title: Enhancing transcription replication conflict targets ecDNA positive cancers

first: Guiping

last: Wang

summary:
Extrachromosomal DNA (ecDNA) presents a major challenge for cancer patients. EcDNA renders tumours treatment-resistant by facilitating massive oncogene transcription and rapid genome evolution, contributing to poor patient survival. At present, there are no ecDNA-specific treatments. Here we show that enhancing transcription replication conflict enables targeted elimination of ecDNA-containing cancers. Stepwise analyses of ecDNA transcription reveal pervasive RNA transcription and associated single-stranded DNA (ssDNA), leading to excessive transcription replication conflicts and replication stress (RS) compared to chromosomal loci. Nucleotide incorporation on ecDNA is markedly slower, and RS is significantly higher in ecDNA-containing tumours regardless of cancer type or oncogene cargo. pRPA2-S33, a mediator of DNA damage repair that binds ssDNA, shows elevated localization on ecDNA in a transcription dependent manner, along with increased DNA double strand breaks, and activation of the S-phase checkpoint kinase, CHK1. Genetic or pharmacological CHK1 inhibition abrogates the DNA replication check point, causing extensive and preferential tumour cell death in ecDNA-containing tumours. We advance a highly selective, potent, and bioavailable oral CHK1 inhibitor, BBI-2779, that preferentially kills ecDNA-containing tumour cells.  In a gastric cancer model containing FGFR2 on ecDNA, BBI-2779 suppresses tumour growth and prevents ecDNA-mediated acquired resistance to the pan-FGFR inhibitor infigratinib, resulting in potent and sustained tumour regression in mice. Transcription replication conflict emerges as a target for ecDNA-directed therapy, exploiting a synthetic lethality of excess to treat cancer.

This SuperSeries is composed of the SubSeries listed below.

This work was delivered as part of the eDyNAmiC team supported by the Cancer Grand Challenges partnership funded by Cancer Research UK (CGCATF-2021/100012 and CGCATF-2021/100025) and the National Cancer Institute (OT2CA278688 and OT2CA278635) to H.Y.C., P.S.M., and V.B. This project was supported by NIH RM1-HG007735 (H.Y.C., W.J.G., C.C.).


title: ScRNA-seq and scATAC-seq analysis of human pluripotent stem cell (hPSC) derived macrophages iMacs

status: Public on May 23 2024

city: Beijing

first: Jie

state: Beijing

address: Tsinghua University

department: Basic Medical Sciences, School of Medicine

country: China

institute: Tsinghua University

type: Expression profiling by high throughput sequencing

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Purpose: to depict the single-cell map and reveal the heterogeneity of hPSC derived macrophages  Methods: Differentiation day 14 iMacs were dissociated to obtain a single-cell suspension. scRNA-seq and scATAC-seq library preparation and sequencing were done by CapitalBio Technology (https://www.capitalbiotech.com/). scRNA-seq, the cell suspension was immediately loaded onto the Chromium Single Cell Controller (10× Genomics) for droplet formation. cDNA and subsequent sequencing library were generated using Chromiun Single Cell 3’ Reagent Kit (v3 Chemistry) (10× Genomics) according to the instruction manual. For sc-ATAC-seq, cell nuclei were extracted with Chromiun Nuclei Isolation Kit (v2 Chemistry) (10x Genomics) and then loaded onto the Chromium Single Cell Controller for droplet formation.  Results: ScRNA-seq and scATAC-seq data of differentiation day 14 iMacs from hPSCs Conclusions: day 14 differentiation culture, which consisted primarily of monocytes and macrophages.

overall_design: Perform scRNA-seq and scATAC-seq of differentiation day 14 iMacs from hPSCs

first: Yaxuan

last: Zhang

first: Xingwu

first: Hui

last: Qiu


title: Enhancing transcription replication conflict targets ecDNA positive cancers [H3K36me3_CHIP]

city: Stanford

address: 256 Campus Dr

country: USA

institute: Stanford University

summary:
Extrachromosomal DNA (ecDNA) presents a major challenge for cancer patients. EcDNA renders tumours treatment-resistant by facilitating massive oncogene transcription and rapid genome evolution, contributing to poor patient survival. At present, there are no ecDNA-specific treatments. Here we show that enhancing transcription replication conflict enables targeted elimination of ecDNA-containing cancers. Stepwise analyses of ecDNA transcription reveal pervasive RNA transcription and associated single-stranded DNA (ssDNA), leading to excessive transcription replication conflicts and replication stress (RS) compared to chromosomal loci. Nucleotide incorporation on ecDNA is markedly slower, and RS is significantly higher in ecDNA-containing tumours regardless of cancer type or oncogene cargo. pRPA2-S33, a mediator of DNA damage repair that binds ssDNA, shows elevated localization on ecDNA in a transcription dependent manner, along with increased DNA double strand breaks, and activation of the S-phase checkpoint kinase, CHK1. Genetic or pharmacological CHK1 inhibition abrogates the DNA replication check point, causing extensive and preferential tumour cell death in ecDNA-containing tumours. We advance a highly selective, potent, and bioavailable oral CHK1 inhibitor, BBI-2779, that preferentially kills ecDNA-containing tumour cells.  In a gastric cancer model containing FGFR2 on ecDNA, BBI-2779 suppresses tumour growth and prevents ecDNA-mediated acquired resistance to the pan-FGFR inhibitor infigratinib, resulting in potent and sustained tumour regression in mice. Transcription replication conflict emerges as a target for ecDNA-directed therapy, exploiting a synthetic lethality of excess to treat cancer.

This work was delivered as part of the eDyNAmiC team supported by the Cancer Grand Challenges partnership funded by Cancer Research UK (CGCATF-2021/100012 and CGCATF-2021/100025) and the National Cancer Institute (OT2CA278688 and OT2CA278635) to H.Y.C., P.S.M., and V.B. This project was supported by NIH RM1-HG007735 (H.Y.C., W.J.G., C.C.).

overall_design: H3K36me3 ChIP-seq of COLO320DM and COLO320HSR-  2 biological replicates each.

first: Natasha

last: Weiser

first: Howard

last: Chang


title: Active DNA demethylation controls transgenerational epigenetic stability

city: West Lafayette

first: Jian-Kang

last: Zhu

state: IN

address: 625 Agriculture Mall Dr.

department: Department of Horticulture and Landscape Architecture

institute: Purdue University

type: Methylation profiling by high throughput sequencing

summary: We examined spontaneously occurring variation in DNA methylation in wild type Col-0 plants, together with nrpd1 and ros1 mutants that are all propagated from single-seed descent for 10 generations.

overall_design: The samples examined are:  Col-0_G1, nrpd1_G1, ros1_G1, Col-0_G10_rep1, Col-0_G10_rep2, Col-0_G10_rep3, Col-0_G10_rep4, Col-0_G10_rep5, nrpd1_G10_rep1, nrpd1_G10_rep2, nrpd1_G10_rep3, nrpd1_G10_rep4, nrpd1_G10_rep5, ros1_G10_rep1, ros1_G10_rep2, ros1_G10_rep3, ros1_G10_rep4, ros1_G10_rep5, Col-0_salt_G10_rep1, Col-0_salt_G10_rep2, Col-0_salt_G10_rep3, Col-0_salt_G10_rep4, Col-0_salt_G10_rep5, nrpd1_salt_G10_rep1, nrpd1_salt_G10_rep2, nrpd1_salt_G10_rep3, nrpd1_salt_G10_rep4, ros1_salt_G10_rep1, ros1_salt_G10_rep2, ros1_salt_G10_rep3, ros1_salt_G10_rep4, ros1_salt_G10_rep5, Col-0_G7, ros1_G7, ros1_salt_G7

first: Kai

last: Tang

first: Xiaohong

first: Shaojun

last: Xie

first: Zhaobo

last: Lang


title: Dynamic astrocytic gene expression profiles after ischemic stroke

first: Song

address: NO.95, Yong'An Road, Xicheng District

institute: Beijing Friendship Hospital, Capital Medical University

summary: Analysis of astrocytic gene expression profiles after ischemic stroke. Stroke is a complicated disease caused by the interaction of multiple celltypes. Results provide new insights into the molecular mechanisms underlying astrocytic activation after ischemic stroke.

overall_design: Astrocytic gene expression profiles of sham, day1, day3, day7, day21, and day60  focal ischemic stroke mice model

first: Yuhualei

last: Pan

first: Chengjie


title: Histone arginine methylation by Prmt5 is required for lung branching morphogenesis through repression of BMP signaling

city: Hefei

first: LU

last: Qingshuang

address: 111 Kowloon Road

department: Institute of Physical Science and Information Technology

institute: Anhui University

summary: Branching morphogenesis is essential for the successful development of a functional lung to accomplish its gas exchange function. Although many studies have highlighted requirements for the bone morphogenetic protein (BMP) signaling pathway during branching morphogenesis, little is known about how BMP signalingis regulated. Here we report that the protein arginine methyltransferase 5 (Prmt5) and symmetric dimethylation at histone H4 arginine 3 (H4R3sme2) directly associate with chromatin of Bmp4 to suppress its transcription. Inactivation of Prmt5 in the lung epithelium results in halted branching morphogenesis, altered epithelial cell differentiation and neonatal lethality. These defects are accompanied by increased apoptosis and reduced proliferation of lung epithelium, as a consequence of elevated canonical BMP-Smad1/5/9 signaling. Inhibition of BMP signaling by Noggin rescues the lung branching defects of Prmt5 mutant in vitro. Taken together, our results identify a novel mechanism through which Prmt5-mediated histone arginine methylation represses canonical BMP signaling to regulate lung branching morphogenesis.

overall_design: In order to further explore the molecular mechanism of cell deletion in ciliated cells after Prmt5 knockout in epithelial cells,We chose the E13.5 period and put Shh; Prmt5 embryo and litter control of the main airway part (including trachea and lung Isolated from the whole lung, RNA was extracted for transcriptome sequencing (RNA-SEQ) analysis.

first: Qiuling

last: Li

last: Jiao


title: Interferons and epigenetic mechanisms in training, priming and tolerance of monocytes and hematopoietic progenitors [ATAC-seq]

status: Public on May 22 2024

city: New York

first: Bikash

last: Mishra

state: NY

address: 515 E 71st ST

institute: Hospital for Special Surgery

summary: Training and priming of innate immune cells involve preconditioning by PAMPs, DAMPs and/or cytokines that elicits stronger induction of inflammatory genes upon secondary challenge. Previous models distinguish training and priming based upon whether immune activation returns to baseline prior to secondary challenge. Tolerance is a protective mechanism whereby potent stimuli induce refractoriness to secondary challenge. Training and priming are important for innate memory responses that protect against infection, efficacy of vaccines, and maintaining innate immune cells in a state of readiness; tolerance prevents toxicity from excessive immune activation. Dysregulation of these processes can contribute to pathogenesis of autoimmune/inflammatory conditions, post-COVID-19 hyperinflammatory states, or sepsis- associated immunoparalysis. Training, priming and tolerance regulate similar ‘signature’ inflammatory genes such as TNF, IL6 and IL1B and utilize overlapping epigenetic mechanisms. We review how interferons (IFNs), best known for activating Jak-STAT signaling and interferon- stimulated genes, also play a key role regulating training, priming and tolerance via chromatin- mediated mechanisms. We present new data on how monocyte-to-macrophage differentiation modulates IFN-g-mediated priming, changes AP-1 and CEBP activity, and attenuates superinduction of inflammatory genes. We present a ‘training-priming continuum’ model that integrates IFN-mediated priming into current concepts about training and tolerance and proposes a central role for STAT1 and IRF1.

overall_design: To investigate priming effect of IFNg on human monocytes vs macrophages, we purified CD14+ monocytes from human buffy coats. Monocytes were cultured overnight at 37°C, 5% CO2 in RPMI-1640 medium supplemented with 10% heat-inactivated defined FBS, penicillin-streptomycin, L-glutamine and 20 ng/ml human M-CSF and treated with indicated stimulation. Macrophages were cultured similary but for 3 days and stimulated with indicated stimulation.

first: Lionel

last: Ivashkiv


title: ACSL4-programmed sphingomyelin metabolism rejuvenates tumor-infiltrating T-cells to enhance the efficacy of anti-PD-1 immunotherapy

city: Nanjing

first: Guilai

last: Liu

state: Jiangsu

address: Longmian road 639

institute: China Pharmaceutical University

summary: To investigate whether the expression of ACSL4 may alter the anti-tumor effecet of T-cells, we conditionally knocked out the Acsl4 gene in mouse T-cells (Acsl4-cKO) as descried in Wang et al. Redox Biol 2022. To further demonstrate the importance of SM in ACSL4-mediated T-cell anti-tumor immunity, we conducted a complementary study by intra-tumoral injection of SM in subcutaneous MC38 tumors inoculated in Acsl4-cKO mice.

overall_design: Tumor-infiltrating CD8+ T-cells  from MC38 inoculted Acsl4-cKO mice treated with vehicle or SM,  or control mice were isolated were isolated by FACS and analyzed using ATAC-seq.

first: Yun

first: Beiying

last: Dai


title: Enhancing transcription replication conflict targets ecDNA positive cancers [RNA-seq]

first: Guiping

last: Wang

overall_design: To compare stable RNA contents in isogenic cell pairs, we performed total RNA profiling in COLO320 DM, COLO320 HSR, GBM39ec and GBM39 HSR, 2 biological replicates each

first: Rui


title: The genes expression in WT and XBAT31 mutant under normal and high temperature at reproductive stage of Arabidopsis.

city: Hang Zhou

first: 霖霖

last: 张

address: Yuhangtang Road No.866

institute: Zhejiang University

summary: To investigate the genes which relay on  XBAT31 under high temperature and are induced by high temperature.

overall_design: 1-12 stage flowers of  Arabidopsis were taken at normal (22℃) and high temperature (38°C ) for 3 hours. There are 3 biological replicates and each replicate has 9 flowers, respectively.

first: Linlin

last: Zhang

first: Qiaoyun

first: Ziwei

last: Yao

first: Jingliang

last: Sun

first: Haiping

last: Lu

first: Jianxiang


title: Deficiency in peroxisomes underlies failures in neonatal hepatic immune cell hematopoiesis and antigen presentation in a severe Zellweger disease model

city: Halifax

first: Francesca

last: Di Cara

state: Nova Scotia

address: 5850/5980 University Avenue

country: Canada

institute: Dalhousie University

summary: Peroxisome Biogenesis disorders (PBDs) represent a group of metabolic conditions that cause severe developmental defects. Peroxisomes are essential metabolic organelles, present in virtually every eukaryotic cell and mediating key processes in immunometabolism. To date, the full spectrum of PBDs remains to be identified, and the impact PBDs have on immune function is unexplored. This study presents a characterization of the hepatic immune compartment of a neonatal PBD mouse model at single-cell resolution to establish the importance and function of peroxisomes in developmental hematopoiesis. We report that hematopoietic defects are a feature in a severe PBD murine model. Finally, we identify a role for peroxisomes in the regulation of the major histocompatibility class II expression and antigen presentation to CD4+ T cells in dendritic cells. This study adds to our understanding of the mechanisms of PBDs and expands our knowledge of the role of peroxisomes in immunometabolism.

overall_design: Liver immune cells from pex2 homozygous null mutant, pex2 heterozygous, and wildtype neonatal mice were isolated by Fluorescence-activeated cell sorting (FACs) according to fluorescence-conjugted anti-CD45 antibody labelling and viability staining and analyzed using scRNAseq.

first: Brendon

middle: D

last: Parsons

first: Daniel

last: Medina-Luna

first: Michal

last: Scur

first: Marinella

last: Pinelli

first: Gayani

middle: S

last: Gamage

first: Rebecca

middle: A

last: Chilvers

first: Yannick

last: Hamon

first: Ibrahim

last: Ahmed

first: Stephane

last: Savary

first: Andrew

middle: P

last: Makrigiannis

first: Nancy

middle: E

last: Braverman

first: Juan

middle: R

last: Alcazar

first: Eicke

last: Latz

first: Tobias

last: Karakach


title: Identifying a perivascular niche supporting lung alveoli epithelial stem cells and metastatic cancer cells

city: Chicago

first: Qian

last: Chen

state: IL

address: 909 S Wolcott avenue, COMRB 4160

department: Department of Pharmacology and Regenerative Medicine

institute: University of Illinois at Chicago

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

overall_design: Refer to individual Series


title: Single cell RNA-seq of bladder tissue

city: Shanghai

first: Fei

last: Su

state: Shanghai

address: 800 Dongchuan Road

department: School of Life Science and Biotechnology

institute: Shanghai Jiao Tong University

summary: We report the application of single cell RNA-seq  for transcript profiling in bladder tissue from Interstitial cystitis/bladder pain syndrome (IC/BPS) with Hunner lesions  and without Hunner lesions and normal tissue.

overall_design:
Examination of 3 bladder tissue from NHIC, 2 HIC and 2 unaffected normal tissue.

>>>Submitter states that raw data will be submitted to the Genome Sequence Archive for Human (GSA-Human)<<<

last: Xiao


title: RNA-seq Series of Bladder Carcinoma Tumors

city: paris

first: Elodie

last: chapeaublanc

address: 26 rue d'ulm

department: UMR 144

country: France

institute: Institut Curie

summary:
Background: Muscle-invasive bladder cancer is a common aggressive disease with unmet clinical needs. Recent work established a set of consensus bladder cancer transcriptomic subtypes that distinguishes the cell identity of bladder cancers for improved diagnosis and treatment. However, how these distinct subtypes are regulated remains unclear. Given the link between super-enhancers and the regulation of cell identity, we hypothesized that epigenetic activation of distinct super-enhancers could drive the transcriptional programs of the various bladder cancer subtypes.
Results: Through integrated RNA sequencing and epigenomic profiling of histone marks (H3K27ac, H3K27me3, H3K9me3) in a diverse panel of 15 primary bladder tumours, seven bladder cancer cell lines, and two primary cultures from normal human urothelia, we established the first integrated epigenetic map of bladder cancer and demonstrate the link between bladder cancer subtype and epigenetic control. Through H3K27ac analysis, we identify the repertoire of activated super-enhancers in bladder cancer that distinguish molecular subtypes. Building on these findings, we reveal the super-enhancer-regulated networks of candidate master transcription factors for Luminal and Basal bladder cancer subgroups. We find that FOXA1, a key pioneer factor in Luminal bladder cancers identified in our Luminal transcription factor network, binds subgroup-specific bladder super-enhancers and correlates with their activation. Furthermore, CRISPR-Cas9 inactivating mutation of FOXA1 triggers a shift from Luminal to Basal cell identity. This shift is accompanied by an overexpression of ZBED2, one of the newly identified transcriptional regulators in the Basal-specific transcription factor network. 5Finally, we show that both FOXA1 and ZBED2 play concordant roles in preventing inflammatory response in bladder cancer cells through STAT2 inhibition and promote cancer cell survival.
Conclusions: Overall, our study provides new data for understanding epigenetic regulation of muscle- invasive bladder cancer and identifies a coregulated network of super-enhancers and associated transcription factors as new potential targets for the treatment of this aggressive disease

overall_design: RNA-seq Series of Bladder carcinoma tumors

first: Hélène

last: Neyret-Kahn

first: François

last: Radvanyi

first: Isabelle

last: Bernard-Pierrot

last: Chapeaublanc

first: Jacqueline

last: Fontugne


title: Mis-splicing derived neoantigens and cognate T cell receptors in splicing factor mutant leukemias

first: Jahan

last: Rahman

state: New York

address: 417 E 68th St

department: Center for Hematologic Malignancies

institute: Memorial Sloan Kettering

summary: Mutations in RNA splicing factors are prevalent across cancers and generate recurrently mis-spliced mRNA isoforms. Here we identified a series of bona fide neoantigens translated from highly stereotyped splicing alterations promoted by neomorphic, leukemia-associated somatic mutations in the splicing machinery. We utilized feature-barcoded peptide-MHC dextramers to isolate neoantigen-specific T cell receptors (TCR) from both healthy donors and patients with leukemia. While circulating neoantigen-specific CD8+ T cells were identified in patients with active disease, they were dysfunctional with reduced inflammatory response gene signatures. In contrast, donor CD8+ T cells with tumor-reactive TCRs were present following curative allogeneic hematopoietic cell transplant. T cells engineered with TCRs recognizing an SRSF2 mutant-induced neoantigen in CLK3 resulted in specific recognition and cytotoxicity of SRSF2 mutant leukemia. These data identify RNA mis-splicing derived neoantigens and neoantigen-specific TCRs across patients and provide proof-of-concept to genetically redirect T cells to public mis-splicing derived neoantigens in myeloid leukemias.

overall_design: We isolated bulk T cells from patient PBMCs via negative selection and stained the T cells with a dextramer pool. We sorted live, CD3+, CD8+ dextramer+ cells and performed RNA-, TCR-, and dextramer feature barcode sequencing (Fig. S5A). For certain samples where dextramer+ cell numbers were limiting, live CD3+ CD8+ dextramer- cells were also sorted, stained with cell hashing antibodies, and doped into the dextramer+ populations. Using this approach, we single-cell profiled CD8+ T-cells from nine samples across five distinct HLA-A*02:01+ SRSF2 mutant myeloid leukemia patients (Fig. 5A, Fig.S5B, and Table S3) yielding 75,343 high-quality T cells.

first: Won

middle: J

last: Kim

first: Edie

last: Cross

first: Emma

last: de Neef

first: Inaki

last: Etxeberria

first: Erich

last: Sabio

first: Eric

last: Wang

first: Sydney

middle: X

last: Lu

first: Andrea

last: Belleville

first: Nina

last: Fox

first: Cynthia

last: Castro

first: Pu

last: Zhang

first: Takeshi

last: Fujino

first: Jennifer

last: Lewis

first: Beatrice

first: Robert

middle: F

last: Stanley

first: Susan

last: Dewolf

first: Ronan

last: Chaligne

first: Priya

last: Koppikar

first: Jeffrey

last: Molldrem

first: Matthew

last: Gigoux

first: Taha

last: Merghoub

first: Anthony

last: Daniyan

first: Benjamin

last: Greenbaum

first: Christopher

last: Klebanoff

middle: K

last: Bradley

first: Omar

last: Abdel-Wahab


title: Mapping of TOP1cc and R-loops im MCF-7 after TOP1 KD and/or RNase H OE [CHiP_DRIP_Seq]

city: Jerusalem

first: Rami

last: Aqeilan

address: Ein Kerem

country: Israel

institute: Hebrew University of Jerusalem

summary: DNA double-stranded breaks (DSBs) pose a significant threat to genomic integrity, and their generation during essential cellular processes like transcription remains poorly understood. In this study, we employed DRIP-seq and ChIP-seq to map the change in TOP1cc and R-loops after the depletion of TOP1 and R-loops, to comprehensively investigate the interplay between transcription, DSBs, Topoisomerase 1 (TOP1), and R-loops. Our findings revealed the presence of DSBs at highly expressed genes enriched with TOP1 and R-loops, indicating their crucial involvement in transcription-associated genomic instability. Depletion of R-loops and TOP1 specifically reduced DSBs at highly expressed genes, uncovering their pivotal roles in transcriptional DSB formation. By elucidating the intricate interplay between TOP1cc trapping, R-loops, and DSBs, our study provides novel insights into the mechanisms underlying transcription-associated genomic instability. Moreover, we establish a link between transcriptional DSBs and early molecular changes driving cancer development. Notably, our study highlights the distinct etiology and molecular characteristics of driver mutations compared to passenger mutations, shedding light on the potential for targeted therapeutic strategies. Overall, these findings deepen our understanding of the regulatory mechanisms governing DSBs in hypertranscribed genes associated with carcinogenesis, opening avenues for future research and therapeutic interventions.

overall_design: TOP1cc ChIP-seq and DRIP-seq for MCF-7 cells, for control cells transfected with scramble siRNA and invected with empty vector (EV), cells after TOP1 knockdown (KD), cells after RNase H overexpression (OE), cells after both manipulations (TOP1 KD + RNase H OE).

first: Osama

last: Hidmi

first: Sara

last: Oster

first: Jonathan

last: Monin


title: Gene expression profiles of HYDROX-cultured human liver organoids compared to those of Matrigel-cultured human liver organoids.

city: Suita

first: Yanran

last: Tong

state: Osaka

address: 1-6, Yamadaoka

department: Graduate School of Pharmaceutical Sciences

country: Japan

institute: Osaka University

summary: Human liver organoids, an in vitro 3D culture system to recapitulate biological tissue, are expected to be used for drug discovery. However, Matrigel, the most widely used extracellular matrix for organoid culture, has concerns about safety and reproducibility since it is murine-derived. Morever, low hepatic functions of human liver organoids compared to primary human hepatocytes is considered a challenge. Herein, we attempted to culture human liver organoids, established from primary (cryopreserved) human hepatocytes (PHH), using HYDROX, a chemically defined 3D nanofiber. While proliferative capacity of human liver organoids was lost by HYDROX-culture, the gene expression level of a hepatocyte marker CYP3A4 and the CYP3A4 metabolic activity in HYDROX-cultured liver organoids were significantly improved, comparable to those of PHH. HYDROX-cultured liver organoids when treated with hepatotoxic drugs such as acetaminophen showed similar cell viability to that of PHH, suggesting that HYDROX-cultured liver organoids could be applied to drug-induced hepatotoxicity test. Furthermore, HYDROX-cultured liver organoids maintained its functions for up to 35 days and could be used to estimate chronic drug-induced hepatotoxicity such as those of fialuridine. Our findings demonstrated that human liver organoids obtained high liver functions by HYDROX-culture, meaning that HYDROX could contribute to drug discovery as a novel biomaterial.

overall_design: Gene expression profiles of HYDROX-cultured human liver organoids (human liver organodids cultured on HYDROX-plates for 7 days) were compared with those of Matrigel-cultured human liver organoids (each sample was replicated three times).

first: Yukiko

last: Ueyama-Toba

first: Jumpei

last: Yokota

first: Junko

last: Enomoto

first: Haruka

last: Yamazaki

first: Masaki

last: Kanai

first: Hayato

last: Matsui

first: Hiroyuki

last: Mizuguchi


title: Synergically enhanced anti-tumor immunity of in vivo CAR by vaccination-elicited antibody-mediated cellular cytotoxicity boosting

city: Shenzhen

first: Chengzhi

last: Zhao

address: No. 1088 Xueyuan Avenue, Nanshan District, Shenzhen, Guangdong

institute: Southern University of Science and Technology

summary: Chimeric Antigen Receptor (CAR) T cell therapy has shown promise in treating hematologic malignancies. However, it is limited to individualized cell therapy and faces challenges, including high costs, extended preparation time, and limited efficacy against solid tumors. Here, we generated circular RNAs (circRNAs) encoding Chimeric Antigen Receptor (CAR) transmembrane proteins, referred to as circRNACAR, which mediated remarkable tumor killing in both T cells and macrophages. In addition, macrophages exhibited efficient phagocytosis of tumor cells and pro-inflammatory polarization induced by circRNACAR in vitro. We demonstrated that circRNACAR, delivered with immunocyte-tropic lipid nanoparticles (LNPs), significantly inhibited tumor growth, improved survival rates and induced a pro-inflammatory tumor microenvironment in mice. Importantly, the combination of circRNAAnti-HER2-CAR and circRNA-based cancer vaccines encoding the corresponding transmembrane HER2 antigen, termed circRNAHER2, exhibited synergistically enhanced anti-tumor activity. Notably, we found that circRNACAR could boost the level of circRNAHER2-elicited antibodies, which could mediate effective killing of HER2+ tumor cells by macrophages, indicating the potential of vaccination-elicited antibodies in developing novel immunotherapy. This proof-of-concept study demonstrated that the combination of circRNA-based in vivo CAR and vaccines, termed in vivo CAR-VAC, holds the potential to become an upgraded off-the-shelf immunotherapy, and also sheds light on the huge potential of vaccination-elicited antibodies in cancer immunotherapy.

overall_design: comparative gene expression profiling analysis of RNA-seq data for tumor-infiltrating immune cells after PBS, circRNAEV or circRNACAR treatment.

first: Yanyan

last: Wang

first: Xinyue

first: Yichi

last: Xu

first: Liang

last: Qu


title: tp53 R217H and R242H Mutant Zebrafish Display Dysfunctional p53 Hallmarks and Recapitulate LFS Phenotypes [RNA-seq]

city: Ottawa

first: Kim

last: Kobar

state: Ontario

address: 401 Smyth Rd

institute: Children's Hospital of Eastern Ontario Research Institute

summary: Li-Fraumeni syndrome (LFS) is a hereditary cancer predisposition syndrome associated with a highly penetrant and diverse tumor spectrum characterized by germline mutations in the TP53 tumor suppressor gene. To better understand how TP53 mutations predispose individuals with LFS to cancer development, we characterized tp53 R217H and R242H zebrafish lines as the first zebrafish p53 hotspot mutants (human R248 and R273H). These mutants have lost several key wildtype p53 functions and recapitulate many LFS phenotypes. Specifically, we have shown that the R217H and R242H alleles result in partial-to-no activation of key p53 target genes, are resistant to apoptosis in a dominant negative manner and exhibit a defective G1 cell-cycle checkpoint in vivo. The loss of these wildtype p53 functions predisposed the fish to develop spontaneous tumors as early as 6 months of age. Tumor histology resembles human sarcomas, a predominant LFS tumor type. tp53 R242H mutants developed tumors earlier with a higher lifetime incidence than tp53 null or R217H mutants, suggesting it is a more aggressive mutation, while the R217H allele may be hypomorphic. Additionally, we observed mutation-specific differences both in the tumor type and sex bias across tp53 null, R217H, and R242H genotypes with associated diverse transcriptomic and DNA methylome profiles, impacting metabolism, cell signalling, and biological macromolecule synthesis and degradation. These tp53 zebrafish mutants demonstrate fidelity to their human counterparts and provide new insights into underlying tumorigenesis mechanisms and kinetics, which may inform more tailored tumor surveillance approaches and novel therapeutic targets in LFS.

overall_design: We generated tp53 zebrafish mutants representing two of the most common mutation residues found in human cancers, tp53 R217H and R242H (human R248 and R273) using CRISPR to knock in the mutations and also obtained a CG1 tp53 null line from the Langenau lab (Ignatius et al., 2018). We also used the transparent casper strain as a comparison to zebrafish with wildtype p53. Of note, the tp53 null fish are in the CG1 background while the casper, tp53 R217H, and tp53 R242H fish are in the AB background. We performed bulk RNA sequencing on homozygous pooled samples with fifty 30 hour post-fertilization (hpf) larvae exposed to 0 or 30 Gy of irradiation at 24 hpf. We had three biological replicate for each genotype and treatment group for a total of 8 different groups with 24 samples total (casper 0 Gy, casper 30 Gy, tp53 null 0 Gy, tp53 null 30 Gy, tp53 R217H 0 Gy, tp53 R217H 30 Gy, tp53 R242H 0 Gy, tp53 R242H 30 Gy).

first: Sergey

middle: V

last: Prykhozhij

first: Jason

middle: N

last: Berman


title: Cell type specific differences in the transcriptomes of adipose derived stem cells and vaginal fibroblasts in patients with pelvic organ prolapse

city: Scottsdale

first: Xianfeng

last: Chen

state: AZ

address: 13400 E Shea Blvd

institute: Mayo Clinic

summary: The purpose of the study was to understand the molecular phenotypes of adipose-derived stem cells (ASCs) and vaginal fibroblasts (VFBs) and whether pelvic organ prolapse (POP) affects their biological properties. We performed RNA sequencing of paired ASCs and VFBs from six patients with POP and six controls (CTRL). We found no differentially expressed genes (DEG) between POP and CTRL in ASCs and VFBs. Less stringent statistical analysis of VFBs transcriptome showed 23 genes with higher and 29 genes with 2-fold lower expression in POP compared to CTRL. Among the latter were five genes involved in the synaptogenesis pathway found to be significant. We were only able to validate POP related differences for VLDLR protein. We found 508 DEG with 4-fold difference between ASCs and VFBs (when  POP and CTRL samples were combined for each cell type). The DEG with higher expression in each cell type formed  distinct functional networks including Homeobox transcription factors, extracellular matrix (ECM) related proteins, growth factors, and others. These data show that cell-specific RNA populations are intrinsically more characteristic than the disease peculiarities and that VLDLR may play a role in POP pathogenesis.

overall_design: The transcriptomes of POP and CTRL in either ASCs or VFBs were compared (DESeq2, FDR <0.05) to identify differentially expressed genes (DEG). Subsequently, the transcriptomes of VFBs were compared between POP and CTRL (non-adjusted p <0.01; 2-fold difference in gene expression) followed by Ingenuity Pathway Analysis on DEG considering that pathways with FDR <0.05 might be pathogenic. The identified DEG were validated at a protein level by immunoblotting. We also performed a pairwise comparison after combining the gene expression data of POP and CTRL for ASCs and VFBs to identify cell type specific DEG and analyzed the functional associations among them (STRING platform).

first: Tchoukalova

last: Yourka

first: Chen

last: Xianfeng


title: The ATHB2 microProtein affects plant development and the shade avoidance response [Iso-seq]

city: Umeå

first: Stephan

last: Wenkel

address: Parallellvägen 21

country: Sweden

institute: Umeå Plant Science Center

summary: Plants display remarkable adaptability to varying light conditions, crucial for their optimal growth and development. Here, we investigate the functional significance of ATHB2-miP, a microProtein arising from an alternative transcript of the ATHB2 gene, in both white light and shade environments, supported by comprehensive phenotyping experiments. ATHB2, a homeodomain-leucine zipper transcription factor, is known for its involvement in growth regulation. By employing genetic, transcriptomic analyses, and phenotyping experiments, we dissect the regulatory role of ATHB2-miP. Transcriptomics unravel genes differentially induced and repressed upon ATHB2-miP expression, elucidating its impact on growth-related pathways. Notably, ATHB2-miP's expression responds to shade conditions, suggesting its role in light-mediated responses. Our research underscores ATHB2-miP's multifaceted contribution to plant growth and development, emphasizing its potential as a pivotal player in light-responsive gene regulation.

overall_design: This submission refers to 3 sets of Iso-Seq data, generated from WT Col-0 Arabidopsis seedlings RNA, grown in white light and treated with 45 and 90 minutes of FR enriched light. We conducted transcriptomics, phenotyping studies, genetic analyses and protein interaction experiments on 1) mutant lines generated by CRISPR-Cas9 where the microProein was deleted 2) a t-knockout line of the full ATHB2 gene and 3) lines overexpressing the microProtein under the control of a 35S promoter 4) a full ATHB2 overexpressor line under the control of a 35S promoter.

first: Wenkel

last: Stephan

first: Ashleigh

last: Edwards

first: Maurizio

last: Chiurazzi


title: Interferons and epigenetic mechanisms in training, priming and tolerance of monocytes and hematopoietic progenitors [RNA-seq]

first: Bikash

last: Mishra

first: Lionel

last: Ivashkiv


title: The S249/T252 site of RB enhanced the function of TRIM24 to activate the mTOR signaling pathway through DUSP2 in prostate cancer

city: Changsha

first: Huaiyuan

last: Liang

address: 183 Renmin Middle Road

institute: Institute of Urological Cancer, Central South University


title: RNA sequencing analysis of TFCP2L1 overexpression in thyroid cancer cells

city: Guangzhou

first: Chuimian

last: Zeng

state: Guangdong

address: No.58, Zhongshan Road II

department: Department of Endocrinology

institute: The First Affiliated Hospital of Sun Yat-sen University

summary: To investigate the genes and pathways regulated by TFCP2L1 in thyroid cancer cells, we established TFCP2L1-overexpressing TPC1 cell line and performed RNA sequencing for further study.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for TPC1-vector cells and TPC1-TFCP2L1 cells

first: Yilin

last: Zhang

first: Yanbing

last: Li

first: Hongyu

last: Guan


title: Single cell RNA sequencing data of brain myeloid cells after experimental stroke

city: Edinburgh

first: Barry

middle: W

last: McColl

address: 49 Little France Crescent

department: Centre for Discovery Brain Sciences

country: United Kingdom

institute: University of Edinburgh

summary: Acute stroke triggers extensive changes to myeloid immune cell populations in the brain that may be targets for limiting brain damage and enhancing repair. Immunomodulatory approaches will be most effective with precise manipulation of discrete myeloid cell phenotypes in time and space. Here, we investigate how stroke alters mononuclear myeloid cell composition and phenotypes at single-cell resolution and key spatial patterns. Our results show that multiple reactive microglial states and monocyte-derived populations contribute to an extensive myeloid cell repertoire in post-stroke brains. We identify important overlaps and distinctions among different cell types/states that involve ontogeny- and spatial-related properties. Notably, brain connectivity with infarcted tissue underpins the pattern of local and remote altered cell accumulation and reactivity. Our discoveries suggest a global but anatomically governed brain myeloid cell response to stroke that comprises diverse phenotypes arising through intrinsic cell ontogeny factors interacting with exposure to spatially organized brain damage and neuro-axonal cues.

overall_design: Distal middle cerebral artery occlusion was performed on three C57BL/6 mice using electrocoagulation. Mononuclear phagocytes (CD45+CD3-CD19-Ly6G-) were isolated from ipsilateral and contralateral brain regions using FACs at day three. Samples obtained from each mouse were individually barcoded with hash-tag oligonucletide-conjugated CD45 antibodies and then multiplexed for 10x Chromium single cell RNA sequencing with feature barcoding technology.

first: Anirudh

last: Patir

first: Jack

last: Barrington


title: A novel type of spinal ependymoma with anaplastic features

city: Hamburg

first: Julia

middle: Elisabeth

last: Neumann

address: Martinistrasse 52

department: ZMNH

country: Germany

institute: University Hospital Hamburg Eppendorf

summary: Ependymoma (EPN) is a type of brain tumour.. Here we present a novel subtype of spinal ependymoma, that shows an anaplastic morphology, OLIG2 and MYCN positvity in immunostaining and a distinct epigenetic and proteomic profile.

overall_design: Bisulphite converted DNA from the 4 samples were hybridised to the Illumina Infinium 850k Human Methylation Beadchip

first: Antonia

last: Gocke

first: Ulrich

last: Schüller


title: RNA sequencing determines nuclear-accumulated Cav3.2iPA on transcriptional regulation of human iPSC-derived sensory neurons

city: Milwaukee

first: Hongwei

last: Yu

state: WI

address: 8701 Watertown plank Road

department: Anesthesiology

institute: Medical College of Wisconsin

summary: By targeting the intrinsically disordered regions (IDR) of calcium channel subtype 3.2 (Cav3.2), we discovered selective T-type/Cav3.2 peptide inhibitors (Cav3.2iPAs) for AAV-mediated peripheral sensory neuron-specific analgesia. We found that Cav3.2iPAs are highly accumulated in the nuclei after expression in human induced pluripotent stem cells-derived sensory neurons (hiPSC-SNs). This study aims to determine whether nuclear-accumulated Cav3.2iPAs disturb the transcriptional regulation of hiPSC-SNs which becomes a safety concern.

overall_design: RNA sequencing compares the transcriptomes in hiPSC-SNs-expressing Cav3.2iPA and hiPSC-SN-expressing Cav3.2NP (control)

first: Angela

last: Mathison


title: Gene expression data from precision cut tumor slices from head and neck or mesothelioma tumors

city: Philadelphia

first: Caitlin

last: Tilsed

state: Pennsylvania

address: 407F 4312 Chestnut St

institute: The University of Pennsylvania

summary: As cancer immunotherapy and precision medicine dynamically evolve, there is greater need for pre-clinical models that can better replicate the intact tumor and its complex tumor microenvironment (TME). Precision-cut tumor slices (PCTS) have recently emerged as an ex vivo human tumor model, offering the opportunity to study individual patient responses to targeted therapies, including immunotherapies. However, little is known about the physiologic status of PCTS and how they can be utilized in view of the intra-tumoral heterogeneity. In this study, we generated PCTS from head and neck cancers (HNC) and mesotheliomas (Meso) and studied their viability in culture, the extent of variability in serial PCTS, and whether tumor immune response can be measured following PCTS T cell activation. We also undertook transcriptomic analyses to understand the changes that occur in the timeframe between PCTS generation and up to 72hr in culture. Our findings suggest that PCTS viability can be tumor-specific and culture conditions-dependent, but an overall viability of up to 72 hrs was observed. Tumor heterogeneity is present and can skew findings if appropriate experimental design and multiple control PCTS are not considered. Activation of endogenous T cells and measurement of immune responses is possible, however, our transcriptomic analyses showed major changes occurring during the first 24hr culture period of PCTS, involving genes related to wound healing, extracellular matrix, hypoxia, and IFNγ-dependent pathways. Our data suggest the PCTS model may be especially useful for studies involving exogenous immune therapies, such as adoptive cell therapy, rather than for the study of the effects of therapy on endogenous immune systems.

overall_design: Tumors were collcted from patients with head and neck or mesothelion and sectioned into thin slices. RNA was collected and sequencing performed on slices fresh after sectioning or after 24, 48 or 72 hours in culture.

first: Trivedi

last: Sumita

first: Tilsed

last: Caitlin

first: Liousa

last: Maria

first: Brotman

last: Joshua

first: Brody

last: Robert

first: Rajasekaran

last: Karthik

first: Singhal

last: Sunil

first: Albelda

last: Steven

first: Klampasta

last: Astero


title: Single-nucleus multi-omics dissection of physical exercise effects on neurocognition

city: Houston

first: Zheng

last: Sun

state: Texas

address: 1329 Moursund St

institute: Baylor College of Medicine


title: Mapping of physiological DNA double stranded breaks in normal breast cells and breast cancer cells [BLISS]

first: Rami

last: Aqeilan

summary: DNA double-stranded breaks (DSBs) pose a significant threat to genomic integrity, and their generation during essential cellular processes like transcription remains poorly understood. In this study, we employed the advanced BLISS techniques to map DSBs, with different genetic manipulations to comprehensively investigate the interplay between transcription, DSBs, Topoisomerase 1 (TOP1), and R-loops. Our findings revealed the presence of DSBs at highly expressed genes enriched with TOP1 and R-loops, indicating their crucial involvement in transcription-associated genomic instability. Depletion of R-loops and TOP1 specifically reduced DSBs at highly expressed genes, uncovering their pivotal roles in transcriptional DSB formation. By elucidating the intricate interplay between TOP1cc trapping, R-loops, and DSBs, our study provides novel insights into the mechanisms underlying transcription-associated genomic instability. Moreover, we establish a link between transcriptional DSBs and early molecular changes driving cancer development. Notably, our study highlights the distinct etiology and molecular characteristics of driver mutations compared to passenger mutations, shedding light on the potential for targeted therapeutic strategies. Overall, these findings deepen our understanding of the regulatory mechanisms governing DSBs in hypertranscribed genes associated with carcinogenesis, opening avenues for future research and therapeutic interventions.

overall_design: In-suspension break labeling in situ and sequencing (sBLISS) for MCF-7 cells, for control cells transfected with scramble siRNA and invected with empty vector (EV), cells after TOP1 knockdown (KD), cells after RNase H overexpression (OE), cells after both manipulations (TOP1 KD + RNase H OE). sBLISS was also performed on MCF-7 cells after E2 treatment, E2 treatment and RNase H OE, and control cells treated with vehicle and infected with EV. Additionally sBLISS was also performed on MCF-7 cells after E2 treatment, E2 treatment and TOP1 KD, and control cells treated with vehicle and transfected with scrambleRNA. sBLISS was also performed on synchronized MCF-7 cells in the G1 cell cycle stage. sBLISS was also performed on HMLE cells, and MCF-10A WT cells as well as MCF-10A RAS transformed cells. all BLISS experiments were replicated in duplicates, and in quadruplicates in HMLE cells.

first: Osama

last: Hidmi

first: Sara

last: Oster

first: Jonathan

last: Monin


title: Alterations in genes associated with cytosolic RNA sensing in whole blood are associated with coronary microvascular disease in SLE

city: Los Angeles

first: Lihong

last: Huo

state: CA

address: 8700 Beverly Blvd, Los Angeles, CA 90048

institute: Cedars-Sinai

summary: Systemic lupus erythematosus (SLE) patients are 90% women and over three times more likely to die of cardiovascular disease than women in the general population. Chest pain with no obstructive cardiac disease is associated with coronary microvascular disease (CMD), where narrowing of the small blood vessels can lead to ischemia, and frequently reported by SLE patients. Using whole blood RNA samples, we asked whether gene signatures discriminate SLE patients with coronary microvascular dysfunction (CMD) on cardiac MRI (n=4) from those without (n=7) and whether any signaling pathway is linked to the underlying pathobiology of SLE CMD.  RNA-seq analysis revealed 143 differentially expressed (DE) genes between the SLE and healthy control (HC) groups, with virus defense and interferon (IFN) signaling being the key pathways identified as enriched in SLE as expected. We next conducted a comparative analysis of genes differentially expressed in SLE-CMD and SLE-non-CMD relative to HC samples. Our analysis highlighted differences in IFN signaling, RNA sensing and ADP-ribosylation pathways between SLE-CMD and SLE-non-CMD. This is the first study to investigate possible gene signatures associating with CMD in SLE, and our data strongly suggests that distinct molecular mechanisms underly vascular changes in CMD and non-CMD involvement in SLE.

overall_design: This study collected whole blood RNA samples from female subjects aged 37 to 57, comprising 11 SLE patients (4 SLE-CMD, 7 SLE-non-CMD) and 10 HC. Total RNA was then used for library preparation and sequencing. Differential gene expression analysis was performed to identify gene signatures associated with CMD in SLE patients.

first: Erica

last: Montano

first: Moumita

last: Bose

first: Caroline

last: Jefferies


title: Effect of TOP1 knockdown and/or RNase H OE on the transcriptional profile of MCF-7 cells [CEL_Seq]

summary: DNA double-stranded breaks (DSBs) pose a significant threat to genomic integrity, and their generation during essential cellular processes like transcription remains poorly understood. In this study, we employed CEL-seq technique to investigate the effect of TOP1 KD and RNase H OE on transcriptional profile, with the different genetic manipulations. Our findings revealed the presence of DSBs at highly expressed genes enriched with TOP1 and R-loops, indicating their crucial involvement in transcription-associated genomic instability. Depletion of R-loops and TOP1 specifically reduced DSBs at highly expressed genes, uncovering their pivotal roles in transcriptional DSB formation. By elucidating the intricate interplay between TOP1cc trapping, R-loops, and DSBs, our study provides novel insights into the mechanisms underlying transcription-associated genomic instability. Moreover, we establish a link between transcriptional DSBs and early molecular changes driving cancer development. Notably, our study highlights the distinct etiology and molecular characteristics of driver mutations compared to passenger mutations, shedding light on the potential for targeted therapeutic strategies. Overall, these findings deepen our understanding of the regulatory mechanisms governing DSBs in hypertranscribed genes associated with carcinogenesis, opening avenues for future research and therapeutic interventions.

overall_design: CEL-seq for MCF-7 cells, for control cells transfected with scramble siRNA and invected with empty vector (EV), cells after TOP1 knockdown (KD), cells after RNase H overexpression (OE), cells after both manipulations (TOP1 KD + RNase H OE).


title: MIR4435-2HG as a novel predictive biomarker of chemotherapy response and death in pediatric B-cell ALL [RNA-seq]

city: New Orleans

first: Jovanny

last: Zabaleta

state: LA

address: 1700 Tulane Ave

department: Interdisciplinary Oncology

institute: LSUHSC

summary: B-cell acute lymphoid leukemias (B-ALL) are the most common neoplastic diseases in children. Survival rates in Hispanics are lower than in non-Hispanic children. It is, therefore, necessary to find predictive and prognostic biomarkers of relapse and death in this population. Our aim was to identify biomarkers of treatment response, which may also predict relapse and death, through identifying differentially expressed and methylated genes between patients who responded or did not respond to induction treatment. DNA and RNA samples were extracted from 28 bone marrow samples from Hispanic children newly diagnosed with B-ALL. mRNA was sequenced using the NextSeq2000 Illumina platform. Bisulfite-treated DNA was annealed to Illumina Infinium EPIC Methylation chips. Gene expression and differential methylation were compared between responders and non-responders at day 15 and at the end of induction chemotherapy. DAPK1, CNKSR3, MIR4435-HG2, CTHRC1, NPDC1, SLC45A3, ITGA6, and ASCL2 were overexpressed and hypomethylated in non-responders. The overexpression of MIR4435-2HG, DAPK1, ASCL2, SCL45A3, CNKSR3, and NPDC1 can predict non-response at day 15 and refractoriness. Additionally, higher expression of MIR4435-2HG increases the probability of non-response, death, and the risk of death. DAPK1, CNKSR3, and MIR4435-2HG are also overexpressed in relapse samples. Finally, MIR4435-2HG overexpression, together with positive minimal residual disease, is associated with poorer survival, and together with high expression of DAPK1 and ASCL2, it could improve the risk classification of patients with normal karyotype. In conclusion, MIR4435-2HG is a potential predictive and prognostic biomarker in children with B-ALL, and its detection at diagnosis could improve survival rates in our patients

overall_design: 26 fresh bone marrow samples.

first: Yulieth

last: Torres-Llanos

first: Nataly

last: Cruz-Rodriguez

first: Sandra

middle: M

last: Quijano

first: Paula

middle: C

last: Guzman

first: Iliana

last: de los Reyes

last: Poveda

first: Ana

last: Infante

first: Liliana

last: Lopez

first: Alba

middle: L

last: Combita


title: MIR4435-2HG as a novel predictive biomarker of chemotherapy response and death in pediatric B-cell ALL [methylation]

type: Methylation profiling by genome tiling array

summary: B-cell acute lymphoid leukemias (B-ALL) are the most common neoplastic diseases in children. Survival rates in Hispanics are lower than in non-Hispanic children. It is, therefore, necessary to find predictive and prognostic biomarkers of relapse and death in this population. Our aim was to identify biomarkers of treatment response, which may also predict relapse and death, through identifying differentially expressed and methylated genes between patients who responded or did not respond to induction treatment. DNA and RNA samples were extracted from 27 bone marrow samples from Hispanic children newly diagnosed with B-ALL. mRNA was sequenced using the NextSeq550 Illumina platform. Bisulfite-treated DNA was annealed to Illumina Infinium EPIC Methylation chips. Gene expression and differential methylation were compared between responders and non-responders at day 15 and at the end of induction chemotherapy. DAPK1, CNKSR3, MIR4435-HG2, CTHRC1, NPDC1, SLC45A3, ITGA6, and ASCL2 were overexpressed and hypomethylated in non-responders. The overexpression of MIR4435-2HG, DAPK1, ASCL2, SCL45A3, CNKSR3, and NPDC1 can predict non-response at day 15 and refractoriness. Additionally, higher expression of MIR4435-2HG increases the probability of non-response, death, and the risk of death. DAPK1, CNKSR3, and MIR4435-2HG are also overexpressed in relapse samples. Finally, MIR4435-2HG overexpression, together with positive minimal residual disease, is associated with poorer survival, and together with high expression of DAPK1 and ASCL2, it could improve the risk classification of patients with normal karyotype. In conclusion, MIR4435-2HG is a potential predictive biomarker in children with B-ALL, and its detection at diagnosis could improve survival rates in our patients.


title: Chromatin accessibility profile at single nuclei level of cells from mouse hippocampus

first: Zheng

last: Sun

summary: We performed single-nucleus RNA-seq and chromatin accessibility analyses (snRNA-seq and ATAC-seq) in the hippocampus of wild-type (WT) mice and mice carrying amyloid precursor protein knock-in mutations (APPNL-G-F) in response to chronic wheel-running exercise.

overall_design: 8-week old female WT and APPNL-G-F mice were put into cages with running wheel for voluntary exercise for 10 weeks. Mice from 4 different conditions (WT_RT, WT_EX, APP_RT, APP_EX) were subjected to single nuclei RNA and ATAC sequencing.

first: Li

last: Wenbo

first: Yumei

last: Li


title: MIR4435-2HG as a novel predictive biomarker of chemotherapy response and death in pediatric B-cell ALL

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

overall_design: Refer to individual Series


title: R-loops and Topoisomerase 1 facilitate formation of transcriptional DSBs at gene bodies of hypertranscribed cancer genes

first: Rami

last: Aqeilan

summary:
DNA double-stranded breaks (DSBs) pose a significant threat to genomic integrity, and their generation during essential cellular processes like transcription remains poorly understood. In this study, we employed advanced techniques to map DSBs, R-loops, and Topoisomerase 1 cleavage complex (TOP1cc) and re-analyzed ChIP-seq and DRIP-seq data to comprehensively investigate the interplay between transcription, DSBs, Topoisomerase 1 (TOP1), and R-loops. Our findings revealed the presence of DSBs at highly expressed genes enriched with TOP1 and R-loops, indicating their crucial involvement in transcription-associated genomic instability. Depletion of R-loops and TOP1 specifically reduced DSBs at highly expressed genes, uncovering their pivotal roles in transcriptional DSB formation. By elucidating the intricate interplay between TOP1cc trapping, R-loops, and DSBs, our study provides novel insights into the mechanisms underlying transcription-associated genomic instability. Moreover, we establish a link between transcriptional DSBs and early molecular changes driving cancer development. Notably, our study highlights the distinct etiology and molecular characteristics of driver mutations compared to passenger mutations, shedding light on the potential for targeted therapeutic strategies. Overall, these findings deepen our understanding of the regulatory mechanisms governing DSBs in hypertranscribed genes associated with carcinogenesis, opening avenues for future research and therapeutic interventions.

This SuperSeries is composed of the SubSeries listed below.


title: Comparative 3D genomic architectures elucidate the function of the adaptive human-specific inversions in human brain evolution

city: Beijing

first: Ying

last: Liu

address: 5 Summer Palace Road, Haidian District, Beijing

department: College of Future Technology

institute: Peking University

summary: Structural variation (SV) is a class of genetic variants involving long stretches of DNA and provides substantial innovations for adaptation and evolution. Investigation of SV profiles and turnover in nonhuman primates can help answer fundamental questions, such as what makes us uniquely human. Here, we present the high-resolution comparative 3D genome organizations of prefrontal cortex (PFC) of adult human and rhesus macaque. Based on a comprehensive SV atlas in rhesus macaque populations, we found that inversions are selectively constrained from the perspective of 3D genome, those inducing dramatic changes in chromosomal conformation tends to be purged by the action of purifying selection, implying the inconspicuous adaptive roles of inversions associated with strong effects. Furthermore, by integrating comparative genomics and multi-omics data across human and rhesus macaque, such as gene structures, regulatory elements and 3D genomic organizations, we highlighted a category of human-specific inversions with strong effects and found that they had undergone rapid fixation and caused significant changes in gene expression in human brain development, which may represent a driving force in human brain evolution. Overall, our findings reveal the great value of comparative 3D Hi-C maps in elucidating the adaptive functions of structural variants.

sample_organism:
Macaca mulatta
Homo sapiens

platform_organism:
Macaca mulatta
Homo sapiens

overall_design: Hi-C performed on prefrontal cortex (PFC) of an adult human and a rhesus macaque

first: Wanqiu

last: Ding

first: Xiangshang

first: Jie

last: Zhang

first: Lu

last: Tian

first: Chuan-Yun


title: Macrophages modulate fibrosis during newt lens regeneration [PWaltl]

city: Baltimore

first: Seth

last: Blackshaw

state: MD

address: 733 N Broadway

department: Neuroscience

institute: Johns Hopkins University

summary: Previous studies indicated that macrophages play a role during lens regeneration in newts, but their function has not been tested experimentally. Here we generated a transgenic newt reporter line in which macrophages can be visualized in vivo. Using this new tool, we analyzed the location of macrophages during lens regeneration. We uncovered early gene expression changes using bulk RNAseq in two newt species, Notophthalmus viridescens and Pleurodeles waltl. Next, we used clodronate liposomes to deplete macrophages, which inhibited lens regeneration in both newt species. Macrophage depletion induced the formation of scar-like tissue, an increased and sustained inflammatory response, an early decrease in iris pigment epithelial cell (iPEC) proliferation and a late increase in apoptosis. Some of these phenotypes persisted for at least 100 days and could be rescued by exogenous FGF2. Re-injury alleviated the effects of macrophage depletion and re-started the regeneration process. Together, our findings highlight the importance of macrophages in facilitating a pro-regenerative environment in the newt eye, helping to resolve fibrosis, modulating the overall inflammatory landscape and maintaining the proper balance of early proliferation and late apoptosis.

overall_design: Bulk RNA-seq was performed using dorsal iris tissue from Pleurodeles waltl over a time course of lens regeneration, including from the intact iris, 1 day post-lentectomy and 4 days post-lentectomy.

first: Georgios

last: Tsissios

first: Anthony

last: Sallese

first: J

last: Perez-Estrada

first: Jared

last: Tangeman

first: Weihao

last: Chen

first: Byran

last: Smucker

first: Sophia

last: Ratvasky

first: Erika

last: Grajales-Esquivel

first: Arielle

last: Martinez

first: Kimberly

last: Visser

first: Alberto

last: Araus

first: Hui

last: Wang

first: Andras

last: Simon

first: Maximina

middle: H

last: Yun

first: Katia

last: Del Rio-Tsonis


title: Macrophages modulate fibrosis during newt lens regeneration [Nvir]

overall_design: Bulk RNA-seq was performed using dorsal iris tissue from Notophthalmus viridescens over a time course of lens regeneration, including from the intact iris, 6 hours post-lentectomy, 1 day post-lentectomy and 4 days post-lentectomy.


title: Gene expression profile at single nuclei level of cells from mouse hippocampus

first: Zheng

last: Sun

first: Li

last: Wenbo

first: Yumei

last: Li


title: Macrophages modulate fibrosis during newt lens regeneration.

first: Seth

last: Blackshaw

sample_organism:
Notophthalmus viridescens
Pleurodeles waltl


title: Upregulation of inflammatory genes and pathways links obesity to severe COVID-19

status: Public on May 21 2024

first: Jay

last: Kolls

address: 1430 Tulane Ave

department: Medicine

institute: Tulane University

summary: Obesity is a risk factor for developing severe COVID-19. However, the mechanism underlying obesity-accelerated COVID-19 remains unclear. Here, we report results from a study in which K18-hACE2 mice were fed an obesity-inducing western diet (WD) for over 3 months before intranasal infection with SARS-CoV2. After infection, the WD-fed K18-hACE2 mice lost more body weight and had more severe lung inflammation than normal chow (NC)-fed mice.  Bulk RNAseq analysis of lungs and adipose tissue revealed that a diverse landscape of various immune cells, inflammatory markers, and pathways are upregulated in obese COVID-19 patients or the WD-fed K18-hACE2 mice when compared with their respective control groups. When compared with infected NC-fed mice in the lung, the infected WD-fed mice had upregulation of IL-6, a well-established marker for severe COVID-19. These results indicate that obesity-accelerated severe COVID-19 caused by SARS-CoV-2 infection in the K18-hACE2 mouse model can be used for dissecting the cellular and molecular mechanisms underlying pathogenesis. Furthermore, in the transcriptome analysis of the lung and adipose tissue obtained from deceased COVID-19 patients, we found upregulation of an array of genes and pathways associated with Inflammation. Both the K18-hACE2 mouse model and human COVID-19 patient data support a link between inflammation and an obesity-accelerated COVID-19 disease phenotype.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Comparitive gene expression analysis was done using bulk RNA-seq data comparing NC (normal chow) and WD (western diet to induce obesity) fed K18-hACE2 mice (adipose tissue and lung tissue) infected with SARS-CoV-2 (WA).

first: Joshua

last: Currey

first: Xuebin

last: Qin

first: Mst

last: Shamima Khatun


title: CDK4/6 inhibits the STING signaling axis through RB1-dependent and RB1-independent pathways in prostate cancer

first: Huaiyuan

last: Liang

summary: The efficacy of immunotherapy in prostate cancer is not satisfactory due to the heterogeneity of this cancer, the “cold” tumor microenvironment and the paucity of neoantigens. The STING-TBK1-IRF3 signaling axis as core molecules of the innate immune system is increasingly recognized as a candidate target for cancer immunotherapy. Inhibition of CDK4/6 has been reported to r increase T-cell activation to enhance antitumor immunity. However, little is known about the relationship between CDK4/6 and innate immune-related STING signaling. In this study, we revealed that combination treatment with CDK4/6 inhibitors and STING agonists is significantly more effective than treatment with STING agonists alone in eliminating prostate cancer cells. Then, we demonstrated that CDK4/6 phosphorylate TBK1 at S527 to inactive the STING signaling pathway independent of RB1 in prostate cancer cell. On the other hand, CDK4/6 phosphorylate RB1 at S249/T252 to promote the interaction of RB1 with TBK1 and diminish the phosphorylation of TBK1 at S172, which also suppresses the activation of the STING pathway. Collectively, we found that CDK4/6 inhibits the STING/TBK1/IRF3 axis through RB1-dependent and RB1-independent pathways and elucidated the detailed mechanism. These insights lay a solid foundation for the synergistic application of CDK4/6 inhibitors with STING activators in prostate cancer patients.

overall_design: Chromatin immunoprecipitation DNA-sequencing (CUTtag) for the prostate cancer after knockdown of RB1 by siRNA, overexpression of RBN and RB(244-255) by plasmids in 22RV1 cells.

first: Xin

last: Jin


title: Functional characterization of a heat shock factor regulated enhancer at coronary artery disease associated locus that controls CALCRL expression in endothelial cells

city: Kuopio

first: Minna

middle: U

last: Kaikkonen

address: Neulaniementie 2

country: Finland

institute: A.I.Virtanen Institute for Molecular Sciences

summary: We performed RNA-seq in TeloHAEC with samples where CALCRL was silenced using siRNA or enhancer-8 deleted using CRISPR-cas9 system to investigate the genes that are differentially expressed under CALCRL knockdown.

overall_design: mRNA expression profiling of teloHAEC cells following si-CALCRL and enhancer-8 CRISPR-Cas9 deletion.

first: Ilakya

last: Selvarajan


title: The S249/T252 site of RB enhanced the function of TRIM24 to activate the mTOR signaling pathway through DUSP2 in prostate cancer [RNA-Seq]

summary: Prostate cancer (PC) is the most common tumour diagnosis and a serious threat to the lives of men worldwide. It is well recognized that androgens play a unique role in prostate cancer and thus endocrine therapy, represented by androgen deprivation therapy (ADT), is a cornerstone of prostate cancer treatment. However, after ADT treatment, it is inevitable that a number of changes will occur leading from hormone sensitive prostate cancer to castration-resistant prostate cancer (CRPC).  RB1 deletion, which is only present in 21% of the population, is the only genomic factor associated with the prognosis of androgen receptor signaling inhibitor therapy. We previously reported that phosphorylated RB has the ability to inhibit p65 in prostate cancer, thereby promoting immune tumour escape and increasing sensitivity to radiotherapy. The N-terminal end of RB also enhances the sensitivity of CDK4/6 inhibitors by recognizing the FXXXV amino acid motif on HDAC5 to down-regulate cell cycle-related genes. Recently, it has been shown that TRIM28 binds to and mediates the ubiquitinated degradation of RB protein. However, the relationship between TRIM24 and RB remains a mystery. In this study, we found that the phosphorylated RB N-terminal can interact with TRIM24 to form a complex that enhances AR signaling. And RB/TRIM24 used DUSP2 as an intermediate bridge to activate the mTOR pathway and promote prostate cancer progression. Finally, we also designed RB-linker-protac, which can attenuate TRIM24 protein levels and inactivate the mTOR signaling pathway, thereby inhibiting prostate cancer. This provides new insights into the treatment of prostate cancer.

overall_design:
Comparative gene expression profiling analysis of RNA-seq data for the prostate cancer after knockdown of TRIM24 and RB1 by siRNA, overexpression of RBN and RB(244-255) by plasmids and inhibition of RB1 by RB Protein hydrolysis targeting chimeras (PROTAC) in 22RV1 cells.
Comparative gene expression profiling analysis of RNA-seq data for the C4-2 cells after kncokdown of TRIM24 and RB1


title: Transcriptome analysis of pancreatic cancer patients with low and high metabolic tumor volume (MTV)

city: Seoul

first: Baek Gil

last: Kim

address: 50-1 Yonsei-Ro

country: South Korea

institute: Yonsei University

summary: Transcriptome wide-comparison between MTV-low and -high groups was performed with three types: 1) cancer to normal site ratio, cancer site, and normal site. Among three types of comparison, the cancer-to-normal site ratio was the most informative. 211 genes were generally upregulated, and 166 genes were downregulated in the MTV-high group compared to the MTV-low group. Of 211 upregulated gens, 44 showed more than a 2-fold difference and statistical significance. On the other hand, of 166 downregulated genes, 56 showed more than a 2-fold difference and statistical significance. In functional annotation analysis, the GO terms of the upregulated genes were enriched to Wnt signaling (GO: 0035567, 0060070, 0060071), sister chromatid cohesion (GO: 0007062), and cell division (GO: 0000278, 0051301) trends (Figure 1E-a), whereas those of the downregulated genes were enriched to rRNA processing (GO: 0006364), translation (GO: 0006412, 0006413), nuclear-transcribed mRNA catabolic process (GO: 0000184) trends (Figure 1E-b). 28 upregulated genes belonged to Wnt signaling or Cell division, while 22 downregulated genes belonged to rRNA processing.

overall_design: Pancreatic cancer patients were classified into low MTV and high MTV groups by 18F-FDG PET/CT scan, and their post-resected tissues were used for total RNA-Seq.

first: Baek

middle: G

first: Sung

last: Lee

first: Yeonsue

last: Jang

first: Suki

last: Kang

first: Chang

first: Nam

last: Cho


title: Uncovering novel ferroptosis regulators in pancreatic ductal adenocarcinoma (PDAC)

first: Dan

last: Su

address: No. 1 Shuaifuyuan, Wangfujing Street, Beijing

department: Department of General Surgery

institute: Peking Union Medical college hospital


title: Uncovering novel ferroptosis regulators in pancreatic ductal adenocarcinoma (PDAC) using parallel Ribo-seq

summary: Pancreatic ductal adenocarcinoma (PDAC) is one of the most intractable malignancies and systematic chemotherapies are the most commonly-used standard treatment for PDAC patients diagonosed at advanced stages. However, drug resistance may arise as early as several weeks after chemotherapy initiation, thus limiting the therapeutic efficacy of chemotherapy reagents. Accumulating evidence has demonstrated that acquired tolerance to apoptotic cell death induced by cytotoxic drugs is one of the critical reasons for the failure of chemotherapy. Ferroptosis was originally identified as a unique form of programmed cell death, which is characterized by shrunk mitochondria accompanied with condensed membranes and decreased crista, in drug screens searching for small molecules that can selectively kill RAS-expressing cancer cells. Unlike apoptosis, ferroptosis is driven by iron-catalyzed excessive lipid peroxidation, ultimately leading to the rupture of plasma membrane and cell death. Recent studies indicated that ferroptosis induction can be a promising therapeutic strategy in cancer treatment since it not only can kill tumor cells directly but also can synergize with chemotherapy ,radiotherapy,targeted therapy and immunotherapy. However, transformed malignant cells can acquired resistance to ferroptosis through adaptative changes in their epigenome, genome, transcriptome, translatome or proteome. Multiple mechanisms, such as decreased coenzyme Q10 production mediated by the upregulation of apoptosis-inducing factor mitochondria-associated 2 (AIFM2/FSP1) and elevated expression of cytoprotective genes as a result of the activation of antioxidant transcription factor NFE2L2/NRF2 (nuclear factor erythroid 2-like 2), have been demonstrated to confer ferroptosis resistance. However, transformed malignant cells can acquired resistance to ferroptosis through adaptative changes in their epigenome, genome, transcriptome, translatome or proteome. Multiple mechanisms, such as decreased coenzyme Q10 production mediated by the upregulation of apoptosis-inducing factor mitochondria-associated 2 (AIFM2/FSP1) and elevated expression of cytoprotective genes as a result of the activation of antioxidant transcription factor NFE2L2/NRF2 (nuclear factor erythroid 2-like 2), have been demonstrated to confer ferroptosis resistance.

overall_design: To uncover novel ferroptosis regulators in PDAC, CFPAC-1 and PANC-1 cells were treated with 10 μM or 1 μM  IKE (imidazole ketone erastin, a derivative of erastin) for 24 h. PDAC cells treated with 0.1% DMSO were used as a parallel control.  After removing dead cells and cell debris, survived PDAC cells were collected for sequencing analysis.

first: Ruobing

last: Wang

first: Chen

last: Ding

first: Taiping

last: Zhang


title: The S249/T252 site of RB enhanced the function of TRIM24 to activate the mTOR signaling pathway through DUSP2 in prostate cancer [ChIP-Seq]

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for the prostate cancer after knockdown of RB1 by siRNA, overexpression of RBN and RB(244-255) by plasmids in 22RV1 cells.


title: Transcriptional profiling of retinal astrocytes identifies a specific marker and points to functional specialization

city: Cambridge

first: William

middle: Jacob

last: Gammerdinger

state: MA

address: Massachusetts Hall

institute: Harvard University

summary: Astrocyte heterogeneity is an increasingly prominent research topic, and studies in the brain have demonstrated substantial variation in astrocyte form and function, both between and within regions. In contrast, retinal astrocytes are not well understood and remain incompletely characterized. Along with optic nerve astrocytes, they are responsible for supporting retinal ganglion cell axons and an improved understanding of their role is required. We have used a combination of microdissection and Ribotag immunoprecipitation to isolate ribosome-associated mRNA from retinal astrocytes and investigate their transcriptome, which we also compared to astrocyte populations in the optic nerve. Astrocytes from these regions are transcriptionally distinct, and we identified retina-specific astrocyte genes and pathways. Moreover, although they share much of the ‘classical’ gene expression patterns of astrocytes, we uncovered unexpected variation, including in genes related to core astrocyte functions. We additionally identified the transcription factor Pax8 as a highly specific marker of retinal astrocytes and demonstrated that these astrocytes populate not only the retinal surface, but also the prelaminar region at the optic nerve head. These findings are likely to contribute to a revised understanding of the role of astrocytes in the retina.

overall_design: We investigated the transcriptional profile of retinal astrocytes by analyzing differential gene expression between highly enriched IP (immunoprecipitated) RNA from these cells and RNA from Input controls. We also compared gene expression with immunoprecipitated RNA from optic nerve head (ONH) and myelinated optic nerve proper (ONP) astrocytes, which form separate populations. Retinal astrocyte RNA from IP samples and Input controls was isolated and used for RNA-seq, then analyzed alongside our existing RNA-seq data for ONH and ONP astrocytes from a prior study; a pair of adapter samples (one each for ONH and ONP astrocytes) were prepared and sequenced alongside the retinal IP and Input samples to control for potential batch effects.

first: Paul

last: Cullen

first: Shannan

last: Ho Sui

first: Arpan

last: Mazumder

first: Daniel

last: Sun


title: Uncovering novel ferroptosis regulators in pancreatic ductal adenocarcinoma (PDAC) using parallel RNA-seq

summary: Pancreatic ductal adenocarcinoma (PDAC) is one of the most intractable malignancies and systematic chemotherapies are the most commonly-used standard treatment for PDAC patients diagonosed at advanced stages. However, drug resistance may arise as early as several weeks after chemotherapy initiation, thus limiting the therapeutic efficacy of chemotherapy reagents. Accumulating evidence has demonstrated that acquired tolerance to apoptotic cell death induced by cytotoxic drugs is one of the critical reasons for the failure of chemotherapy. Ferroptosis was originally identified as a unique form of programmed cell death, which is characterized by shrunk mitochondria accompanied with condensed membranes and decreased crista, in drug screens searching for small molecules that can selectively kill RAS-expressing cancer cells. Unlike apoptosis, ferroptosis is driven by iron-catalyzed excessive lipid peroxidation, ultimately leading to the rupture of plasma membrane and cell death. Recent studies indicated that ferroptosis induction can be a promising therapeutic strategy in cancer treatment since it not only can kill tumor cells directly but also can synergize with chemotherapy ,radiotherapy,targeted therapy and immunotherapy. However, transformed malignant cells can acquired resistance to ferroptosis through adaptative changes in their epigenome, genome, transcriptome, translatome or proteome. Multiple mechanisms, such as decreased coenzyme Q10 production mediated by the upregulation of apoptosis-inducing factor mitochondria-associated 2 (AIFM2/FSP1) and elevated expression of cytoprotective genes as a result of the activation of antioxidant transcription factor NFE2L2/NRF2 (nuclear factor erythroid 2-like 2), have been demonstrated to confer ferroptosis resistance. However, transformed malignant cells can acquired resistance to ferroptosis through adaptative changes in their epigenome, genome, transcriptome, translatome or proteome. Multiple mechanisms, such as decreased coenzyme Q10 production mediated by the upregulation of apoptosis-inducing factor mitochondria-associated 2 (AIFM2/FSP1) and elevated expression of cytoprotective genes as a result of the activation of antioxidant transcription factor NFE2L2/NRF2 (nuclear factor erythroid 2-like 2), have been demonstrated to confer ferroptosis resistance. Therefore, unravelling the underlying mechanisms responsible for reduced sensitivity to ferroptosis inducers, such as erastin or RSL3, may pave the way for designing effective therapeutic regimens for PDAC patients.


title: BET inhibition reforms the immune microenvironment and alleviates T-cell dysfunction in chronic lymphocytic leukemia

city: Omaha

first: Dalia

last: El-Gamal

state: NE

address: 986805 NE Med Center BCC 4.12.396

department: Eppley Institute

institute: University of Nebraska Medical Center

summary: Redundant tumor microenvironment (TME) immunosuppressive mechanisms and epigenetic maintenance of terminal T-cell exhaustion greatly hinder functional anti-tumor T-cell responses in chronic lymphocytic leukemia (CLL). Bromodomain and extra-terminal (BET) proteins regulate key pathways contributing to CLL pathogenesis and TME interactions, including T-cell function and differentiation. Herein, we report that blocking BET protein function alleviates immunosuppressive networks in the CLL TME and repairs inherent T-cell defects. The pan-BET inhibitor OPN-51107 reduced exhaustion-associated cell signatures, fostering improved T-cell proliferation and effector function in patient samples and murine models of CLL. These findings establish the immunomodulatory action of BET-i on CLL T-cells and support the use of BET inhibitors in treatment strategies for CLL.

overall_design: In order to assay the global effect of the BET inhibitor OPN-51107 on the chronic lymphocytic leukemia tumor microenvironment, Eμ-TCL1 adoptive transfer mice were treated with OPN-51107 or vehicle control for 3 weeks. Murine splenic cells and peripheral blood mononuclear cells (PBMCs) were then evaluated for gene expression changes using the NanoString PanCancer IO 360™ Panel.

first: Audrey

last: Smith

first: Sydney

middle: A

last: Skupa

first: Alexandria

middle: P

last: Eiken

first: Timothy

middle: E

last: Reznicek

first: Elizabeth

last: Schmitz

first: Nolan

last: Williams

middle: Y

last: Moore

first: Christopher

last: D'Angelo

first: Avyakta

last: Kallam

first: Matthew

last: Lunning

first: R

last: Bociek

first: Julie

last: Vose

first: Eslam

last: Mohamed

first: Anna

last: Mahr

last: Denton

first: Ben

last: Powell

first: Gideon

last: Bollag

first: M

last: Rowley


title: Iron controls the development of airway hyperreactivity by regulating ILC2 metabolism and effector function

first: Benjamin

last: Hurrell

address: 1450 Biggy Street

institute: USC

summary: Group 2 innate lymphoid cells (ILC2s) rapidly induce a type 2 inflammation in the lungs in response to allergens. Here, we focused on the role of iron – a critical nutritional trace element – on ILC2 function and asthma pathogenesis. In the lungs, transferrin receptor 1 (TfR1) is rapidly upregulated and functional during ILC2 activation, while blocking transferrin uptake reduces ILC2 expansion and activation. Iron deprivation reprograms ILC2 metabolism, inducing a HIF-1a-driven upregulation of glycolysis and inhibition of oxidative mitochondrial activity. Consequently, in vivo iron chelation or induction of hypoferremia notably reduces the development of airway hyperreactivity in experimental models of ILC2-driven allergic asthma. Human circulating ILC2s rapidly induce TfR1 during activation, while iron deprivation reduces effector functions. Finally, we found a negative relation between circulating ILC2 TfR1 expression and airway function in cohorts of patients with asthma. Collectively, our studies define cellular iron as a critical regulator of ILC2 function.

overall_design: Activated lung ILC2s were FACS sorted and cultured with an iron chelator for 18h after which culture was supplemented or not with iron for an additional 8 hours.


title: Deciphering lineage specification during early embryogenesis in mouse gastruloids using integrative multi-layered proteomics [scRNA-seq]

city: Nijmegen

first: Suzan

last: Stelloo

address: Geert Grooteplein Zuid 28

department: Molecular Biology

country: Netherlands

institute: Radboud University

summary: Gastrulation is a critical stage of embryonic development during which the three germ layers are established. Deciphering the molecular mechanisms underlying this proces from a protein perspective remains a significant challenge. To address this, we employed a multilayered mass spectrometry-based proteomics approach to investigate the global dynamics of (phospho)protein expression during differentiation of ESCs towards gastruloids – an in vitro model of gastrulation-stage embryogenesis. Our findings revealed that many proteins exhibited temporal expression with unique expression profiles corresponding to the three germ layers. Additionally, we profiled enhancer interaction landscapes in ESCs and gastruloids using p300 proximity labeling, which revealed numerous gastruloid-specific transcription factors and chromatin remodelers. Subsequent degron based perturbations combined with scRNA-seq revealed a critical role for Zeb2 in regulating mouse and human somitogenesis. Overall, this study provides a rich resource for developmental and synthetic biology communities endeavoring to understand mammalian embryogenesis.

overall_design: Somitic gastruloids were generated as previously described (Protoc. Exch. https://doi.org/10.1038/protex.2018.094, van den Brink et al., Nature 2020).  Somitic gastruloids expressing Zeb2-degron tag were treated with 100nM dTAG-13 or DMSO at 72 and 96h during gastruloid formation.

first: Michiel

last: Vermeulen


title: Transplanted pluripotent stem cell derived-islets engraft superior to human islets

city: Helsinki

first: Jouni

middle: Antero

last: Kvist

state: Uusimaa

address: Haartmaninkatu 8

department: Research Program for Molecular Neurology

institute: University of Helsinki

summary: To investigate gene expression differences between human stem cell-derived islets and human islets before transplantation. We performed gene expression profiling analysis using data obtained from RNA-seq of 5 different batches of stem cell-derived islets and human islets from 7 donors.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for stem cell-derived islets (sample A-E) and human islet donors (sample I-VII)).

first: Julia

last: Thorngren

first: Anja

last: Brboric

first: Svitlana

last: Vasylovska

first: Daisy

last: Hjelmqvist

first: Gunilla

middle: T

last: Westermark

first: Jonna

last: Saarimäki-Vire

first: Diego

last: Balboa

first: Timo

last: Otonkoski

first: Per-Ola

last: Carlsson

first: Joey

last: Lau


title: CD100 stimulation on primary hepatocyte

city: Xi'an

first: Chao

last: Fan

address: #169, Changle Road, Xincheng District

institute: The Fourth Military Medical University

summary: To determine the effects of CD100 stimulation on hepatocyte, we have employed mRNA microarray expression profiling as a discovery platform to identify genes with the potential to response to CD100 stimulation, for evaluating the CD100 effects on the metabolism of hepatocyte. The primary hepatocyte, but not the hepatocellular cell lines, was used for the CD100 stimulation experiment ex vivo, and different expressing genes was identified that distinguished among MOCK, CD100 stimulated and TNF-a-stimulated (as a positive control) groups.

overall_design: The hepatocytes (10E6 each well) were stimulated by 1μg BSA (MOCK), 1μg CD100 or 200ng TNF-a (as positive control). The gene expression of MOCK group, CD100-stimulated group and TNF-a-stimulated group were measured by microarray at 24 hours after exposure to different stimulators. Three independent experiments were performed at each time using the primary hepatocytes purchased from Zenobio (Research Triangle, Raleigh-Durham, NC, US).


title: Reanalysis of peripheral blood gene expression of IFIH1 rs1990760 variants in critically-ill COVID-19 patients with and without corticosteroids treatment [third-party re-analysis of GSE177025]

city: La Jolla, CA 92093

first: Pradipta

last: Ghosh

address: 9500 Gilman Drive (MC 0651)

department: Departments of Medicine and Cellular and Molecular Medicine

institute: University of California San Diego

summary: Peripheral blood gene expression analysis of IFIH1 rs1990760 variants in critically-ill COVID-19 patients with and without corticosteroids treatment

relation:
Reanalysis of: GSM5370588
Reanalysis of: GSM5370589
Reanalysis of: GSM5370590
Reanalysis of: GSM5370591
Reanalysis of: GSM5370592
Reanalysis of: GSM5370593
Reanalysis of: GSM5370594
Reanalysis of: GSM5370595
Reanalysis of: GSM5370596
Reanalysis of: GSM5370597
Reanalysis of: GSM5370598
Reanalysis of: GSM5370599
Reanalysis of: GSM5370600
Reanalysis of: GSM5370601
Reanalysis of: GSM5370602
Reanalysis of: GSM5370603
Reanalysis of: GSM5370604
Reanalysis of: GSM5370605
Reanalysis of: GSM5370606
Reanalysis of: GSM5370607
Reanalysis of: GSM5370608

overall_design:
Re-analysis of GSE177025
Basecalling was performed using the default Ion Torrent software
trimming was performed using the fastp software v0.20.1 and the parameters -f 3, -r, -M 15
Counts were calculated using the pseudoalignment tool salmon v1.4.0 with the default parameters for mapping-based quantification and the hg38 salmon partial index
Reads were loaded in R using the tximport library
Genome_build: hg38 (GRCh38)
The 21n_expr.txt includes log2 of TPM values


title: Deciphering lineage specification during early embryogenesis using integrative multi-layered proteomics [RNA-Seq]

summary: Gastrulation is a critical stage in embryonic development during which the germ layers are established. The advances and increasing availability of sequencing technologies led to the identification of gene regulatory programs that control the emergence of the germ layers and its derivatives. However, proteome-based studies of early mammalian development are scarce. To overcome this, we utilized gastruloids and a multilayered mass spectrometry-based proteomics approach to investigate the global dynamics of (phospho)protein expression during gastruloid differentiation. Our findings revealed that many proteins exhibit temporal expression with unique expression profiles corresponding to the three germ layers, which we also validated using single-cell proteomics technology. Notably, detected temporal protein expression dynamics in gastruloids aligns with equivalent mouse embryo stages. Additionally, we profiled enhancer interaction landscapes using P300 proximity labeling, which revealed numerous gastruloid-specific transcription factors and chromatin remodelers. Subsequent degron-based perturbations combined with scRNA-seq identified a critical role for ZEB2 in regulating mouse and human somitogenesis. Overall, this study provides a rich resource for developmental and synthetic biology communities endeavoring to understand mammalian embryogenesis.

overall_design: Mouse gastruloids were generated as previously described (Protoc. Exch. https://doi.org/10.1038/protex.2018.094, van den Brink et al., Nature 2020). Human somitoids were generated following the protocol by Sanaki-Matsumiya et al., Nature Communications 2022)


title: An age-specific differentiation path from hematopoietic stem cells contributes to exacerbated thrombosis

city: Santa Cruz

first: Camilla

last: Forsberg

state: California

address: 1156 High Street

institute: Institute for the Biology of Stem Cells, University of California Santa Cruz

summary: To investigate the transcriptome of age-specific MkPs compared to canonical MkPs

overall_design: mRNA levels were examined in Young or Old Lin-cKit+Sca1-CD150+CD41+GFP+ or Tom+ cells, or  Old Lin-cKit+Sca+-CD150+Flk2-Tom+ cells FACS sorted from the bone marrow of Young or Old FlkSwitch mice

first: D

last: Poscablo

first: E


title: In vitro Naïve B-cell Class Switching Stimulation with ATM-inhibitor and DNA-PKcs-inhibitor

city: Leiden

first: Hailiang

last: Mei

address: Einthovenweg 20

department: Sequencing Analysis Support Core

institute: Leiden University Medical Center

summary: Analyze Class switch recombination of naïve B-cells in vitro mimicking Ataxia Telangiectasia using a small molecule inhibitor (KU-55933). We measure switched IgG and IgG memory B-cells, and analyzed transcriptomics (this data).

overall_design: Healthy control PBMC's cultured and stimulated with medium containing IL10, IL21, anti-IgM and CD40L. Different conditions were compared: unstimulated, stimulated, stimulated with DNA-PK-inhibitor, and stimulated with ATM-inhibitor.

first: Sanami

middle: S

last: Takada

first: Tom

last: Kuipers

first: Thomas

last: Weitering


title: Mono and biallelic inactivation of Huntingtin gene in patient-specific iPS cells reveal HTT roles in striatal development and in neuronal functions impaired in Huntington’s disease

city: Corbeil-Essonnes

first: MARGOT

last: JARRIGE

address: 28, Rue Henri Desbruères

institute: I-Stem/INSERM U861

summary: Mutations in the Huntingtin (HTT) gene cause Huntington disease (HD), a neurodegenerative disorder. While HTT is known to be involved, as a scaffold protein, in several cellular functions, its normal and pathogenic functions during the development of the human forebrain remain poorly understood. To investigate the roles of wild-type and mutant HTT alleles during human neural and striatal development we inactivated HTT alleles in a series of isogenic clones of a human induced pluripotent stem cell (iPSC) line derived from a patient with HD. We show that the loss of wild-type, mutant, or both HTT isoforms did not affect the pluripotency of iPSCs or their transition into neural cells. However, we observed that HTT loss caused division impairments in forebrain neuro-epithelial cells. Moreover, HTT-/- derivatives displayed impaired maturation of medium spiny neurons (MSNs) particularly in the acquisition of DARPP32 expression, a key functional marker of MSNs. Finally, we discovered that MSNs and cortical neurons derived from HTT-/- clones exhibited higher levels of baseline DNA damage and lower BDNF axonal transport, two cellular dysfunctions previously described in HD neurons.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for Mono and biallelic inactivation of Huntingtin gene in patient-specific iPS cells

first: Morgane

last: Louessard

first: Michel

last: Cailleret

first: Margot

last: Jarrige

first: Julie

last: Bigarreau

first: Sophie

last: Lenoir

first: Frédéric

last: Saudou

first: Nicole

last: Deglon

first: Anselme

last: Perrier


title: Calcineurin-NFAT signaling controls the neutrophils´ ability of chemoattraction upon fungal infection.

city: Brno

first: Ioanna

last: Papatheodorou

address: Studentska

department: Center for Translational Medicine

country: Czech Republic

institute: International Clinical Research Center (ICRC), St.Anne’s Hospital

summary: Calcineurin-nuclear factor of activated T-cell (CN-NFAT) inhibitors are widely clinically used drugs for immunosuppression but besides their required T-cell response inhibition, they also undesirably affect innate immune cells. Disruption of innate immune cell function can explain the observed susceptibility of CN-NFAT inhibitors-treated patients to opportunistic fungal infections. Neutrophils play a crucial role in the innate immunity pathogen defense, while however the effect of CN-NFAT inhibitors on neutrophil function is poorly described. Thus, we tested the response of human neutrophils to fungal pathogens, namely Candida albicans and Aspergillus fumigatus in the presence of CN-NFAT inhibitors. We report that the NFAT pathway members are expressed in neutrophils and mediates part of the neutrophil response to pathogens. Upon pathogen exposure, neutrophils underwent profound transcriptomic changes. Importantly, genes involved in the regulation of the immune response and chemotaxis, including the chemokines CCL2 , CCL3, and CCL4 were significantly upregulated. The presence of CN-NFAT inhibitors attenuated the expression of these chemokines and impaired the ability of neutrophils to chemoattract other immune cells. Our results amend knowledge about the impact of CN-NFAT inhibition in human neutrophils.

overall_design:
In order to investigate the effect of CN-NFAT inhibitors on neutrophil function we treated neutrophils from healthy donors with fungal pathogens with or without prior pretreatment with CN-NFAT inhibitor Cyclosporine A.
Differential gene expression analysis from RNAseq data from neutrophils treated with CN-NFAT inhibitor Cyclosporine A and/or fungal pathogens.

first: Ondřej

last: Vymazal

first: Ivana

last: Andrejčinová

first: Veronika

last: Bosáková

first: Gianluca

last: Vascelli

first: Kamila

last: Bendíčková

first: Teresa

last: Zelante

first: Marcela

last: Hortová-Kohoutková

first: Jan

last: Frič


title: Revisiting the potency of Tbx2 expression in transforming outer hair cells into inner hair cells at multiple ages in vivo

first: Zhiyong

last: Liu

address: 7 Science Park Road ZGC Life Science Park

institute: National Institute of Biological Sciences, Beijing

summary: The mouse auditory organ cochlea contains two types of sound receptors: inner hair cells (IHCs) and outer hair cells (OHCs). Tbx2 is expressed in IHCs but repressed in OHCs, and neonatal OHCs that misexpress Tbx2 transdifferentiate into IHC-like cells. However, the extent of this switch from OHCs to IHC-like cells and the underlying molecular mechanism remain poorly understood. Furthermore, whether Tbx2 can transform fully mature adult OHCs into IHC-like cells is unknown. Here, our single-cell transcriptomic analysis revealed that in neonatal OHCs misexpressing Tbx2, 85.6% of IHC genes, including Slc17a8, are upregulated, but only 38.6% of OHC genes, including Ikzf2 and Slc26a5, are downregulated. This suggests that Tbx2 cannot fully reprogram neonatal OHCs into IHCs. Moreover, Tbx2 also failed to completely reprogram cochlear progenitors into IHCs. Lastly, restoring Ikzf2 expression alleviated the abnormalities detected in Tbx2+ OHCs, which supports the notion that Ikzf2 repression by Tbx2 contributes to the transdifferentiation of OHCs into IHC-like cells. Our study evaluates the effects of ectopic Tbx2 expression on OHC lineage development at distinct stages of either male or female mice and provides molecular insights into how Tbx2 disrupts the gene-expression profile of OHCs. This research also lays the groundwork for future studies on OHC regeneration.Significance Statement Elucidation of the molecular and genetic mechanisms governing the determination and stability of cochlear inner hair cells (IHCs) and outer hair cells (OHCs) should provide valuable insights into the regeneration of damaged IHCs and OHCs. Here, we conditionally overexpress Tbx2 in vivo in cochlear sensory progenitors, neonatal OHCs, or adult OHCs. Our results show that Tbx2 overexpression alone can partially destabilize the OHC fate but cannot fully convert OHCs into IHCs. Specifically, we demonstrate that Ikzf2 repression due to Tbx2 overexpression is one of the key pathways disrupting the OHC fate.

overall_design: The smart-seq based single-cell RNA-seq approach was applied to wild type cochlear outer hair cells (OHCs) at P30 as well as the converted IHCs (cIHCs) at P42. The cIHCs are the cochlear OHCs expressing ectopic Tbx2 and transdifferentiating into IHCs. The wild type OHCs are obtained from the Slc26a5-CreER/+; Ai9/+ mice at P30, whereas the cIHCs are from theSlc26a5-CreER/+; Rosa26Tbx2/+ mice at P42. Briefly, both wild type OHCs and cIHCs express Tdtomato and are manually picked under the fluorescent microscope. In this study, 32 wild type OHCs at P30 were de-novo picked. Together with the 17 OHCs at P30 reported in our previous study, we totally have 49 OHCs. All the 67 cIHCs were  de-novo picked in this study.

first: Zhenghong

last: Bi

first: Minhui

last: Ren

first: Yu

last: Zhang

first: Shunji

last: He

first: Lei

last: Song

first: Xiang

last: Li