title: Interleukin-10/CSF1R fusion protein rejuvenates the exhausted T cells for executing antitumor activity IV

last: Chen

summary: Bulk RNA sequence (RNA-seq) has been used to explore the effect of IL10/CSF1R fusion protein modulation in tumor microenvironment.  Four groups were analyzed which contain Ctrl, CSF1R, IL10 and BF10.  MTC-Q1 oral cancer cells were orthotopic injection into C57BL/6J mice. tumor tissue has been isolated (~ 100 mm3) and total RNA were extracted by Trizol, followed by cDNA were sequenced by Illumina NovaSeq6000.

overall_design: Examination of therapeutic modulation of IL10/CSF1R fusion protein within tumor microenvironment in mice cancer model.  RNA-seq of tumor tissues from orthotopic cancer tissue after four dose treatments.

first: Yao-Wen


title: Effect on gene expression profilng of BTSCs following HDAC1/2 inhibition with romidepsin.

status: Public on Jul 09 2023

city: Calgary

first: Ravinder

last: Bahia

state: Alberta

address: 3330 Hospital drive NW

department: Cell Biology and Anatomy

country: Canada

institute: University of Calgary

type: Expression profiling by high throughput sequencing

summary: To investigate the roles of HDAC1/2 in regulating the transcriptional programs associated with BTSC stem cell, growth and differentiation features, we performed RNA-seq using samples obtained from romidepsin treated versus DMSO vehicle treated GBM BTSCs.

overall_design: Comparative gene expression profiling of RNA-seq data obtained from romidepsin treated versus DMSO vehicle treated BT67 cells (3 biological replicated).

middle: K

first: Xiaoguang

last: Hao

first: Rozina

last: Hassam

first: Orsolya

last: Cseh

first: Danielle

middle: A

last: Bozek

first: H

last: Luchman

first: Samuel

last: Weiss


title: Interleukin-10/CSF1R fusion protein rejuvenates the exhausted T cells for executing antitumor activity V

city: Taipei

first: JuPei

address: No. 155, Sec. 2, Linong St. Beitou Dist.

country: Taiwan

institute: National Yang Ming Chiao Tung University

summary: Bulk RNA sequence (RNA-seq) has been used to explore the effect of IL10/CSF1R fusion protein modulation in tumor microenvironment.  Four groups were analyzed which contain Ctrl, CSF1R, IL10 and BF10.  MTC-Q1 oral cancer cells were orthotopic injection into C57BL/6J mice. tumor tissue has been isolated (~ 100 mm3) and total RNA were extracted by Trizol, followed by cDNA were sequenced by Illumina NextSeq 550.

last: Chang

first: Muh-Hwa

last: Yang


title: A biallelic multiple nucleotide length polymorphism explains functional causality at the 5p15.33 prostate cancer risk locus

status: Public on Jul 11 2023

city: Budapest

first: Andras

last: Furedi

state: HU

address: Magyar tudosok krt 2

department: Molecular Life Sciences

country: Hungary

institute: HUN-REN RCNS

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: RNA-seq analysis on the liver of darkness rats treated with Limosilactobacillus reuteri

city: Shanghai

first: Shang

last: Li

address: 845 Lingshan Road

country: China

institute: Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University

summary: We have previously demonstrated that the arrhythmic expressions of circadian clock genes due to constant darkness induce glycometabolic and reproductive hallmarks of polycystic ovary syndrome (PCOS) in rats. Limosilactobacillus reuteri (L.reuteri) is a promising dietary intervention for host dysmetabolism, while its potential effect on circadian dysrhythmia-induced PCOS remains elusive. Here, we evaluated the amelioration of L.reuteri regimen on constant darkness-induced PCOS-like rats through detecting hepatic gene expression profiles by RNA-seq.

overall_design: The control group was under a circadian rhythm of 12:12 h light-dark cycle (lights on at 7:30 a.m. and off at 7:30 p.m.), orally given with saline once a day. The darkness group and the darkness+L.reuteri group were respectively given with saline or L.reuteri (1010 CFU/ml, 100 mg/d) by daily oral gavage during 8-week constant darkness housing. RNA was isolated from 24 liver samples of 14-week-old female rats from the above 3 groups.

first: Junyu

last: Zhai

first: Yanzhi

last: Du


title: Restoring Tumor Immunogenicity with Dendritic Cell Reprogramming [ATAC-seq]

status: Public on Jul 07 2023

city: Lund

first: Ilia

last: Kurochkin

state: Skane

address: BMC A12, Sölvegatan 17

country: Sweden

institute: Lund University

summary: Decreased antigen presentation contributes to the ability of cancer cells to evade the immune system. We used the minimal gene regulatory network of type 1 conventional dendritic cells (cDC1) to reprogram cancer cells into professional antigen- presenting cells (tumor-APCs). Enforced expression of the transcription factors PU.1, IRF8, and BATF3 (PIB) was sufficient to induce cDC1 phenotype in 36 cell lines derived from human and mouse hematological and solid tumors. Within 9 days of reprogramming, tumor-APCs acquired transcriptional and epigenetic programs associated with cDC1 cells. Reprogramming restored the expression of antigen presentation complexes and costimulatory molecules on the surface of tumor cells, allowing the presentation of endogenous tumor antigens on MHC-I, and facilitating targeted killing by CD8+ T cells. Functionally, tumor-APCs engulfed and processed proteins and dead cells, secreted inflammatory cytokines, and cross- presented antigens to naïve CD8+ T cells. Human primary tumor cells could also be reprogrammed to increase their capability to present antigen and to activate patient-specific tumor-infiltrating lymphocytes. In addition to acquiring improved antigen presentation, tumor-APCs had impaired tumorigenicity in vitro and in vivo. Injection of in vitro generated melanoma-derived tumor-APCs into subcutaneous melanoma tumors delayed tumor growth and increased survival in mice. Antitumor immunity elicited by tumor-APCs was synergistic with immune checkpoint inhibitors. Our approach serves as a platform for the development of immunotherapies that endow cancer cells with the capability to process and present endogenous tumor antigens.

overall_design: Population ATAC-seq profiling of T98G human cancer cell line after transduction with polycystronic vectors encoding PU.1, IRF8 and BATF3 at day 3, 5, 7 and 9. ATAC-seq profilling of eGFP-transduced T98G cell line and freshly isolated peripheral blood Dendritic cells type 1 (HLA-DR+CD11C+CD141+) were used as controls.

first: Olga

last: Zimmermannova

first: Carlos-Filipe

last: Pereira


title: Transcriptome-wide RNA binding analysis of C9orf72 poly(PR) dipeptides

status: Public on Jul 10 2023

city: London

first: Adrian

last: Isaacs

address: Cruciform Building, Gower Street

department: Neurodegenerative Diseases

country: United Kingdom

institute: University College London

summary: An intronic GGGGCC repeat expansion in C9orf72 is a common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. The repeats are transcribed in both sense and antisense directions to generate distinct dipeptide repeat proteins, of which poly(GA), poly(GR) and poly(PR) have been implicated in contributing to neurodegeneration. Poly(PR) binding to RNA may contribute to toxicity, but analysis of poly(PR)-RNA binding on a transcriptome-wide scale has not yet been carried out. We therefore performed crosslinking and immunoprecipitation (CLIP) analysis in human cells to identify the RNA binding sites of poly(PR). We found that poly(PR) binds to nearly 600 RNAs, with the sequence GAAGA enriched at the binding sites. In vitro experiments showed that poly(GAAGA) RNA binds poly(PR) with higher affinity than control RNA and induces phase-separation of poly(PR) into condensates. These data indicate that poly(PR) preferentially binds to poly(GAAGA)-containing RNAs, which may have physiological consequences.

overall_design: Genome-wide iiCLIP interrogation of polyPR, intronic GGGGCC repeat expansion in C9orf72, binding to RNA.

first: Rubika

last: Balendra

first: Igor

last: Ruiz de los Mozos

first: Hana

middle: M

last: Odeh

first: Idoia

last: Glaria

first: Carmelo

last: Milioto

first: Katherine

last: Wilson

first: Agnieszka

last: Ule

first: Martina

last: Hallegger

first: Laura

last: Masino

first: Stephen

last: Martin

first: Rickie

last: Patani

first: James

last: Shorter

first: Jernej


title: Metabolic Programs of T Cell Tissue Residency Empower Tumor Immunity [RNA-seq]

city: La Jolla

first: Miguel

last: Reina-Campos

state: California

address: 9420 Athena Circle

country: USA

institute: La Jolla Institute for Immunology

summary: Tissue-resident memory CD8 T cells (TRM) offer fast, robust, and long-term protection at sites of re-infection1. Tumor-infiltrating lymphocytes (TIL) with characteristics of TRM maintain enhanced effector functions, predict responses to immunotherapy, and accompany better prognoses2,3. Thus, an improved understanding of the metabolic strategies that enable tissue residency could inform new approaches to empower T cell responses in tissues and solid tumors. To systematically define the basis for the metabolic reprogramming supporting TRM differentiation, survival, and function, we leveraged in vivo functional genomics, untargeted metabolomics, and transcriptomics of virus-specific memory CD8 T cell populations. We found that memory CD8 T cells deployed a range of adaptations to tissue residency, including a marked reliance on non-steroidal products of the mevalonate/cholesterol pathway, such as Coenzyme Q (CoQ), driven by increased activity of the transcription factor Srebp2. This metabolic adaptation was most pronounced in the small intestine (SI), where TRM interface with dietary cholesterol and maintain a heightened state of activation4, and was shared by functional TIL in diverse tumor types in mice and humans. Enforcing CoQ synthesis through Fdft1 deletion or Pdss2 overexpression promoted mitochondrial respiration, memory formation upon viral infection, and enhanced antitumor immunity. In sum, through a systematic exploration of TRM metabolism, we reveal how these programs can be leveraged to empower CD8 T cell memory formation in the context of acute infections and enhance antitumor immunity.

overall_design: RNAseq of sorted P14 CD8 T cells from mice at day 7 after infection with LCMV. P14 CD8 T cells were isolated from the spleen, liver, kidney and small intestine. All samples but spleen were gated on intravascular-associated negative cells based on in vivo CD8a stain prior to analysis. Every replicate has cells pooled from two mice.

first: Ananda

last: W Goldrath


title: Single cell RNAseq of peripheral blood mononuclear cells from children with mitochondrial disease

status: Public on Jul 06 2023

city: Bethesda

first: Peter

last: McGuire

state: MD

address: 10 Center Dr

institute: NHGRI

summary: Mitochondria are nearly ubiquitous in all cells, suggesting that mitochondrial dysfunction may extend to the immune system.  In this study, we utilized Single-Cell RNA sequencing (scRNA-seq) to characterize cellular heterogeneity and the pathologic underpinnings of mitochondrial disease in peripheral blood mononuclear cells from pediatric patients. Transcriptomes of individual cells from affected and healthy tissue samples revealed marked reductions in select populations involved in the humoral immune response, especially antigen presenting cells, B cell and plasma populations, with sparing of T cell populations. MTRNR2L8, a marker of bioenergetic stress, was significantly elevated in populations that were most depleted. mir4485, a miRNA contained in the intron of MTRNR2L8, was co-expressed. mir4485 was found to have a role in promoting survival by modulating apoptosis. The results highlight distinct molecular phenotypes and identify previously unrecognized in mitochondrial diseases.

overall_design:
Case-control single cell RNAseq
contributor: NISC Comparative Sequencing Program

>>>Submitter states: Raw data not available due to patient privacy concerns. Raw data will be submitted to dbGaP.<<<

middle: J

first: Payal

last: Banerjee


title: Nicotinamide mononucleotide (NMN) mediates alleviation of poly(I:C) induced inflammatory response in human primary cell cultures

city: Tokyo

first: Anton

last: Kratz

address: Saisei Ikedayama Bldg. 5-10-25 Higashi Gotanda Shinagawa

country: Japan

institute: The Systems Biology Institute

summary: NMN is the direct precursor of the nicotinamide adenine dinucleotide (NAD+) and is considered a key component to increase NAD+ levels and mitochondrial activity in cells. In this study, based on transcriptome analysis, we showed that NMN alleviates poly(I:C)-induced inflammatory response in two types of human primary cell cultures, human pulmonary microvascular endothelial cells (HPMEC) and human coronary artery endothelial cells (HCAEC). The major inflammatory mediators including IL-6 and PARP family members are grouped into co-expressed gene modules and significantly down-regulated under NMN-dosage in poly(I:C)-induced conditions in both cell types. The Bayesian network analysis of module hub genes predicted common genes including eukaryotic translation initiation factor 1B (EIF1B) and distinct genes such as platelet-derived growth factor binding molecules in HCAEC, which potentially regulate the identified inflammation modules. These results have suggested the robust regulatory mechanism of NMN to alleviate inflammatory pathways, which may open up the possibility of a new role for NMN replenishment in chronic or acute inflammation treatment.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for HPMEC and HCAEC, with and without Poly(I:C) treatment and with three dosages of NM  N (0.1, 1, 10 mM), with matching controls.

first: Hitomi

last: Sano

first: Taiko

last: Nishino

first: Haruna

last: Imamura

first: Yuki

last: Yoshida

first: Noriaki

last: Shimizu

first: Hiroaki

last: Kitano

first: Ayako

last: Yachie


title: Demethylase-independent roles of LSD1 in regulating enhancers and cell fate transition

first: Jiwei

last: Chen

address: 130 DongAn Road

institute: Fudan University


title: Demethylase-independent roles of LSD1 in regulating enhancers and cell fate transition [RNA-seq]

summary: Metazoan enhancers are decorated by mono-methylation (me1) of the lysine 4 residue on histone H3 (H3K4), a mark deposited by methyltransferases MLL3/MLL4 and removed by the lysine-specific histone demethylase 1A (LSD1 or KDM1A) via its flavin adenine dinucleotide (FAD)-dependent amine oxidase activity. As a component of histone deacetylases HDAC1/2-containing complex CoREST, LSD1 is required for animal development, and is implicated in Kabuki Syndrome-like congenital diseases and multiple types of cancer. Although prior research has investigated the demethylase function of LSD1 extensively, the mechanisms underlying LSD1’s role in development and diseases remain enigmatic. Here, we have utilized genetic, epigenetic, genomic, and cell biology approaches to dissect the role of LSD1 and its demethylase activity in gene regulation and cell fate transition. Surprisingly, the catalytic inactivation of LSD1 only has a mild impact on gene expression whereas the loss of LSD1 protein de-represses enhancers globally. Moreover, LSD1 deletion, rather than its catalytic inactivation, causes defects in spontaneous differentiation, the transition from naive to primed pluripotency, embryoid body formation, and cardiomyocyte differentiation. Interestingly, deletion of LSD1 increases H3K27ac levels and binding of P300 to LSD1-targeted enhancers. We further show that the gain in the level of H3K27ac catalyzed by P300/CBP, not the loss of CoREST complex components from chromatin, contributes to the transcription de-repression of LSD1 targets and differentiation defects caused by LSD1 loss. Taken together, our study demonstrates a demethylase-independent role of LSD1 in regulating enhancers and cell fate transition, providing insight into the treatment of diseases driven by LSD1 mutations and misregulation.

overall_design: Analyzing the role of LSD1 and its demethylase activity in gene regulation

first: Chen

last: Jiwei

first: Cao

last: Kaixiang


title: A group of WRKY transcription factors form a protein complex with PHD-containing proteins to repress the transcription of stress responsive genes

city: Beijing

first: xinjian

last: He

address: No. 7, zhongguancun life science park road, changping district

institute: National Institute of Biological Sciences


title: Coordination of widespread transcription at enhancers and target genes through BRD4 (ChIP-Rx)

city: Innsbruck

first: Univ.-Prof. Dr. Andreas

last: Mayer

address: Innrain 80/82

department: Institute for Genomics and RNomics

country: Austria

institute: Biocenter, Medical University Innsbruck

summary: Gene transcription by RNA polymerase II (Pol II) is under the control of promoters and distal regulatory elements known as enhancers. Enhancers are themselves transcribed by Pol II which correlates with their activity. How enhancer transcription is regulated and coordinated with transcription at target genes has remained unclear. Here, we developed a high-sensitive native elongating transcript sequencing approach, called HiS-NET-seq, to provide an extended high-resolution view on transcription, especially at lowly transcribed regions such as enhancers. HiS-NET-seq uncovers new transcriptional enhancers in human cells. An integrated multi-omics analysis shows that transcription at most enhancers depends on the BET protein BRD4. We provide evidence for a direct BRD4-mediated coupling of Pol II transcription at enhancers and target genes. Specifically, BRD4 links elongation activation at enhancers and genes by maintaining their proximity. These studies reveal that transcription at enhancers and target genes is coordinated at the level of elongation.

overall_design: Pol II occupancy profiling using standard NET-seq or HiS-NET-seq data for K562 WT cells and upon BRD4-specific degradation in K562 dTAG-BRD4 cells.

first: Annkatrin

last: Bressin

first: Olga

last: Jasnovidova

first: Mirjam

last: Arnold

first: Elisabeth

last: Altendorfer

first: Filip

last: Trajkovski

first: Thomas

first: Joanna

middle: E

last: Handzlik

first: Denes

last: Hnisz

first: Andreas


title: Interleukin-10/CSF1R fusion protein rejuvenates the exhausted T cells for executing antitumor activity III

first: JuPei

summary: Bulk RNA sequence (RNA-seq) has been used to explore the effect of IL10/CSF1R fusion protein modulation in tumor microenvironment.  Four groups were analyzed which contain Ctrl, CSF1R, IL10 and BF10.  MTC-Q1 oral cancer cells were orthotopic injection in C57BL/6 mice. CD45-negatve cells has been isolated and RNA were extracted by Trizol, followed by cDNA were sequenced by Illumina NextSeq 550.

overall_design: Examination of therapeutic modulation of IL10/CSF1R fusion protein within tumor microenvironment in mice cancer model.  RNA-seq of CD45-negatve cells from orthotopic cancer tissue after four dose treatments.

first: Yao-Wen

last: Chang

first: Muh-Hwa

last: Yang


title: Coordination of widespread transcription at enhancers and target genes through BRD4


title: Demethylase-independent roles of LSD1 in regulating enhancers and cell fate transition [ChIP-seq]

first: Jiwei

overall_design: ChIP-seq was performed to deterimine the genome-wide occupancy of chromatin proteins and histone modifications in WT and LSD1 mutants.


title: Interleukin-10/CSF1R fusion protein rejuvenates the exhausted T cells for executing antitumor activity II

summary: Bulk RNA sequence (RNA-seq) has been used to explore the effect of IL10/CSF1R fusion protein modulation in tumor microenvironment.  Four groups were analyzed which contain Ctrl, CSF1R, IL10 and BF10.  4T1 breast cancer cells were orthotopic injection into Balb/c mice. tumor tissue has been isolated (~ 100 mm3) and total RNA were extracted by Trizol, followed by cDNA were sequenced by Illumina NextSeq 550.


title: Interleukin-10/CSF1R fusion protein rejuvenates the exhausted T cells for executing antitumor activity

summary: Single cell sequence (SC-seq) has been used to explore the effect of IL10/CSF1R fusion protein modulation in tumor microenvironment.  Four groups were analyzed which contain Ctrl, CSF1R, IL10 and BF10.  MTC-Q1 oral cancer cells were orthotopic injection in C57BL/6 mice. CD45+ immune cells has been isolated, analyzed and sequenced by dropped based scRNA-seq, cDNA were sequenced by Illumina Novaseq 6000.

overall_design: Examination of therapeutic modulation of IL10/CSF1R fusion protein within tumor microenvironment in mice cancer model.  SC-seq of CD45+ immune infiltrated cells from orthotopic cancer tissue after four dose treatments.


title: Characterization of an Eye Field-like State during Optic Vesicle Organoid Development

city: Edinburgh

first: Liusaidh

middle: Jane

last: Owen

address: Crewe Road South

department: MRC HGU

institute: University of Edinburgh


title: Characterization of an Eye Field-like State during Optic Vesicle Organoid Development [ATAC-seq]

summary: Specification of the eye field (EF) within the neural plate marks the earliest detectable stage of eye development. Experimental evidence, primarily from non-mammalian model systems, indicates that the stable formation of this group of cells requires the activation of a set of key transcription factors (TFs). This critical event is challenging to probe in mammals and, quantitatively, little is known regarding the regulation of the transition of cells to this ocular fate. Using optic vesicle organoids to model the onset of the EF, we generate timecourse transcriptomic data allowing us to identify dynamic gene-expression programs that characterise this cellular-state transition. Integrating this with chromatin accessibility data suggests a direct role of canonical EFTFs in regulating these gene-expression changes, and highlights candidate cis-regulatory elements through which these TFs act. Finally, we begin to test a subset of these candidate enhancer elements, within the organoid system, by perturbing the underlying DNA sequence and measuring transcriptomic changes during EF activation.

overall_design: ATAC sequencing was performed on a timecourse through optic vesicle organoid differentiation grown from Rx-GFP mouse ESCs. A sample was collected every 24 hours from day 0 (defined as the day organoids are seeded) to day 3 and at day 5. On day 5, cells were sorted using FACS on the basis of GFP expression to produce 2 samples for this timpoint. GFP has been knocked into the Rax locus in this cell line, meaning GFP positive samples correspond to cells expressing Rax and committed to an eye field lineage.

first: Jacqueline

last: Rainger

first: Hemant

last: Bengani

first: Fiona

last: Kilanowski

first: David

middle: R

last: FitzPatrick

first: Andrew

middle: S

last: Papanastasiou


title: Restoring Tumor Immunogenicity with Dendritic Cell Reprogramming [mouse RNA-Seq]

first: Ilia

last: Kurochkin

overall_design: Population mRNAseq profiling on Lewis Lung Carcinoma (LLC) and B16 melanoma murine cancer cell lines after transduction with polycystronic vectors encoding PU.1, IRF8 and BATF3 at day 3, 5, 7 and 9. mRNA profilling of eGFP-transduced LLC and B16 were used as controls (d0).

first: A

middle: G

last: Ferreira

first: E

last: Ascic

first: I

last: Caiado

first: O

last: Zimmermannova

first: F

last: Pereira


title: Hearing of malaria mosquitoes is modulated by a beta-adrenergic-like octopamine receptor and serves as insecticide target

status: Public on Jul 05 2023

first: Marta

last: Andrés

address: 332 Grays Inn Rd

department: Ear Institute

summary: Malaria mosquitoes acoustically detect their mating partners within large swarms that form transiently at dusk. Indeed, male malaria mosquitoes preferably respond to female flight tones during swarm time. This phenomenon implies a sophisticated context- and time-dependent modulation of mosquito audition, the mechanisms of which are largely unknown. Using transcriptomics, we identify a complex network of candidate neuromodulators regulating mosquito hearing in the species Anopheles gambiae. Among them, octopamine stands out as auditory modulator during swarm time. In-depth analysis of octopamine auditory function shows that it affects the mosquito ear on multiple levels: it modulates the tuning and stiffness of the flagellar sound receiver and controls the erection of antennal fibrillae. We show that two α- and β-adrenergic-like octopamine receptors drive octopamine’s auditory roles and demonstrate that the octopaminergic auditory control system can be targeted by insecticides. Our findings highlight octopamine as key for mosquito hearing and mating partner detection, and as a potential novel target for mosquito control.

overall_design: To investigate expression, and circadian pattern of expression of transcripts in Anopheles gambiae mosquito ears we conducted the following: First, we entrained mosquitoes, in environmental incubators, to 12H:12H light/dark cycle with 1-hour ramped light transition to simulated dawn and dusk (ZT0-ZT1 and ZT12-ZT13, respectively; ZTX is the formalised notation of an entrained circadian cycle's phase). Temperature and relative humidity were set constantinside the incubators throughout the circadian day, at 28 °C and 80% respectively. Mosquitoes were exposed to the circadian entrainment in the incubators for three days before performing any experiment. Then, on day four of circadian entrainment, 35 male and 35 female mosquitoes were removed from the incubators at six different circadian time points (ZT0, ZT4, ZT8, ZT12, ZT16, ZT20), and we collected their Johnston's organs (JO; the mosquito's "internal ear"). This procedure was repeated 3 times, resulting in three biological replicates of JO samples at each circadian time point. Finally, we performed RNA sequencing of the samples, and conducted transcript expression, differential expression (between males and females), and circadian expression analyses on the sequenced data.

first: Marcos

last: Georgiades

first: Alexandros

middle: C

last: Alampounti

first: Jason

last: Somers

first: Matthew

middle: P

last: Su

last: Ellis

first: Judit

last: Bagi

first: Scott

last: Tytheridge

first: Watson

last: Ntabaliba

first: Sarah

last: Moore

first: Joerg

middle: T

last: Albert


title: First-line durvalumab and tremelimumab with chemotherapy in RAS-mutated metastatic colorectal cancer: a phase 1b/2 trial [RNA-Seq]

city: DIJON

first: François

last: Ghiringhelli

address: 1 rue du Professeur Marion

country: France

institute: UMR INSERM 1231

summary: While patients with microsatellite instable, metastatic colorectal cancer (CRC) benefit from immune checkpoint blockade, chemotherapy with targeted therapies remains the only therapeutic option for microsatellite stable (MSS) tumors. The single arm, phase IB/II MEDITREME trial evaluates safety and efficacy of durvalumab plus tremelimumab in combination with mFOLFOX6 chemotherapy in first line, in 57 patients with RAS-mutant unresectable mCRC. Safety was primary objective of phase IB and no safety issue was observed. The primary objective of phase II was efficacy in terms of 3-month PFS in MSS patients, which was met with a 3 month PFS of 90.7% [95% CI: 79.2-96%]. For secondary objective, response rate was 64.5%, median PFS was 8.2 months [95% CI: 5.9–8.6] and overall survival was not reached in MSS patients.  Higher tumor mutational burden and a lower degree of genomic instability in responder patients. Integrated transcriptomic analysis underlined that high immune signature, and low epithelio-mesenchymal transition were associated with better outcome. Immunomonitoring showed induction of neoantigen and NY-ESO1 and TERT blood tumor specific T cell response associated with better PFS. The combination of durvalumab-tremelimumab with mFOLFOX6 was tolerable with promising clinical activity in MSS mCRC.

overall_design: We performed gene expression profiling analysis using RNA-seq data from 34 patients splited into responders and non-responders

last: Ghiringelli

first: Marion

last: Thibaudin

first: Caroline

last: Truntzer


title: Postnatal cecal and cecal content luminal miRNA signatures in C57BL/6-VAF/Elite® mice

status: Public on Jul 08 2023

city: Toronto

first: Elena

last: Comelli

state: Ontario

address: 1 king's college circle, MSB5354

department: Nutritional Sciences

institute: University of Toronto

summary: The interplay between the intestinal microbiota and host is critical to intestinal ontogeny and homeostasis. MicroRNAs (miRNAs) may be an underlying link. Intestinal miRNAs are microbiota-dependent and when shed in the lumen, affect resident microorganisms. Yet, longitudinal relationships between intestinal tissue miRNAs, luminal miRNAs, and luminal microorganisms have not been elucidated, especially in early life. Here, we investigated the postnatal cecal miRNA and microbiota populations, their relationship, and their impact on intestinal maturation in specific and opportunistic pathogen free mice; we also assessed if they can be modified by an intervention with allochthonous probiotic lactobacilli. We report that cecal and cecal content miRNA and microbiota signatures are temporally regulated, correlated, and modifiable by probiotics with implications for intestinal maturation. These findings help with understanding causal relationships within the gut ecosystem and provide a basis for preventing and managing their alterations in diseases throughout life.

overall_design: C57BL/6-VAF/Elite® 6-8-week-old female mice were randomized to the control (n=7, receiving sterile drinking water) or probiotics (n=6; receiving a probiotic mixture of Lacticaseibacillus rhamnosus R0011 and L. helveticus R0052 (ratio 95:5) in the drinking water) group starting one week before breeding. On Day 7 (D7), mice were bred harem-style. Pregnant dams were housed individually and continued receiving sterile water with or without probiotics throughout the pregnancy until parturition at D28 (postnatal day (PND) 0). The litters were followed longitudinally; offspring were euthanized at PND14, PND21, and PND36, and cecum and its content (paired) were collected for total RNA extraction and miRNA analysis. For the control group, a total of 12 samples were analyzed: 1 male offspring/dam/PND (n = 4 male offspring/PND).  For the probiotics group, a total of 12 samples were analyzed across all dams: n = 3, n = 5, and n = 4 male offspring for PND14, PND21, and PND36, respectively.

first: Amel

last: Taibi

first: Tomas

last: Tokar

first: Giorgio

last: Gargari


title: Landscape of immune checkpoint blockade-induced immune alterations in the microenvironment of primary and metastatic brain tumors

city: Los Angeles

first: Lu

last: Sun

address: 10833 Le Conte Avenue

institute: University of California, LA

summary: Brain tumors reside in an immune privileged microenvironment where inflammatory processes are heavily regulated. Nevertheless, our recent study showed a significant immune infiltration in recurrent glioblastoma (rGBM) treated with neoadjuvant PD-1 checkpoint blockade therapy. Compared to GBM, the intracranial immunotherapy response of brain metastases (BrM) has been much less studied. Therefore we studied the effect of pre-surgical immune checkpoint blockade (ICB) on the immune compartments of BrM, using multiplex immunofluorescence, mass cytometry and single-cell RNA sequencing. ICB induced much higher T cell infiltration in BrM compared to rGBM. It also caused the loss of a unique perivascular macrophage subset (CD206+LYVE1+), the absence of which associated with the enhanced T cell infiltration in BrM. These T cells produced high levels of IFN-γ that elicited and interferon response transcriptional program in the tumor-associated myeloid cells.  T cell-myeloid interactions were also strengthened by ICB in BrM and might have led to the terminal exhaustion of the infiltrating T cells. Our results emphasize the importance of adopting novel immune-boosting approaches to rejuvenate the systemic T cell pool so as to improve tumor control of brain metastases.

overall_design: We processed a total of 18 single cell transcriptomes of metastatic brain tumors with/without prior immune checkpoint blockade treatment. One of the sample, LB3935T, was excluded from the combined counts matrix and subsequent integrated analysis, due to low cell number (<=100 cells).

first: Jenny

last: Kienzler

first: Jeremy

last: Reynoso

first: Alexander

last: Lee

first: Geoffrey

last: Owens

first: Richard

last: Everson

first: Linda

last: Liau

first: Willy

last: Hugo

first: Won

last: Kim

first: Robert

last: Prins


title: Depletion of pyruvate kinase (PK) activity causes glycolytic intermediate imbalances and reveals a PK-TXNIP regulatory axis [RNA-seq]

city: Oberschleißheim

last: Schneider

address: Ingolstädter Landstraße 1

department: Institute of Functional Epigenetics

country: Germany

institute: Helmholtz Zentrum München

summary:
Objective
Cancer cells convert more glucose into lactate than healthy cells, what results in their growth advantage. Pyruvate kinase (PK) is a key rate limiting enzyme in this process, what makes it a promising potential therapeutic target. However, currently it is still unclear what consequences the inhibition of PK has on cellular processes. Here, we systematically investigate the consequences of PK depletion for gene expression, histone modifications and metabolism.
Methods
Epigenetic, transcriptional and metabolic targets were analysed in different cellular and animal models with stable knockdown or knockout of PK.
Results
Depleting PK activity reduces the glycolytic flux and causes accumulation of glucose-6-phosphate (G6P). Such metabolic perturbation results in stimulation of the activity of a heterodimeric pair of transcription factors MondoA and MLX but not in a major reprogramming of the global H3K9ac and H3K4me3 histone modification landscape. The MondoA:MLX heterodimer upregulates expression of thioredoxin-interacting protein (TXNIP) – a tumour suppressor with multifaceted anticancer activity. This effect of TXNIP upregulation extends beyond immortalised cancer cell lines and is applicable to multiple cellular and animal models.
Conclusions
Our work shows that actions of often pro-tumorigenic PK and anti-tumorigenic TXNIP are tightly linked via a glycolytic intermediate. We suggest that PK depletion stimulates the activity of MondoA:MLX transcription factor heterodimers and subsequently, increases cellular TXNIP levels. TXNIP-mediated inhibition of thioredoxin (TXN) can reduce the ability of cells to scavenge reactive oxygen species (ROS) leading to the oxidative damage of cellular structures including DNA. These findings highlight an important regulatory axis affecting tumour suppression mechanisms and provide an attractive opportunity for combination cancer therapies targeting glycolytic activity and ROS-generating pathways

overall_design: Differential gene expression analysis of RNA-seq data between HEK293 and MCF-7 cells with PKM enzyme knocked down and their wild-type (WT) equivalents

first: Anna

last: Nieborak

first: Saulius

last: Lukauskas


title: Human prostate organoid generation and the identification of prostate development drivers using inductive rodent tissues

city: West Lafayette

first: Nadia

middle: Marie

last: Atallah

state: IN

address: 201 S. University St

department: Center for Cancer Research

institute: Purdue University

summary: Mechanistic links between development and disease can be exploited to provide clues to the cellular signalling pathways that might become re-activated during adult pathogenesis such as those affecting the prostate. However, the mechanisms underpinning prostate development require further characterisation. Previously, our group developed novel methods to produce prostate organoids using induced pluripotent stem cells (iPSCs). Here, we show that the iPSCs can be fully differentiated into prostate organoids using neonatal rodent seminal vesicle mesenchyme (SVM) in vitro. Additionally, we elucidated molecular drivers of prostate induction through RNA-sequencing analyses of the rat urogenital sinus (the embryonic precursor of the prostate gland) as well as the neonatal seminal vesicles. We identified candidate drivers evident in the inductive mesenchyme and epithelium involved with prostate specification. Our top candidates included Spx, Trib3, Snai1, Snai2, Nrg2, and Lrp4. This work lays the foundations to further interrogate the reactivation of developmental genes in adulthood, leading to prostate disease.

overall_design: Tissues from rats known to induce prostate development were isolated. RNA-sequencing of the tissues identified candidate drivers of prostate development. Co-culture of seminal vesicles with human iPSCs in vitro generated prostate organoids.

first: Parmveer

last: Singh

last: Lanman

first: Hannah

middle: L

last: Kendall

first: Laura

last: Wilson

first: Ryan

last: Long

first: Omar

last: Franco

first: Adriana

last: Buskin

first: Colin

last: Miles

first: Simon

middle: W

last: Hayward

first: Rakesh

last: Heer

first: Craig

middle: N

last: Robson


title: Depletion of pyruvate kinase (PK) activity causes glycolytic intermediate imbalances and reveals a PK-TXNIP regulatory axis [ChIP-seq]

overall_design: ChIP with reference exogenous genome (ChIP-Rx) for histone modifications H3K9ac, H3K4me3 as well as transcription factors MondoA and MLX in wild-type (WT) HEK293 cells and HEK293 cells with PKM enzyme knocked down.


title: A Pilot Study Of Immunological and Microbiome Profiling Reveals Distinct Inflammatory Profiles For Smokers and Electronic Cigarette Users [re-analysis]

city: Columbus

first: Peter

last: Shields

state: OH

address: 420 West 12th Ave

institute: The Ohio State University

summary:
Background: Smoking increases pulmonary inflammation, but an effect by electronic cigarette (EC) vaping is less understood. We previously reported smokers (SM) had increased lung immune cell counts and inflammatory gene expression in bronchial epithelial cells compared to EC users and never-smokers (NS). Here we report association of smoking and vaping with immune cell subtypes and gene expression in bronchoalveolar lavage.

Methods: SM, EC users, and NS underwent bronchoscopy (n=28).  RNASeq and the CIBERSORT computational algorithm were used to determine immune cell subtypes, along with inflammatory gene expression and microbiome metatranscriptomics. Correlations and associations were assessed across and within the tobacco-use groups, corrected for false discovery rates.

Results: Classification of macrophage subtypes revealed a 2-fold increase in M0 macrophages for smokers and EC users relative to never-smokers, with a concordant decrease in M2 macrophages. There were 68, 19, and 1 significantly differentially expressed inflammatory genes (DEG) (FDR<0.05; log2-fold change>1) between SM/NS, SM/EC users, and EC users/NS respectively. CSF-1 and GATA3 expression correlated positively and inversely with M0 and M2 macrophages respectively. Correlation profiling for DEG showed distinct lung profiles for each participant group. There were 3 bacteria genera-DEG correlations and 3 bacteria genera-macrophage subtype correlations (Diffcorr>0.5 and FDR<0.1).

Conclusions:  In this pilot study, smoking and EC use were associated with an increase in undifferentiated M0 macrophages, but smokers differed from EC users and NS for inflammatory gene expression. The preliminary data support the hypothesis that smoking and EC use have toxic lung effects  influencing inflammatory responses, but not via changes in the microbiome.

overall_design:
Inflammatory gene expression profiling by high throughput sequencing (RNA-seq)

The re-analyzed GSE180785 Samples (28 out of 54) have been re-accessioned for the convenient retrieval of the complete raw data from SRA.

Please note that, during the lung microbiome analysis, one subject (102B(T2351_PS16)) was removed from the analysis for being an outlier, and thus the bacteria genus csv only contains 27 data column.

first: Kevin

last: Ying

first: Ewy

last: Mathé


title: Demethylase-independent roles of LSD1 in regulating enhancers and cell fate transition [scRNA-seq]

first: Jiwei

last: Chen

overall_design: WT and LSD1KO  day 6 EBs were dissociated with TrpLE and cells wer analyzed using scRNA-seq

first: Chen

last: Jiwei

first: Cao

last: Kaixiang


title: Alterations in lysosomal gene expression and lipid homeostasis in patient-derived liver organoids with acid sphingomyelinase deficiency

status: Public on Jul 04 2023

city: Madrid

first: GEMA

last: GOMEZ-MARIANO

address: Carretera de Majadahonda Pozuelo

country: Spain

institute: ISCIII

summary: We generated hepatic organoids from Nieman Pick Type B patients with the aim to investigate their lipid status.  To discover genes related to lipid metabolism likely involved in lipid deposits.

overall_design: Comparative gene expression analysis of RNA-seq data of hepatic organoids derived from Niemann Pick Type B (ASMD-B)

first: Gema

last: Gómez-Mariano

first: Sara

last: Pérez-Luz

first: Beatriz

last: Martínez-Delgado


title: Coordination of widespread transcription at enhancers and target genes through BRD4 (His-NET-Seq)

first: Univ.-Prof. Dr. Andreas

last: Mayer

first: Annkatrin

last: Bressin

first: Olga

last: Jasnovidova

first: Mirjam

last: Arnold

first: Elisabeth

last: Altendorfer

first: Filip

last: Trajkovski

first: Thomas

last: Kratz

first: Joanna

last: Handzlik

first: Denes

last: Hnisz

first: Andreas


title: Distinct roles for TUP1 and CYC8 in the regulation of gene transcription in yeast

city: Dublin 2

first: Alastair

last: Fleming

address: College Green

department: Microbiology Dept.

country: Ireland

institute: Trinity College Dublin

summary: The Tup1-Cyc8 complex in Saccharomyces cerevisiae was one of the first global co-repressors of gene transcription discovered. The model for Tup1-Cyc8 function proposes that Tup1p mediates the repression activity, whilst Cyc8p recruits the complex to target genes. However, despite years of study, a full understanding of the contribution of Tup1p and Cyc8p to complex function is lacking. We examined TUP1 and CYC8 single and double deletion mutants and show that CYC8 represses more genes than TUP1, and there are genes subject to (i) unique repression by TUP1 or CYC8, (ii) redundant repression by TUP1 and CYC8, and (iii) there are genes at which de-repression in a cyc8 mutant is dependent upon TUP1, and vice-versa. We also reveal that Tup1p and Cyc8p can make distinct contributions to commonly repressed genes possibly via specific interactions with histone deacetylases. Furthermore, we show that Tup1p and Cyc8p can be found independently of each other to negatively regulate gene transcription and can persist at active genes to negatively regulate on-going transcription. Together, these data suggest that Tup1p and Cyc8p can associate with inactive and active genes to mediate distinct negative and positive regulatory roles when functioning within, and possibly out with the complex.

overall_design: We carried out RNA-Seq analysis to compare the transcriptomes of the tup1 and cyc8 single mutants and the tup1 cyc8 double mutant strain, to wt. Three replicate RNA samples are included for each strain.

first: Brenda

last: Lee

middle: B


title: Renal transcriptome of the desert adapted Jaculus jaculus during water restriction and rehydration

city: Bristol

first: Benjamin

middle: Thomas

last: Gillard

address: Dorothy Hodgkin Building

department: Bristol Medical School

institute: University of Bristol

summary: Jaculus jaculus were split into three experimental groups: control (water access ad libitum), dehydrated (11 days water restriction), and rehydrated (7 days water restriction followed by 3 days water ad libitum). Kidney samples were extracted and sequenced to investigate changes in gene expression during osmotic challenge in a desert adapted model.

overall_design: Whole Jaculus jaculus left kidney samples from control (n=4), dehydrated (n=4), rehydrated (n=5) groups

first: Nabil

last: Amor

first: Fernando

last: Alvira-Iraizoz

first: Audrys

last: Pauza

first: Colin

last: Campbell

first: Michael

last: Greenwood

first: Abdulaziz

last: Alagaili

first: David

last: Murphy


title: Metabolic regulation of CTCF expression and chromatin association dictates starvation response in mice and flies

city: Odense M

first: Babukrishna

last: Maniyadath

address: Campusvej 1

department: Department of Biochemistry and Molecular Biology

country: Denmark

institute: University of Southern Denmark

summary: Temporal and coordinated control of gene expression is vital to mount appropriate physiological responses, especially in the context of dynamic metabolic transitions. Despite this, the interplay between chromatin architectural proteins and metabolism in regulating transcription is largely unknown. Here, we demonstrate that CTCF, the transcriptional regulator and boundary insulating protein, is itself regulated by metabolic inputs and its expression is associated with fed/fasted states. Although CTCF is well established as a key determinant of gene expression programs, our results indicate that its loci-specific functional diversity is associated with physiological plasticity during fed-fast cycles. Intriguingly, CTCF ChIP-seq and its differential expression point towards a paradoxical and yet tuneable binding to chromatin, as a function of hepatic metabolic status. Given emerging literature that indicates loci-/affinity-dependent functions of CTCF, we illustrate a physiological setting for such disparate association that is possibly mediated by the abundance of a non-coding RNA, Jpx. Further, meta-analyses of fed-fast dependent global transcriptional response and CTCF occupancy hint at the crucial role of CTCF in dictating hepatic gene expression and metabolism. Importantly, altering CTCF expression influenced the temporal cascade of metabolic gene expression. Heterologous perturbation of CTCF, to levels that mimic physiological oscillation, impacted mitochondrial energetics and lipidome profiles in hepatocytes. In addition to demonstrating the significance of CTCF expression and its differential chromatin association, our study also highlights the interplay between metabolism and chromatin organizers.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for boundary insulator CTCF in ad-libitum fed and 24-hours starved mice liver.

first: Devashish

last: Sen

first: Subhash

last: Khatri

first: Arnab

last: Chakraborty

first: Neelay

last: Mehendale

first: Snigdha

last: Nadagouda

first: Arshdeep

last: Kaur

first: Siddhesh

last: Kamat

first: Ullas

last: Kolthur-Seetharam

first: Shreyam

last: Chowdhury

first: U

last: Sandra


title: Coordination of widespread transcription at enhancers and target genes through BRD4 (HiChIP)

first: Univ.-Prof. Dr. Andreas

last: Mayer

overall_design: Comparative analysis of 3D interactions using HiChIP data upon BRD4-specific degradation in K562 dTAG-BRD4 cells.

first: Annkatrin

last: Bressin

first: Olga

last: Jasnovidova

first: Mirjam

last: Arnold

first: Elisabeth

last: Altendorfer

first: Filip

last: Trajkovski

first: Thomas

middle: A

last: Kratz

first: Joanna

last: Handzlik

first: Denes

last: Hnisz

first: Andreas


title: Endothelial CD34 expression and regulation of immune cell response in-vitro

city: Paris-cedex 10

first: Antonio

middle: José

last: Alberdi

state: Ile-de-France

address: 1, Av Claude Vellefaux

department: Institut de REcherche Saint Louis

institute: Université Paris Cité

summary:
Endothelial cells cover the lining of different blood vessels and lymph nodes, and have major functions including the transport of blood, homeostasis, inflammatory responses, control of infiltration of circulating cells into the tissues, and formation of new blood vessels. Therefore, understanding these cells is of major interest. The morphological features, phenotype and function of endothelial cells varies according to the vascular bed examined. The sialomucin, CD34, is widely used as an endothelial marker. However, CD34 is differentially expressed on endothelial cells in different organs and in pathological conditions.  Little is known about regulation of endothelial CD34 expression or function. This marker is also strongly modified in-vitro in endothelial cell models, including human umbilical vein endothelial cells (HUVEC) and endothelial progenitor cells (EPC). We have therefore selected CD34high and CD34low endothelial cell subpopulations and examined their CD34 expression and function.
Results: Transcriptomic analysis showed that CD34 gene and protein expressions are highly correlated, that CD34high cells are less proliferative, and have a stronger expression of IL-33 and Angiopoietin 2, compared to CD34low cells. Higher secretion levels of IL-33 by CD34high HUVECs was confirmed by ELISA. Finally, when endothelial cells were allowed to interact with peripheral blood mononuclear cells (PBMCs), CD34high endothelial cells induced a stronger proliferation of regulatory T lymphocytes (Tregs) compared to CD34low cells whereas expansion of other CD4+-T cell subsets was equivalent. These results suggest that CD34 expression by endothelial cells in-vitro associates with their ability to proliferate and with an immunogenic ability that favours the tolerogenic response.

overall_design: Gene expression analysis were performed on CD34high and CD34low endothelial colony-forming cells (ECFs).

first: Lousineh

last: Arakelian

first: Julien

last: Lion

first: Guillaume

last: Churlaud

first: Rezlene

last: Bargui

first: Briac

last: Thierry

first: Evelyne

last: Mutabazi

first: Patrick

last: Bruneval

first: Christelle

last: Doliger

first: Jérôme

last: Larghero

first: Nuala

last: Mooney


title: RTEL1 is involved in transcriptional gene silencing and inhibition of heterochromatic DNA methylation

city: Clermont-Ferrand

first: Olivier

last: Mathieu

state: Auvergne

address: 28 Place Henri Dunant

department: UMR 6293 - iGReD

institute: CNRS

type: Methylation profiling by high throughput sequencing

summary: Transcriptional gene silencing is an essential mechanism for controlling the expression of genes and transposable elements (TEs) and is commonly associated with high levels of DNA methylation. Thus, release of silencing is frequently observed in mutants for genes involved in DNA methylation maintenance. However, studies in Arabidopsis thaliana have revealed that transcriptional repression of some DNA-methylated genes and TEs also relies on repressors that operate through pathways largely independent of DNA methylation. One such pathway appears linked to DNA replication and DNA repair and mutations in factors involved in these processes such as the polymerases alpha, delta and epsilon destabilize silencing. Here, we isolate a new mutant allele of the helicase REGULATOR OF TELOMERE ELONGATION 1 (RTEL1) gene from a genetic screen for TGS-deficient mutants. RTEL1 is highly conserved among eukaryotes and plays an important role in the resolution of DNA secondary structures during DNA replication. We show that rtel1 causes release of TGS without loss of DNA methylation or global changes in chromatin accessibility. However, many heterochromatic loci show increased levels of DNA methylation and chromatin accessibility in rtel1 mutants. Our findings reveal a function for RTEL1 in maintaining silencing and in preventing DNA hypermethylation of heterochromatin.

overall_design: RNA-seq of wild-type, rtel1, mom1 and rtel1 mom1 mutant plants; ATAC-seq of wild-type and rtel1 seedlings; ChIP-seq for H3K27me3 in wild-type and rtel1 seedlings.

first: Amy

last: Hesketh

first: Margaux

last: Olivier

first: Marie-Noëlle

last: Pouch-Pélissier

first: Thierry

last: Pélissier

first: David

last: Latrasse

first: Ying

last: Huang

first: Moussa

last: Benhamed


title: Characterization of an Eye Field-like State during Optic Vesicle Organoid Development [RNA-seq]

first: Liusaidh

last: Owen

overall_design: RNA sequencing was performed on a timecourse through optic vesicle organoid differentiation grown from Rx-GFP mouse ESCs. Samples were collected in biological triplicates every 24 hours from day 0 (defined as the day organoids are seeded) to day 5. At days 4 and 5 cells were sorted using FACS on the basis of GFP expression to produce 2 samples for each of these timpoints. GFP has been knocked into the Rax locus in this cell line, meaning GFP positive samples correspond to cells expressing Rax and committed to an eye field lineage.

middle: J

first: Jacqueline

last: Rainger

first: Hemant

last: Bengani

first: Fiona

last: Kilanowski

last: FitzPatrick

first: Andrew

last: Papanastasiou


title: Characterizing control of memory CD8 T cell differentiation by BTB-ZF transcription factor Zbtb20 using single cell RNA-sequencing (scATAC-Seq)

status: Public on Jul 03 2023

city: Lebanon

first: Edward

last: Usherwood

state: NH

address: Rubin Building, Level 7, 710-53

department: Microbiology and Immunology

institute: Geisel School of Medicine at Dartmouth College

summary: Members of the BTB-ZF transcription factor family regulate the immune system. Our lab identified that family member Zbtb20 contributes to the differentiation, recall responses and metabolism of CD8 T cells. Here, we report a characterization of the transcriptional and epigenetic signatures controlled by Zbtb20 at single-cell resolution during the effector and memory phases of the CD8 T cell response. Without Zbtb20, transcriptional programs associated with memory CD8 T cell formation were upregulated throughout the CD8 T response. A signature of open chromatin was associated with genes controlling T cell activation, consistent with the known impact on differentiation. Additionally, memory CD8 T cells lacking Zbtb20 were characterized by open chromatin regions with overrepresentation of AP-1 transcription factor motifs and elevated RNA- and protein-level expression of the corresponding AP-1 components. Finally, we describe motifs and genomic annotations from the DNA targets of Zbtb20 in CD8 T cells identified by CUT&RUN. Together, these data establish the transcriptional and epigenetic networks contributing to the control of CD8 T cell responses by Zbtb20.

overall_design: 4 samples of murine OT-I CD8 T cells responding to Listeria-actA-OVA infection. Each sample is the result of pooling OT-I cells from the splenocytes of four individual mice. Of these samples, 2 were collected 10 days post-infection and 2 were collected 35 days post-infection. At each time point, 1 sample is WT OT-I cells and 1 sample is Zbtb20 KO OT-I cells.

first: Nicholas

middle: K

last: Preiss

first: Owen

middle: M

last: Wilkins


title: Depletion of pyruvate kinase (PK) activity causes glycolytic intermediate imbalances and reveals a PK-TXNIP regulatory axis.

first: Robert

last: Schneider

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

overall_design: Refer to individual Series


title: DNA Methylome and 3D Chromatin Landscape of Cell Types Across the Mouse Brain [6H, 7A]

city: La Jolla

first: Joseph

middle: R

last: Ecker

state: CA

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

summary: Illustrating the cellular architecture of the mammalian brain is critical to understanding its diverse functions and complex animal behaviors. With support from NIMH and NINDS as part of the BRAIN Initiative Cell Census Network (BICCN), the Center for Epigenomics of the Mouse Brain Atlas (CEMBA) applied single nucleus methylation sequencing in 117 brain funtional regions across the whole adult male P56 C57BL/6J mouse brain. In each major brain regions, we identified distinct cell clusters and formed them in a hierarchical taxonomy. These clusters include known primary brain cell types and possible sub-types. We use these data to identify the epigenomic characteristics and to define specific regulatory elements for each cell cluster. We integrate the single cell methylome with single cell gene expression and chromatin accessibility profiles from the same brain region to further identify potential enhancers and their corresponding genes for several neuronal cell types. By brain region spacial comparisons, we found spatial specificity among excitatory neuronal clusters within the same cortical layer. We identify brain-region-specific transcription factors that may regulate cortical region development of excitatory neurons. These data define the landscape of cell type and spatial heterogeneity in the mouse brain and the underlining regulatory epigenomic basis.

overall_design: Apply snmC-seq2 to 117 brain functional regions to identify epigenomic landcape of the adult mouse brain, this subseries consists of one single nucleus methylation sequencing experiment of brain region (6H_200312, 6H_200319, 7A_191008, 7A_191017).

first: Hanqing

last: Liu

first: Anna

last: Bartlett

first: Jacinta

last: Lucero

first: Julia

last: Osteen

first: Jordan

last: Altshul

first: Mia

last: Kenworthy

first: Rosa

last: Castanon

last: Nery

first: Peter

last: Berube

first: Huaming

last: Chen

first: Michael

last: Nunn

first: Jesse

last: Dixon

first: M

last: Behrens


title: Characterizing control of memory CD8 T cell differentiation by BTB-ZF transcription factor Zbtb20 using single cell RNA-sequencing (scRNA-Seq)

first: Edward

last: Usherwood

last: Preiss


title: Histone deacetylation and cytosine methylation are required for the normal compartmentalization of heterochromatin in the genome organization of Neurospora crassa

city: Colorado Springs

first: Andrew

middle: David

last: Klocko

state: Colorado

address: 278 Centennial Hall, 1420 Austin Bluffs Pkwy

department: Chemistry and Biochemistry

institute: University of Colorado Colorado Springs


title: SMAD4 maintains the Fluid Shear Stress set point to protect against Arterial-Venous Malformations

city: Mannheim

first: Roxana

last: Ola

state: Baden-Württemberg

address: Ludolf-Krehl-Str. 13 - 17

institute: Medical Faculty Mannheim, Heidelberg University

summary: Vascular networks form, remodel and mature under the influence of both fluid shear stress (FSS) and soluble factors. Physiological FSS promotes and maintains vascular stability via synergy with Bone Morphogenic Protein 9 (BMP9) and BMP10. Conversely, mutation of the BMP receptors ALK1, Endoglin or the downstream effector SMAD4 leads to Hereditary Hemorrhagic Telangiectasia (HHT), characterized by fragile and leaky arterial-venous malformations (AVMs). But how endothelial cells (EC) integrate FSS and BMP signals in vascular development and homeostasis, and how mutations give rise to vascular malformations is not well understood. Here, we aimed to elucidate the mechanism of synergy between fluid shear stress and SMAD signaling in vascular stability and its failure in HHT. We have now found that loss of Smad4 increases ECs’ sensitivity to flow by lowering the FSS set point with resulting AVMs exhibiting features of excessive flow-mediated morphological responses. Mechanistically, loss of Smad4 disinhibits flow-mediated Klf4-Tie2-PI3K/Akt signaling leading to increased EC proliferation and loss of arterial identity due to Klf4-mediated repression of cyclin dependent Kinase (CDK) inhibitors, CDKN2A and CDKN2B. Thus, AVMs caused by Smad4 deletion are characterized by chronic high flow remodeling with excessive EC proliferation and loss of arterial identity as triggering events.

overall_design: Gene expression profiling high throughput sequencing of HUVECS transfected with Control or SMAD4 siRNA in the presence or absence of 12dynes/cm2 fluid shear stress

first: Kuheli

last: Banerjee

first: Yanzhu

last: Lin

first: Johannes

last: Gahn

first: Julio

last: Cordero

first: Purnima

last: Gupta

first: Islam

last: Mohamed

first: Mariona

last: Graupera

first: Gergana

last: Dobreva

first: Martin

last: Schwartz


title: Gene expression profile at single cell level of breast cancer cells or stromal cells from a 3D culture model and cells forming mammary tumor in mouse breast cancer model

city: Atlanta

first: Manoj

last: Bhasin

state: GA

address: 615 Michael Street, Whitehead Biomedical Research Building

department: Department of Pediatrics

institute: Emory University

summary: Single-cell transcriptomics to investigate shows a reprogramming of awakened cancer cells including several survival cues such as stemness, chemoresistance, as well as an altered tumor microenvironment with augmented pro-tumor immune signaling. IL-6 plays a role in cancer cell proliferation, whereas G-CSF mediates tumor immunosuppression. Pathways and differential expression analyses confirmed MEK as the key regulatory molecule in cancer cell outgrowth and survival.

overall_design: For 3D in vitro experiments, we isolated cancer cells and/or stromal cells with or without chemotherapy and analyzed using scRNAseq. For in vivo experiments, we harvested and digested the mammary fat pad (mfp) or mammary tumors isolated from mice treated with vehicle or docetaxel (DTX) and analyzed using scRNAseq.

first: Ramya

last: Ganesan

first: Upaasana

last: Krishnan


title: Transcriptome profiling of EGFP and TEADi expressing podocytes

city: Freiburg

first: Manuel

last: Rogg

state: Baden Wuerttemberg

address: Breisacher Straße 115a

department: Institute of Surgical Pathology

institute: University Medical Center Freiburg

summary: EGFP (control) and TEADi expressing podocytes were generated by lentivirus transduction to analyze transcriptional signaling by YAP/TAZ-TEAD. Tetracycline-inducible TEADi is a GFP-tagged inhibitor of the interaction of YAP1 and TAZ with TEAD transcription factors. pInducer20 EGFP-TEADi was a gift from Ramiro Iglesias-Bartolome (Addgene plasmid # 140145 ; http://n2t.net/addgene:140145 ; RRID:Addgene_140145). TEADi was previously described: YAP1/TAZ-TEAD transcriptional networks maintain skin homeostasis by regulating cell proliferation and limiting KLF4 activity. Yuan Y., Park J., Feng A., Awasthi P., Wang Z., Chen Q., Iglesias-Bartolome R.. Nat Commun 11, 1472 (2020). 10.1038/s41467-020-15301-0. Transcriptome profiling (RNA-Sequencing) and differential gene expression analysis of 3 independent replicates per genotype was performed. TEADi podocytes exhibit transcriptional changes compared to WT cells including downregulation of prominent YAP target genes like CTGF, CYR61 and ANKRD1.

overall_design: mRNA sequencing analysis of EGFP and TEADi expressing immortalized human podocyte cell lines (3 replicates per genotype).

first: Christoph

last: Schell


title: Cell type and stage specific transcriptional, chromatin and cell-cell communication landscapes in the mammary gland (ATAC-Seq).

city: Bruxelles

first: Yura

last: Song

address: 808, route de Lennik

country: Belgium

institute: Université Libre de Bruxelles

summary: The mammary gland (MG) is composed of three main epithelial lineages, the basal cells (BC), the estrogen receptor (ER) positive luminal cells (ER+ LC), and the ER negative LC (ER- LC). Defining the cell identity of each lineage and how it is modulated throughout the different stages of life is important to understand how these cells function and communicate throughout life.  Here, we used transgenic mice specifically labelling ER+ LC to isolate with high purity the 3 distinct cell lineages of the mammary gland and defined their expression profiles and chromatin landscapes by performing bulk RNAseq and ATACseq of these isolated populations at different stages of life. Our analysis identified conserved genes, ligands and transcription factor (TF) associated with a specific lineage throughout life as well as genes, ligands and TFs specific for a particular stage of the MG. In summary, our study identified genes and TF network associated with the identity, function and cell-cell communication of the different epithelial lineages of the MG at different stages of life.

overall_design: FACS isolated ER+ LC, ER- LC and BC from mammary gland at puberty, adulthood or mid pregnancy

first: Alexandra

last: Van Keymeulen

first: Marco

last: Fioramonti

first: Cédric

last: Blanpain


title: Histone deacetylation and cytosine methylation are required for the normal compartmentalization of heterochromatin in the genome organization of Neurospora crassa [ChIP-Seq]

first: Andrew

last: Klocko

summary: Chromosomes must correctly fold in eukaryotic nuclei for proper genome function. Eukaryotic organisms hierarchically organize their genomes: in the fungus Neurospora crassa, chromatin fiber loops compact into Topologically Associated Domain (TAD)-like structures that are anchored by heterochromatic region aggregates. However, insufficient information exists on how histone post-translational modifications, including acetylation, impact genome organization. In Neurospora, the HCHC (HDA-1, CDP-2, HP1, CHAP) complex deacetylates heterochromatic regions including centromeres: loss of individual HCHC members increases centromeric acetylation and cytosine methylation. Here, we evaluate the role of the HCHC complex on genome organization using chromosome conformation capture with high-throughput sequencing (Hi-C) in Δcdp-2 or Δchap deletion strains. CDP-2 loss increases interactions between intra- and inter-chromosomal heterochromatic regions, while removal of CHAP decreases heterochromatic region compaction. Individual HCHC mutants exhibit different histone PTM patterns genome-wide: in Dcdp-2, heterochromatic H4K16 acetylation is increased, yet some heterochromatic regions lose H3K9 trimethylation, which locally increases inter-heterochromatin contacts; CHAP loss produces minimal acetylation changes but increases H3K9me3 enrichment in heterochromatin. Furthermore, deletion of the DIM-2 DNA methyltransferase in a Δcdp-2 background causes extensive genome disorder, as heterochromatic-euchromatic contacts increase despite additional H3K9me3 enrichment. Our results highlight how enhanced cytosine methylation ensures heterochromatic compartmentalization when silenced regions are acetylated.

overall_design: We analyzed genome organization of mutants of a heterochromatin-specific histone deacetylase complex (the HCHC complex) found in the filamentous fungus Neurospora crassa by chromosome conformation capture by paired-end high-throughput sequencing (Hi-C); Hi-C libraries were generated with the four-base cutters DpnII and MseI, which monitor euchromatic and heterochromatic regions of the Neurospora genome, respectively, due to the location of the recognition sequences of these restriction enzymes. All HCHC mutant Hi-C data was compared to wild type Hi-C data, previously reported in Rodriguez et al. (Pubmed ID: 35244156). We also examined the histone post-translational modification enrichment of wild type and HCHC mutant strains, specifically looking at the acetylation of lysine 9 on histone H3, the acetylation of lysine 16 on histone H4, and the trimethylation of lysine 9 on histone H3.

first: Ashley

last: Scadden

first: Alayne

last: Graybill

first: Clayton

last: Hull-Crew

first: Tiffany

last: Lundberg

first: Nickolas

last: Lande

middle: D


title: Transcriptome profiling of EPB41L5 WT and KO podocytes

summary: We generated EPB41L5 WT and KO podocytes to model human glomerular kidney disease in vitro. Immortalized human podocytes were genome edited applying the CRISPR/Cas9 technique. Two monoclonal cell lines were generated for WT and KO genotypes (WT-1, WT-2, KO-1 & KO-2).  Transcriptome profiling (RNA-Sequencing) and differential gene expression analysis was performed in duplicate for each cell line. EPB41L5 KO podocytes exhibit transcriptional changes compared to WT cells. Analyzed cell were previously described: Schell C, Rogg M, Suhm M, et al. The FERM protein EPB41L5 regulates actomyosin contractility and focal adhesion formation to maintain the kidney filtration barrier. Proc Natl Acad Sci U S A. 2017;114(23):E4621-E4630. doi:10.1073/pnas.1617004114

overall_design: mRNA sequencing analysis of two EPB41L5 WT and two EPB41L5 KO immortalized human podocyte cell lines (2 replicates; 2x WT-1, 2x WT-2, 2x KO-1, 2x KO-2).


title: Cell type and stage specific transcriptional, chromatin and cell-cell communication landscapes in the mammary gland (RNA-Seq).


title: Retinoic acid dependent spermatogonial differentiation in mice

city: San Antonio

first: Brian

middle: P.

last: Hermann

state: TX

address: 1 UTSA Circle

department: Department of Biology

institute: University of Texas at San Antonio

summary: Spermatogonial differentiation is a developmental process that is essential for spermatogenesis, but the molecular and cellular changes that germ cells must undergo to transition from undifferentiated spermatogonia to differentiating spermatogonia remain largely undefined. Retinoic acid (RA) is necessary and sufficient for spermatogonial differentiation. Using the postnatal mouse testis, we examine the transcriptome changes that accompany spermatogonial differentiation. Spermatogenesis was synchronized by administration of potent and selective RA synthesis inhibitor; as a result, testes contained only undifferentiated spermatogonia. Then, the inhibitor was discontinued, and mice were given a single dose of exogenous RA to initiate spermatogonial differentiation. We measured transcriptomes in FACS-enriched germ cells either before RA administration, when the cells correspond to Aal spermatogonia (and a minor contribution of spermatogonial stem cells) or at two points after RA administration, when the cells correspond to A1 or A3 differentiating spermatogonia. The results of this study reveal the full transcriptome changes accompanying spermatogonial differentiation in the mouse.

overall_design: WIN 18,446 treatment through P10 blocked normal spermatogonial differentiation and thus testes at P11 were filled with STRA8-/KIT- undifferentiated stem and progenitor spermatogonia. At P11, a single injection of exogenous RA induced all spermatogonia (save for the spermatogonial stem cells, or SSCs) to differentiate. RA is extremely labile (half-life in mice <0.5 hr), and thus is not expected to persist in the testis past P12. Twenty-four hrs later in these ‘RA-sufficient’ testes, nearly all spermatogonia were STRA8+/KIT+ type A1 differentiating spermatogonia that continued differentiating (A3). This study is paired with GSE194376.

first: Oleksandr

last: Kirsanov

first: Taylor

last: Johnson

first: Bryan

last: Niedenberger

last: Malachowski

first: Emma

last: Gilbert

first: Debajit

last: Bhowmick

first: Douglas

last: Ozdinler

last: Gray

first: Guang

last: Hu

first: Kelsey

last: Fisher-Wellman

first: Anukriti

last: Singh

first: Christopher

last: Geyer


title: Cell type and stage specific transcriptional, chromatin and cell-cell communication landscapes in the mammary gland

first: Yura

last: Song


title: Deletion of KvB2 leads to loss of muscle function in young and old mice

status: Public on Jul 01 2023

city: Tampa

first: Feng

last: Cheng

state: FL

address: 12901 Bruce B Downs Blvd MDC 30

department: Department of Pharmaceutical Science

institute: University of South Florida

summary: To investigate the skeletal muscle function and aging phenotype of KvB2 null mice we utilized the young and old KvB2 KO mice

overall_design: We performed the analyses of body weights and hindlimb limb muscles weights, forelimb grip strength and hindlimb muscle EDL myofiber force frequency relationship, muscle TA myofiber cross-section area and tissue fibrosis, muscle EDL sarcomere length, and Gastrocnemius muscles gene expression analysis through RNA Sequencing and RT-qPCR validation of  the KvB2 WT and KO genotypes

first: Ravikumar

last: Manickam

first: Jazmine

last: Virzi

first: Anish

last: Potti

first: David

last: Russ

first: Srinivas

last: Tipparaju


title: Retinoic acid is dispensable for initiation and completion of male meiosis in mice

first: Brian

last: Hermann

summary: Meiosis is a specialized form of cell division essential for sexual reproduction. The unique cellular events of meiosis have been extensively studied in model eukaryotes, including mammals. In mammals, the molecular and cellular changes that germ cells must undergo to transition from mitotic spermatogonia to meiotic spermatocytes remain largely undefined. Retinoic acid (RA) has been proposed as the meiosis inducing substance (MIS) required for female and male meiotic initiation. However, evidence for this presumption is based on studies of fetal gonads – meiosis is initiated in fetal ovaries but suppressed in fetal testes. Critically, the requirement for RA in meiosis has never been formally tested because germ cell differentiation and meiotic initiation in the fetal ovary are temporally inseparable. Using the postnatal mouse testis, where differentiation and meiotic initiation are temporally distinct (8.6 days apart), we test the hypothesis that RA is dispensable for initiation and progression through meiosis. Complementary in vivo experimental models were designed to address the hypothesis. In the first, “RA-sufficient” (RA-suf) model, spermatogenesis was synchronized by administration of potent and selective RA synthesis inhibitor; as a result, testes contained only undifferentiated spermatogonia. Then, the inhibitor was discontinued, and mice were given a single dose of exogenous RA to initiate spermatogonial differentiation. Germ cells completed differentiation, entered and completed meiosis, and formed haploid gametes at predicted timepoints. In the second, “RA-deficient” (RA-def) model, mice were treated as above, except the inhibitor was administered continuously. The lack of testicular RA in this model was confirmed in multiple ways, including assessment of RA-responsive gene expression, in vitro serum-free cultures, and dose-response experiments. To our surprise, germ cells in RA-def testes completed differentiation, underwent meiotic DNA replication, completed the landmark cytological processes of meiosis, and formed postmeiotic haploid spermatids, all on the predicted timeline of the RA-suf model. To define the effect of RA-deficiency on gene expression during meiotic initiation, large, highly enriched populations of premeiotic spermatogonia from RA-suf mice, and meiotic preleptotene spermatocytes from both models were FACS-isolated and subjected to bulk RNA-seq. Interestingly, analyses revealed few significant differences in expression of characteristic meiotic genes at the mRNA level. However, genes that were significantly different between RA-suf and RA-def preleptotene spermatocytes were associated with spermiogenesis functions. Taken together, our data reveal that, after initiating spermatogonial differentiation, RA is dispensable for initiation, progression through, and completion of meiosis in the male germline.

overall_design: In this study, we utilized an in vivo model of synchronized steady-state spermatogenesis . Synchronization was accomplished by dosing mice with the potent and highly selective RA synthesis inhibitor WIN 18,446 beginning at postnatal day (P)1, prior to endogenous RA signaling and when testes contain only prospermatogonia, the precursors of spermatogonia. Daily WIN 18,446 treatment through P10 blocked normal spermatogonial differentiation and thus testes at P11 were filled with STRA8-/KIT- undifferentiated stem and progenitor spermatogonia. At P11, a single injection of exogenous RA induced all spermatogonia (save for the spermatogonial stem cells, or SSCs) to differentiate. RA is extremely labile (half-life in mice <0.5 hr), and thus is not expected to persist in the testis past P12. Twenty-four hrs later in these ‘RA-sufficient’ testes, nearly all spermatogonia were STRA8+/KIT+ type A1 differentiating spermatogonia that proceeded through the remaining stages (A2-4, In, B) of differentiation and entered meiosis on P19 as STRA8+/DMRT1- preleptotene spermatocytes. These spermatocytes then proceeded predictably through meiosis to form round spermatids by P30.

first: Oleksandr

last: Kirsanov

last: Johnson

last: Niedenberger

first: Benjamin

last: Hale

first: Jiajia

last: Wang

first: Karen

last: Schindler

middle: P


title: HDAC3 and HDAC8 PROTAC dual degrader reveals roles of histone acetylation in gene regulation

city: Gainesville

first: Daiqing

last: Liao

state: Florida

address: 2033 Mowry Road

department: Anatomy and Cell Biology

institute: University of Florida

summary: RNA-seq experiments were performed to assess effects of degradation of HDAC3 and HDAC8 on transcriptome

overall_design: Cells were treated with control (DMSO), an HDACi (UF010), PROTACs (YX968, YX26862) that degrade HDAC3/HDAC8, or a negative control (YX968-NC) for various lengths of time

first: Yufeng

last: Xiao


title: Genome-wide DNA methylation profiling of colorectal tumours from MLH1 epimutation carriers

city: Melbourne

first: Jihoon

middle: Eric

last: Joo

state: Vic

address: 305 Grattan Street

department: Clinical Pathology

country: Australia

institute: University of Melbourne

type: Methylation profiling by genome tiling array

summary: Genome-wide DNA methylation analysis of colorectal tumour samples from 44 FFPE tumour and 15 FFPE normal mucosa samples was performed to invesgitate genome-wide DNA methylation signatures that can distinguish MLH1 epimutation carrier CRCs. This was further applied to resolve clinically challenging CRCs including MLH1 promoter VUS carriers and MLH1 methylated EOCRCs

overall_design: Genomic DNA  derived from 44 FFPE tumour samples and 15 FFPE normal mucosa samples was bisulfite converted and hybridised to the Illumina Infinium EPIC human methylation beadchips. The methylation data was preprocessed using minfi Bioconductor package and underwent Functional normalisation. Probes with detection P >0.05 were removed. Beta values were calculated using minfi.

middle: E

first: Khalid

last: Mahmood

first: Romy

last: Walker

first: Peter

last: Georgeson

first: Mark

last: Clendenning

first: John

last: Hopper

first: Finlay

last: Macrae

last: Jenkins

first: Christophe

last: Rosty

first: Ingrid

last: Winship

first: Daniel

last: Buchanan


title: Histone deacetylation and cytosine methylation are required for the normal compartmentalization of heterochromatin in the genome organization of Neurospora crassa [Hi-C]

first: Andrew

last: Klocko

first: Ashley

last: Scadden

first: Alayne

last: Graybill

first: Clayton

last: Hull-Crew

first: Tiffany

last: Lundberg

first: Nickolas

last: Lande


title: Integrative genome-wide analysis of PBMCs confirmed SOD1-specific functionality of ATF3 in ALS blood

city: Ulm

first: Karin

last: Danzer

state: Baden-Wurttemberg

address: University of Ulm Campus, Building N24, R3306 Albert-Einstein-Allee 11

department: Department of Neurology

institute: University of Ulm / DZNE, Ulm

summary: Amyotrophic Lateral Sclerosis (ALS) is characterized by a progressive loss of upper and lower motor neurons, leading to progressive weakness and atrophy of limb, bulbar and respiratory muscles. Since genetics alone fails to explain the etiology of ALS, the investigation of the epigenome might provide novel insights in disease mechanisms. Among all the different histone modifications the repressive trimethylation of lysine 27 on histone H3 (H3K27me3) is of particular interest in the context of ALS for several reasons. In this study, we interrogate the epigenetic landscape in peripheral blood mononuclear cells (PBMCs) of fALS patient with either chromosome 9 open reading frame 72 (C9orf72) or superoxide dismutase 1 (SOD1) mutation and aimed to identify key epigenetic footprints of the disease. To this end, we used an integrative approach that combines chromatin immunoprecipitation targeting H3K27me3 (ChIP-Seq) with the matching gene expression data to gain new insights into the impact of chromatin remodeling on ALS-related molecular mechanisms. In particular, multiple lines of evidence in this work indicated that one of the hub molecules that modulates changes in PBMC transcriptome in SOD1-positive ALS cases is ATF3. We also reported a potential suppression of SNAP25, with ATF3 involved as well, in SOD1-positive ALS blood.

overall_design: Comparative H3K27me3 occupancy analysis in peripheral blood mononuclear cells (PBMCs) of familial ALS patients with either C9orf72 (n = 3) or SOD1 (n = 2) mutation compared to healthy control (n = 3), where n denotes the number of biological replicates per sample group.

first: Volkan

last: Yazar

first: Julia

last: Kuhlwein

first: Antje

last: Knehr

first: Veselin

last: Grozdanov

first: Arif

last: Ekici

first: Albert

middle: C

last: Ludoplph


title: Characterizing control of memory CD8 T cell differentiation by BTB-ZF transcription factor Zbtb20 using single cell RNA-sequencing (CUT&RUN)

first: Edward

last: Usherwood

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: 8 total samples. 4 samples are murine OT-I CD8 T cells transduced to express Zbtb20 with an N-terminal 3XFLAG tag. 4 samples are HEK 293 cells transfected to express Zbtb20 with an N-terminal 3XFLAG tag.

first: Nicholas

last: Preiss

first: Owen

last: Wilkins


title: Droplet-based bisulfite sequencing for high-throughput profiling of single-cell DNA methylomes

city: Blacksburg

first: Chang

last: Lu

state: VA

address: 235 Goodwin Hall, 635 Prices Fork Road, Virginia Tech

department: Chemical Engineering

institute: Virginia Tech

summary: We present a high-throughput and droplet-based bisulfite sequencing library preparation platform. Our protocol allows production of BS library of 2,000-10,000 single cells within 2 days. In addition to validation using mixed cell lines, we also used the technology to profile various cell types in mouse and human brain samples.

overall_design:
We used our developed Drop-BS platform to profile single-cell methylomes on one human-mouse mixed sample, four cell line samples, one mouse brain PFC sample, and two human brain PFC samples. In the “Human_mouse_species_mixing”, we mixed equal amounts of GM12878 nuclei and mouse brain nuclei as the input nuclei to prepare a sequencing library using Drop-BS. We used GM12878 cell line, HEK293 cell line, MCF7 cell line separately to prepare three cell line Drop-BS samples. The Drop-BS library of “Three_cell_lines_mixing” was prepared using a mixture of the three with equal portions. We extracted nuclei from mouse, the first human brain PFC tissue, and the second human brain PFC tissue to prepare the “Mouse_brain” library, “Human_brain” library, and “Human_brain2” library respectively.

>>> Submitter states that raw data for the Human_brain sample will be submitted to dbGaP due to patient privacy concerns.<<<

first: Qiang

last: Zhang


title: Three-dimensional molecular cartography of human cerebral organoids revealed by double-barcoded spatial transcriptomics

city: Evry

first: Marco Antonio

last: MENDOZA-PARRA

address: 2 rue Gaston Crémieux

institute: CNRS UMR8030

summary: Spatially-resolved transcriptomics methodologies are revolutionizing our understanding of complex tissues, but their elevated costs represent still a bottle-neck for their democratization. In this work we present a low-cost strategy for manufacturing molecularly double-barcoded DNA arrays, enabling large-scale spatially-resolved transcriptomics studies.  We applied this technique to spatially resolve gene expression in several human brain organoids, including the reconstruction of a 3-dimensional view from multiple consecutive sections, revealing gene expression divergencies throughout the tissue.

overall_design: Spatial transcriptomics assays based on the use of in-house manufactured double-barcoded DNA arrays.

first: Aude

last: Bramoulle

first: Antoine

last: Aubert

first: François

last: Stüder

first: Julien

last: Moehlin

first: Lucie

last: Madrange

first: Frank

last: Yates

first: Jean-Philippe

last: Deslys

first: Marco

last: Mendoza-Parra

first: Gwendoline

last: Lozachmeur


title: Proximity-dependent recruitment of Polycomb Repressive Complexes by the lncRNA Airn [CHART-Seq]

city: Chapel Hill

first: Joseph

middle: Mauro

last: Calabrese

state: NC

address: 120 Mason Farm Rd

department: Pharmacology

institute: UNC Chapel Hill

summary: In the developing mouse embryo, expression of the lncRNA Airn induces gene silencing and recruits Polycomb Repressive Complexes (PRCs) to varying extents over a 15 megabase domain, but the mechanisms remain unclear. Using high-resolution approaches, we show in mouse trophoblast stem cells that Airn expression induces long-range changes to chromatin architecture centered around a network of CpG islands occupied by PRCs and SMC1A/Cohesin. Extent of contact with Airn predicts underlying intensity of Polycomb-directed chromatin modifications and Airn-induced changes to chromatin architecture. Deletion of individual CpG islands alternately increase or decrease gene silencing and PRC activity over the domain in a manner predicted by changes in chromatin architecture. We conclude that Airn is a potent repressive lncRNA whose effects are controlled by an equilibratory network of DNA regulatory elements that modulate its frequency of contact with target chromatin.

overall_design: Capture Hybridization Analysis of RNA Targets coupled with sequencing (CHART-Seq) in TSCs and ESCs.

first: Aki

last: Braceros

first: J


title: Proximity-dependent recruitment of Polycomb Repressive Complexes by the lncRNA Airn

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

overall_design: Refer to individual Series


title: Simultaneous Mapping of DNA Binding and Nucleosome Positioning with SpLiT-ChEC

city: Eugene

first: Laura

middle: Emily

last: McKnight

state: Oregon

address: 1318 Franklin Blvd Room 273 Onyx Bridge

department: Institute of Molecular Biology

institute: University of Oregon

summary: The organization of chromatin – including the positions of nucleosomes and the binding of other proteins to DNA – helps define transcriptional profiles in eukaryotic organisms. While techniques like ChIP-Seq and MNase-Seq can map protein-DNA and nucleosome localization separately, assays designed to simultaneously capture nucleosome positions and protein-DNA interactions can produce a detailed picture of the chromatin landscape. Most assays that monitor chromatin organization and protein binding rely on antibodies, which often exhibit nonspecific binding, and/or the addition of bulky adducts to the DNA- binding protein being studied, which can affect their expression and activity. Here, we describe SpyCatcher Linked Targeting of Chromatin Endogenous Cleavage (SpLiT-ChEC), where a 13-amino acid SpyTag peptide, appended to a protein of interest, serves as a highly-specific targeting moiety for in situ enzymatic digestion. The SpyTag/SpyCatcher system forms a covalent bond, linking the target protein and a co-expressed MNase-SpyCatcher fusion construct. SpyTagged proteins are expressed from endogenous loci, whereas MNase-SpyCatcher expression is induced immediately before harvesting cultures. MNase is activated with high concentrations of calcium, which primarily digests DNA near target protein binding sites. By sequencing the DNA fragments released by targeted MNase digestion, we found that this method recovers information on protein binding and proximal nucleosome positioning. SpLiT-ChEC provides precise temporal control that we anticipate can be used to monitor chromatin under various conditions and at distinct points in the cell cycle.

overall_design: Enzymatic digestion of genomic DNA by targeting MNase-SpyCatcher through covalent linkage to tagged proteins via Spytag fusion

first: Orion

middle: G

last: Banks

first: Michael

last: Harms

first: Jeffrey

middle: N


title: Proximity-dependent recruitment of Polycomb Repressive Complexes by the lncRNA Airn [RNA-Seq]

overall_design: Total RNA sequencing (RNA-Seq) in TSCs


title: Single-cell analysis of Primordial Germ Cell-like cells (hPGCLC) (female and male) using 10X genomics

city: Leiden

first: Susana

last: Chuva de Sousa Lopes

address: Einthovenweg 20

department: Anatomy and Embryology

country: Netherlands

institute: Leiden University Medical Center

summary: Current human primordial germ cell like cells (hPGCLCs) differentiation methods from pluripotent stem cells (PSCs) are inefficient, and it is challenging to generate sufficient hPGCLCs to optimize the next steps to achieve in vitro gametogenesis. We present a new differentiation method that uses diluted basement membrane extract (BME) and low BMP4 concentration to induce efficient hPGCLC differentiation, in scalable 2D cell culture.  We show that BME overlay potentiates BMP/SMAD signaling, induced lumenogenesis and increases expression of key hPGCLC progenitor markers such as TFAP2A, EOMES, GATA3 and CDX2. These findings highlight the importance of factors in the BME in hPGCLC differentiation, and demonstrate the potential of the BME-overlay method as a new model for interrogating the formation of PGCs and amnion in humans.

overall_design: Four hiPSC lines were used for differentiation to hPGCLC  and were used for single cell cluster analysis.

first: S

first: I

last: Moustakas

first: A

last: Overeem

first: Y

middle: W

last: Chang


title: Terminal differentiation of villus-tip enterocytes is governed by distinct  members of Tgfb superfamily

city: Zurich

first: Tomas

last: Valenta

address: Winterthurerstrasse 190

department: Institute of Molecular Life Sciences

country: Switzerland

institute: University of Zurich

summary: The protective and absorptive functions of the intestinal epithelium rely on differentiated enterocytes in the villi. The differentiation of enterocytes is orchestrated by sub-epithelial mesenchymal cells producing distinct ligands along the villus axis, in particular Bmps and Tgf. Here we show that individual Bmp ligands and Tgf drive distinct enterocytic programs specific to villus zonation. Bmp4 is expressed mainly from the centre to the upper part of the villus, and it activates preferentially genes connected to lipid uptake and metabolism. In contrast, Bmp2 is produced by villus-tip mesenchymal cells, and it influences the adhesive properties of villus-tip epithelial cells and the expression of antimicrobial peptides. Hence, Bmp2 promotes the terminal enterocytic differentiation at the villus-tip. Additionally, Tgf induces an epithelial gene expression programs similar to that triggered by Bmp2. The inhibition of Bmp receptor type I in vivo and using intestinal organoids lacking Smad4 revealed that Bmp2-driven villus-tip program is activated by a canonical Smad-dependent mechanism. Finally, we established an organoid cultivation system that enriches for villus-tip enterocytes and thereby better mimics the cellular composition of the intestinal epithelium. Altogether our data suggest that not only Bmp gradient, but also the activity of individual Bmp drives specific enterocytic programs.

overall_design: Intestinal crypts were initially seeded in ENR medium (without any Bmp/Tgfb). After 48 h the debris was removed: old medium was removed and matrigel domes containing organoids were broken with 1 ml plastic tip with cut-off tip and transferred into 15 ml tube containing 10 ml of ice-cold PBS. The crypts were incubated 10 min on ice, sedimented and then upper 8 ml of PBS containing cell debris was sucked out and replaced by fresh PBS. Organoids were centrifuged (300 g, 4 °C, 5 min) and put into fresh matrigel domes. Recombinant Bmps (500 ng/ml each) were added for 24.   Mesenchymal cells from duodenum of PdgfraH2BeGFP mouse were isolated as described previously8,9. The duodenal tissue was harvested, opened longitudinaly and rinsed. The tissue was gently rocked for 30 minitues at room temperature in Gentle Cell Dissociation Reagent to remove epithelial cell. After the detachment of epithelium remaining pieces were digested for 35 minutes in DMEM supplelemented with 1 mg/ml collagenase D and 0,3 mg/ml dispase in thermomixer at 37°C with gentle shaking (60 rpm in horizonally oriented 50ml Falcon tube). Afterwards cells were filtered through 70m cell strainer and seeded. 24 h after seeding non-adherent cells were washed out. The cells were cultivated in MesenCult medium. The medium was changed 3 times a week. The mesenchymal cells were immortalised by transduction of (HPV16)-E6 gene using retroviral particles. For the retroviral production PlatinumE cells were transfected with pMXs-EF1-Puro retroviral construct containing (HPV16)-E6 insert re-cloned from p1322HPV-16-E6 vector. Lipofectamine 3000 and OptiMEM medium were used according to manufacturers protocol. Transfection medium was changed for cultivation medium 6 h after transfection. Viral particles were produced for 72 h. Retroviral partciles were concentrated overnight at 4 °C by RetroX concentrator and used for infection of mesenchymal cells by spinoculation (30 °C, 450 g ,1 h). 72 hours after the spinoculation the medium with viral particles was changed and cells were cells were selected with puromycin (1 μg/ml) and further expanded. The immortalized cells were sorted based on internal eGFP fluorescence intensity to eGFPlow eGFPhigh and further separatelly expanded. Occassionally,the cells were retreated with puromycin. The cells were passaged with TripleExpres solution.  Total RNA was isolated using RNeasy Micro Kit (Qiagen 74004).Sequencing libraries were prepared from total RNA using the KAPA mRNA Hyperprep Kit, followed by size distribution analysis in the Agilent 2100 Bioanalyzer using the High Sensitivity DNA Kit (Agilent). Libraries were sequenced in two runs of the Illumina NextSeq 500 instrument (Illumina, USA) using a 75nt single-end configuration. The complete processing of isolated RNA was done by Genomics and Bioinformatics core facility at Institute of Molecular Genetics of the Czech Academy of Sciences.

first: Linda

last: Berkova

first: Hassan

last: Fazilaty

first: Jan

last: Kubovciak

first: Monika

last: Stastna

first: Dusan

last: Hrckulak

first: Martina

last: Vojtechova

last: Brügger

first: George

last: Hausmann

first: Vladimir

last: Korinek

first: Konrad

last: Basler


title: Gene expression profiles in gastric cancer cells with TM4SF1-AS1 knockdown

city: Sapporo

first: Hiromu

last: Suzuki

state: Hokkaido

address: S1, W17, Chuo-ku

department: Department of Molecular Biology

institute: Sapporo Medical University

summary: To clarify the function of a long noncoding RNA TM4SF1-AS1 in gastric cancer (GC), we carried out a gene expression microarray analysis using GC cell lines HSC45 transfected with a siRNA targeting TM4SF1-AS1 or a control siRNA. Gene ontology analysis revealed that genes associated with interferon signaling and immune response were significantly enriched among downregulated genes.

overall_design: HSC45 cells were transfected with a siRNA targeting TM4SF1-AS1 or a control siRNA. Ninety-six hours after transfection, total RNA was extracted and gene expression profiles were analyzed.

first: Hiroshi

last: Kitajima


title: Effect of visible light on the transcriptome of Clostridium autoethanogenum during gas fermentation

city: Wuhan

first: Pier-Luc

last: Tremblay

state: Hubei

address: 122 Luoshi Road

department: School of Chemistry, Chemical Engineering and Life Science

institute: Wuhan University of Technology

summary: The expression profile of C. autoethanogenum DSM 10061 grown autotrophically with H2:CO:CO2 under visible light at an intensity of 4200 lux versus the expression profile of C. autoethanogenum DSM 10061 grown autotrophically in the dark

overall_design: Illumina RNA-seq of total RNA extracted from C. autoethanogenum wild type grown for 48 hours with H2:CO:CO2 (35:50:15) in Tanner modified medium under visible light at an intensity of 4200 lux or in the dark

first: Yu

last: Kang

first: Tian

last: Zhang


title: Chromatin organization in 3D distinguishes the influences of dMes-4/NSD and Hypb/dSet2 in protecting genes from H3K27me3 silencing [Hi-C]

city: Toulouse

first: Olivier

last: Cuvier

address: 118 route de Narbonne

department: LBME

institute: CNRS

summary: Maintenance of gene expression programs requires cell-type specific barcoding of genomes through euchromatin interspaced with facultative heterochromatin domains where PRC1/2 hinder chromatin accessibility thereby repressing genes inside such domains. At heterochromatin-euchromatin borders, regulation of heterochromatin spreading may depend on transcriptional activity, histone modifiers, and chromatin barrier insulators or the borders of topological associated domain (TADs). Here we show that depletion of H3K36 di- or tri-methyl histone methyl transferases dMes-4/NSD or Hypb/dSet2 induces H3K27me3 spreading at H3K27me3 borders, accounting for the repression of hundreds of genes upon depletion of these enzymes. dMes-4/NSD influences genes within active chromatin hubs and flanked by H3K27me3 borders demarcated by a TAD border, unlike dHypb/dSet2 that protects genes near H3K27me3 borders in absence of a TAD border. Insulator mutants that disrupt 3D chromatin hubs recapitulate only the H3K27me3 spreading observed upon depletion of dMes-4/NSD, and not of Hypb/dSet2. Hi-C data demonstrate how dMes-4/NSD directly block the propagation of the long-range interactions marking the inactive TADs, onto neighbor active regions, unlike Hypb/dSet2. Our data thus show a division of labor between dMes-4/NSD and Hypb/dSet2 to protect genes against H3K27me3 silencing, highlighting a direct influence of H3K36me marks on the demarcation of H3K27me3 domains within repressive TADs.

overall_design: We used the technology of Hi-C to quantify long-range interactions and chromosome conformation in order to describe topological domains in presence or absence of H3K36 HMTs.Chromatin extracted from cells in wild-type or after a knock-down of HypB or Mes4

first: David

last: Depierre

first: Charlène

last: Perrois

last: Fosseprez