title: Screening for genes that promote copper toxicity in a cell model of Wilson disease [HepG2 cells ATP7B-KO]

city: Pozzuoli

first: Diego

last: Carrella

state: NAPOLI

address: Via Campi Flegrei 34

country: Italy

institute: TIGEM

summary: To find new genes that might be targeted for reduction of Copper (Cu) toxicity in Wilson disease we employed genome-wide shRNA screening in ATP7B-KO HepG2 cells, a bona fide WD cell model. Cells were screened using the Dharmacon™ Decode™ Pooled Lentiviral shRNA Screening Library targeting 18205 human genes (Dharmacon #RHS6083). The Lentiviral RNAi screening library is made of 10 pools of 9570 GIPZ short hairpin RNAs (shRNAs) packaged into high-titer lentiviral particles. Genomic DNA was isolated from the cells and read counts for each shRNAs were obtained by deep sequencing to determine the abundance of each shRNA in the Cu-treated cells compared to controls. This analysis allowed the identification of genes whose downregulation promoted survival of ATP7B-KO cells upon Cu overload. To exclude that some shRNAs would have been responsible for ATP7B-KO survival by suppressing generic proapoptotic genes, we run the same screening with the proapoptotic drug vinblastine instead of CuCl2. Genes, whose suppression allowed cell to resist Cu but not vinblastine were considered as Cu-specific hits and through this approach we identified PrP as a suitable target for reduction of Cu toxicity in WD. Indeed, we found that Cu accumulation in ATP7B-KO cells stimulates expression of PrP, which brings to  toxic Cu overload while PrP suppression significantly reduces Cu accumulation and toxicity in ATP7B-deficient cells. In sum our data highlight an important regulatory role of PrP in copper metabolism and as a therapeutic tool for Wilson disease.

overall_design: HepG2 cells ATP7B-KO, available in our lab, were infected with the Dharmacon™ Decode™ Pooled Lentiviral shRNA library composed of 10 pools. Each individual pool was cultured and left untreated or treated with 0.5mM CuCl2 and divided into three biological replicate.

first: Raffaella

last: Petruzzelli

first: Federico

last: Catalano


title: Screening for genes that promote copper toxicity in a cell model of Wilson disease

status: Public on Dec 09 2024

type: Expression profiling by high throughput sequencing

overall_design: HepG2 cells ATP7B-KO, available in our lab, were infected with the Dharmacon™ Decode™ Pooled Lentiviral shRNA library composed of 10 pools. Each individual pool was cultured and left untreated or treated with 60uM vinblastine and divided into three biological replicate.

first: D

first: R

first: F


title: Spatial Transcriptomics of Brains Infected with VEEV

city: Livermore

first: Aimy

last: Sebastian

state: CA

address: 7000 East Ave

department: BBTD

country: USA

institute: LLNL

summary: Venezuelan equine encephalitis virus (VEEV) is a neurotropic alphavirus that causes neurological disease in both humans and equine. In this study we paired spatial transcriptomics and a mouse model of VEEV to generate a transcriptomic profile of infected or control brains at 6 days post infection. We identified regionally distinct myeloid and lymphocyte populations, contributing to our understanding of the neuroinflammation caused by VEEV.

overall_design: Spatial transcriptomics was performed on sections of brain prepared from either C3H/HeN mice infected intranasally with VEEV TC-83 at 6 days post-infection or healthy controls.

first: Margarita

middle: V

last: Rangel

first: Dina

middle: R

last: Weilhammer


title: Neurodevelopmental and behavioral defects in congenital heart disease [ChIP-seq]

city: Pittsburgh

first: Hisato

last: Yagi

state: PA

address: 530 45th street, Rangos Res Center 8162

institute: University of Pittsburgh

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Hypoplastic left heart syndrome (HLHS) is a severe congenital heart disease associated with microcephaly and poor neurodevelopmental outcomes. Here we show that the Ohia HLHS mouse model, with mutations in Sap130, a chromatin modifier, and Pcdha9, a cell adhesion protein, also exhibits microcephaly associated with mitotic block and increased apoptosis leading to impaired cortical neurogenesis. Transcriptome profiling, DNA methylation, and Sap130 ChIPseq analyses all demonstrated dysregulation of genes associated with neurogenesis, cognitive impairment and autism. This involved perturbation of REST transcriptional regulation of neurogenesis, disruption of CREB signaling regulating synaptic plasticity, and defects in neurovascular coupling mediating cerebral blood flow. Adult mice harboring either the Pcdha9 mutation, which showed normal brain anatomy, or forebrain-specific Sap130 deletion via Emx1-Cre, which showed microcephaly, both demonstrated learning and memory deficits and autism-like behavior. These novel findings provide mechanistic insights indicating the adverse neurodevelopment in HLHS may involve cell autonomous/nonautonomous defects and epigenetic dysregulation.

overall_design: Chromatin immunoprecipitation of Sap130 was performed with rabbit anti-Sap130 antibody (A302-491A: Bethyl laboratories, 2ug)

first: Lo

last: Cecilia

first: Gabriel

last: George

first: Yagi

last: Hisato

first: Bais

last: Abha


title: Single-cell transcriptomic profiling of brain immune infiltration following Venezuelan equine encephalitis virus infection

summary: Neurotropic alphaviruses such as Venezuelan equine encephalitis virus (VEEV) are critical human pathogens that continually expand to naïve populations and for which there are no licensed vaccines or therapeutics. The neuropathology of VEEV has been attributed to the immune response in the brain yet the underlying mechanisms and specific immune cell populations involved are not fully elucidated. In this study, a murine model of lethal VEEV infection is paired with single-cell RNA sequencing to transcriptionally profile the immune response longitudinally in the brain following infection. Infection-induced immune changes in the brain was also compared to changes in peripheral blood mononuclear cells (PBMCs). The results define an inflammatory response involving transcriptionally distinct subpopulations of activated microglia and infiltrating proinflammatory myeloid populations and cytotoxic lymphocytes. This study advances our understanding of the immune mechanisms underlying viral encephalitis toward the goal of identifying new therapeutic targets.

overall_design: Single-cell RNA sequencing was performed on  immune cells isolated from the brains and blood of C3H/HeN mice infected intranasally with VEEV TC-83 at 2, 4, and 6 days post infection or from brains of uninfected control mice.


title: Proteomic study identifies Aurora-A mediated regulation of alternative splicing through multiple splicing factors

status: Public on Dec 10 2024

city: Frederick

first: Thomas

last: Gonatopoulos-Pournatzis

state: Maryland

address: 1050 Boyles Street

institute: National Cancer Institute

summary: The cell cycle regulator Aurora-A kinase presents an attractive target for cancer therapies, though its inhibition is also associated with toxic side effects. To gain a more nuanced understanding of Aurora-A function, we applied shotgun proteomics to identify 407 specific protein partners, including several splicing factors. Supporting a role in alternative splicing, we found that Aurora-A localizes to nuclear speckles, the storehouse of splicing proteins. Aurora-A interacts with and phosphorylates splicing factors both in vitro and in vivo, suggesting that it regulates alternative splicing by modulating the activity of these splicing factors. Consistently, Aurora-A inhibition significantly impacts the alternative splicing of 505 genes, with RNA motif analysis revealing an enrichment for Aurora-A interacting splicing factors. Additionally, we observed a significant positive correlation between the splicing events regulated by Aurora-A and those modulated by its interacting splicing factors. An interesting example is represented by CLK1 exon 4, which appears to be regulated by Aurora-A through SRSF3. Collectively, our findings highlight a broad role of Aurora-A in the regulation of alternative splicing.

overall_design: RNA sequencing was performed in HEK293T cells either with Aurora-A inhibition (24 hours) or depleted of endogenous splicing factors (48 hours) using RNAi (non-targeting siRNAs were used as control) to explore a functional link between Aurora-A and key splicing regulators.

first: Arun

middle: P

last: Damodaran

first: Olivia

last: Gavard

first: Jean-Philippe

last: Gagné

first: Malgorzata

middle: E

last: Rogalska

first: Amit

middle: K

last: Behera

first: Estefania

last: Mancini

first: Giulia

last: Bertolin

first: Thibault

last: Courtheoux

first: Bandana

last: Kumari

first: Justine

last: Cailloce

first: Agnès

last: Mereau

first: Guy

middle: G

last: Poirier

first: Juan

last: Valcárcel

first: Erwan

last: Watrin

first: Claude

last: Prigent


title: A 5-transcript signature for discriminating viral and bacterial etiology in pediatric pneumonia

city: Santiago de Compostela

first: Alberto

last: Gómez-Carballa

state: A coruña

address: Choupana sn

department: Pediatrics

country: Spain

institute: GENVIP research group

summary: Pneumonia stands as the primary cause of death among children under five, with its onset attributed to a broad spectrum of microorganisms. Diagnosis poses an ongoing challenge, relying on clinical or microbiological criteria, often resulting in delayed and inaccurate treatment and unnecessary therapy. Our research focuses on identifying host transcriptomic biomarkers in the blood of children affected by viral and bacterial pneumonia, alongside healthy controls. The main goal is to establish a gene-expression signature enhancing disease diagnosis and management. We conducted an analysis of a total of 192 whole blood samples, comprising 38 controls and 154 viral and bacterial pneumonia patients recruited through the EUCLIDS clinical network. Our investigation identified 5,486 differentially expressed genes (DEGs) when comparing blood RNA from pneumonia patients with healthy controls. Functional enrichment analysis highlighted pathways related to the immune system response, encompassing neutrophil degranulation, humoral immune response, and various inflammatory pathways. In the comparative analysis of gene expression between viral and bacterial pneumonia patients, we identified 272 DEGs. Gene set analysis revealed a significant difference in pathway enrichment for the immune response, contingent on the over-regulation of gene sets in viral or bacterial pneumonias. Furthermore, we identified a 5-transcript host signature specifically designed to distinguish between viral and bacterial pediatric pneumonia (FAM20A, BAG3, TDRD9, MXRA7 and KLF14; AUC: 0.95 [0.88–1.00]), pseudo-validated in a cohort including probable bacterial and viral patients (AUC: 0.87 [0.77–0.97]). This signature holds the potential to enhance the accuracy of previously described general transcript-based signatures for viral and bacterial infections.

overall_design: Children with pneumonia (n = 154) and age and sex matched healthy controls (n = 38) were selected from the EUCLIDS database for transcriptome analysis in whole blood samples collected in PAXgeneTM and Tempus™ RNA tubes. In a subsequent analysis, pneumonia samples were further classified using the algorithm developed by Herberg et al. C-reactive protein (CRP) and neutrophil threshold criteria were employed where available in definitive bacterial (DB) and definitive viral (DV), respectively (see 30 for details on clinical characteristics of the cohort and definitions on DB and DV).Thus, DB (n = 40) and DV (n = 9) pneumonias were selected to study the differences in transcriptome depending on the etiology of the infection

first: Sandra

last: Viz-Lasheras

first: Dominic

last: Habgood-Coote

first: Myrsini

last: Kaforou

first: Antonio

last: Salas


title: Bulk RNA-seq analysis of ATF3 kncokdown on HRMECs

city: Suita

first: Chihiro

last: Ueda

address: l2-2, Yamadaoka

department: Ophthalmology

country: Japan

institute: Osaka University

summary: To investigate the role of ATF3 in vascular endothelial cells, gene expression profiling analysis was performed using RNA-seq data from human retinal microvascular endothelial cells (HRMECs) transfected with ATF3 siRNA.

overall_design: HRMECs were treated with negative control or ATF3 siRNA for 24 hours and comparative gene expression profiling analysis was performed using RNA-seq data.

first: Susumu

last: Sakimoto

first: Masahito

last: Yoshihara

first: Kohji

last: Nishida


title: Title: Neurodevelopmental and behavioral defects in congenital heart disease

first: Hisato

last: Yagi

overall_design: Expression profiling using RNA-seq of 8 samples collected from brain of HLHS mice (n=3 at E13.5-E14.5) and their littermate controls (n=5 at E13.5-E14.5).

first: Lo

last: Cecilia

first: Gabriel

last: George

first: Yagi

last: Hisato

first: Bais

last: Abha


title: A high-fidelity CRISPR-Cas13 system improves abnormalities associated with C9ORF72-linked ALS/FTD

status: Public on Dec 08 2024

city: Urbana

first: Sijia

last: Zhang

state: IL

address: 1406 W Green St

department: Bioengineering

institute: Gaj Lab

summary: An abnormal expansion of a GGGGCC hexanucleotide repeat in the C9ORF72 gene is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two debilitating neurodegenerative disorders driven in part by gain-of-function mechanisms involving transcribed forms of the repeat expansion. By utilizing a Cas13 variant with reduced collateral effects, we developed a high-fidelity RNA-targeting CRISPR-based system for C9ORF72-linked ALS/FTD. When delivered to the brain of a transgenic rodent model, this Cas13-based platform effectively curbed the expression of the GGGGCC repeat-containing RNA without affecting normal C9ORF72 levels, which in turn decreased the formation of RNA foci and reversed transcriptional deficits. This high-fidelity Cas13 variant possessed improved transcriptome-wide specificity compared to its native form and mediated efficient targeting in motor neuron-like cells derived from a patient with ALS. Our results lay the foundation for the implementation of RNA-targeting CRISPR technologies for C9ORF72-linked ALS/FTD.

overall_design: Analyzed the differentially expressed genes for RfxCas13d and a high-fidelity RfxCas13d variant when programmed to target human C9ORF72. Experiments were conducted in induced motor neurons and in triplicates or more. Each replicate is an independent biological replicate.

first: Tristan

middle: X

last: McCallister

first: Colin

last: Lim

first: William

middle: M

last: Terpstra

first: Maria

middle: A

last: Zeballos C.

first: Jackson

last: Powell

first: Thomas

last: Gaj


title: Follistatin secreted by myocytes from hiPSCs results in cardiomyocyte cytokinesis in large mammal hearts with postinfarction LV remodeling

city: Birmingham

first: Thanh

middle: Minh

last: Nguyen

state: AL

address: 1670 University Blvd, Volker Hall G094

department: Department of Biomedical Engineering

institute: University of Alabama at Birmingham

summary: In this work, we generated a new human induced-pluripotent stem cells derived-cardiomyocytes (hiPSC-CM); in this new hiPSC-CM cell line, Cyclin D2 (CCND2), a cell-cycle activator, was overexpressed, and class I-II of major histocompatibility complex were knocked down. This new cell line (named KO/OEhiPSC-CM) and the wildtype hiPSC-CM (name WThiPSC-CM) were transplanted into pig hearts with ischemic-reperfusion (I/R). The objective is to examine engraftment, cardiac function change, and how the host (pig) heart responded to the transplantation. Surprisingly, while the pig hearts receiving KO/OEhiPSC-CM showed significant cardiac function 4 weeks after I/R, the KO/OEhiPSC-CM grafts were undetectable. Analyzing the single-nuclei RNA sequencing (SnRNAseq)  data of these pig hearts showed that 1) the host cardiomyocyte actively reentered cell-cycle, which was among the key factors for cardiac function improvement; 2) this effect was mediated via HIPPO/YAP signaling pathway; and 3) HIPPO/YAP signaling pathway activity was a result of Follistatin (FST) secreted by the transplanted KO/OEhiPSC-CM.

overall_design: KO/OEhiPSC-CM and WThiPSC-CM spheroids were differentiated in shaking flasks, purified, characterized, and intramyocardially injected into pigs after I/R; control animals were injected with basal medium. One and for week after I/R, SnRNAseq data from 3 pig groups (receiving KO/OEhiPSC-CM, WThiPSC-CM, and control-basal medium) at the I/R border zone were collected using the 10XGenomics workflow.

first: Yuhua

last: Wei

first: Lei

last: Ye

first: Gregory

last: Walcott

first: Jianyi


title: N6-methyladenosine regulates metabolic remodeling in kidney aging through transcription regulator GLIS1

city: shenyang

first: xiuli

last: Guo

address: No. 155 Nanjing North Street, Heping District, Shenyang, Liaoning Province

country: China

institute: The First Hospital of China Medical University

type:
Expression profiling by array
Other

summary: Aged-related kidney impairment that involves tubular epithelial cell senescence and renal fibrosis become a severe threaten for worldwide public health. m6A-mRNA epitranscriptomic microarray was performed to reveal the targeted genes with abnormal m6A modification in aged human kidney tissues.

overall_design: Control group and experimental group,Biological replicates: 3control replicates, 3 experimental replicates.

first: Li

last: Xu

first: Hongyuan

last: Yu

first: Xinwang

last: Zhu

first: Yonghong

first: Hang

last: Mei

first: Ping

last: He

first: Min

last: Wang

first: Yue

last: Liu

first: Fanming

last: Jiang

first: Yu

last: Sun


title: NEDD4L mediates intestinal epithelial cell ferroptosis to restrict inflammatory bowel diseases [colonic tissues without isolation]

city: Hangzhou

first: Wenlong

last: Lin

state: Zhejiang

address: 66 Yuhang Tang Road

institute: Zhejiang University

summary: To explore the underlying mechanisms of NEDD4L in regulating colitis, colonic tissues from DSS-treated Nedd4lf/f VillinCre (IEC KO)  mice and Nedd4lf/f (WT) littermates were subjected to RNA-sequencing analysis.

overall_design: Gene expression profiling analysis of RNA-seq data from colonic tissues on the 7th day from the Nedd4lf/fVillinCre and Nedd4lf/f mice administered 2 % DSS .

first: Jingjing

last: Liang


title: GILS1 alleviates cell senescence and renal fibrosis through PGC-1α-mediated mitochondrial quality control in kidney aging

summary: Mitochondria take part in a network of intracellular processes that regulate homeostasis. Mitochondrial dysfunction is implied as a crucial factor in age-related chronic kidney disease. Gli-like transcription factor 1 (GLIS1) is enriched in physiology aging kidneys and decreased in aged human kidney that was confirmed in our previous research, however, the correlation between mitochondrial disturbances and GLIS1-driven kidney aging are not clearly clarified. Thus, we investigated the role of GLIS1 in the homeostasis of mitochondrial quality control both in vivo and in vitro. The lower expression of GLIS1 was identified in natural and accelerated aged models, accompanied by the mitochondrial quality control (MQC) dysfunctions, including enhanced mitochondrial fission, reduced mitochondrial biogenesis and mitophagy. After the induction of kidney aging, adeno-associated virus carrying GLIS1 over-expression or vector were delivered to the mice.

overall_design:
8-week-old C57BL/6 mice were administered subcutaneous injections of D‐gal (500mg/kg/day) for 8 weeks, which is a well-established aging model both in vitro and in vivo
we then performed gene expression profiling analysis using data obtained from RNA-seq of 6 different tissue of kidney.

first: Huimeng

last: Qi

first: Shuo

last: Chen


title: Epigenetic modulation through BRD8 impairs conversion of primed EpiSCs to naïve ESCs [ATAC-Seq]

city: Shenzhen

first: Andrew

last: Hutchins

address: 1088 Xueyuan Rd

department: Bioinformatics

institute: Southern University of Science and Technology

summary: Epigenetic control of cell type is a critical determinant in cell type stability and permissive differentiation. However, the epigenetic control mechanisms are not well understood. Here, we explore the epigenetic control of the conversion of primed mouse EpiSCs to naïve mouse ESCs as a mimic for early stages of embryonic development. Whilst the conversion of naïve to primed state is relatively permissive and mimics a normal developmental program, the reverse process, primed to naïve is inefficient in the absence of transgenes, which suggests the presence of potent epigenetic barriers that resist this artificial dedifferentiation. Here, we identify the histone acetylation reader BRD8 as a one-way barrier that blocks the conversion of primed to naïve cells. BRD8 works by maintaining histone acetylation on promoters and transcribed gene bodies and it’s absence promotes the formation of heterochromatin.

overall_design: RNA-seq of the EpiSCs undergoing a primed to naïve conversion at the indicated days. The EpiSCs were transfected with shRNAs targetting Brd8 or Igf2bp2.

first: Xiuling

last: Fu


title: the effect of knockout Brd7 on 4T07-TGL cells under in vitro condition [scRNA-Seq]

status: Public on Dec 06 2024

city: Houston

first: Junfeng

last: Zhang

state: TX

address: 2130 W. Holcombe Blvd

department: Department of Cancer Biology

institute: The University of Texas MD Anderson Cancer Center

summary: Metastasis is a major cause of mortality in cancer and is likely influenced by epigenetic and gene regulatory factors. Using an in vivo screen, we demonstrated that several subunits of the polybromo-associated BAF (PBAF) complex, particularly BRD7, were required for breast cancer metastatic dormancy in lung. BRD7 loss induced metastatic outgrowth, along with modifications in epigenomic landscapes and upregulated oncogenic signaling. Breast cancer cells harboring BRD7 loss also reprogrammed the surrounding immune microenvironment, downregulating MHC-1 expression and promoting a pro-metastatic cytokine profile. Flow cytometric and single-cell analyses revealed increased levels of pro-tumorigenic neutrophils, CD8+ exhausted T cells, and CD4+ stress response T cells in lungs harboring BRD7-deficient metastases. Finally, attenuating this immunosuppressive milieu by neutrophil depletion, neutrophil extracellular trap (NET) inhibition, or immune checkpoint therapy abrogated metastatic outgrowth. These findings, for the first time, implicate BRD7 and PBAF in metastasis, pointing to targetable underlying mechanisms involving specific immune cell compartments.

overall_design: To interrogate the effects of BRD7 loss on specific immune microenvironmental constituents,we performed single cell RNAseq on Cd45+ cells sorted from dissociated lungs removed from naïve 6 week-old mice or similarly aged littermates 14 days after injection with either control or BRD7-KO 4T07 cells.


title: TGFBI is upregulated in ectopic endometrial stromal cells and under hypoxic conditions in endometriosis.

status: Public on Dec 07 2024

city: tartu

first: Maire

last: Peters

department: Dept. of Obstetrics and gynecology

country: Estonia

institute: University of Tartu

summary: The objective of the study was to compare the transcriptome of the primary eutopic and ectopic endometrial stromal cells isolated from paired eutopic endometrium and peritoneal lesions from women with endometriosis (N = 5) exposed to normoxic (20% O2) or hypoxic (1% O2) conditions for 48 hours. Induced expression of TGFBI was found in ectopic cells and ectopic & eutopic cells exposed to hypoxia at both RNA and protein (secreted to the culture media) levels. TGFBI gene and protein expression ex vivo in euoptic endometrium of women with endometriosis and controls (women without endometriosis) was found to be increased in the proliferative phase of menstrual cycle compared to secretory phase.

overall_design: Compare the expression of mRNAs in lysates of differently treated eutopic and ectopic endometrial stromal cells

first: Meruert

last: Sarsenova

first: Nageswara

last: Boggavarapu

first: Keiu

last: Kask

first: Vijayachitra

last: Modhukur

first: Külli

last: Samuel

first: Pille

last: Soplepmann

first: Kristina

last: Gemzell-Danielsson

first: Parameswaran

middle: L

last: Lalitkumar

first: Andres

last: Salumets

first: Darja

last: Lavogina


title: the effect of knockout Brd7 on 4T07-TGL cells under in vitro condition

overall_design: To study the function of PBAF subunit Brd7 in Breast cancer metastatic dormancy, we established Brd7 knockout 4T07-TGL cell lines by CRISPR/Cas9.  To ascertain BRD7-dependent enhancer profiles in our 4T07 isogenics, we performed ChIP-seq for H3K27ac, a canonical enhancer mark, in our KO (Brd7-KO1, Brd7-KO2) and control (4T07-TGL-sgCtr) lines.


title: Three-dimensional chromatin reorganization during CD8+ T cell activation is mediated by TRIM28 [CUT&Tag]

city: Beijing

first: Ruifeng

last: Li

address: haidian

institute: Tsinghua University

summary: T cell activation is accompanied by extensive changes in epigenome. However, whether and how high-ordered chromatin organization is involved in CD8+ T cell activation is unclear. Here, we showed extensive changes in the three-dimensional genome during CD8+ T cell activation, associated with changes in gene transcription. Moreover, we identified Tripartite motif containing 28 (TRIM28) as an essential mediator in re-organization of spatial chromosomal interactions. Trim28 deficiency impaired CD8+ T cell activation in vitro and in vivo, due to its regulation on autocrine IL-2 production. Mechanistically, TRIM28 bound to genes associated with CD8+ T cell activation and promoted the formation of chromosomal loops. At the loop anchor regions, TRIM28 is likely recruited by CTCF, and required for binding of RNA Pol II and cohesin to form active chromosomal structure. These results thus identify a critical role of TRIM28-dependent chromatin topology in gene transcription during CD8+ T cell activation.

overall_design: CUT&Tag for SMC1 of Trim28fl/fl vs Trim28-/- CD8+ T cells activated in vitro.

first: Kun

last: Wei

first: Chen

last: Dong


title: Glia detect and transiently protect against dendrite substructure disruption

city: New York

first: Katherine

middle: Celina

last: Varandas

state: New York

address: 1230 York Ave

institute: The Rockefeller University

summary: Glia assess axon structure to modulate myelination and axon repair. Whether glia similarly detect dendrites and their substructures is not well understood. Here, we show that glia monitor the integrity of dendrite substructures and transiently protect their perturbation. We demonstrate that disruption of C. elegans sensory neuron dendrite cilia elicits acute glial responses, including increased accumulation of glia-derived extracellular matrix around cilia, changes in gene expression, and alteration of secreted protein repertoire. DGS-1, a 7-transmembrane domain neuronal protein, and FIG-1, a multifunctional thrombospondin-domain glial protein, are required for glial detection of cilia integrity, physically interact, and exhibit mutually-dependent localization to and around cilia, respectively. Glial responses to dendrite cilia disruption transiently protect against damage. Thus, our studies uncover a homeostatic, protective, dendrite glia signaling interaction regulating dendrite substructure integrity.

overall_design: To investigate the AMsh glia responses to loss of associated dendritic cilia, we profiled AMsh glia from F16F9.3pro::dsRed animals (dsRed+) and all other cells (dsRed-) in wild type and cilia mutant [che-2(e1033)] animals.

middle: C

first: Shai

last: Shaham


title: NEDD4L mediates intestinal epithelial cell ferroptosis to restrict inflammatory bowel diseases and colorectal tumorigenesis

first: Wenlong

last: Lin

first: Jingjing

last: Liang


title: Epigenetic modulation through BRD8 impairs conversion of primed EpiSCs to naïve ESCs [RNA-Seq]

first: Andrew

last: Hutchins

first: Li

last: Sun

first: Xiuling

last: Fu


title: Single-cell RNA-seq of peripheral blood links cell-type-specific regulation of splicing to autoimmune and inflammatory diseases [scRNAseq_JP_RIK]

status: Public on Dec 05 2024

city: Singapore

first: Boxiang

last: Liu

address: 12 Science Drive 2, MD1 - Tahir Foundation Building, Level 5

country: Singapore

institute: National University of Singapore

summary: Alternative splicing is pivotal in the genetics of complex traits, but a detailed understanding requires relevant cell types from diverse genetic ancestries. Here, we described cell-type-specific, sex-biased, and ancestry-biased alternative splicing in ~1M peripheral blood mononuclear cells from 474 healthy donors from the Asian Immune Diversity Atlas. We identified widespread sex- and ancestry-biased differentially splicing, most of which are cell-type-specific. We identified 11,577 independent cis-sQTLs, 607 trans-sQTLs, and 107 dynamic sQTLs. Colocalization between cis-eQTL and trans-sQTL revealed a cell-type-specific regulatory relationship between hnRNPLL and PTPRC. We observed a strong enrichment of cis-sQTL effects in autoimmune and inflammatory disease heritability. Specifically, we functionally validated an Asian-specific sQTL disrupting the 5’ splice site of TCHP exon four to putatively modulate the risk of Graves’ disease in East Asian populations. Our work highlights the critical impact of ancestral diversity and provides a roadmap to dissect splicing mechanisms in complex diseases at single-cell resolution.

overall_design: The raw data contains PBMCs from healthy Asian donors and Caucasians. Asian donors are from Singaporean Chinese, Singaporean Indian, Singaporean Malay, Japanese, and Koreans. Caucasian samples are used as control samples for each batch. Raw genotype and phenotype data associated with this study are available in dbGaP under accession phs003848.v1.p1. Processed expression data is provided in GEO.

first: Chi

last: Tian

first: Yuntian

last: Zhang

first: Yihan

last: Tong

first: Kian Hong

last: Kock

first: Kyung Yeon

last: Han

first: Yoshinari

last: Ando

first: Woong-Yang

last: Park

first: Jay W.

last: Shin

first: Shyam

last: Prabhakar


title: The effect of knockout Brd7 on 4T07-TGL cells under in vitro condition

first: Junfeng

overall_design: BRD7 and the PBAF complex had been widely implicated in regulating chromatin accessibility. To study the effect of brd7 loss on chromatin accessibility, we performed ATAC-seq on  BRD7-knockdown 4T07 lines.


title: NKp46 regulates type 1 innate lymphoid cell proliferation and function and anti-acute myeloid leukemia activity

city: Duarte

first: Zhenlong

last: Li

address: 1500 East Duarte Road

institute: City of hope

summary: NKp46 is a critical regulator of natural killer (NK) cell immunity, but its function in non-NK innate immune cells remains unclear. Here, we show that NKp46 is indispensable for expressing IL-2 receptor-α (IL-2Rα) by non-NK liver-resident type-1 innate lymphoid cells (ILC1s). Deletion of NKp46 reduces IL-2Rα on ILC1s by downregulating NF-κB signaling, thus impairing ILC1 proliferation and cytotoxicity in vitro and in vivo. Likewise, the binding of anti-NKp46 antibody to NKp46 triggers the activation of NF-κB, the expression of IL-2Rα, interferon- γ (IFN-γ), tumor necrosis factor (TNF), proliferation, and cytotoxicity. Functionally, NKp46 expressed on mouse ILC1s interacts with tumor cells through cell–cell contact, increasing ILC1 production of IFN-γ and TNF, and enhancing cytotoxicity. In a mouse model of acute myeloid leukemia (AML), deletion of NKp46 impairs the ability of ILC1s to control tumor growth and reduces survival. This can be reversed by injecting NKp46+ ILC1s into NKp46 knock-out mice. Human NKp46+ ILC1s exhibit stronger cytokine production and cytotoxicity than their NKp46 counterparts, suggesting that NKp46 plays a similar role in humans. These findings identify an NKp46–NF-κB–IL-2Rα axis and suggest that activating NKp46 with an anti-NKp46 antibody, may provide a potential strategy for anti-tumor innate immunity.

overall_design: Comparison of the differences between Ncr1⁺/⁺ ILC1s and Ncr1gfp/gfp ILC1s

first: Jianhua

last: Yu


title: Loss of Hoxa5 function affects Hox gene expression in different biological contexts

city: Québec

first: Lucie

last: Jeannotte

state: QC

address: 2260, boulevard Henri Bourassa

department: Biologie moléculaire, Biochimie médicale & Pathologie

country: Canada

institute: Université Laval

summary: Hoxa5 plays numerous roles in development, but its downstream molecular effects are mostly unknown. We applied bulk RNA_seq assays to characterize the transcriptional impact of the loss of Hoxa5 gene function in seven different biological contexts, including developing respiratory and musculoskeletal tissues that present phenotypes in Hoxa5 mouse mutants. This global analysis revealed few shared transcriptional changes, suggesting that HOXA5 acts mainly via the regulation of context_specific effectors. However, Hox genes themselves appeared as potentially conserved targets of HOXA5 across tissues. Notably, a trend toward reduced expression of HoxA genes was observed in Hoxa5 null mutants in several tissue contexts. Comparative analysis of epigenetic marks along the HoxA cluster in lung tissue from two different Hoxa5 mutant mouse lines revealed limited effect of either mutation indicating that Hoxa5 gene targeting did not significantly perturb the chromatin landscape of the surrounding HoxA cluster. Combined with the shared impact of the two Hoxa5 mutant alleles on phenotypes and Hox expression, these data argue against the contribution of local cis effects to Hoxa5 mutant phenotypes and support the notion that the HOXA5 protein acts in trans in the control of Hox gene expression.

overall_design: To explore the global transciptional  changes resulting from the loss of Hoxa5 gene function across space and time, we applied bulk RNA sequencing (RNA-seq) to tissues from the developing respiratory and musculoskeletal systems that present severe phenotypes in Hoxa5 mutant mice (Hoxa5tm1Rob, Jeannotte et al., 1993). For trachea (E15.5), lungs (E12.5 and E15.5), diaphragm (E15.5) and somites (E12.5), four biological replicates were used for each genotype. For somites (E10.5), four wild-type (wt) and three mutants were used while for interscapular brown adipose tissue (iBAT; E18.5), five samples were used for each genotype.

first: Béatrice

last: Frenette

first: Josselin

last: Gueno

first: Nicolas

last: Houde

first: Kim

last: Truchon

first: Jennifer

middle: H

last: Mansfield


title: USP5 stabilizes YTHDF1 to control cancer immune surveillance through mTORC1-mediated phosphorylation

city: Chongqing

first: Tianshu

last: Fang

state: China

address: 临江路74号（重医附二院渝中院区）

institute: 重医附二院渝中院区第二临床学院

summary: N6-methyladenosine (m6A) binding protein YTHDF1 is frequently upregulated in various cancers and its depletion enhances the efficacy of immune checkpoint blockade (ICB) therapy. This study reveals that USP5 interacts with YTHDF1, preventing its K11-linked polyubiquitination, thereby stabilizing YTHDF1 and promoting its oncogenic properties. In response to insulin, mTORC1 phosphorylates USP5, facilitating its dimerization and subsequent binding to YTHDF1, while the CUL7-FBXW8 complex promotes its degradation. Notably, YTHDF1 or USP5 deficiency increases PD-L1 expression and impairs immune response gene expression, contributing to immune evasion. Combining USP5 inhibitors with anti-PD-1 therapy enhances antitumor T-cell immunity and improves tumor regression in mouse models. Thus, USP5 may serve as a biomarker for stratifying patients for anti-PD-1 therapy, suggesting a novel strategy of combining USP5 inhibition with PD-(L)1 blockade to enhance cancer treatment efficacy.

overall_design: RNA immunoprecipitation sequencing (RIP-seq) for Ythdf1 in Hepa 1-6 Wid type and Usp5 Knockdown  cell lines.

last: Shao

first: Lei

last: Xi

first: Yangfan

last: Lv

first: Muhammad

last: Idris

first: Lin

first: Ya

last: Cao

first: Jingyi

last: Xiang

first: Xi

last: Xu

first: Belinda

last: Ong

first: Qiongyi

first: Xu

last: Peng

first: Xiaoyan

last: Yue

first: Feng

first: Chungang

last: Liu


title: Effect of 4.25% PD fluid treatment on the peritoneum of MR1+/+ C57BL/6J and MR1-/- C57BL/6J mice

first: Yuxiang

last: Sun

state: Texas

address: One Baylor Plaza, Houston, TX, 77030, United States

institute: Baylor College of Medicine

summary: Mr1 knockout (Mr1−/−) mice, in which thymic cells lose the ability to promote the development of mature Mucosal-associated invariant T (MAIT) cells, induced systemic MAIT cell depletion. MAIT cells often exhibit altered functions during fibrosis and inflammatory diseases. Here we used intraperitoneal injections of 4.25% PD fluid (0.1 mL/g) daily for six weeks to mimic peritoneal dialysis (PD) related fibrosis in mice, and investigated whether Mr1 knockout could ameliorated PD-induced changes in peritoneal transcriptome. We analyzed bulk RNA-seq of peritoneums from Mr1+/+ + PBS group (n=3), Mr1+/+ + PD group (n=3), Mr1-/- + PBS group (n=2), and Mr1-/- + PD group (n=2), and demonstrated that Mr1 knockout could ameliorated PD-induced activation in fibrosis, hyperglycolysis, and mTORC1 signalings. In addition, the activation scores of these pathways are positively correlated with each other.

overall_design: A cohort of Mr1+/+ and Mr1-/- C57BL/6 male mice received intraperitoneal injection of PBS or 4.25% PD fluid of the same volume for 6 weeks. After modeling, mice were euthanized, and peritoneums were collected for sqeuencing.

first: Peng

last: Hui

first: Hu

last: Zhaoyong


title: Human macrophage response to the emerging enteric pathogen Aeromonas veronii: inflammation, apoptosis, and downregulation of histones

city: Sydney

first: Li

last: Zhang

state: NSW

address: Botany Street

department: School of Biotechnology and Biomolecular Sciences

country: Australia

institute: University of New South Wales

summary: The macrophage cell (THP-1 derived macrophages) gene responses to Aeromonas veronii and Escherichia coli were investigated by comparative differential expression analysis

overall_design: Comparative gene expression profiling analysis of RNA-seq data for THP-1 derived macrophages treated with Aeromonas veronii and Escherichia coli and without treatment was performed


title: Hepatic transcriptome analysis of WT and PRDX1Cys52Ser mutant mice on CDAHFD

city: TIANJIN

first: Yin

last: Wen

state: TIANJIN

address: Address:No.6 North Huanrui Rd, Beichen District, Tianjin, P.R China

department: NHC Key Laboratory of Hormones and Development

institute: Chu Hsien-I Memorial Hospital & Tianjin Institute of Endocrinology, Tianjin Medical University

summary: In this study, we investigated how the peroxidase peroxiredoxin 1 protects against NASH. One previous study showed that PRDX1 cys52 was mutated to ser52 and consequently PRDX1’s peroxidase activity was impaired. Surprisingly, PRDX1Cys52Ser mice showed robust resistance to diet-induced NASH. To this end, We sought to understand how PRDX1Cys52Ser mutant confers resistance to NASH although impairing PRDX1’s peroxidase activity. We fed WT and PRDX1Cys52Ser mice a CDAHFD for 8 weeks and collected liver samples for RNA sequencing analysis. As revealed by gene-set enrichment analysis (GSEA) and KEGG pathway enrichment analysis, there was an enrichment of genes involved in inflammation, fiberosis and JAK-STAT signaling.

overall_design: We fed 8-week-old male mice a CDAHFD for 8 weeks and collected their liver samples for RNA sequencing. 3 mice per genotype (WT or PRDX1 Cys52Ser) were used.

first: Wen

last: Yin

first: Zhonghao

last: Bai


title: Chromatin conformation, gene transcription, and nucleosome remodeling as an emergent system

city: Evanston

first: vadim

last: backman

address: 2170 Campus Drive, Silverman Hall 3627

institute: Northwestern University

summary: In single cells, variably sized nanoscale chromatin structures are observed but it is unknown if these form a cohesive framework that regulates RNA transcription. Herein we demonstrate that the human genome is an emergent, self-assembling system. Conformationally-defined heterogenous, nanoscopic packing domains form by the interplay of transcription, nucleosome remodeling, and loop extrusion. We show that packing domains are not topologically associated domains. Instead, packing domains exist across a structure-function life-cycle that couples heterochromatin and transcription in situ, explaining how heterochromatin enzyme inhibition can produce a paradoxical decrease in transcription by destabilizing domain cores. Applied to development and aging, we show the pairing of heterochromatin and transcription at myogenic genes that could be disrupted by nuclear swelling. In sum, packing domains represent a new foundation to explore the interactions of chromatin and transcription at the single cell level in human health.

overall_design: To investigate the relationship between higher order genome organization and terminal cellular differentiation,  we took advantage of plastic myoblast cells and differentiated myotube cells. After treating 2 technical replicates of myoblasts with differentiation media, we generated Hi-C for each set of samples using the Qiagen Epitect Hi-C Kit (Qiagen, #59971). We processed each replicate using the HiC Nextflow pipeline and generated .cool files for each replicate. Replicates were merged to maximize resolution

first: Lucas

last: Carter

first: Andrew

last: Skol

first: Kyle

last: MacQuarrie


title: Epigenetic modulation through BRD8 impairs conversion of primed EpiSCs to naïve ESCs [ChIP-Seq]

last: Hutchins

first: Xiuling

last: Fu


title: human genome-wide silencers

city: shenzhen

first: Guoli

last: Li

address: penfei street

institute: Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences

summary: Silencers, the yin to enhancers' yang, play a pivotal role in fine-tuning gene expression throughout the genome. However, despite their recognized importance, comprehensive identification of these regulatory elements in the genome is still in its early stages. We developed a method called Ss-STARR-seq to directly determine the activity of silencers in the whole genome. In this study, we applied Ss-STARR-seq to human cell lines K562, LNCaP, and 293T, and identified 134,171, 137,753, and 125,307 silencers on a genome-wide scale, respectively, these silencers function in various cells in a cell-specific manner. Silencers exhibited a substantial enrichment of transcriptional-inhibitory motifs, including REST, and demonstrated overlap with the binding sites of repressor transcription factors within the endogenous environment. Interestingly, H3K27me3 did not reflect silencer activity but facilitated the silencer's inhibitory role on gene expression. Additionally, the silencer did not have any significant histone markers at the genome-wide level. Our findings unveil that aspect-silencers not only transition into enhancers throughout diverse cell lines but also achieve functional conversion with insulators. Regarding to biological effects, knockout experiments underscored the functional redundancy and specificity of silencers in regulating gene expression and cell proliferation. In summary, this study pioneers the elucidation of the genome-wide silencer landscape in human cells, delineates their global regulatory features, and identifies specific silencers influencing cancer cell proliferation.

first: Xiusheng

last: Zhu


title: Single-cell RNA-seq of peripheral blood links cell-type-specific regulation of splicing to autoimmune and inflammatory diseases [scRNAseq_KR_SGI]

first: Boxiang

first: Chi

last: Tian

first: Yuntian

first: Yihan

last: Tong

first: Kian Hong

last: Kock

first: Kyung Yeon

last: Han

first: Yoshinari

last: Ando

first: Woong-Yang

last: Park

first: Jay W.

last: Shin

first: Shyam

last: Prabhakar


title: Tissue origin of endothelial cells determines immune system modulation and regulation of HIF1α-, TGFβ- and VEGF- signaling

city: Hamburg

first: Andreas

last: Neueder

address: Falkenried 94

department: Center for Molecular Neurobiology (ZMNH)

country: Germany

institute: University Medical Center Hamburg-Eppendorf

summary: Dysfunction of the blood retinal barriers and/or proliferation of retinal and choroidal endothelial cells are caused by late stages of diabetic retinopathy (DR) and neovascular age-related macular degeneration (nAMD). To elucidate endothelial-derived pathomechanisms in DR and nAMD, we established immortalized mouse cell lines of retinal (REC),  choroidal (ChEC) and brain (BEC)endothelial cells. We compared the functional and transcriptomic state of these cells depending on their tissue origin. Intriguingly, activation of the wingless-type MMTV integration site (Wnt)/β-catenin signaling pathway restored barrier properties in BEC and REC, but increased permeability of ChEC. Transcriptome profiling showed that the immune system and pathways such as hypoxia-inducible factor (HIF) 1/2α-, transforming growth factor (TGF) β- and vascular endothelial growth factor (VEGF) were differentially regulated among endothelial cells. These findings significantly increase the understanding of the vascular biology of endothelial cells, highlighting the fact that depending on their tissue origin, their contribution to vascular pathologies such as DR or nAMD may vary.

overall_design: Immortalised endothelial cells of different origin were sequenced by the nanopore long-read technology.

first: Robin

last: Heiden

first: Laura

last: Hannig

first: Jakob

middle: S

last: Bernhard

first: Mario

last: Vallon

first: Anja

last: Schlecht

first: Nico

last: Hofmann

first: Süleyman

last: Ergün

first: Franziska

last: Hoschek

first: Maximilian

last: Wagner

first: Carola

middle: Y

last: Förster

first: Barbara

last: Braunger


title: Air-liquid_interface_Caco-2_culture_with_vasointestinal_peptide_ mimicks_ gut_ mucosal_ barrier_ function_ in_ permeability_ and_bacterial infection

status: Public on Dec 04 2024

city: Utrecht

first: Karin

last: Strijbis

state: Utrecht

address: Yalelaan 1

department: Biomolecular Health Sciences

country: Netherlands

institute: Utrecht University

summary: The intestinal mucosal barrier is a dynamic system that allows nutrient uptake, stimulates healthy microbe-host interactions, and prevents invasion by pathogens. The mucosa consists of epithelial cells connected by cellular junctions that regulate the passage of nutrients covered by a mucus layer that plays an important role in host-microbiome interactions. Mimicking the intestinal mucosa for in vitro assays, in particular the generation of a viscous mucus layer, has proven to be challenging. Here, we present a novel in vitro intestinal culture model that produces a robust mucus layer and is based on the widely used intestinal Caco-2 cell line. We investigated the effects of air-liquid interface (ALI) culturing compared to liquid-liquid interface (LLI) on Caco-2 cells grown under low-glucose conditions in Transwell plates. In addition, we determined the impact of vasointestinal peptide (VIP) on mucus secretion, epithelial barrier properties and microbe-mucus interactions. A combination of ALI culturing with VIP addition led to formation of a robust mucus layer on the apical surface of the Caco-2 confluent layer in which the intestinal secreted mucin MUC2 was a major component. RNAseq analysis demonstrated upregulation of unique gene clusters in response to ALI and VIP conditions, but the ALI-VIP combination treatment resulted in a significant upregulation of multiple mucin genes and proteins including MUC2, MUC13 and MUC17. Expression of tight junction proteins was significantly altered in the ALI-VIP condition leading to increased permeability to small molecules, more closely reflecting an intestinal epithelium permissive for nutrient uptake. In infection experiments with commensal Lactobacillus plantarum, pathogenic Salmonella enterica serovar Enteritidis, or enterotoxigenic Escherichia coli (ETEC), the ALI-VIP mucus layer separated the bacteria from the underlying epithelium. In conclusion, ALI-VIP culture of Caco-2 cells provides an attractive in vitro model to study the function of the intestinal mucosal barrier and pathogenic and commensal microbe-host interactions.

overall_design: To investigate the impact of ALI culturing and VIP on mucus production and epithelial barrier properties of Caco-2 cells. We demonstrate that the combined ALI-VIP cultures lead to high upregulation of secreted and transmembrane mucins and an altered tight junction component expression, allowing the uptake of small molecules. We then performed gene expression profiling analysis using data obtained from bulk RNA-seq of 4 different conditions after 14 days of differentiation. Comparative gene expression analysis of RNA-seq data for Caco-2 cells and its different culture and treatment conditions (LLI-NT, LLI-VIP, ALI-NT and ALI-VIP)

first: Evelien

last: Floor

first: Jinyi

last: Su

first: Maitrayee

last: Chatterjee

first: Elise

last: Kuipers

first: Noortje

last: IJssennagger

last: Giordano

first: Silvia

last: Mihăilă

first: Daphne

last: Stapels

first: Yvonne

last: Vercoulen


title: Priming of Antiviral Innate Immune Responses in Specific Alveolar Macrophage Subsets by Cholesterol Deprivation

city: Kumamoto

first: Ken

last: Takashima

state: Kumamoto

address: 1-1-1, Honjo, Chuo-ku

department: Department of Immunology

institute: Kumamoto University

summary: Cholesterol metabolism is associated with antiviral innate immune responses; however, its underlying mechanism is not fully elucidated. In this study, we performed a chemical screening to isolate small chemicals that affect the activities of the cytoplasmic viral RNA sensor RIG-I and found that statins, which inhibit cholesterol synthesis, markedly enhanced RIG-I-dependent type I interferon (IFN) production following viral infections. Restriction of cholesterol synthesis induced the expression of noncanonical type I IFNs, such as IFN-ω and IFN-α16, in the SREBP1 transcription factor-dependent manner. Produced noncanonical type I IFNs subsequently increased the expression of RIG-I via the type I IFN receptor, thereby augmenting RIG-I-dependent cytokine expression following viral infection. Administering statins augmented RIG-I-dependent cytokine expression in the mouse lung, and a mouse obesity model exhibited reduced RIG-I response in the lung compared to wild-type mice. Single-cell transcriptome analyses revealed a subset of alveolar macrophages that increase the RIG-I expression in response to the restriction of cholesterol synthesis in vivo. This study uncovered an alveolar macrophage population responding to cholesterol metabolism and priming antiviral innate immune responses.

overall_design: To investigate the impact of cholesterol metabolism in lung tissue, we injected Pitavastatin, which inhibits cholesterol biosynthesis, into mice and harvest their lung tissue. On the other hands, we also harvest lung tissue from Ob/Ob mice. And then, we constructed the sequence library for single cell-RNA seq.

first: Tasuku

last: Nishimura

first: Takahisa

last: Kouwaki

first: Hiroyuki

last: Oshiumi


title: The global RNA-binding protein RbpB is a regulator of polysaccharide utilization in Bacteroides thetaiotaomicron

city: Würzburg

first: Alexander

middle: J.

last: Westermann

address: Josef-Schneider-Straße 2/D15

institute: Helmholtz Institute for RNA-based Infection Research (HIRI), Helmholtz Centre for Infection Research (HZI),

summary: Paramount to human health, symbiotic bacteria in the gastrointestinal tract rely on the breakdown of complex polysaccharides to thrive in this sugar-deprived environment. Gut Bacteroides are metabolic generalists and deploy dozens of polysaccharide utilization loci (PULs) to forage diverse dietary and host-derived glycans. The expression of the multi-protein PUL complexes is tightly regulated at the transcriptional level. However, how PULs are orchestrated at translational level in response to the fluctuating levels of their cognate substrates is unknown. Here, we identify the RNA-binding protein RbpB and a family of noncoding RNAs as key players in post-transcriptional PUL regulation. We demonstrate that RbpB interacts with numerous cellular transcripts, including a paralogous noncoding RNA family comprised of 14 members, the FopS (family of paralogous sRNAs). Through a series of in-vitro and in-vivo assays, we reveal that FopS sRNAs repress the translation of SusC-like glycan transporters when substrates are limited—an effect antagonized by RbpB. Ablation of RbpB in Bacteroides thetaiotaomicron compromises colonization in the mouse gut in a diet-dependent manner. Together, this study adds to our understanding of RNA-coordinated metabolic control as an important factor contributing to the in-vivo fitness of predominant microbiota species in dynamic nutrient landscapes.

overall_design: Identification of RNAs co-purified with 3x-FLAG-tagged RbpB by cross-linking immunoprecipitation and sequencing (CLIP-seq) and non-crosslinked samples as controls. Two biological replicates

first: Ann-Sophie

last: Rüttiger

first: Daniel

last: Ryan

first: Luisella

last: Spiga

first: Vanessa

last: Lamm-Schmidt

first: Gianluca

last: Prezza

first: Sarah

last: Reichardt

first: Madison

last: Langford

first: Lars

last: Barquist

first: Franziska

last: Faber

first: Wenhan

last: Zhu

middle: J


title: Cohesin phosphorylation accelerates DNA damage repair and reduces translocations

city: bethesda

first: Seolkyoung

last: Jung

state: MD

address: 10 Center Dr

department: NIAMS

institute: NIH

summary: we describe and clarify several mechanisms whereby Cohesin contributes to DNA repair. Upon acute Cohesin depletion, we observed delayed DNA repair and increased frequency of translocations. These translocations were directly correlated to contact frequency between damaged domains. We found that phosphorylation of the Cohesin subunit SMC1A is required to direct specific Cohesin recruitment at DSBs, without affecting its distribution in the undamaged genome. Moreover, Cohesin depletion caused a significant increase in translocations in the 53BP1 deficient background, indicating that Cohesin functions to suppresses 53BP1 independent end joining mechanisms. Our work sheds light on a previously unappreciated mechanisms whereby gCohesin is loaded at DSBs and actively extrudes loops between the break and nearest natural TAD boundary. This process modulates the 3D proximity of broken ends to accelerate repair speeds and prevent long range end joining events during NHEJ.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for Rad21, Nipbl and Mre11 in HCT116 cells in different genotypic background w/wo DSB and HiC analysis w/wo auxin in DSB.

first: Michael

last: Fedkenheuer

first: Yafang

last: Shang

first: Kevin

first: Solji

last: Park

first: Davide

last: Mazza

first: Hua

last: Tan

first: Dali

last: Zhong

first: Robin

last: Sebastian

first: Sushil

last: Jaiswal

first: Jan

last: Wisneski

first: Vittorio

last: Sartorelli

first: John

last: O'Shea

first: Laura

last: Elnitski

first: Andre

last: nessenzweig

first: Fei-Long

last: Meng

first: Rafael

last: Casellas

first: Mirit

last: Aladjem


title: The effect of knockout Brd7 on 4T07-TGL cells under in vitro condition [RNA-seq]

city: Houston

first: Junfeng

last: Zhang

institute: The University of Texas MD Anderson Cancer Center

summary: Metastasis is a major cause of mortality in cancer and is likely influenced by epigenetic and gene regulatory factors. Using an in vivo screen, we demonstrated that several subunits of the polybromo-associated BAF (PBAF) complex, particularly BRD7, were required for breast cancer metastatic dormancy in lung. BRD7 loss induced metastatic outgrowth, along with modifications in epigenomic landscapes and upregulated oncogenic signaling. Breast cancer cells harboring BRD7 loss also reprogrammed the surrounding immune microenvironment, downregulating MHC-1 expression and promoting a pro-metastatic cytokine profile. Flow cytometric and single-cell analyses revealed increased levels of pro-tumorigenic neutrophils, CD8+ exhausted T cells, and CD4+ stress response T cells in lungs harboring BRD7-deficient metastases. Finally, attenuating this immunosuppressive milieu by neutrophil depletion, neutrophil extracellular trap (NET) inhibition, or immune checkpoint therapy abrogated metastatic outgrowth. These findings, for the first time, implicate BRD7 and PBAF in metastasis, pointing to targetable underlying mechanisms involving specific immune cell compartments.

overall_design: To study the function of PBAF subunit Brd7 in Breast cancer metastatic dormancy, we established Brd7 knockout 4T07-TGL cell lines by CRISPR/Cas9.  RNAseq was performed for 2 Brd7 knockout clones (Brd7-KO1, Brd7-KO2) compared with the control cells (4T07-TGL-sgCtr), each group was in triplicate.


title: Satellite cell dynamics is impaired in chicken Wooden Breast myopathy via altered Syndecan-4 function

city: Stony Brook

first: Addolorata

last: Pisconti

address: 100 Nicolls Rd

department: Biochemistry and Cell Biology

institute: Stony Brook University

summary: Introduction: Skeletal muscle satellite cells (MuSCs, or stem cells) play a crucial role in muscle development, maintenance, and regeneration, serving both hypertrophic support and regenerative myogenesis. Syndecans (SDCs) act as communication bridges within the muscle microenvironment, regulating interactions with extracellular matrix components, and contributing significantly to tissue repair and inflammation. Specifically, syndecan-4 (SDC4) is involved in muscle regeneration at multiple stages. Methods: This study delves into the emerging challenge of wooden breast (WB) myopathy and its connection to SDC4. Our hypothesis proposes that disruptions in MuSC dynamics through SDC4 contribute to the increased incidence of breast myopathies observed in growing broilers. To test our hypothesis, non-affected and affected broilers were systematically selected, and the characteristics of WB myopathy were studied both in vitro and in vivo. SDC4 overexpression in MuSCs and blocking peptides corresponding to the SDC4 ectodomain were used for investigation of the role of SDC4 in muscle development and its shedding levels. Results and Discussion: Examination in vivo of affected muscles revealed smaller fibers and changes in metabolic pathways. In vitro studies unveiled disrupted proliferation of MuSCs in WB myopathy, accompanied by downregulation of several muscle markers. Investigation of a potentially role of SDC4 in the pathogenesis of WB myopathy revealed a decreased tendency in SDC4 gene expression and increased shedding of its ectodomain. Moreover, we showed that SDC4 overexpression is linked to decreased proliferation in MuSCs and affected myogenesis. We detected the impairment proliferation of WB-affected MuSCs, revealing critical insights into the dysfunctional state of these cells in myopathy. Additionally, by treating MuSCs with blocking peptides derived from the SDC4 ectodomain, we identified altered proliferation.  Taken together, this work contributes valuable knowledge on the molecular mechanisms underlying WB myopathy and role of SDC4 in this chicken myopathy.

overall_design: Primary myoblaststs from male chickens (Gallus gallus) of the Ross-308 breed, 36 days old, were isolated from the pectoralis muscle and expanded in culture prior to being lysed and processed for RNA-sequencing.

first: Lucie

last: Pejskova

first: Mona

last: Pedersen

first: Sissel

last: Ronning


title: CLEC16A in astrocytes promotes mitophagy and limits pathology in a multiple sclerosis mouse model [Single Cell Dataset]

status: Public on Dec 03 2024

city: Boston

first: Zhaorong

last: Li

address: 60 Fenwood Road

institute: Dana Farber Cancer Institute

summary:
Astrocytes promote neuroinflammation and neurodegeneration in multiple sclerosis (MS) through cell-intrinsic activities and their ability to recruit and activate other cell types. Here, in a genome-wide CRISPR-based forward genetic screen investigating regulators of astrocyte proinflammatory responses, we identified the C-type lectin domain-containing 16A gene (CLEC16A), linked to MS susceptibility, as a suppressor of nuclear factor-κB (NF-κB) signaling. Gene and small-molecule perturbation studies in mouse primary and human embryonic stem cell-derived astrocytes in combination with mitochodrial, metabolic and multiomic analyses established that CLEC16A promotes mitophagy, limiting mitochondrial dysfunction and the accumulation of mitochondrial products that activate NF-κB, the NLRP3 inflammasome and gasdermin D. Astrocyte-specific Clec16a inactivation increased NF-κB, NLRP3 and gasdermin D activation in vivo, worsening experimental autoimmune encephalomyelitis, a mouse model of MS. Moreover, we detected disrupted mitophagic capacity and gasdermin D activation in astrocytes in samples from individuals with MS. These findings identify CLEC16A as a suppressor of astrocyte pathological responses and a candidate therapeutic target in MS.
Scheduled for publication in Nature Neuroscience
DOI: 10.1038/s41593-025-01875-9

overall_design: For single cell RNA-sequencing, CNS cells from n=3 Control EAE and n=3 CLEC16AGFAP EAE (Day 23) were hashed before FACS sorting using multiplex 3’ Cell Plex Set Kit A (#100261, 10x Genomics).  After sorting and washing,  astrocytes from 6 samples were pooled and loaded into one well of a 10X Chromium Next GEM Chip G (10X Genomics). Single-cell libraries were further generated using the Single Cell 3’ Reagent Kit v3.1 chemistry (10x Genomics) using Chromium Next GEM Single Cell 3’ GEM, Library & Gel Bead Kit v3.1 (PN-1000128) according to the manufacturer’s protocol. The generated scRNA-seq libraries were sequenced by NOVA-Seq S1 at the Broad Institute.

first: Atsushi

last: Kadowaki

first: Michael

last: Wheeler

first: Brian

last: Andersen

first: Hong-Gyun

last: Lee

first: Tomer

last: Illuz

first: Francisco

last: Quintana


title: Evolution of myeloid-mediated immunotherapy resistance in prostate cancer

city: San Francisco

first: Aram

last: Lyu

state: CA

address: 513 Parnassus Avenue, HSE 301

institute: University of California San Francisco

summary: Patients with advanced metastatic castration-resistant prostate cancer (mCRPC) are refractory to immune checkpoint inhibitors (ICIs), partly because there are immunosuppressive myeloid cells in tumours. However, the heterogeneity of myeloid cells has made them difficult to target, making blockade of the colony stimulating factor-1 receptor (CSF1R) clinically ineffective. Here we use single-cell profiling on patient biopsies across the disease continuum and find that a distinct population of tumour-associated macrophages with elevated levels of SPP1 transcripts (SPP1hi-TAMs) becomes enriched with the progression of prostate cancer to mCRPC. In syngeneic mouse modelling, an analogous macrophage population suppresses CD8+ T cell activity in vitro and promotes ICI resistance in vivo. Furthermore, Spp1hi-TAMs are not responsive to anti-CSF1R antibody treatment. Pathway analysis identifies adenosine signalling as a potential mechanism for SPP1hi-TAM-mediated immunotherapeutic resistance. Indeed, pharmacological inhibition of adenosine A2A receptors (A2ARs) significantly reverses Spp1hi-TAM-mediated immunosuppression in CD8+ T cells in vitro and enhances CRPC responsiveness to programmed cell death protein 1 (PD-1) blockade in vivo. Consistent with preclinical results, inhibition of A2ARs using ciforadenant in combination with programmed death 1 ligand 1 (PD-L1) blockade using atezolizumab induces clinical responses in patients with mCRPC. Moreover, inhibiting A2ARs results in a significant decrease in SPP1hi-TAM abundance in CRPC, indicating that this pathway is involved in both induction and downstream immunosuppression. Collectively, these findings establish SPP1hi-TAMs as key mediators of ICI resistance in mCRPC through adenosine signalling, emphasizing their importance as both a therapeutic target and a potential biomarker for predicting treatment efficacy.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: To understand the evolution of the mechanisms underlying myeloid-mediated immunotherapy resistance and to identify targets for enhancing therapeutic efficacy in prostate cancer, we performed single-cell profiling of patient biopsies from cases of localized, metastatic hormone-sensitive prostate cancer, or mCRPC. We then reverse translated these findings in a syngeneic mouse model (MyC-CaP) of HSPC and CRPC, conducting multi-omic single-cell assessment and studies to assess tumor efficacy and gain mechanistic insights.

first: Zenghua

last: Fan

first: Matthew

last: Clark

first: Averey

last: Lea

first: Diamond

last: Luong

first: Ali

last: Setayesh

first: Alec

last: Starzinski

first: Rachel

last: Wolters

first: Marcel

last: Arias-Badia

first: Kate

last: Allaire

first: Kai

last: Wu

first: Vibha

last: Gurunathan

first: Laura

last: Valderrábano

first: Richard

last: Miller

first: Xiao

last: Wei

first: Rebecca

last: Heist

first: Eliezer

last: Van Allen

first: Lawrence

last: Fong


title: High resistance barrier and prophylactic protection in preclinical models of SARS-CoV-2 with two siRNA combination

first: Emily

last: Wong

address: 1800 Owens St

institute: VIR BIOTECHNOLOGY, INC.

summary: RNA interference is a natural antiviral mechanism that could be harnessed to combat SARS-CoV-2 infection by targeting and destroying the viral RNA. We identified potent lipophilic small-interfering RNA (siRNA) conjugates targeting highly conserved regions of the SARS-CoV-2 outside of the spike-encoding region capable of achieving ≥3-log viral reduction. Serial passaging studies demonstrated that a two-siRNA combination prevented development of resistance compared to a single-siRNA approach. Viral resistance to single siRNA treatment occurred due to emergence of point mutations at critical positions required for siRNA-mediated target binding and cleavage, which led to a loss of siRNA efficacy. With a two-siRNA combination, emergence of mutations within the siRNA binding site was abolished. When delivered intranasally, two-siRNA combination protected Syrian hamsters from weight loss and lung pathology by viral infection upon prophylactic administration but not following onset of infection. Together, the data support potential utility of RNAi as a prophylactic approach with high resistance barrier to counteract SARS-CoV-2 emergent variants and complement vaccination. Most importantly, given that the siRNAs can be rapidly developed from a new pathogen sequence, this strategy has implications as a new type of preventive medicines that may protect against future coronavirus pandemics.

sample_organism: Severe acute respiratory syndrome coronavirus 2

overall_design: To determine the barrier to drug resistance for the lead siRNAs, we used in vitro viral serial passaging method. SARS-CoV-2 virus was passaged five times in the presence of either single-siRNA (COV-siRNA1 or COV-siRNA2) or the two-siRNA combination (COV-siRNA1+2) at 5x, 10x, and 20x of EC50 determined from earlier experiments.

first: Anna

last: Sahakyan

first: Dana

last: Bohan

first: Roberto

last: Spreafico


title: Three-dimensional chromatin reorganization during CD8+ T cell activation is mediated by TRIM28 [CUT&Tag CTCF]

city: Beijing

first: Ruifeng

last: Li

address: haidian

institute: Tsinghua University

summary: T cell activation is accompanied by extensive changes in epigenome. However, whether and how high-ordered chromatin organization is involved in CD8+ T cell activation is unclear. Here, we showed extensive changes in the three-dimensional genome during CD8+ T cell activation, associated with changes in gene transcription. Moreover, we identified Tripartite motif containing 28 (TRIM28) as an essential mediator in re-organization of spatial chromosomal interactions. Trim28 deficiency impaired CD8+ T cell activation in vitro and in vivo, due to its regulation on autocrine IL-2 production. Mechanistically, TRIM28 bound to genes associated with CD8+ T cell activation and promoted the formation of chromosomal loops. At the loop anchor regions, TRIM28 is likely recruited by CTCF, and required for binding of RNA Pol II and cohesin to form active chromosomal structure. These results thus identify a critical role of TRIM28-dependent chromatin topology in gene transcription during CD8+ T cell activation.

overall_design: CUT&Tag for TRIM28 of in WT and CTCF-deficient CD8+ T cells in vitro.

first: Kun

first: Chen

last: Dong


title: Three-dimensional chromatin reorganization during CD8+ T cell activation is mediated by TRIM28

overall_design: Hi-C experiment of naïve and Trim28fl/fl vs Trim28-/- CD8+ T cells activated in vitro.


title: Viral oncogene manipulates YY1 to alter host 3D genome organization and promote B cell proliferation

city: Minneapolis

first: Chong

last: Wang

state: MINNESOTA

institute: University of Minnesota

summary: YY1 plays essential roles in regulating enhancer-promoter interactions. Here we report that Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNALP), essential for EBV immortalization of naïve B lymphocytes(NBLs), manipulates YY1 to alter enhancer-promoter interactions during transformation. EBNALP colocalized with TOP2B and was tethered to DNA through DPF2 in lymphoblastoid cell lines(LCLs). EBNALP inactivation led to reduced looping at enhancers including CCND2 identified by H3K27ac HiChIP, with reduced TOP2B, DPF2, and YY1 DNA binding. EBNALP interacted with YY1 in LCLs by immune precipitation and colocalized by immunofluorescence. DPF2 CRISPR knock out reduced EBNALP and YY1 DNA binding. CRISPRi repression of CCND2 enhancers reduced CCND2 TSS H3K27ac and CCND2 expression. EBNALP also repressed enhancer looping at enhancers including CASP1. EBNALP inactivation led to increased enhancer looping with increased TOP2B, DPF2, and YY1 binding these enhancer sites.DNA accessibility increased at these sites following EBNALP inactivation by ATAC-seq. H3K9me3 was enriched at EBNALP repressed enhancer sites in LCLs.These data identified novel mechanisms through which EBVmanipulates YY1’s function to rewire host DNA elements and reorganize the host genome to facilitate EBV mediated growth transformation.

overall_design: To further study the mechanism through which EBNALP alters host genome organization and transcriptional regulation.  Here we performed H3K27AC HiChIP, ATAC-seq, RNA-seq and DPF2, YY1 Cut&Run after WT or EBNALP mutant EBV infection of naive B cells. CRISPRi was used to validate those function links.

first: Merrin

last: Leong

first: Weiyue

last: Ding

first: Bo

last: Zhao

first: Mingxiang

last: Teng


title: Three-dimensional chromatin reorganization during CD8+ T cell activation is mediated by TRIM28 [ATAC-Seq]

address: Haidian District

department: Institute for Immunology and School of Medicine

overall_design: ATAC-seq analysis of Trim28fl/fl vs Trim28-/- CD8+ T cells naïve and activated in vitro.


title: Cell integrity limits ploidy in budding yeast

city: Cambridge

first: Charles

middle: Arthur

last: Whittaker

state: MA

institute: Koch Institute

summary: Evidence suggests that increases in ploidy have occurred frequently in the evolutionary history of organisms, and can serve adaptive functions to specialized, somatic cells in multicellular organisms (Edgar & Orr-Weaver, 2001; Orr-Weaver, 2015; Van De Peer et al., 2017). However, the sudden multiplication of all chromosome content may present physiological challenges to the cells in which it occurs. Experimental studies have associated increases in ploidy with reduced cell survival and proliferation (Andalis et al., 2004; Fujiwara et al., 2005). To understand the physiological challenges that suddenly-increased chromosome content imposes on cells, we used S. cerevisiae cells to ask how much chromosomal DNA cells may contain, and what determines this limit. We generated polyploid cells using two distinct methods causing cells to undergo endoreplication and identified the maximum ploidy of these cells, 32-64C. We found that physical determinants that alleviate or exacerbate cell surface stress increase and decrease the limit to ploidy, respectively. We also used these cells to investigate gene expression changes associated with increased ploidy, and identified the repression of genes involved in ergosterol biosynthesis. We propose that ploidy is inherently limited by the impacts of growth in size, which accompany whole genome duplication, to cell surface integrity.

overall_design:
Gene expression profiling of two yeast strains that double in ploidy with each cell cycle. Genome duplication was induced in mad1Δ bub2Δ (MB) and cdc24-1 swe1Δ (CS) cells and gene expression was profiled in three replicate samples of each strain, every four hours, for 16 hours total.
Reference samples, of constant DNA content, were included to control for effects of each strain’s genotype on gene expression with experimental time (MB cells not treated with nocodazole; CS cells grown at 25C).
Additional reference samples, consisting of cell cycle-arrested cells, were included to control for effects of enlarged cell size on gene expression with experimental time (WT cells treated with nocodazole; cdc24-1 cdc20-1 swe1Δ cells grown at 37C).
Mean DNA content was measured for each sample and samples were used, in two tests, to identify changes in gene expression relative to DNA content.

first: Juliet

last: Barker

first: Stephen

last: Bell

first: Andrew

last: Murray

first: Angelika

last: Amon


overall_design: 1. To globally analyze the genes regulated by TRIM28, we performed RNA-seq analysis of Trim28fl/fl vs Trim28-/- CD8+ T cells activated in vitro. 2. To globally analyze the genes regulated by IL-2, we performed RNA-seq analysis of untreated vs anti-IL-2-treated CD8+ T cells in the presence of anti-CD3 and anti-CD28 for 3 days in vitro

last: Wei


title: Centromere positioning orchestrates telomere bouquet formation and the initiation of meiotic differentiation

city: Salamanca

first: Alberto

last: Pineda-Santaella

address: Pintor Fernando Gallego

institute: IBFG

summary: Accurate gametogenesis requires the establishment of the telomere bouquet, an evolutionarily conserved, 3D chromosomal arrangement. In this spatial configuration, telomeres temporarily aggregate at the nuclear envelope during meiotic prophase, which facilitates chromosome pairing and recombination. The mechanisms governing the assembly of the telomere bouquet remain largely unexplored, primarily due to the challenges in visualizing and manipulating the bouquet. Here, using Schizosaccharomyces pombe as a model system to elucidate telomere bouquet function, we reveal that centromeres, traditionally perceived as playing a passive role in the chromosomal reorganization necessary for bouquet assembly, likely serve as the key initiators of telomere bouquet formation. We demonstrate that centromeres have the remarkable capability to induce telomere mobilization, which is even sufficient to trigger bouquet formation and the meiotic transcription program in mitotic cells. This discovery highlights the finely tuned control exerted over long-distance heterochromatic regions and underscores a pivotal step in the mechanism of eukaryotic telomere bouquet formation.

overall_design: In order to assess the effect of centromere declustering on general gene expression, we performed RNA-seq of asynchronic log-phase cultures in rich medium of cells of wt and sad1.2 csi1Δ backgrounds.

last: Jiménez-Martín

first: Rebeca

last: Martín-García

first: Daniel

last: León-Periñán

first: Antonia

last: Terrizano

first: Rafael

middle: R

last: Daga

first: Sigurd

last: Braun

first: Alfonso

last: Fernández-Álvarez


title: Three-dimensional chromatin reorganization during CD8+ T cell activation is mediated by TRIM28 [ChIP-Seq]

first: Chen

last: Dong

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChlP-seg) for Trim28, CTCF as well as the histone modifications H3K27ac and H4K16ac of Trim28fl/fl vs Trim28-/- CD8+ T cells activated in vitro.

first: Kun

first: Ruifeng

last: Li


title: SMAD4 lossdifferentlyshapespancreatictumorswithdistinct KRASmutations

status: Public on Dec 02 2024

city: CAMBRIDGE

first: Giulia

last: Biffi

state: Cambridgeshire

address: Cancer Research UK, Robinson Way

country: United Kingdom

institute: University of Cambridge

summary: We compared KPC Smad4 WT and Smad4 KO tumours from orthotopically-grafted organoid-derived mouse models

overall_design: 8 samples in total - 3 KPC Smad4 WT (ROSA) and 5 KPC Smad4 KO murine PDAC tumours

first: Eloise

last: Lloyd

first: Muntadher

last: Jihad


city: Bunkyo-ku

first: Yu

last: Yamashita

state: Tokyo

address: Yushima 1-5-45

department: Cell Signaling

institute: Institute of Science Tokyo

summary: We examined a global mRNA expression in primary osteoblasts from wild-type and Fam102a knockout mice.

overall_design: The calvarial cells isolated from the calvarial bone of wild-type or Fam102a knockout newborn mice and were cultured in osteogenic medium. After 7 days, total RNA was extracted and analyzed for gene expression.

first: Mikihito

last: Hayashi

first: Takehito

last: Ono

first: Tomoki

last: Nakashima


title: Next Generation Sequencing from control and CD97 knockdown of glioblastoma stem cells

city: Kaifeng

first: Xiong

last: Jin

state: Henan

address: Jin Ming Avenue

department: School of Life Sciences & School of Pharmacy

institute: Henan University

summary: Glioblastoma (GBM) stem cells (GSCs) are largely associated with a poor prognosis of GBM. Although the importance of identifying and validating molecular markers in GBM has been emphasized, single markers covering all GSCs are lacking. Through in vitro antibody screening, in this study, we demonstrated that CD97 identifies GSCs as a novel surface marker. Compared to conventional well-known GSC markers, such as CD133, CD44 and CD15, only CD97 was notably detected in all tested patient-derived GSCs. GBM patients expressing high levels of CD97 experienced worse clinical outcomes, and were predictive of poor overall survival in the public database. CD97 is required for GSC proliferation and self-renewal and was shown to reduce mouse survival with aggressive tumor progression in an orthotopic xenograft model. A comparative transcriptomic analysis revealed that CD97 triggers mTORC2 signaling through Akt (S473) phosphorylation, enhancing the expression of downstream genes, including ARHGAP1, BZW1, and BZW2. Moreover, inhibition of mTORC2 signaling with the pharmacological inhibitor JR-AP2-011 suppressed GSC proliferation, self-renewal, and downstream gene expression. Furthermore, sorting of CD97-enriched cells from patient-derived GSCs revealed that CD97 promotes GSC self-renewal and tumorigenicity via mTORC2/AKT signaling. Thus, our findings suggest that CD97 may be a general GSC enrichment marker in GBM and that the CD97-related pathway might serve as a therapeutic target for GBM.

overall_design: mRNA profiles of control and CD97 knockdown glioblastoma stem cells

first: Jinlong

last: Yin

first: Sun Il

last: Choi


title: Single cell RNA-sequencing of M002 oHSV-treated murine glioblastoma

city: Birmingham

first: Jianmei

middle: Wu

last: Leavenworth

state: AL

address: 1670 University Boulevard

department: Neurosurgery

institute: UAB

summary: In this study, we evaluated the effects of M002 oncolytic herpes simplex virus (oHSV) in a syngeneic intracranial glioblastoma stem cell model. We harvested tumor-bearing mouse brain tissue 45 days post-injection of M002 oHSV and performed single-cell RNA-sequencing (scRNA-seq).

overall_design: Cells were isolated from glioma tumor-bearing brain tissue using fluorescence-activated cell sorting (FACS) using CD45- and CD45+ fractions and analyzed using scRNAseq.

first: Sadashib

last: Ghosh

first: Jianmei Wu


title: Non-canonical roles of CFH in retinal pigment epithelial cells revealed by dysfunctional rare CFH variants

city: Biberach

first: Petr

last: Taus

address: Birkendorfer Straße 65

institute: Boehringer Ingelheim

summary: Complement Factor H (CFH) common genetic variants have been associated with age-related macular degeneration (AMD). While most previous in vitro RPE studies focused on the common p.His402Tyr CFH variant, we characterized rare CFH variants that are highly penetrant for AMD using induced pluripotent stem cell derived retinal pigment epithelium (iPSC-RPE). Our results show that lower FH levels were detected in AMD RPE, which potentially disrupted canonical and non-canonical roles of FH. Specifically, AMD RPE displayed higher inflammation rate and a reduced set of differentially expressed genes compared to control RPE upon A2E and blue light challenge. Additionally, cholesterol efflux and POS phagocytosis defects, dysregulated complement levels, larger sub-RPE deposits and increased mitochondrial stress were observed in AMD RPE. Thus, our study reveals new non-canonical roles for FH in regulating important RPE functions.

overall_design: RPE cells were exposed to 5 µM A2E for three days followed by blue light exposure for 10 minutes on the fourth day. On the fifth day supernatant and cells were collected for ELISA and RNAseq respectively. Control cells were treated with DMSO and exposed to light or DMSO without light. Untreated samples were also cultured under the same conditions as A2E and DMSO treated samples and collected as controls at the end of the experiment.

first: Seba

last: Almedawar

first: Sofie

last: Brink


title: H3K56 acetylation regulates chromatin maturation following DNA replication

city: New York

first: Zhiguo

last: Zhang

state: NY

address: 1130 St. Nicholas Avenue

department: Pediatric and Genetics and Development

institute: Columbia University

summary: Following DNA replication, the newly reassembled chromatin is disorganized and must mature to its steady state to maintain both genome and epigenome integrity. However, the regulatory mechanisms governing this critical process remain poorly understood. Here, we show that histone H3K56 acetylation (H3K56ac), a mark on newly-synthesized H3, facilitates the remodeling of disorganized nucleosomes in nascent chromatin, and its removal at the subsequent G2/M phase of the cell cycle marks the completion of chromatin maturation. In vitro, H3K56ac enhances the activity of ISWI chromatin remodelers, including yeast ISW1 and its human equivalent SNF2h. In vivo, a deficiency of H3K56ac in nascent chromatin results in the formation of closely packed di-nucleosomes and/or tetra-nucleosomes. In contrast, abnormally high H3K56ac levels disrupt chromatin maturation, leading to genome instability. These findings establish a central role of H3K56ac in chromatin maturation and reveal a mechanism regulating this critical aspect of chromosome replication.

overall_design: We synchronized yeast cells (Rtt109, Chd1 and Isw1 depletion and H3K56R mutation in WT and mcm2-3A background cells ) at G1 and released into early S phase in the presence of BrdU. We then performed BrdU immunoprecipitation using anti-BrdU antibodies following single-strand DNA library preparation and sequencing (ssSeq). we also performed ChIP and eSPAN experiments followed by single-strand DNA sequencing (ssSeq) for H3K4me3, H3K56ac. The sequencing tag was mapped to both Watson (red) and Crick (blue) strands of the reference genome.

first: Shoufu

last: Duan


title: Insulin signaling regulates R2 retrotransposon expression to orchestrate transgenerational rDNA copy number maintenance

first: Amelie

last: Raz

address: 455 Main St

institute: Whitehead Institute for Biomedical Research

summary: Preserving a large number of essential yet highly unstable ribosomal DNA (rDNA) repeats is critical for the germline to perpetuate the genome through generations. Spontaneous rDNA loss must be countered by rDNA copy number (CN) expansion. Germline rDNA CN expansion is best understood in Drosophila melanogaster, which relies on unequal sister chromatid exchange (USCE) initiated by DNA breaks at rDNA. The rDNA-specific retrotransposon R2 responsible for USCE-inducing DNA breaks is typically expressed only when rDNA CN is low to minimize the danger of DNA breaks; however, the underlying mechanism of R2 regulation remains unclear. Here we identify the insulin receptor (InR) as a major repressor of R2 expression, limiting unnecessary R2 activity. Through single-cell RNA sequencing we find that male germline stem cells (GSCs), the major cell type that undergoes rDNA CN expansion, have reduced InR expression when rDNA CN is low. Reduced InR activity in turn leads to R2 expression and CN expansion. We further find that dietary manipulation alters R2 expression and rDNA CN expansion activity. This work reveals that the insulin pathway integrates rDNA CN surveying with environmental sensing, revealing a potential mechanism by which diet exerts heritable changes to genomic content.

overall_design: 50 testes were hand dissected from 1 to 5-day-old flies in 1x PBS and transferred immediately into tubes with cold 1x PBS on ice. Tissue dissociation was performed. Cell viability and density was determined on a hemocytometer using Trypan Blue stain and DIC imaging. Cells were processed for library preparation using the 10 X Genomics Chromium Controller and Chromium Single Cell Library and Gel Bead Kit following standard manufacturer’s protocol. Amplified cDNA libraries were quantified by bioanalyzer, size selected by AMPure beads, and sequenced on a NovaSeq SP.

first: Jonathan

middle: O

last: Nelson

first: Alyssa

last: Slicko

first: Yukiko


title: RNA-sea of Smad4 wild-type or Smad4 knockout KvPC PDAC organoids from monocultures or co-cultures with pancreatic stellate cells and RNA-seq of flow-sorted PSCs from the same co-cultures.

first: Giulia

last: Biffi

summary:
RNA-sequencing of flow-sorted Smad4 wild-type (WT) or Smad4 knockout (KO) KvPC (i.e. KRASG12V; p53 mutant) pancreatic ductal adenocarcinoma (PDAC) organoids from monocultures or co-cultures with pancreatic stellate cells (PSCs) and RNA-sequencing of flow-sorted PSCs from the same co-cultures.
We investigated changes in transcriptome in Smad4 KO KvPC PDAC organoids compared to Smad4 WT KvPC PDAC organoids in monoculture or in co-culture with PSCs. We also investigated the changes in transcriptome in PSCs co-cultured with Smad4 WT or Smad4 KO KvPC PDAC organoids.

overall_design: 35 samples in total. Comparative gene expression profiling analysis of RNA-seq data for flow-sorted Smad4 WT or Smad4 KO KvPC PDAC organoids in monoculture or co-cultured with PSCs, and of co-cultured PSCs.

first: Eloise

last: Lloyd

first: Muntadher

last: Jihad


title: GABAB receptor modulation of membrane excitability in human pluripotent stem cell-derived sensory neurons by baclofen and α-conotoxin Vc1.1

status: Public on Dec 01 2024

city: Wollongong

first: Dominic

last: Kaul

address: Northfields Ave

institute: University of Wollongong

summary: GABAB receptor (GABABR) activation is known to alleviate pain by reducing neuronal excitability, primarily through inhibition of high voltage-activated (HVA) calcium (CaV2.2) channels and potentiating G protein-coupled inwardly rectifying potassium (GIRK) channels. While the analgesic properties of small molecules and peptides have been primarily tested on isolated murine dorsal root ganglion (DRG) neurons, emerging strategies to develop, study, and characterise human pluripotent stem cell (hPSC)-derived sensory neurons present a promising alternative. In this study, hPSCs were efficiently differentiated into peripheral DRG-induced sensory neurons (iSNs) using a combined chemical and transcription factor-driven approach, via a neural crest cell intermediate. Molecular characterisation and transcriptomic analysis confirmed the expression of key DRG markers such as BRN3A, ISLET1, and PRPH, in addition to GABABR and ion channels including CaV2.2 and GIRK1 in iSNs. Functional characterisation of GABABR was conducted using whole-cell patch clamp electrophysiology, assessing neuronal excitability under current clamp conditions in the absence and presence of GABABR agonists baclofen and α-conotoxin Vc1.1. Both baclofen (100 mM) and Vc1.1 (1 mM) significantly reduced membrane excitability by hyperpolarizing the resting membrane potential, and increasing the rheobase for action potential firing. In voltage-clamp mode, baclofen and Vc1.1 inhibited HVA Ca2+ channel currents, which were attenuated by the selective GABABR antagonist CGP 55845. However, modulation of GIRK channels by GABABRs was not observed in the presence of baclofen or Vc1.1, suggesting that functional GIRK1/2 channels were not coupled to GABABRs in hPSC-derived iSNs. This study reports is the first to report GABABR modulation of membrane excitability in iSNs by baclofen and Vc1.1, highlighting their potential as a future model for studying analgesic compounds.

overall_design: In preparation for iSN differentiation, cell culture plasticware was first coated with 10 μg/ml Poly-D-Lysine (Sigma–Aldrich) for 30 min at room temperature, then coated with Matrigel overnight at 37 °C. CD271+ NCCs were then seeded at a density of approximately 8.0 ´ 103 cells/cm2 on 12 mm circular glass coverslips for patch-clamp studies, 2.6 ´ 104 cells/cm2 on 13 mm circular glass coverslips for immunocytochemistry, or 2.6 ´ 104 cells/cm2 directly on plasticware for transcriptomic analyses. iSN differentiation was completed following previously published methods with some modifications (Hulme et al., 2020) St Clair-Glover et al, 2024). Briefly, cells were cultured in neuronal medium containing Neurobasal™ medium containing 1% N2, 1% B-27 minus vitamin A, 1% Insulin-transferrin-Selenium-A, and 1% GlutaMAX™ (all from Gibco, Waltham, MA, USA). Throughout iSN differentiation, the media was supplemented with 10 μM Y-27632, 10 ng/ml BDNF, 10 ng/ml GDNF, 10 ng/ml NT-3, and 10 ng/ml b-NGF (all from STEMCELL Technologies), with media changes  conducted every 2-3 days. For the initial 96 h after plating (day 6-10), NGN2 expression was induced by the application of doxycycline at 2 µg/mL (Sigma-Aldrich, D9891). From day 14, cells were phased into BrainPhys™ neuronal medium (BrainPhys™ Neuronal Medium, 0.02% NeuroCult™ SM1 without Vitamin A, 0.01% N2 Supplement-A [all from STEMCELL Technologies]) for maturation, with a ratio of 25:75, 50:50, 75:25, 100:0 (BrainPhys™: neuronal media) at each media change. Where required, proliferating cells were removed by adding anti-mitotic agent, cytosine β-D-arabinofuranoside at 2.5 μM (AraC, Sigma-Aldrich), for 48 h between days 16-18

first: Mitchell

last: St Clair-Glover

first: Arsalan

last: Yousuf

first: Mirella

last: Dottori

first: David

last: Adams


title: RNA-sea of Smad4 wild-type or Smad4 knockout KPC PDAC organoids from monocultures or co-cultures with pancreatic stellate cells and RNA-seq of flow-sorted PSCs from the same co-cultures.

summary:
RNA-sequencing of flow-sorted Smad4 wild-type (WT) or Smad4 knockout (KO) KPC (i.e. KRASG12D; p53 mutant) pancreatic ductal adenocarcinoma (PDAC) organoids from monocultures or co-cultures with pancreatic stellate cells (PSCs) and RNA-sequencing of flow-sorted PSCs from the same co-cultures.
We investigated changes in transcriptome in Smad4 KO KPC PDAC organoids compared to Smad4 WT KPC PDAC organoids in monoculture or in co-culture with PSCs. We also investigated the changes in transcriptome in PSCs co-cultured with Smad4 WT or Smad4 KO KPC PDAC organoids.

overall_design: 44 samples in total. Comparative gene expression profiling analysis of RNA-seq data for flow-sorted Smad4 WT or Smad4 KO KPC PDAC organoids in monoculture or co-cultured with PSCs, and of co-cultured PSCs.


title: Targeting CERT sensitizes AML Cells to FLT3 Inhibitors through activating GRP78-ATF6-CHOP Axis

city: GuangZhou

first: Xiaofan

last: Sun

address: 613 Huangpu Avenue, Tianhe District

institute: The First Affiliated Hospital of Jinan University

summary: Ceramide Transfer Protein (CERT) determines the ratio of ceramide and Sphingomyelin in cells. Targeting CERT sensitizes solid cancer cells to chemotherapy. However, the role of CERT in AML remains elusive. Knockdown CERT or its inhibitor HPA-12  inhibited growth and promoted apoptosis in AML cells carrying FLT3-ITD mutation. HPA-12 alone or with the FLT3 inhibitor Crenolanib synergistically inhibited proliferation and triggered apoptosis in FLT3-ITD mutated AML cells in vitro and in vivo. Interestingly, co-treatment of HPA-12 and Crenolanib is effective in FLT3-ITD+ and FLT3-TKD+ AML patients. The synergistic effects were mediated by the endoplasmic reticulum (ER) stress-GRP78/ATF6/CHOP axis and mitophagy. Our data suggest a possible strategy to enhance the efficacy of FLT3 inhibitors in AML.

overall_design: both including DMSO, 48 h HPA-12 and Creno at the indicated concentrations alone or in combination treatment groups

first: Sun

last: Xiaofan

first: Li

last: Yue

first: Du

last: Juan

first: Zeng

last: Hui


title: Mitochondrial homeostasis of endocrine organ navigates nutrient restriction checkpoint

city: Guangzhou

first: Suning

last: Liu

state: Guangdong

address: No.55, West of Zhongshan Avenue, Tianhe District

institute: South China Normal University

summary: Steroid hormones are powerful endocrine regulators, but little is known about the integration of their production with environmental conditions. Here, we use Drosophila prothoracic gland (PG) to examine how insect steroid hormone production navigates a developmental checkpoint critical for juvenile-to-adult transition. Extensive transcriptome analysis of the PGs revealed that starvation during pre-nutrient restriction checkpoint (NRC) downregulated substantial mitochondria-related genes. We then discovered that pre-NRC starvation reduced prothoraciotropic hormone signaling, insulin signaling, and TORC1 activity in PG cells, which prevented mitochondrial fragmentation and import of Disembodied, a key steroidogenic enzyme. Ultimately, this starvation caused severe mitophagy and proteasome dysfunction, blocking ecdysteroidgenesis and metamorphosis. By contrast, post-NRC starvation did not impair mitochondrial homeostasis in PG cells, but reduced Sit expression and induced moderate autophagy to promote ecdysteroidgenesis, leading to precocious metamorphosis. This study constitutes a paradigm for exploring how steroid hormones themselves are controlled in response to environmental stress like starvation.

overall_design: The RGs of control or starved w1118 larvae were dissected in PBS in batches of 10–40 at a time, and pooled into three biological replicates for every treatment. About 1,200 RGs were pooled per sample, which yielded ~1-5 μg of RNA required for sequencing. Total RNA was extracted using the miRNeasy Micro Kit (QIAGEN, Germany), then stored at ~80 °C. RNA quality, especially DNA contamination, was monitored on 1.5% agarose gels. RNA concentration was measured using NanoDrop 2000 (Thermo Fisher Scientific, USA). RNA integrity was assessed using the RNA Nano 6000 Assay Kit of the Agilent Bioanalyzer 2100 system (Agilent Technologies, USA). High-quality RNA samples from Drosophila RGs were sent to Biomarker Technologies Corporation (Beijing, China) for construction of cDNA libraries and sequencing.


title: Loss of an oligodendrocyte specific silencer element provides a framework to assess the pathogenic impact of lamin B1 structural variants

city: Pittsburgh

first: Sarah

last: Hainer

state: PA

address: 4249 Fifth Ave

department: Department of Biological Sciecnes

institute: University of Pittsburgh

summary: The role of non-coding regulatory elements and how they might contribute to tissue type specificity of disease phenotypes is poorly understood. Autosomal Dominant Leukodystrophy (ADLD) is a fatal, adult-onset, neurological disorder that is characterized by extensive CNS demyelination. Most cases of ADLD are caused by tandem genomic duplications involving the lamin B1 gene (LMNB1) while a small subset are caused by genomic deletions upstream of the gene. Utilizing data from recently identified families that carry LMNB1 gene duplications but that do not exhibit demyelination, ADLD patient tissues, CRISPR modified cell lines and mouse models, we have identified a novel silencer element that is lost in ADLD patients and that specifically targets overexpression only to oligodendrocytes. This element consists of CTCF binding sites that mediate three-dimensional chromatin looping involving the LMNB1 and the recruitment of the PRC2 repressor complex.  Loss of the silencer element in ADLD identifies a previously unknown role for silencer elements in tissue specificity and disease causation.

overall_design: CUT&RUN experiments in human fibroblast cells and oligodendrocytes from Mus musculus

first: Bruce

last: Nmezi

first: Guillermo

last: Bey

first: Talia

middle: D

last: Oranburg

first: Kseniia

last: Dudnyk

first: Santana

last: Lardo

first: Nathan

last: Herdman

first: Anastasia

last: Jacko

first: Sandy

last: Rubio

first: Emanuel

last: Alcocer

first: Julia

last: Kofler

first: Dongkyeong

last: Kim

last: Rankin

first: Emma

last: Kivuva

first: Nicholas

last: Gutowski

first: Katherine

last: Schon

first: Jelle

last: van den Ameele

first: Patrick

middle: F

last: Chinnery

first: Sérgio

middle: B

last: Sousa

first: Filipe

last: Palavra

first: Camilo

last: Toro

first: Filippo

last: Vairo

first: Jonas

last: Saute

first: Lisa

last: Pan

first: Murad

last: Alturkustani

first: Robert

last: Hammond

first: Francois

last: Gros-Louis

first: Michael

last: Gold

first: Yungki

last: Park

first: Genvieve

last: Bernard

first: Raili

last: Raininko

first: Jian

last: Zhou

first: Quasar

last: Padiath