethers consuming nutrient deficient treatments and those consuming a ME restricted treatment, despite these wethers expressing behaviours indicative of satiated and hungry states, respectively. In addition, genes involved with glycolysis (TPI1), the citric acid cycle (CS, OGDH, GLUD1, GOT1) and oxidative phosphorylation (COX5A, ATP5MC1, ATP5F1B, ATP5MC3) were downregulated in the ARC of wethers fed a nutrient deficient (LCP-LP-UMEI) relative to the non-deficient (HCP-HP-UMEI) treatment. In summary, a model for voluntary feed intake restriction was established to determine genome-wide molecular changes in the hypothalamus of young ruminants.

overall_design: Tissue samples collected from the arcuate nucleus (ARC), lateral hypothalmus (LHA) and ventromedial hypothalamus (VMH) of 40 Merino sheep (7 months old) fed one of five dietary treatments (n = 8/treatment) for 63 days. Sample ARC_35 was not sequenced and is not included.

first: Nicholas

last: Hudson

first: Stephen

middle: T

last: Anderson

first: Dennis

middle: P

last: Poppi

first: Simon

last: Quigley


title: Calorie Restriction Outperforms Bariatric Surgery in Reversing Obesity-Driven Tumor Progression in a Mouse Model of Triple Negative Breast Cancer-Study2_tumor

first: michael

last: coleman

summary: lean control, obese, and formerly obese C57BL6N mice which underwent weight loss via vertical sleeve gastrectomy or calorie restriction were injected with E0771 cells and tumor growth was monitored

overall_design: 6 E0771 tumors from each group were analyzed

first: Michael

last: Coleman

last: Hursting


title: Extra-tumoral analysis of GBM bearing murine brains

city: Chicago

first: April

last: Bell

state: IL

address: 2053 N Mozart St Apt 3F

institute: Northwestern University

summary: Young (8-12 weeks) and old (80-90 weeks) mice were engrafted with 50,000 murine GL261 cells. Extra-tumoral tissue was isolated on Day 21 post intracranial injection and sent for RNA sequencing analysis. Tissue from tumor naïve young and old mice was included as .controls

overall_design: 8-12 week old and 8-90 week old C57BL/6 mice were engrafted with 50,000 murine GL261 cells. Naïve young and old mice were included as controls. Extra-tumoral tissue was isolated on Day 21 post intracranial engraftment.

first: Erik

last: Ladomersky

first: Derek

last: Wainwright


title: Calorie Restriction Outperforms Bariatric Surgery in Reversing Obesity-Driven Tumor Progression in a Mouse Model of Triple Negative Breast Cancer-Study1_tumor

summary: lean control, obese, and formerly obese C57BL6N mice which underwent weight loss via vertical sleeve gastrectomy or low fat diet were injected with E0771 cells and tumor growth was monitored

overall_design: 5-6 E0771 tumors from each group were analyzed


title: Longitudinal global transcriptomic profiling of preclinical systemic sclerosis reveals molecular changes associated with disease progression

first: Shervin

last: Assassi

state: TX

address: 6431 Fannin St

institute: UTHouston

summary:
Objective: To investigate peripheral blood cell (PBCs) global gene expression profile of SSc at its preclinical stage (PreSSc) and to characterize the molecular changes associated with progression to a definite disease over time.
Material and methods: Clinical data and PBCs of 33 participants with PreSSc and 16 healthy controls (HCs) were collected at baseline and follow-up (mean 4.2 years). Global gene expression profiling was conducted by RNA sequencing and a modular analysis was performed.
Results: Comparison of baseline PreSSc to HCs revealed 2889 differentially expressed genes. Interferon signalling was the only activated pathway among top over-represented pathways. Moreover, 10 modules were significantly decreased in PreSSc samples (related to lymphoid lineage, cytotoxic/NK cell, and erythropoiesis) in comparison to HCs. At follow-up, 14 subjects (42.4%) presented signs of progression (evolving PreSSc) and 19 remained in stable preclinical stage (stable PreSSc). Progression was not associated with baseline clinical features or baseline PBC transcript modules. At follow-up stable PreSSc normalized their down-regulated cytotoxic/NK cell and protein synthesis modules while evolving PreSSc kept a down-regulation of cytotoxic/NK cell and protein synthesis modules. Transcript level changes of follow-up vs baseline in stable PreSSc vs evolving PreSSc showed 549 differentially expressed transcripts (336 up and 213 down) with upregulation of the EIF2 Signalling pathway.
Conclusions: Participants with PreSSc had a distinct gene expression profile indicating that molecular differences at a transcriptomic level are al_x0002_ready present in the preclinical stages of SSc. Furthermore, a reduced NK signature in PBCs was related to SSc progression over time.

overall_design:
Whole blood global gene expression profiling was performed in baseline samples of patients with preclinical stages of systemic sclerosis and in their samples obtained on follow-up visit  (mean 4.2 years after baseline visit), as well as in matched healthy control samples.
Patients who kept preclinical features on follow-up visit, were labelled as ‘stable PreSSc’ while those progressing to definite SSc were labelled as ‘evolving PreSSc’.
Samples that have the same Pair number come from the same patient (one from baseline and one from follow-up). Healthy controls do not have follow-up samples

first: Chiara

last: Bellocchi

first: Lorenzo

last: Beretta

first: Xuan

last: Wang

first: Marka

last: Lyons

first: Maurizio

last: Marchini

first: Vincenzo

last: Carbonelli

first: Nicola

last: Montano


title: Intratumoral dose-specific GM-CSF modulates breast tumor oxygenation and anti-tumor immunity through macrophages

city: Morgantown

first: Gangqing

last: Hu

state: West Virginia

address: 64 Medical Center Drive

department: MicroBiology, Immunology, and Cell Biology

institute: West Virginia University

summary: Background: Granulocyte-Macrophage Colony Stimulating Factor (GM-CSF) is a hematopoietic growth factor and adjuvant in cancer immunotherapy via stimulation of dendritic cells/APCs. However, GM-CSF has yielded inconsistent results and its role regarding in vivo modulation of macrophages remains underexplored. We previously demonstrated that 100ng “high-dose” intratumor (IT) GM-CSF ablated tumor blood vessels and worsened tumor hypoxia after 3 weeks through tumor-associated macrophage (TAM) soluble VEGFR-1 production in PyMT murine breast cancer. Here, we investigate a role for “low-dose” IT GM-CSF on tumor oxygen and the impact on immunotherapy response, TAMs/myeloid cells, and TILs relative to “high-dose”.    Methods: We performed IT injections of dose-specific GM-CSF or saline controls and then evaluated phenotypic effects after 3 weeks. We used Electron Paramagnetic Resonance Oximetry to measure and image in vivo tumor oxygen in real-time, and fluorescent immunohistochemistry to assess tumor blood vessels. IT GM-CSF doses were tested in priming PyMT tumors for sensitization to PD1. We performed RNA Sequencing of TAM and CD8 TIL to observe transcriptional changes coupled with flow cytometry of peripheral blood monocytes, tumor myeloid, and TIL populations in immunology cold" PyMT tumors response to dose-optimized GM-CSF. Lastly, we assessed and compared effects of IT GM-CSF on TAM and TIL in an immunologically “hot” 4T1 breast cancer model.    Results: 5ng IT GM-CSF significantly increased PyMT tumor oxygen without augmenting tumor growth and promoted tumor vessel health via increased pericyte coverage. Priming of PyMT tumors with 5ng IT GM-CSF (“low-dose”, hypoxia reduced) sensitized “cold” PyMT tumors to PD1, but this synergy was not observed with 100ng (“high-dose”, hypoxia exacerbated). Immunologically, 5ng GM-CSF did not increase monocyte mobilization or alter phenotypic marker expression on TAMs, but reduced hypoxic and inflammatory transcriptional programs in macrophages and CD8 TIL isolated from PyMT tumors. 100ng increased infiltration of myeloid cells and TAMs but these TAMs had reduced MHCII expression, suggesting support of immune-suppressive TAM under hypoxia. Some tumors exhibited increased CD8 polyfunctionality either dose suggesting mild CD8 TIL priming. On the other hand, 100ng in “hot” 4T1 tumors resulted in increased TAM MHCII and other immunostimulatory molecules with moderate increases in CD8 TIL polyfunctionality and exhausted PD1hiTIM3+ phenotype, indicating that GM-CSF may have opposing effects on macrophage modulation based on tumor immunological status.

overall_design: 11 samples analyzed, with 2-3 replicates per cell type per treatment group (5ng GM-CSF) or saline control. Control samples are "saline treated". Macrophage "saline1" consists of 2 biological replicates pooled together due to low cell yield from 2 individual tumors.

first: Nicole

last: Mihalik

first: Lei

first: Ryan

last: Percifield

first: Edwin

last: Wan

first: Timothy

last: Eubank


title: High concentration of adrenaline blocks key cell cycle genes and exhibits antifibrotic and vasoconstrictor effects in glaucoma surgery

city: London

first: Cynthia

last: Yu-Wai-Man

address: Westminster Bridge Road

country: United Kingdom

institute: King's College London

summary: To investigate the effects of adrenaline on fibrosis in glaucoma surgery. Human Tenon’s fibroblasts (HTFs) were treated with adrenaline (0%, 0.0005%, 0.01%) for 24 hours and RNA-Sequencing was performed on the Illumina NextSeq2000. We carried out detailed gene ontology, pathway, disease and drug enrichment analyses.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for HTFs treated with different concentrations of adrenaline (0%, 0.0005%, 0.01%).


title: Effects of Rift Valley fever virus infection on gene expression and alternative splicing in HEK 293 cells

city: Missoula

first: J Stephen

last: Lodmell

state: MT

address: 32 campus drive

department: Biological Sciences

institute: The University of Montana

summary: Rift Valley fever virus (RVFV) is an important human and livestock pathogen. To better understand the molecular virology and mechanisms of pathogenesis in human HEK293 cells during RVFV MP-12 strain infection, we used high-throughput mRNA sequencing technology to identify and analyze differentially expressed genes and mRNA splicing patterns triggered by infection or by expression of RVFV nucleocapsid protein. Here we supply the results of our RNA-seq analysis of RVFV-infected cells and cells transfected with RVFV nucleocapsid protein expressing plasmids. Some of the results were published in: "Transcriptome profiling in Rift Valley fever virus infected cells reveals modified transcriptional and alternative splicing programs" by Katherine E Havranek, Luke Adam White, Jean-Marc Lanchy, J Stephen Lodmell. PLoS One. 2019 May 28;14(5):e0217497. PMID: 31136639 PMCID: PMC6538246.

overall_design: To investigate the impact of Rift Valley Fever Virus (RVFV) infection on human gene expression and alternative splicing patterns, we infected HEK cells with RVFV MP12 strain and harvested total RNA two days post-infection. The transcript expression levels and alternative splicing events were measured by the analysis of 150-bp Illumina sequencing paired-reads of the purified polydenylated RNAs. We also analyzed, using the same Illumina sequencing technique, the impact heterologous expression of the viral RNA-binding N protein on host gene expression levels and alternative splicing events. The HEK293 cells used in that experiment were transfected with a viral N protein expression plasmid and the total RNA was harvested two days after the transfection and the purified polydenylated RNAs were processed for Illumina sequencing.

first: Katherine

last: Havranek

first: Luke

last: White

first: Jean-Marc

last: Lanchy

first: J


title: Casein kinase 1 and 2 phosphorylate Argonaute proteins to regulate miRNA-mediated gene silencing

city: Québec

first: VIVEK

middle: NILESH

last: SHAH

state: Québec

address: 9 Rue McMahon

department: Département de biologie moléculaire, de biochimie médicale et de pathologie

country: Canada

institute: Université Laval

type: Non-coding RNA profiling by high throughput sequencing

summary: MicroRNAs (miRNAs) together with Argonaute (AGO) proteins form the core of the RNA-induced silencing complex (RISC) to regulate gene expression of their target RNAs post-transcriptionally. Argonaute proteins are subjected to intensive regulation via various post-translational modifications that can affect their stability, silencing efficacy and specificity for targeted gene regulation. We report here that in Caenorhabditis elegans, two conserved serine/threonine kinases - casein kinase 1 alpha 1 (CK1A1) and casein kinase 2 (CK2) - regulate a highly conserved phosphorylation cluster of 4 Serine residues (S988:S998) on the miRNA-specific AGO protein ALG-1. We show that CK1A1 phosphorylates ALG-1 at sites S992 and S995, while CK2 phosphorylates ALG-1 at sites S988 and S998. Furthermore, we demonstrate that phospho-mimicking mutants of the entire S988:S998 cluster rescue the various developmental defects observed upon depleting CK1A1 and CK2. In humans, we show that CK1A1 also acts as a priming kinase of this cluster on AGO2. Altogether, our data suggest that phosphorylation of AGO within the cluster by CK1A1 and CK2 is required for efficient miRISC-target RNA binding and silencing.

overall_design:
To evaluate the effects on global miRNA levels after CK1A1 and CK2 depletion in wild-type animals by HT-sequencing
The control RNAi is control treatment. And the kin-19 RNAi corresponds to CK1A1 depletion. CK1A1 is the ortholog of kin-19 in C elegans. And, kin-3 RNAi and kin-10 RNAi correspond to CK2 depletion. As CK2 is made of two subunits, kin-3 (catalytic) and kin-10 (regulatory) in C. elegans.

first: Vivek

middle: N

last: Shah

first: Julia

last: Neumeier

first: Miguel

last: Quévillon  Huberdeau

first: Daniela

last: Zeitler

first: Astrid

last: Bruckmann

first: Gunter

last: Meister

first: Martin

last: Simard


title: DNA-PKcs is a key transcriptional and splicing regulator of AR-Vs in advanced prostate cancer

status: Public on Sep 11 2023

city: Newcastle upon Tyne

first: Beth

last: Adamson

address: Paul O'Gorman Building, Framlington Place

department: Translational and Clinical Research Institute

institute: Newcastle University

summary: To investigate the role of DNA-PKcs and RBMX in the regulation of androgen receptor variants we used the CWR22Rv1-AR-EK and CWR22Rv1 cell line in which DNA-PKcs or RBMX has been knocked down by siRNA or DNA-PKcs was inhibited with 1  µM NU7441, NU5455 or AZD7648. We then performed gene expression analysis using data obtained from RNA-seq at two time points (72 hours knockdown and 24 hours drug treatment).

overall_design: Comparative gene expression profiling analysis of RNA-seq data for CWR22Rv1-AR-EK and CWR22Rv1 cells.

last: Gaughan

first: Nicholas

last: Brittain


title: ZNF683 marks a CD8+ T cell population associated with anti-tumor immunity following anti-PD-1 therapy for Richter syndrome [TCR-seq]

city: Copenhagen

first: William

middle: Winston

last: Agace

address: Blegdamsvej 3B

department: Skin Immunology Research Center

country: Denmark

institute: Copenhagen University

summary: Unlike many other hematologic malignancies, Richter syndrome (RS), an aggressive B cell lymphoma originating from indolent chronic lymphocytic leukemia, is responsive to PD-1 blockade. To discover the determinants of response, we analyzed single-cell transcriptome data generated from 17 bone marrow samples longitudinally collected from 6 RS patients. Response was associated with intermediate exhausted CD8 effector/effector memory T cells marked by high expression of the transcription factor ZNF683, determined to be evolving from stem-like memory cells and divergent from terminally exhausted cells. This signature overlapped with that of tumor-infiltrating populations from PD-1 responsive solid tumors. ZNF683 was found to directly target key T cell genes (TCF7, LMO2, CD69) and impact pathways of T cell cytotoxicity and activation. Analysis of pre-treatment peripheral blood from 10 independent RS PD-1 treated patients, as well as solid tumor PD-1 treated patients, supported an association of ZNF683high T cells with response.

overall_design: rhTCRseq

first: Erin

last: Parry

first: Stephanie

last: Deng

first: Camilla

middle: K

last: Lemvigh

first: Catherine

last: Wu


title: ZNF683 marks a CD8+ T cell population associated with anti-tumor immunity following anti-PD-1 therapy for Richter syndrome

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

overall_design: Refer to individual Series


title: Transcriptomic Profile of the Bovine Endosalpinx & Endometrium to Identity Putative Embryokines

city: Gainesville

first: Peter

last: Hansen

state: Florida

address: 2250 Shealy Drive

department: Department of Animal Sciences

institute: University of Florida

summary: Although the oviduct and uterus regulate early embryonic development, understanding of the specific maternally-derived molecules that signal to the embryo is incomplete. Objectives of this study were to characterize expression of genes encoding for growth factors, cytokines, and other cell-signaling ligands in the endosalpinx and endometrium of the bovine female and analyze spatial changes in gene expression throughout the reproductive tract.

overall_design: Five adult nulliparous Holstein heifers were synchronized using a 5-day CIDR-Synch protocol. On day 2 after ovulation, animals were slaughtered and samples were collected from the ampulla, upper uterine horn, mid uterine horn, and uterine body of both the contralateral and ipsilateral horns relative to ovulation.

first: Quinn

last: Hoorn

first: Maria

middle: B

last: Rabaglino

first: Tatiane

last: Maia

first: Masroor

last: Sagheer

first: Dailin

last: Fuego


title: Macrophage AMPK β1 activation by PF-06409577 reduces the inflammatory response, cholesterol synthesis, and atherosclerosis in mice

city: Hamilton

first: Dongdong

last: Wang

state: Ontario

address: 1200 Main Street West

department: Department of Medicine

institute: McMaster University

summary: Abstract: Atherosclerotic cardiovascular disease (CVD) is the leading cause of death in the developed world and is characterized by both chronic low-grade inflammation and dyslipidemia. AMP-activated protein kinase (AMPK) inhibits cholesterol synthesis and dampens inflammation. While activation of AMPK has been shown to have beneficial effects on aspects of atherosclerosis, to date, only compounds that indirectly and non-specifically activate the kinase (i.e. metformin, canagliflozin, salsalate) or have low oral and cellular bioavailability (i.e. A769662) have been examined. In the current study, we have tested the effects of an orally bioavailable and potent activator of AMPK, PF-06409577, that selectively targets AMPKb1-containing complexes which are predominately expressed in the murine liver and immune cells, including macrophages. We found that daily oral gavage with PF-06409577 reduced levels of atherosclerotic plaque in two  models of atherosclerosis through a mechanism dependent on the AMPKb1 isoform in myeloid cells. These observations were not associated with changes in hepatic or plasma lipid levels but instead was associated with reductions in inflammatory pathways in macrophages. In cultured bone marrow-derived macrophages, PF-06409577 activated AMPK and reduced inflammatory pathways including the NF-κB pathway as well as fatty acid and cholesterol synthesis; effects requiring the AMPKβ1 isoform. These data indicate that pharmacologically targeting macrophage AMPKβ1 may be a promising strategy for reducing atherosclerosis.

overall_design: Murine Bone Marrow derived Macrophages from Wildtype and AMPKbeta1 knockout mice were treated with 10uM PF-06409577 or Veh (DMSO) for 6 hours to assess the effects of AMPK activation on inflammation and metabolism.

first: Emily

last: Day

first: Logan

last: Townsend

first: Sonia

last: Rehal

first: Battsetseg

last: Batchuluun

first: Marisa

last: Morrow

first: Rachel

last: Lu

first: Lucie

last: Lundenberg

first: Jessie

middle: H

first: Eric

middle: M

last: Desjardins

first: Tyler

last: Smith

first: Amogelang

last: Raphenya

first: Andrew

middle: G

last: McArthur

first: Morgan

last: Fullerton

first: Gregory

last: Steinberg


title: Effect of PME-1 knockout on MEF gene expression

city: yamaguchi

first: Shunta

last: Ikeda

state: yamaguchi

department: Joint faculty of veterinary medicine

institute: Yamaguchi University

summary: We investigated the role of PME-1 (protein phosphatase methylesterase-1) as a specific methylesterase for the critical cancer suppressor, protein phosphatase 2A (PP2A), and its potential contributions to the development and malignancy of various cancers. Notably, no gene expression database related to PME-1 deficiency had been previously reported. To address this knowledge gap, we conducted an analysis of the impact of PME-1 on gene expression using mouse embryonic fibroblasts (MEFs) isolated from both PME-1 wild-type (WT) and knockout (KO) mice.

overall_design: PME-1 KO cells and their controls (WT) were collected 24 hours after seeding in DMEM (high glucose) medium and subjected to RNA-seq analysis.

first: Koichi

last: Sato

first: Takashi

last: Ohama


title: Effect of prostaglandin E2 (PGE2) treatment on human dermal lymphatic endothelial cells (HDLECs)

city: New York

first: Timothy

middle: Cragin

last: Wang

state: New York

address: 1130 Saint Nicholas Avenue, ICRC

institute: Columbia University

summary: To determine how PGE2 treatment affects HDLECs, we treated cultured HDLECs with either a vehicle or 500 nM dmPGE2 for 16 hours

overall_design: Comparative gene expression profiling analysis of RNA-seq data for HDLEC cell line with or without dmPGE2

first: Jiang

last: Zhengyu

first: Waterbury

last: Quin


title: Systemic and intrinsic functions of ATRX in glial cell commitment and CNS myelination

first: Sarfraz

last: Shafiq

address: Victoria Hospital, 800 commissioners road east

department: Anatomy and Cell Biology

institute: University of Western Ontario

summary: Total forebrain RNA (10 μg) was isolated from three P17 pairs of littermate-matched AtrxFoxG1Cre and control mice using the RNeasy Mini kit (Qiagen Cat# 74104). cRNA was generated and hybridized to an Affymetrix Mouse Genome 430 2.0 Array at the London Regional Genomics Center (London, Canada). Probe signal intensities were generated using GCOS1.4 (Affymetrix Inc., Santa Clara, CA) using default values for the Statistical Expression algorithm parameters and a Target Signal of 150 for all probe sets and a Normalization Value of 1. Gene level data was generated using the RNA preprocessor in GeneSpring GX 7.3.1 (Agilent Technologies Inc., Palo Alto, CA). Data were then transformed (measurements less than 0.01 set to 0.01), normalized per chip to the 50th percentile, and per gene to control samples. Probe sets representing Atrx transcripts were removed (10 sets). The remaining probe sets were filtered by fold change ≥1.5 between control and AtrxFoxG1Cre samples, and by confidence level of P<0.05. Significantly overrepresented GO categories were determined using the Enrichr web platform

overall_design: 3 controls and 3 ATRX FOXG1 Cre Knockouts

first: Megan

middle: E

last: Rowland

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last: Jiang

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last: Ghahramani

first: Miguel

last: Pena-Ortiz

first: Vanessa

last: Dumeaux

first: Nathalie

last: Bérubé