title: Argonaute-siRNA loading via the RNA binding protein RDE-4 in C. elegans

city: Fort Collins

first: Taiowa

last: Montgomery

state: CO

address: 1878 Campus Delivery

department: Department of Biology

institute: Colorado State University

summary: Small RNAs associate with Argonaute proteins to control gene expression, impacting a wide range of cellular processes, such as antiviral defense, transposon silencing, and development. Plants and animals typically have several classes of small RNAs, along with multiple Argonautes. These Argonautes often confer distinct functionality to the various classes of small RNAs. But how small RNAs are selectively loaded into the appropriate Argonaute is not well understood. Small interfering RNAs (siRNAs) and microRNAs (miRNAs) are two classes of small RNAs commonly generated from double-stranded RNA (dsRNA) precursors by the endoribonuclease Dicer, although not all siRNAs or miRNAs are produced in this manner. dsRNA-derived siRNAs are often cleaved from fully base-paired precursors, whereas miRNAs typically originate from partially base-paired hairpins.  In C. elegans, Dicer/DCR-1 processing of siRNAs and a related dsRNA-derived small RNA class known as 26G-RNAs is mediated by the dsRNA-binding protein RDE-4.  In the study associated with this data, we used small RNA high-throughput sequencing to show that RDE-4 also promotes loading of siRNAs but not miRNAs into the Argonaute RDE-1, but not ALG-1, and loading of 26G-RNAs into the Argonaute ERGO-1, but not into the two paralogous Argonautes ALG-3 and ALG-4.  Although ALG-3/4 associated 26G-RNAs do not require RDE-4 for Argonaute loading, the levels of these small RNAs are strongly reduced in rde-4 mutants. Therefore, RDE-4 functions in both dsRNA processing and selective loading of small RNAs into specific Argonautes. Our findings reveal a role for RDE-4 as a critical determinant of small RNA loading specificity and provide insight into the mechanisms by which small RNAs are selectively paired with their corresponding Argonaute proteins.

overall_design: High-throughput small RNA sequencing was performed on rde-4 and rde-1 mutant strains, non-mutant controls, and transgenic strains expressing GFP or HA fusions to Argonaute proteins. The animals were either left untreated or subjected to RNAi targeting oma-1 and/or nrfl-1 ord dcr-1. RNA was extracted from cell lysates or Argonaute co-immunoprecipitates from C. elegans cultured at 20°C. The resulting small RNA data was analyzed using the tinyRNA pipeline.


title: Structural and regulatory determinants of flagellar motility in Rhodobacterales – The archetypal flagellum of Phaeobacter inhibens DSM 17395

status: Public on Mar 21 2025

city: Třeboň

first: Jürgen

last: Tomasch

address: Novohradská 237

country: Czech Republic

institute: Center Algatech, Institute of Microbiology of the Czech Academy of Science

type: Expression profiling by high throughput sequencing

summary: Transcriptomes of P. inhibens DSM 17493 wild-type and transposon mutations of the three genes of the CtrA phosphorelay are compared.

overall_design: P. inhibens ctrA::Tn, cckA::Tn and chpT::tn are compared to wild type DSM 17493

first: Jörn

last: Petersen


title: Self-regulating gene therapy ameliorates phenotypes and overcomes gene dosage sensitivity in a mouse model of Rett syndrome

status: Public on Mar 19 2025

city: Edinburgh

first: Paul

last: Ross

address: Hugh Robson Building. George Square

country: United Kingdom

institute: University of Edinburgh

summary: Conventional methods of gene transfer lead to inconsistent transgene expression within cells. This variability can be problematic, particularly in conditions like Rett syndrome (RTT), a neurological disorder caused by mutations in the MECP2 (methyl-CpG binding protein 2) gene, because overexpression of MECP2 can also cause adverse effects. To address these challenges, we devised a gene regulation system called Expression Attenuation via Construct Tuning (EXACT), which employs a self-contained, microRNA-based feed-forward loop that not only ensures more consistent transgene expression but also protects against excessive expression. Through cell-based screening assays, we demonstrated the ability of the EXACT circuit to modulate the expression of full-length human MeCP2. Compared to a conventional construct, an EXACT-MECP2 construct exhibited a narrower range of cellular protein expression. Furthermore, the degree of regulation by the EXACT circuit increased with higher transgene doses in vitro and in wild-type mice and mice modeling RTT. Based on cellular and in vivo testing, we identified an optimal configuration for the adeno-associated virus serotype 9 (AAV9) construct for self-regulated MECP2 gene therapy, designated NGN-401. Delivery of NGN-401 to neonatal male Mecp2-/y hemizygous mice via intracerebroventricular injection resulted in prolonged survival and amelioration of RTT-like phenotypes compared to vehicle-treated animals. NGN-401 was also well tolerated by female Mecp2+/- mice and healthy nonhuman primates, in contrast to a conventional construct, which caused toxicity. The results from this study underpin a first-in-human pediatric study of NGN-401 in RTT (ClinicalTrials.gov, NCT05898620).

overall_design: RNA-seq profiling of HEK293A, HepG2, or SH-SY5Y cells transfected with either a plasmid expressing mNeonGreen or one expressing mNeonGreen plus a miRNA (EXACT1).

middle: D

first: Ralph

last: Hector

first: Stuart

middle: R

last: Cobb


title: A RAS(ON) multi-selective inhibitor combination therapy triggers long-term tumor control through senescence-associated tumor-immune equilibrium in preclinical models of PDAC

status: Public on Apr 07 2025

city: Redwood City

first: Ximo

last: Pechuan-Jorge

state: CA

address: 700 Saginaw Drive

country: USA

institute: Revolution Medicines

summary: Pharmacological inhibition of oncogenic RAS represents an attractive strategy to target pancreatic ductal adenocarcinoma (PDAC), an almost ubiquitously RAS-driven disease.  However, initial responses to targeted monotherapy inhibition of active RAS can be followed by relapses, potentially driven by the persistence of drug-tolerant tumor cells. To target these ‘persister’ cells, we investigated strategies to increase their immune visibility in mouse models of PDAC. We show that combining a RAS(ON) multi-selective inhibitor with the CDK4/6 inhibitor palbociclib drives persister cells into a senescent-like state, which coincides with improved tumor control and substantial remodeling of the tumor microenvironment. Combining RAS(ON) and CDK4/6 inhibition with a CD40 agonist results in durable regressions and CD4 T cell-dependent tumor-immune equilibrium. Our studies reveal a combinatorial approach that circumvents resistance to RAS(ON) inhibitor monotherapy in preclinical models and demonstrate a mechanism by which therapy-induced senescence can be reinforced by the immune system, resulting in durable tumor control.

overall_design: Mice (6-10 week old C57Bl/6N female) bearing orthotopic pancreatic tumors derived from KPC cells. Treated with vehicle, Palbociclib, RMC-7977, RMC-7977 + Palbociclib, or RMC-7977 + Palbociclib + FGK4.5 for 7 or 14 days. Tumors processed to isolate cells for scRNA-seq.

first: Caroline

last: Broderick

first: Riccardo

last: Mezzadra

first: Exequiel

last: Sisso

first: Felix

last: Mbuga

first: Rashi

last: Raghulan

first: Almudena

last: Chaves-Perez

first: Amanda

last: Kulick

first: Lingyan

last: Jiang

first: Jingjing

first: Yu-Ji

last: Ho

first: Janelle

last: Simon

first: Eric

last: Chan

last: Rosiek

first: Aveline

last: Filliol

first: Ronan

last: Chaligné

first: Elisa

last: de Stanchina

first: Andrea

last: Schietinger

first: Mallika

last: Singh

first: Scott

middle: W

last: Lowe


title: Development of an Anti-CD99 Antibody Enables Targeting of Diffuse Midline Glioma

status: Public on Mar 28 2025

city: AURORA

first: Etienne

last: Danis

department: Cancer Center

institute: University of Colorado, Denver - Anschutz Medical Campus

summary: This article highlights the development of an antibody targeting CD99 in diffuse midline gliomas (DMG). 1. RNA-seq: We performed bulk RNA sequencing of HSJD-GBM01 cells transduced with H3.3K27M and its paired isogenic H3.3-WT cell lines to find the genes altered due to the transduction with the K27M Mutation compared to isogenic H3-WT controls. 2. RNA-seq: We also performed bulk RNA sequencing of SU-DIPG04 cells transduced with short-hair pin RNA (shRNA) targeting CD99 and with transduced with non-target controls to find the genes altered due the knockdown of CD99 compared to those transduced with non-target controls.

overall_design: 1. RNA-seq samples were obtained from HSJD-GBM01 glioblastoma cells transduced with H3.3K27M mutation and isogenic H3-WT controls. 2. RNA-seq samples were obtained from SU-DIPG04 DMG cells transduced with shRNA targeting CD99 and with non-target controls.

first: Sujatha

last: Venkataraman

first: Bridget

last: Sanford

first: Kenneth

last: Jones

first: Dong

last: Wang

first: Ilango

last: Balakrishnan

first: Krishna

last: Madhavan

first: Rajeev

last: Vibhakar


title: Abemaciclib reduces glioblastoma stemness by targeting the GSK3b-mediated transcriptional regulation of CD44 and TCF7L2.

status: Public on Mar 12 2025

city: Taipei

first: Tsung-Han

last: Hsieh

address: XinYi dist.,WuXing Street, No.250.

country: Taiwan

institute: Taipei Medical University

summary: Glioblastoma multiforme (GBM) is the most common and deadliest glioma, originating from astrocytic progenitors or stem cells and affecting individuals of all ages. Current treatments include surgery, radiation therapy, chemotherapy, and combinations of these modalities. Temozolomide, the first-line chemotherapy for GBM, improves survival when combined with radiation but has limited success due to the tumor's inherent resistance to therapies. Cancer stem cells (CSCs), which exhibit self-renewal and differentiation capabilities, contribute to therapeutic resistance, making them a promising target for GBM therapy. Abemaciclib, a CDK4/6 inhibitor, effectively reduces tumor growth and sphere formation in GBM, although its exact mechanism remains unclear. Our data revealed that Abemaciclib significantly inhibited tumor sphere formation in U87MG and KNS42 cells by downregulating epithelial-mesenchymal transition (EMT)-associated genes, including CD44 and TCF7L2. Notably, silencing CD44 and TCF7L2 also led to a marked reduction in sphere formation. Phospho-kinase array analysis revealed that Abemaciclib reduces pGSK3β and pCREB in sphere cells. Western blotting confirmed that Abemaciclib diminishes pGSK3β-Y216 in parental cells, suggesting reduced kinase activity. Pharmacological inhibition of GSK3β also decreased CD44 and TCF7L2 expression. In an orthotopic animal model, Abemaciclib inhibited tumor formation, with reduced levels of pRB, pGSK3β, Ki67, and CD44 confirmed by immunohistochemistry. Analysis of TCGA and CGGA datasets showed that the mesenchymal GBM subtype (MES-GBM), associated with poor prognosis, has higher EMT gene expression. Treatment of MES-like LN229 cells with Abemaciclib decreased EMT-related genes. These findings suggest that Abemaciclib disrupts GSK3β-mediated EMT pathways, impairing CSC self-renewal and offering a potential therapeutic strategy for GBM.

overall_design: GBM U87MG cells cultured in a stem-cell-specific medium and treated with Abemaciclib for 14 days and collected the RNA to perform mRNA-sequencing.


title: Using DNA Affinity Purification sequencing (DAP-seq) to identify in vitro binding sites of transcription factors potentially involved in aromatic degradation

city: Madison

first: Kevin

middle: S

last: Myers

state: WI

address: 5127 WEI, 1552 University Ave

department: Great Lakes Bioenergy Research Center

institute: University of Wisconsin - Madison

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The genome-wide binding sites of 44 transcription factors from the aromatic metabolizing Alphaproteobacterium Novosphingobium aromaticivorans were identified using DNA Affinity Purification sequencing (DAP-seq). We report 32 of these transcription factors have at least one area of enrichment. These data will be valuable for better understanding of aromatic metabolism.

overall_design: In vitro DNA binding sites of 44 transcription factors from Novosphingobium aromaticivorans are examined using DAP-seq

first: Myers

last: Kevin

first: Linz

middle: M

last: Alexandra

first: Baumgart

middle: A

last: Leo

first: Zhang

last: Yu

first: Blaby

last: Ian

first: Mewalal

last: Ritesh

first: Ng

last: Vivian

first: Noguera

last: Daniel

first: Donohue

middle: J

last: Timothy


title: Tumor-associated neutrophil precursors impair homologous DNA repair and promote sensitivity to PARP-inhibition.

status: Public on Mar 18 2025

city: Bellinzona

first: Marco

last: Bolis

address: Via Francesco Chiesa

country: Switzerland

institute: IOR Institute of Oncology Research

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

overall_design: Refer to individual Series


title: Epidermal loss of PRMT5 leads to the emergence of an atypical basal keratinocyte-like cell population and defective skin stratification [multiome]

status: Public on Mar 11 2025

city: Iowa City

first: Eric

last: Van Otterloo

state: Iowa

address: 801 Newton Rd

institute: University of Iowa

summary: During skin development, ectoderm-derived cells undergo precisely coordinated proliferation, differentiation, and adhesion to yield stratified epidermis. Disruptions in these processes can result in congenital anomalies including ectodermal dysplasia and harlequin ichthyosis. Protein Arginine Methyl Transferase 5 (PRMT5)—an enzyme responsible for methylating arginine residues in histones and other proteins—maintains progenitor status in germ and limb bud cells. Similarly, in vitro evidence suggests that PRMT5 prevents differentiation of basal keratinocytes, leading us to hypothesize that PRMT5 preserves the stem-cell phenotype of keratinocytes in vivo. To test this possibility, we generated conditional knockout (cKO) mice lacking Prmt5 in early ectoderm (E7.5), impacting the entire epidermis. Prmt5 cKOs exhibited gross skin defects, compromised skin barrier function, and reduced postnatal viability. Histological analyses revealed significant defects in epidermal stratification, without alterations in apoptosis or proliferation. Single-cell RNA and ATAC-seq analysis identified an atypical population of basal keratinocyte-like cells in Prmt5 cKOs, that exhibited a senescence-like program, characterized by increased Cdkn1a (p21), elevated senescence-associated secretory phenotype (SASP) molecules (Igfbp2), and decreased developmental transcription factor (Trp63) expression. Our findings suggest that PRMT5 prevents basal keratinocyte senescence by repressing Cdkn1a, shedding light on the epigenetic regulation of basal keratinocyte maintenance and senescence in congenital skin disorders.

overall_design: single cell RNAseq analysis of E15.5 control and Prmt5 cKO back skin samples Bulk RNAseq analysis of E15.5 control and Prmt5 cKO back skin samples

first: Nicole

last: Recka


title: The kinase GSK-3 alters the repertoire of RNA binding proteins present on lipid metabolism transcripts leading to altered expression in the C. elegans nervous system

status: Public on Mar 07 2025

city: Bloomington

first: Ananya

last: Mahapatra

address: 316 E University St

institute: Indiana University, Bloomington

summary: Tissue-specific regulation of gene expression is essential for multicellular organisms, and RNA binding proteins play central roles in these molecular processes. To determine how the Caenorhabditis elegans RNA binding protein ADR-1 regulates tissue-specific gene expression, we profiled the binding targets of ADR-1 in neural cells and assessed the effects of ADR-1 RNA binding on neural gene expression. We identified a cohort of neural transcripts that facilitate lipid synthesis and are directly regulated by ADR-1 RNA binding. To identify cellular factors that influence ADR-1 binding, a forward genetic screen was performed, revealing that the serine/threonine protein kinase, glycogen synthase kinase-3 (GSK-3), inhibits ADR-1 binding to the cohort. Further investigation revealed that RNA binding protein VIG-1, promotes ADR-1 binding to the cohort in a GSK-3-dependent manner. Together, we reveal that interplay between kinases and RNA binding proteins regulates expression of lipid metabolism genes within neural cells, potentially impacting stress resistance and longevity.

overall_design: High throughput sequencing of neural ADR-1 immunoprecipitated samples and lysates in the presence and absence of adr-2; high throughput sequencing of neural cells isolated from adr-2(-) and adr-2(-);dsRBD1 mutant animals

first: Heather

last: Hundley


title: The m6A epitranscriptome of the HepAD38 cell line

city: Fuzhou

first: Kunqi

last: Chen

address: Fujian Medical University

country: China

institute: Fujian Medical University

summary: m6A modification appears in HepAD38 cell line

overall_design: To investigate whether m6A modification is involved in evolution, we performed MeRIP-seq on HepAD38 cell line


title: The epitranscriptome of the HepAD38 cell line

summary: Epitranscriptomic modifications appear in HepAD38 cell line

overall_design: To investigate whether epitranscriptomic modifications is involved in evolution, we performed RIP-seq on HepAD38 cell line


title: Tumor-associated neutrophil precursors impair homologous DNA repair and promote sensitivity to PARP-inhibition [RNA-seq]

summary: Tumor evolution is one of the major mechanisms responsible for acquiring therapy-resistant and more aggressive cancer clones. Whether the tumor microenvironment through immune-mediated mechanisms might promote the development of more aggressive cancer types is crucial for the identification of additional therapeutical opportunities. Here, we identified a novel subset of tumor-associated neutrophils, defined as tumor-associated neutrophil precursors (PreNeu). These PreNeu are enriched in female highly proliferative hormone-dependent breast cancers and impair DNA repair capacity.  Mechanistically, succinate secreted by tumor-associated PreNeu inhibit homologous recombination, promoting error-prone DNA repair through non-homologous end-joining regulated by PARP-1. Consequently, breast cancer cells acquire genomic instability promoting tumor editing and progression. Selective inhibition of these pathways induces increased tumor cell killing in vitro and in vivo. Tumor-associated PreNeu score correlates with copy number alterations in highly proliferative hormone-dependent tumors from breast cancer patients. Treatment with PARP-1 inhibitors counteract the pro-tumoral effect of these neutrophils.

overall_design: MCF-7 cells, plated in EMEM medium supplemented with 10% heat- inactivated charcoal-stripped FBS (CS-FBS), were treated with either conditioned medium (CM) obtained from PreNeu (cm-PreNeu;), from LOX-1+ neutrophils (cm-LOX1), or control (∅).

first: Arianna

last: Calcinotto

first: Siddhartha

last: Mukherjee

first: Cindy

last: Garda

first: Letizia

last: Boffa

first: Angela

last: Elia

first: Matteo

last: Massara


title: Single cell transcriptomic landscape of pediatric B-cell acute lymphoblastic leukemia: dissection of transcriptional heterogeneity and B-cell developmental state

city: MEMPHIS

first: QINGSONG

last: GAO

state: Tennessee

address: 262 DANNY THOMAS PLACE

institute: ST JUDE CHILDREN'S RESEARCH HOSPITAL

summary: Genomic analyses of bulk ALL samples have improved our understanding of the genetic basis and risk stratification of B-cell acute lymphoblastic leukemia (B-ALL), but do not directly examine intratumor heterogeneity or enable inference of leukemia developmental state and cell of origin. In this study we profiled 89 B-ALL samples by single-cell RNA-seq (scRNA-seq) (10X Genomics 5’v2) to investigate transcriptional heterogeneity and developmental states of leukemic cells.

overall_design:
We profiled 89 B-ALL samples by single-cell RNA-seq (scRNA-seq) (10X Genomics 5’v2) and compared them to a scRNA-seq reference map of normal human B-cell development.
**Submitter states that clinical description will be provided in the corresponding publication as supplemental tables, if relevant to the study's conclusion**
**Raw data were submitted to EGA under accession number EGAS00001007512**

first: Ilaria

last: Iacobucci

first: Charles

last: Mullighan


title: The 5-methylcytosine DNA glycosylase ROS1 prevents paternal genome hypermethylation in Arabidopsis endosperm

city: Cambridge

first: Elizabeth

last: Hemenway

state: MA

address: 455 Main St

institute: Whitehead Institute for Biomedical Research

type: Methylation profiling by high throughput sequencing

summary: DNA methylation patterning is a consequence of opposing activities of DNA methyltransferases and DNA demethylases. In many species, reproduction is a period of significant epigenome lability. In flowering plants, two distinct female gametes, the egg cell and the central cell, are fertilized, producing the embryo and the endosperm, respectively, of the seed. The endosperm is an unusual tissue, exemplified by triploidy and reduced DNA methylation. In Arabidopsis thaliana, a 5-methylcytosine DNA glycosylase, DME, demethylates regions in the central cell genome, leading to methylation differences between maternally- and paternally-inherited endosperm genomes after fertilization. Expression of DME in the central cell is required for gene imprinting, or parent-of-origin specific gene expression, in endosperm. DME is part of a four member gene family in Arabidopsis that includes ROS1, DML2 and DML3. It is unknown whether any of the other DNA glycosylases are required for endosperm methylation patterning. Using whole-genome methylation profiling, we identify ROS1 target regions in the endosperm. We show that ROS1 prevents hypermethylation of paternally-inherited alleles in the endosperm at regions that lack maternal or paternal-allele methylation in wild-type. Thus, ROS1 promotes epigenetic symmetry between parental genomes in the endosperm by preventing paternal genome hypermethylation. We propose that ROS1 and DME act in a parent-of-origin-specific manner at shared endosperm targets, and consider possible implications for the evolution of imprinting mechanisms.

overall_design: Whole-genome Enzymatic Methyl-Seq (EM-seq) performed on DNA extracted from 3C and 6C endosperm nuclei collected 7 days after pollination (DAP) and 2C/4C leaf nuclei collected from young leaf tissue.

first: Mary

last: Gehring


title: Single-cell and spatial transcriptomics uncovers the role of senescent vascular cells in pathological arterial remodeling during murine atherosclerosis

status: Public on Mar 05 2025

city: Baltimore

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

institute: NIA-IRP, NIH

summary: Studying cardiovascular senescence is crucial in understanding the diverse manifestations of disease-related changes in the cardiovascular system and their implications for overall health and disease.  To systematically investigate the heterogeneity of senescent vascular cells in atherosclerosis, we employed the senescence reporter mouse p16TdTomato+/-, injected with an adeno-associated virus expressing PCSK9 to induce atherosclerosis when fed a high-fat diet (HFD).  We then treated this mouse with the senolytic drug ABT-737 or vehicle.  Whole-aorta single-cell RNA-sequencing (scRNA-seq) and Gene Set Enrichment Analysis (GSEA) using the SenMayo panel, uncovered 10 cell clusters displaying increased senescent features that were reduced by treatment with ABT-737.  Interestingly, these clusters do not express ‘classical’ senescence markers such as p16 and p21, but were senescent based on SenMayo GSEA, indicating that the senescence transcriptomic signature in this niche requires deeper characterization.  Using unbiased subclustering of the top four potentially senescent clusters, we identified subsets of cells increasing by HFD and reduced by ABT-737 treatment.  Accordingly, unique transcripts associated with each subpopulation, such as Spp1, Ctsb, and Tnfrsf11b, implicated these cells in senescence-related signaling pathways.  We then validated these findings by spatial transcriptomic profiling, locating senescent cells in the cap and core of brachiocephalic arteries from atherosclerotic mice.  Our results uncover a new vascular-specific transcriptomic signature that may be exploited for therapeutic targeting in age-related vascular diseases.

overall_design: Mice of both sexes were entered in the study at 8 weeks of age, when all animals received 3x1010 viral genomes of PCSK9 AAV8 via retro-orbital injection.  Two weeks later, normal chow was maintained or replaced with an atherogenic diet (42% Fat, 0.2% cholesterol, Harlan atherogenic diet TD.88137).  After 16 weeks, a subset of mice on atherogenic diet were administered ABT-737 via intra peritoneal (IP) injection twice per week for an additional 8 weeks while maintaining the atherogenic diet.  All mouse experiments, including the import, housing, experimental procedures, and euthanasia, were performed strictly under an Animal Study Proposal (ASP #474-LGG-2023) and associated amendments, reviewed and approved by the Animal Care and Use Committee (ACUC) of National Institute on Aging (NIA).

first: Krystyna

last: Mazan-Mamczarz

first: Dimitrios

last: Tsitsipatis

first: Angelica

middle: E

last: Carr

first: Bennett

middle: G

last: Childs

first: Carla

last: Dos Santos

first: Carlos

last: Anerillas

first: Brigette

last: Romero

first: Jordan

last: Gregg

first: Marc

last: Michel

first: Rachel

last: Munk

first: Jennifer

middle: L

last: Martindale

first: Yulan

last: Piao

first: Jinshui

last: Fan

first: Maria

middle: O

last: Hernandez

first: Noemi

last: Kedei

first: Madeline

middle: F

last: Wong

first: Olga

last: Fedorova

first: Mona

last: Batish

first: Darren

last: Baker

first: Myriam

last: Gorospe

first: Allison

middle: B

last: Herman


title: Coordinated regulation by lncRNAs results in tight lncRNA–target couplings [ChIRP-Seq]

city: Houston

first: Hua-Sheng

last: Chiu

state: TX

address: 1102 Bates Avenue

institute: Baylor College of Medicine

summary: The determination of long non-coding RNA (lncRNA) function is a major challenge in RNA biology with applications to basic, translational, and medical research. Our efforts to improve the accuracy of lncRNA-target inference identified lncRNAs that coordinately regulate both the transcriptional and post-transcriptional processing of their targets. Namely, these lncRNAs may regulate the transcription of their target and chaperone the resulting message until its translation, leading to tightly coupled lncRNA and target abundance. Our analysis suggested that hundreds of cancer genes are coordinately and tightly regulated by lncRNAs and that this unexplored regulatory paradigm may propagate the effects of non-coding alterations to effectively dysregulate gene expression programs. As a proof-of-principle we studied the regulation of DICER1—a cancer gene that controls microRNA biogenesis—by the lncRNA ZFAS1, showing that ZFAS1 activates DICER1 transcription and blocks its post-transcriptional repression to phenomimic and regulate DICER1 and its target microRNAs.

overall_design: To identify ZFAS1-bound DNA regions (RNA-DNA interactions), we performed Chromatin Isolation by RNA Purification followed by DNA sequencing (ChIRP-Seq) in two cell lines, ECC-1 and NCI-H460. For each cell line, we generated two datasets: an input control and a ZFAS1 pulldown. We then identified DNA regions enriched in the ZFAS1 pulldown relative to the input control. These enriched regions were processed and output as standard narrowPeak format files.

first: Sonal

last: Somvanshi

first: Chung-Te

last: Chang

first: Eric

middle: d

last: de Lavergne

first: Pieter

last: Mestdagh

first: Pavel

last: Sumazin


title: Polyadenylation at RNA polymerase III loci underlies the accumulation of noncoding retrotransposons during herpesviral infection

status: Public on Mar 10 2025

city: Berkeley

first: Britt

last: Glaunsinger

address: 565 Li Ka Shing Center

department: Department of Plant and Microbial Biology

institute: University of California, Berkeley

summary: The cellular accumulation of short non-coding RNAs (ncRNAs) transcribed by RNA Polymerase III (Pol III) is a hallmark of various cellular stressors and inflammatory-associated diseases. Yet, the mechanisms driving the accumulation of these RNAs are largely undefined. Infection with several DNA viruses is known to significantly alter the cellular Pol III transcriptome, leading to the induction of a class of non-coding retrotransposons known as short interspersed nuclear elements (SINE) and tRNA genes. Here, we sought to define the mechanisms driving Pol III transcribed ncRNA abundance during viral infection, using the murine herpesvirus MHV68 as a model. Our findings reveal that while the expression of Pol III transcripts, such as the murine-specific family of B2 SINE ncRNAs and pre-tRNAs, significantly increase during MHV68 infection, Pol III genomic occupancy is enhanced at a much fewer subset of B2 SINE and tRNA genes. Using DNA motif analyses and a convolutional neural network (CNN) based model, we identified non-promoter sequence elements within B2 SINE genes that distinguish infection-induced loci. We found that infection-induced B2 SINE genes are enriched for signal sequences that confer polyadenylation, and endogenous B2 SINE ncRNA polyadenylation depends on mRNA cleavage and polyadenylation (CPSF) machinery. We discovered that mRNA CPSF components are recruited to sites of Pol III transcription in response to MHV68 infection in a manner dependent on Pol III occupancy. Chromatin-associated B2 SINE ncRNAs are also bound by the CPSF complex, suggesting an RNA-dependent, co-transcriptional polyadenylation of B2 SINE ncRNAs. This uncovers an inducible, coupled relationship between Pol III transcription and mRNA-like polyadenylation of ncRNAs. Also, CPSF recruitment to Pol III genes is not restricted to murine genomes but also occurs at human SINE and tRNA genes, suggesting that this previously unknown coupled relationship may be a widespread feature of Pol III transcription.

overall_design: ChIP-seq of Polr3A in mock-treated or MHV68-infected murine NIH3T3 fibroblasts. ChIP-seq of CPSF30 in murine NIH3T3 fibroblasts treated with either a control siRNA or Brf1 siRNA followed by either mock treatment or infection with MHV68.

first: Azra

last: Lari

first: Sahil

last: Shah

first: Xiawen

last: Mao

first: Priyanka

last: Sanghrajka

first: John

last: Karijolich

first: Liana

last: Lareau


title: Longitudinal single-cell immunoprofiling links durable CAR T response to sustained activation and clonotypic expansion of the native cytotoxic T cell repertoire

status: Public on Mar 04 2025

city: Boston

first: Dimitra

last: Karagkouni

address: Brookline 330

institute: Beth Israel Deaconess Medical Center, Harvard Medical School

summary: CD19-directed chimeric antigen receptor (CAR) T-cell therapy has transformed outcomes for patients with relapsed/refractory large B-cell lymphoma (LBCL), yet the mechanisms underlying durable remission remain incompletely understood. While CAR T-cell persistence is associated with response, long-term remission can occur despite rapid CAR T clearance, suggesting the involvement of additional immune mechanisms. To investigate the role of the native T-cell repertoire in shaping response durability, we performed single-cell RNA and TCR sequencing (scRNA-seq/scTCR-seq) on longitudinal peripheral blood samples from LBCL patients treated with axicabtagene ciloleucel (axi-cel) in the ZUMA-1 trial. We compared immune landscapes and clonotypic dynamics among patients achieving durable remission (>1 year), those experiencing early relapse (<6 months), and those with refractory disease. Patients with long-term remission exhibited increased cytotoxic, proinflammatory, and proliferative native T-cell subsets, while early relapse was associated with immunoregulatory populations that may suppress T-cell activation. TCR profiling revealed robust clonotypic expansion of native cytotoxic T cells post-infusion in durable responders, with expansion patterns strongly predicting clinical outcomes. Notably, TCR screens did not identify known viral targets, suggesting tumor-specific immunity may mediate ongoing remission. These findings propose native T-cell clonotypic expansion as a key determinant of durable response to CAR T therapy and highlight its predictive potential for long-term clinical outcomes.

overall_design:
PBMC samples were collected from 32 LBCL patients at leukapheresis, as well as at 4 weeks, 6 months, and 12 months post-CAR T infusion. These samples were analyzed using scRNA-seq and scTCR-seq.

***Submitters state that this submission includes only processed data files due to patient privacy restrictions. Raw sequencing data can be provided following a request for a Data Use Agreement with Kite Pharma, Inc., in accordance with participating patient consent. Kite is committed to sharing clinical trial data with external medical experts and scientific researchers in the interest of advancing public health. Anonymized individual patient data can be provided for studies of Kite or Gilead compounds approved in the US and the EU with a marketing authorization date on or after 1 January 2014 to qualified researchers based on submitted curriculum vitae and reflecting no conflicts of interest. Approval of such requests is at Kite’s discretion and is dependent on the nature of the request, the merit of the research proposed, the availability of the data, and the intended use of the data. Data requests should be sent to medinfo@kitepharma.com and will be addressed within 60 days.

first: Giulia

last: Cheloni

first: Yered

last: Pita-Juarez

first: Daniela

last: Torres

first: Eleni

last: Kanata

first: Jessica

last: Liegel

first: Zachary

last: Avigan

first: Isabella

last: Saldarriaga

first: Georges

last: Chedid

first: Kathrine

last: Rallis

first: Brodie

last: Miles

first: Gayatri

last: Tiwari

first: Jenny

last: Kim

first: Mike

last: Mattie

first: Jacalyn

last: Rosenblatt

first: Ioannis

last: Vlachos

first: David


title: Profiling transcriptome composition and dynamics within nuclear compartments using SLAM-RT&Tag

city: Seattle

first: Jorja

last: Henikoff

state: WA

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

institute: Fred Hutchinson Cancer Research Center

summary: Nuclear compartments are membrane-less nuclear regions that are enriched in functionally related molecules. RNA is a major component of many nuclear compartments, but the identity and dynamics of transcripts within nuclear compartments are poorly understood. Here, we applied Reverse Transcribe & Tagment (RT&Tag) to identify the transcript populations of Polycomb domains and of nuclear speckles within the nucleus. We also developed SLAM-RT&Tag, which combines RNA metabolic labeling with RT&Tag, to quantify transcript dynamics within nuclear compartments. Intriguingly, exceptionally long genes are transcribed adjacent to Polycomb domains, and are transiently associated with chromatin. In contrast, diverse incompletely spliced mature transcripts migrate to nuclear speckles, and are generally transient within speckles. Speckle transcripts are retained longer when splicing is inhibited, suggesting post-transcriptional processing is coupled to speckle dynamics. Our findings argue that nuclear speckles act as quality control checkpoints that transiently confine incompletely spliced transcripts and facilitate their post-transcriptional splicing.

overall_design: We developed SLAM-RT&Tag, which combines RNA metabolic labeling with RT&Tag, to quantify transcript dynamics within nuclear compartments.

first: Nadiya

last: Khyzha

first: Steven

first: Kami

last: Ahmad


title: Dynamic responses to rejection in the transplanted human heart revealed through spatial transcriptomics

city: Phoenix

first: Nicholas

last: Banovich

state: AZ

address: 445 North Fifth Street

department: Integrated Cancer Genomics

institute: TGen - Translational Genomics Research Institute

summary: Allograft rejection following solid-organ transplantation is a major cause of graft dysfunction and mortality. Current approaches to diagnosis rely on histology, which exhibits wide diagnostic variability and lacks access to molecular phenotypes that may stratify therapeutic response. Here, we leverage image-based spatial transcriptomics at sub-cellular resolution in longitudinal human cardiac biopsies to characterize transcriptional heterogeneity in 62 adult and pediatric heart transplant (HT) recipients during and following histologically-diagnosed rejection. Across 28 cell types, we identified significant differences in abundance in CD4+ and CD8+ T cells, fibroblasts, and endothelial cells across different biological classes of rejection (cellular, mixed, antibody-mediated). We observed a broad overlap in cellular transcriptional states across histologic rejection severity and biological class and significant heterogeneity within rejection severity grades that would qualify for immunomodulatory treatment. Individuals who had resolved rejection after therapy had a distinct transcriptomic profile relative to those with persistent rejection, including 216 genes across 6 cell types along pathways of inflammation, IL6-JAK-STAT3 signaling, IFNα/IFNγ response, and TNFα signaling. Spatial transcriptomics also identified genes linked to long-term prognostic outcomes post-HT. These results underscore importance of subtyping immunologic states during rejection to stratify immune-cardiac interactions following HT that are therapeutically relevant to short- and long-term rejection-related outcomes.

overall_design:
To characterize the cellular and molecular heterogeneity in human heart transplant rejection, we generated a spatial transcriptomics data set measuring expression of 477 genes across 195 0.6 um diameter heart tissue cores from 49 adult and 13 pediatric heart transplant recipients using the 10X Genomics Xenium platform. Among patients, 67.7% had acute cellular rejection (n = 42), 11.3% had antibody-mediated rejection (n = 7), and had 21% mixed cellular-antibody mediated rejection (n = 13). This data set also includes biopsies from before (n  = 86, 44.1%) and after (n = 109, 55.9%) treatment.

UPDATE: [Aug-19-2025] The GSE290577_heart_spatial.rds file was replaced.

first: Kaushik

last: Amancherla

first: Angela

last: Taravella Oill

first: Xavier

last: Bledsoe

first: Arianna

last: Williams

first: Nelson

last: Chow

first: Shilin

last: Zhao

first: Quanhu

last: Sheng

last: Bearl

first: Robert

last: Hoffman

first: Jonathan

middle: N

last: Menachem

first: Hasan

middle: K

last: Siddiqi

first: Douglas

last: Brinkley

first: Evan

last: Mee

first: Niran

last: Hadad

first: Vineet

last: Agrawal

first: Jeffrey

last: Schmeckpepper

first: Aniket

last: Rali

first: Stacy

last: Tsai

first: Eric

last: Farber-Eger

first: Quinn

last: Wells

first: Jane

last: Freedman

first: Nathan

last: Tucker

first: Kelly

middle: H

last: Schlendorf

last: Gamazon

first: Ravi

middle: V

last: Shah


title: Hepatic stellate cells control liver zonation and function via R-spondin 3

status: Public on Mar 03 2025

city: New York

first: Atsushi

last: Sugimoto

state: NY

address: 1130 Saint Nicholas Avenue

institute: Columbia University

summary: Hepatic stellate cells (HSCs) exert a central pathogenic role in the development of liver fibrosis. However, the fibrosis-independent and homeostatic functions of these specialized liver pericytes remain poorly understood. Here, we demonstrate that genetic depletion of HSCs alters Wnt target gene expression in hepatocytes and liver zonation, leading to profound alterations of regeneration, cytochrome P450 metabolism and injury. HSC-selective deletion of Rspondin-3, a modulator of Wnt signalling with high enrichment in HSCs, phenocopies the effects of HSC depletion on hepatocyte gene expression, zonation, regeneration and cytochrome P450-mediated detoxification and increases alcohol-associated and metabolic dysfunction-associated steatotic liver injury. Rspondin-3 expression decreases with HSC activation and is associated with disease progression in patients with alcohol-associated and metabolic dysfunction-associated steatotic liver disease. These hitherto unknown protective and hepatocyte-regulating functions of HSCs via Rspondin-3, resembling the R-spondin-expressing stromal niche of other organs, should be integrated into therapeutic concepts for liver fibrosis.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Single nucleus RNA sequencing of human liver cells from healthy controls (n=6) and patients with alcohol-associated cirrhosis (n=4), alcohol-associated hepatitis (n=5), MASLD (n=3), MASH fibrosis (n=4) or MASH cirrhosis (n=4). Single nucleus RNA sequencing of mouse liver cells from LratCre+ TdTomhomo (n=1) and Cre-negative Rspo3 floxed (n=1). Set 1 of single nucleus RNA sequencing of mouse liver cells from LratCre-positive Rspo3 floxed (n=1) and LratCre-negative Rspo3 floxed (n=1)

first: Qiuyan

last: Sun

last: Schwabe


title: Gene expression profile of kidney and lung cells at 24 hours after the onset of renal bilateral ischemia-reperfusion injury

status: Public on Feb 28 2025

city: St. Louis

first: Andreas

last: Herrlich

state: Missouri

address: 660 South Euclid Avenue

department: Medicine

institute: Washington University

summary: Remote lung injury after acute kidney inkury (AKI) is a common issue in humans and is lethal especially in critically-ill patients. We investigated kidney-lung interactions following AKI using single-cell RNA sequencing (scRNA-seq) of kidney and lung in a murine AKI model.

overall_design: Single cell suspensions from lungs and kidneys were generated from mice with AKI (bilateral ischemia reperfusion injury) or control (sham surgery). Samples from 4 animals were pooled in each condition, subsequently processed, sequenced, and analyzed.


title: Cannabichromene: Integrative modulation of apoptosis, ferroptosis, and endocannabinoid signaling in pancreatic cancer therapy

status: Public on Mar 20 2025

city: Chuncheon

first: Sehyun

last: Chae

address: 1 Kangwondaehakgil

department: Division of Chemical Engineering and Bioengineering

country: South Korea

institute: Kangwon National University

summary: Cannabichromene (CBC: C21H3O2, M.W.: 314.46 g) is a non-psychotropic phytocannabinoid derived from Cannabis sativa (Hemp) and has been increasing attention for its potential therapeutic properties. CBC has demonstrated strong anti-inflammatory effects in animal models with edema through non-CB receptor mechanisms; however, there are still required on pharmacological studies in cancer models. In this study, we investigated the molecular mechanisms on anti-cancer activity of CBC in human pancreatic cancer cells. Through mRNA-seq analysis, we observed that many genes involved in cell death pathways were upregulated and also downregulated by CBC treatment, including ferroptosis-related genes such as HMOX1. We confirmed the functional validation of apoptosis and ferroptosis induction following CBC treatment using a variety of molecular assays. In addition, we found that CBC treatment preferentially increased the expression of TRPV1 and CB2 receptors. The cell death was reversed by inhibitor treatment on TRPV1 and CB2, suggesting that receptor expression is necessary for induction of apoptotic cell death. Finally, we confirmed the consistent regulation on apoptosis, ferroptosis, and the endocannabinoid receptors during tumor growth inhibition by CBC treatment in in-vivo xenograft models. Therefore, we propose that CBC exhibits pharmacological activity via integrative modulation for multiple cell death pathways in the pancreatic cancer therapy.

overall_design: RNA profiles of human pancreatic cancer cells (MIA PaCa-2 and PANC-1) treated with PBS or cannabichromene (CBC)

first: Keun-Cheol

last: Kim

first: Young-Taek

last: Oh

first: Yu-Na

last: Hwang

first: Se-Hyun


title: EWS::FLI cooperatively binds at GGAA microsatellites via DBD-a4 helix in TTC-466 [Micro-C]

status: Public on Feb 27 2025

city: Columbus

first: Ariunaa

last: Bayanjargal

state: OH

address: 700 Children's Dr

institute: Nationwide Childrens Hospital

summary: Multi-omics study of DBD-a4 helix of EWS::FLI to understand the underlying mechanism of transcriptional regulation in Ewing sarcoma cell line: TTC-466

overall_design: In TTC-466 cells, we knock-down endogenous EWSR1::ERG with shRNA  (KD) and rescued with FLAG tagged DBD and DBD+ constructs. Cells are then harvested for RNA-Seq (3 bio reps), CUT&Tag (3 bio reps for CTCF, 4 bio reps for H3K27ac, 4 bio reps for FLAG), and Micro-C libraries (2 bio reps).

first: Emily

middle: R

last: Theisen

first: Julia

last: Selich-Anderson

first: Jesse

middle: C

last: Crow


title: Targeted gene editing and near-universal cDNA insertion of CYBA and CYBB as a treatment for chronic granulomatous disease

status: Public on Feb 24 2025

city: Aarhus

first: Jacob

middle: Giehm

last: Mikkelsen

address: Hoegh-Guldbergs Gade 10

department: Department of Biomedicine

country: Denmark

institute: Aarhus University

summary: Chronic granulomatous disease (CGD) is a severe inborn error of immunity (IEI) caused by defects in the NADPH oxidase. The best current treatment option for patients with CGD, allogeneic hematopoietic stem cell (HSC) transplantation, is associated with severe adverse effects such as graft-versus-host disease, highlighting a need for improved treatment options based on transplantation of autologous ex vivo gene-edited HSCs. Here, we generate CRISPR/Cas9-based gene editing strategies for the correction of two CGD-causing variants; CYBA c.287+1G>T (p22phox) causing autosomal CGD and CYBB c.252G>A (NOX2) causing X-linked CGD. We find that rAAV6 outperforms ssODNs and IDLVs as HDR repair templates in CD34+ hematopoietic stem and progenitor cells (HSPCs), and we optimize gene editing strategies further by including mRNA-encoded inhibitors/effectors. In addition, we develop a near-universal gene editing strategy for X-CGD by targeted integration of a truncated CYBB cDNA, covering 86% of X-CGD patients, and show functional ROS production in edited cells. We find prevalent off-target editing and chromosomal translocations that severely decreased the ability of gene edited HSPCs to engraft in immunodeficient mice, however, by limiting off-target editing through the use of a high-fidelity Cas9, we can show that we can substantially rescue the multilineage engraftment potential of the gene edited HSPCs. To further improve safety, we finally we develop a novel paired D10A Cas9n gene editing approach targeting CYBB. We demonstrate that using this D10A Cas9 approach, we can retain high on-target efficacy while we do not detect any off-target editing or translocations between on- or off-target cleavage sites. Collectively, we bring new insights into to safety and off-target effects of gene editing approaches targeting CYBA and CYBB, highlighting key challenges of these approaches while offering a potentially curative D10A Cas9n-based treatment option for patients with CGD with improved safety.

overall_design: DISCOVER-Seq was carried out as previously described. Briefly, 2x107 K562 cells were nucleofected with either mock, sg3/WT-Cas9 RNPs or sg3/HiFi-Cas9. 12h after nucleofection, the cells were crosslinked in 1% formaldehyde for 15 minutes, washed in PBS and pelleted. The pellets were subsequently resuspended in 10 mL LB1 and incubated for 10 minutes on ice before being spun down at 1000xG for 3 minutes at 4°C. The pellets were then resuspended in 10 mL LB2 and incubated on ice for 5 minutes, before being spun down and resuspended in 1 mL LB3. The nuclear extracts were then sonicated using a Q800 sonicator (Qsonica) (Amplitude: 70%, Pulse Rate: 15s on/45s off, Total Sonication On Time: 40 min, Temperatur: 4oC). Hereafter, the samples were spun down at 17.000xG for 15 minutes to pellet debris. The supernatant was mixed with 1.85 mL LB3 and 150 µL 20% Triton X-100 and mixed with anti-MRE11-bound Dynabeads™ Protein A (Thermo Fisher Scientific). The beads were then incubated, rotating at 4°C for 16 hours, after which the beads were washed 5 times in RIPA buffer and once in TBS before resuspending in 200 µL elution buffer. The crosslink was subsequently reversed by 6 hours of incubation at 65°C. Hereafter, 100 µL TE buffer was added to samples before being treated with RNAse A (Thermo Fisher Scientific) and hereafter Proteinase K (Thermo Fisher Scientific). The DNA was then purified from the samples using the MinElute PCR Purification Kit (Qiagen). The samples were library-prepped using the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs) and sequenced with 50 million 150 bp PE at the AUH NGS core on a NovaSeqTM 6000. Reads were mapped using BWA and analyzed using BLENDER2 with default filtering options.

first: Jonas

last: Wolff

first: Thomas

middle: W

last: Skov


title: Effect of normal and PDK1-deficient human pluripotent stem cells on gene expression during cardiomyocyte differentiation.

status: Public on Feb 20 2025

city: Beijing

first: Amina

middle: Hospital Capital Medical

last: Saleem

state: Chaoyang

address: 2Anzhen Road

department: Pathology and Pathophysiology

institute: Capital Medical University

summary: Cardiac remodeling is the primary factor for the development of ischemic heart failure, which can result from various cardiomyopathies. Pyruvate Dehydrogenase Kinase1 (PDK1) is one of the components of AGC kinase family that maintain mitochondrial metabolism. We here report a PDK1-deficient human cardiac myocyte (CM) model that mimicked the human PDK1 homozygous frameshift mutation and determined the effects of PDK1 dysfunction and its underlying mechanism. PDK1 gene knockout did not affect the pluripotency and differentiation efficiency of hiPSCs. Myocardial cells with a PDK1 gene knockout showed abnormal metabolism, increased oxidative stress levels, decreased cell viability, and increased apoptosis. In addition, lentivirus transfection significantly improved the mitochondrial metabolism in the PDK1-deficient human myocardial model. Taken together, our data provide a PDK1-deficient human cardiomyocyte model that exhibits abnormal mitochondrial metabolism, this model represents an important tool to gain insight into the mechanism of action of metabolism disorders resulting in myocardial remodeling, elucidate the gene-phenotype relationship of PDK1 deficiency, and facilitate drug screening.

overall_design: To investigate the function of PDK1 in the pathogenesis of myocardial remodeling, we established a PDK1-deficient human cardiac myocyte (CM) model. We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different cells at one time point. Comparative gene expression profiling analysis of RNA-seq data for cells.

first: Wei

last: Mingyu


title: Fif1/2/3 and Rsc1 regulates the development of Penicillium oxalicum fungi and the expression of cellulolytic genes

status: Public on Feb 19 2025

city: Qingdao 266237

first: Kexuan

last: Ma

state: China

address: No. 72 Binhai Road

department: State Key Laboratory of Microbial Technology

institute: Shandong University

summary: The goals of this study are to compare different gene expressions for Penicillium oxalicum wild type strain (WT), Fif1/2/3 and Rsc1 deletion strain (Δfif1/2/3 and Δrsc1) under conditions of 2% glucose or 1% bran + 1% microcrystalline cellulose as carbon source. The results of correlation analysis between each sample showed that the gene expression level of Δfif1/2/3 and Δrsc1 was significantly different from that of the WT strain. The deletion of Fif1/2/3 and Rsc1 significantly down-regulates the expression levels of sporulation process genes. And the absence of Fif1/2/3 and Rsc1 can affect the expression of cellulase-encoding genes.

overall_design: Examination of differential gene expressions by digital gene expression tag profiling in Penicillium oxalicum wild type strain, Fif1/2/3 and Rsc1 deletion mutant strains in 18h in modifeid Mendal’s medium with 2% glucose or 1% bran + 1% microcrystalline cellulose as carbon sources. Three replicates were included for each samples.

first: Yuqi

last: Qin


city: Tianjin

first: Jie

last: Li

address: Wei Jin Road Nankai District No. 94

department: Department of Cell Biology and Genetics

institute: Nankai University

summary:
We found that H2AK119ub could regulate 2 cell genes, Usp17l could also activate 2c gene, and Usp17l could remove ubiquitination modification of H2AK119ub. Therefore, in order to determine whether Usp17l affects 2c gene transcription by regulating H2AK119ub, we knockdown Usp17l. The enrichment of H2AK119ub/H3K27me3 was detected by CUT&Tag .
We also sorted out Zscan4 positive and negative cells, and detected the concentration of H3K27me3 by cut-tag

overall_design: CUT&Tag of H2AK119ub/H3K27me3  was performed in control and Usp17l Knockdown cells.CUT&Tag of H3K27me3  was performed in  Zscan4 positive and negative cells

first: Panpan

last: Shi

first: Xukun

last: Lu

first: Kairang

last: Jin

first: Linlin

last: Liu

first: Dai

last: Heng

first: Jiao

last: Yang

first: Huiyu

first: Xiaoyan

last: Sheng

first: Xiaoying

last: Ye


title: Tumor-associated neutrophil precursors impair homologous DNA repair and promote sensitivity to PARP-inhibition [ChIP-seq]

city: Bellinzona

first: Marco

last: Bolis

address: Via Francesco Chiesa

institute: IOR Institute of Oncology Research

summary: Tumor evolution is one of the major mechanisms responsible for acquiring therapy-resistant and more aggressive cancer clones. Whether the tumor microenvironment through immune-mediated mechanisms might promote the development of more aggressive cancer types is crucial for the identification of additional therapeutical opportunities. Here, we identified a novel subset of tumor-associated neutrophils, defined as tumor-associated neutrophil precursors (PreNeu). These PreNeu are enriched in female highly proliferative hormone-dependent breast cancers and impair DNA repair capacity.  Mechanistically, succinate secreted by tumor-associated PreNeu inhibit homologous recombination, promoting error-prone DNA repair through non-homologous end-joining regulated by PARP-1. Consequently, breast cancer cells acquire genomic instability promoting tumor editing and progression. Selective inhibition of these pathways induces increased tumor cell killing in vitro and in vivo. Tumor-associated PreNeu score correlates with copy number alterations in highly proliferative hormone-dependent tumors from breast cancer patients. Treatment with PARP-1 inhibitors counteract the pro-tumoral effect of these neutrophils.

overall_design: ChIP-Seq of the histone modification H3K9me3 in MCF-7 cells subjected to CS-FBS and Succinate 2mM (Cat. S9512, Sigma-Aldrich) or control.

first: Arianna

last: Calcinotto

first: Siddhartha

last: Mukherjee

first: Cindy

last: Garda

first: Letizia

last: Boffa

first: Angela

last: Elia

first: Matteo

last: Massara


title: EPAS1 induction drives myocardial degeneration in Desmoplakin-cardiomyopathy [RNA-seq]

status: Public on Feb 15 2025

city: Utrecht

first: Cornelis

middle: J.

last: Boogerd

address: Uppsalalaan 8

department: Developmental Biology and Stem Cell Research

country: Netherlands

institute: Hubrecht Institute

summary: Arrhythmogenic cardiomyopathy (ACM) is frequently attributed to desmosomal mutations, such as those in the desmoplakin (DSP) gene. Patients with DSP- cardiomyopathy are predisposed to myocardial degeneration and arrhythmias. Despite advancements, the underlying molecular mechanisms remain incompletely understood, thus limiting therapeutic options. Here, we employed spatial transcriptomics on an explanted heart from a patient with a pathogenic DSP variant. Our transcriptional analysis revealed endothelial PAS domain-containing protein 1 (EPAS1) as a potential regulator of mitochondrial homeostasis in stressed cardiomyocytes. Elevated EPAS1 levels were associated with mitochondrial dysfunction and hypoxic stress in both human-relevant in vitro ACM models and additional explanted hearts with genetic cardiomyopathy. Collectively, cardiomyocytes bearing pathogenic DSP variants exhibit mitochondrial dysfunction, increased apoptosis, and impaired contractility, which are linked to the increased EPAS1 levels. These findings implicate EPAS1 as a key regulator of myocardial degeneration in DSP-cardiomyopathy, which expand to other forms of ACM.

overall_design: To elucidate roles of EPAS1 in cardiomyopathy, EPAS1 was suppressed in WT iPSC-derived cardiomyocytes using siRNA, or EPAS1 was overexpressed using AAV6 in DSP p.Arg1113X/WT iPSC-derived cardiomyocytes.

first: Eirini

last: Kyriakopoulou

first: Eva

last: van Rooij

middle: J


title: Heme Biosynthesis Regulates BCAA Catabolism and Thermogenesis in Brown Adipose Tissue

status: Public on Feb 18 2025

city: Madison

first: Andrea

last: Galmozzi

address: 6151 WIMR, 1111 Highland Av, WIMR I, room 6151

institute: UW Madison SMPH

summary: The distinctive color of brown adipose tissue (BAT) is attributed to its high content of heme-rich mitochondria. Despite this, the mechanisms by which BAT regulates intracellular heme levels remain largely unexplored. Here, we demonstrate that heme biosynthesis is the primary source of heme in brown adipocytes. Inhibiting heme biosynthesis results in an accumulation of the branched-chain amino acids (BCAAs) valine and isoleucine, due to a heme-associated metabolon that channels BCAA-derived carbons into heme biosynthesis. Heme synthesis-deficient brown adipocytes display reduced mitochondrial respiration and lower UCP1 levels compared to wild-type cells. While exogenous heme supplementation can restore intracellular heme levels and mitochondrial function, UCP1 downregulation persists. This sustained UCP1 suppression is linked to epigenetic regulation induced by the accumulation of propionyl-CoA, a byproduct of disrupted heme synthesis. Finally, disruption of heme biosynthesis in BAT impairs thermogenic response and, in female, but not male, mice, hinders the cold- induced clearance of circulating BCAAs in a sex-hormone-dependent manner. These findings establish adipose heme biosynthesis as a key regulator of thermogenesis and sex-dependent BCAA homeostasis.

overall_design: Upload contains multiple RNAseq datasets derived from cultured adipocytes or harvested BAT samples. Experiment 1-Differentiation time course of immortalized brown (B9), primary brown (pBAT), and primary white (pWAT) adipocytes harvested at day 0, 2, 4, or 6 of differentiation (associated with Ext. Fig 1).  Experiment 2- pBAT adipocytes differentiated in the presence of heme synthesis inhibitor succinylacetone (SA), heme-arginate (HA), or both (associated with Ext. Fig. 2 and 4), Experiment 3-BAT samples collected from WT and brown adipocyte-specific Alas1 KO (BAKO) mice (associated with Fig. 6, Ext. Fig. 9).

first: Dylan

last: Duerre

first: Julia

last: Hansen

first: Hoang

last: Bui


title: The kinase GSK-3 alters the repertoire of RNA binding proteins present on lipid metabolism transcripts leading to altered expression in the C. elegans nervous system

city: Bloomington

first: Ananya

last: Mahapatra

address: 316 E University St

institute: Indiana University, Bloomington

summary: Tissue-specific regulation of gene expression is essential for multicellular organisms, and RNA binding proteins play central roles in these molecular processes. To determine how the Caenorhabditis elegans RNA binding protein ADR-1 regulates tissue-specific gene expression, we profiled the binding targets of ADR-1 in neural cells and assessed the effects of ADR-1 RNA binding on neural gene expression. We identified a cohort of neural transcripts that facilitate lipid synthesis and are directly regulated by ADR-1 RNA binding. To identify cellular factors that influence ADR-1 binding, a forward genetic screen was performed, revealing that the serine/threonine protein kinase, glycogen synthase kinase-3 (GSK-3), inhibits ADR-1 binding to the cohort. Further investigation revealed that RNA binding protein VIG-1, promotes ADR-1 binding to the cohort in a GSK-3-dependent manner. Together, we reveal that interplay between kinases and RNA binding proteins regulates expression of lipid metabolism genes within neural cells, potentially impacting stress resistance and longevity.

overall_design: High throughput sequencing of neural ADR-1 immunoprecipitated samples and lysates in the presence and absence of adr-2; high throughput sequencing of neural cells isolated from adr-2(-) and adr-2(-);dsRBD1 mutant animals

first: Heather

middle: A

last: Hundley


title: The relationship between radiosensitivity and expression of micro RNAs in murine osteoblastic cell exposed to high dose rate alpha particles

city: Hirosaki

first: Satoru

last: Monzen

state: Aomori

address: Hon-cho 66-1

country: Japan

institute: Hirosaki University

summary: Osteoblastic cells in the bone marrow produce the bone matrix, participate in mineralization of bone and regulate the balance of calcium and phosphate ions in the developing bone. In addition, the radiosensitive hematopoietic stem/progenitor cells form a niche with osteoblastic cells to maintain immature state and self-renewal in the bone marrow microenvironment. In the field of radiotherapy, internal radiotherapy using radioactive isotopes that emit alpha particles is performed, and antitumor effect is expected by radioisotopes (such as 223-Ra) when the primary cancer cells metastasize to bone marrow. A major problem in this treatment is that the efficacy and side effects form show a high degree of individual variability, and the response of a patient is very difficult to predict. The mechanisms of the variability are not known, but it is possible that some contributing factors are the alpha radiation-induced modulation of osteoblastic cell activity. However, the characteristics of micro-RNA expressions in alpha-irradiated osteoblastic cells remain unclear. The purpose of this study is to clarify the pattern of miRNA expressions in osteoblast derived from freshly bone marrow cells exposed to alpha particle radiation. Murine bone marrow cells from both femurs were differentiated to osteoblastic cells using cell culture medium and exposed to 0-1 Gy of alpha radiation. The 24 miRNAs were significantly up- or down-regulated: mmu-let-7a-5p, mmu-let-7b-5p, mmu-let-7c-5p, mmu-miR-1224-5p, mmu-miR-125b-5p, mmu-miR-296-5p, mmu-miR-331-3p, mmu-miR-3473b, mmu-miR-3960, mmu-miR-3963, mmu-miR-466i-5p, mmu-miR-494-3p, mmu-miR-5126, mmu-miR-6360, mmu-miR-6366, mmu-miR-6378, mmu-miR-6769b-5p, mmu-miR-6937-5p, mmu-miR-7005-5p, mmu-miR-7042-5p, mmu-miR-7047-5p, mmu-miR-709, mmu-miR-8110, mmu-miR-8117). These results indicate that exposure of osteoblastic cells to alpha particles leads to unique expression of miRNAs and it may give the cell radiosensitive environment.

overall_design: The pattern of micro RNA expressions in osteoblastic cell derived from freshly bone marrows exposed to 0.5 Gy or 1 Gy alpha particles (0.22 Gy/min) was analyzed using micro RNA microarray. The data analysis had been performed triplicate samples.

first: Andrzej

last: Wojcik

first: Mitsuru

last: Chiba

first: Yasushi

last: Mariya


title: Histone H4 lysine 20 monomethylation is not a mark of transcriptional silencers

first: Stephen

last: Gisselbrecht

address: 77 Avenue Louis Pasteur, Rm 468

department: Division of Genetics, Department of Medicine

institute: Brigham and Women's Hospital

summary: Silencers are cis-regulatory elements that down-regulate the expression of target genes. While thousands of silencers have been identified experimentally, a predictive chromatin signature has not been found. H4K20me1 was previously reported to be enriched in silencers, but through reanalysis we found that this enrichment is marginal and was primarily driven by the biased selection of elements used in the screen. We generated H4K20me1 ChIP-seq in Drosophila S2 cells and confirmed that H4K20me1 does not mark transcriptional silencers but instead is associated with active transcription. Transcriptional silencers remain a poorly annotated and difficult to predict class of cis-regulatory elements.

sample_organism:
Drosophila melanogaster
Homo sapiens

platform_organism:
Drosophila melanogaster
Homo sapiens

overall_design: ChIP-seq of the histone modification H4K20me1 (with 3 different commercial antibodies) and of H3K27me3 in Drosophila S2 cells, with input chromatin provided as control, and ChIP-seq of H4K20me1 with 2 of those 3 commercial antibodies from human K562 cells, with input chromatin provided as control. ChIP-seq peaks are compared to published locations of elements shown to have silencer activity in these two cell types.

middle: S

first: Julian

last: Segert

first: Nikola

last: Mizgier

first: Karen

last: Adelman

first: Martha

middle: L

last: Bulyk


title: Adenoviral Vectored Neoantigen Vaccine Augments Hyperexpanded CD8+ T Cell Control of Tumor Challenge in Mice

status: Public on Feb 14 2025

first: Dan

last: Barouch

address: 3 Blackfan Cir

department: CVVR

institute: Beth Israel Deaconess Medical Center

summary: Neoantigens are promising immunogens for cancer vaccines and are traditionally delivered as adjuvanted peptide vaccines. Our goal was to understand how an adenoviral vectored neoantigen vaccine would induce tumor immunity compared to a peptide neoantigen vaccine. We generated adenovirus serotype 26 (Ad26) vaccines encoding MC38-specific neoantigens and compared them to an adjuvanted peptide MC38 neoantigen vaccine. The single-shot Ad26 vaccines induced greater neoantigen specific IFN- CD8+ T cell immune responses than the two-shot adjuvanted peptide vaccine in mice, and Ad26.VP22.7Epi also provided superior protective efficacy compared to the peptide vaccine following tumor challenge. Ad26.VP22.7Epi induced a robust immunodominant CD8+ T cell response against the Adpgk neoantigen, while the peptide vaccine induced lower responses against both Adpgk and Reps1 neoantigens. Tumor infiltrating lymphocytes (TILs) from both vaccine groups were analyzed using scRNA-seq and TCR-seq. Vaccinated mice showed increased CD8+ T cell infiltration, with the peptide vaccine inducing more infiltrating CD8+ T cells than the Ad26.VP22.7Epi vaccine. However, Ad26.VP22.7Epi induced CD8+ T cells showed more upregulation of T cell maturation, activation, and Th1 pathways compared to peptide vaccine induced CD8+ T cells, suggesting improved functional T cell quality. TCR-seq of these TILs also demonstrated that Ad26.VP22.7Epi generated larger T cell hyperexpanded clones compared to the peptide vaccine. These results suggest that the Ad26.VP22.7Epi vaccine led to improved tumor control compared with the peptide vaccine due to increased T cell hyperexpansion and functional activation. Our data suggest that future cancer vaccine development strategies should focus on inducing functional hyperexpanded CD8+ T cell responses and not only maximizing tumor infiltrating CD8+ T cell numbers.

overall_design: In this study, we generated several candidate Ad26 vectors and tested immune responses systemically and within the tumor microenvironment in mice following challenge with two different cancer cell lines. We then used scRNA-seq and TCR-seq to identify differences in tumor infiltrating lymphocytes (TILs) in mice vaccinated with Ad26 or long peptide vaccines to help future cancer vaccine design.

first: Gabriel

last: Dagotto

first: Alessandro

last: Colarusso

first: Robert

last: Patio

first: David

last: Li

first: Tochi

last: Anioke

first: Victoria

last: Giffin

first: Ruoran

last: Guan

first: Trisha

last: Anand

first: Esther

last: Mbiwan

first: Malika

last: Aid


title: TRAF3 regulates TFH cell differentiation during infections (RNA-Seq)

city: Iowa City

first: Noah

last: Butler

state: IA

address: 4-350 BSB, 51 Newton Road

department: Department of Microbiology and Immunology

institute: University of Iowa

summary: Comparison of transcriptome between WT and tamoxifen-inducible deletion of TRAF3 (iTraf3−/−)  SMARTA or PbTII TFH cells during the infections by LCMV armstrong and Plasmodium yoelii, respectively.

overall_design: WT and tamoxifen-inducible deletion of TRAF3 (iTraf3−/−) SMARTA or PbTII cells were transferred to recipient mice infected with LCMV armstrong or Plasmodium yoelii. On day 8 post-infection, WT or iTraf3−/− PbTII CXCR5+SLAMlo TFH cells were sorted into Trizol or RP10 medium. RNA-seq was performed to investigate the mechanism whereby TRAF3 regulates TFH cell differentiation.

first: Peng

last: Shao


title: Impact of CDK7 inhibition across a range of inhibitors in the context of heat shock

city: Boulder

first: Taylor

last: Jones

state: CO

address: 3415 Colorado Ave

institute: University of Colorado Boulder

summary: Here we evaluate the effect of CDK7 inhibition in the context of heat shock across three distinct inhibitors. Two inhibitors are Syros compounds SY-5609 and SY-5102. The final inhibitor is 3MBPP1 in a CDK7 analog sensitive cell line. We find that overall the same gene programs are impacted.

overall_design: RNA-seq in OV90 cells to evaluate the effects of CDK7 inhibition under HS stimulated conditions. There are 3 biological replicates for each condition. Inhibitors here include SY-5102 and 3MBPP1 (in an OV90 CDK7-analog sensitive cell line).

first: Jenna

last: Rimel


title: Establishment of bovine extraembryonic endoderm stem cells enables efficient blastoid formation

status: Public on Feb 12 2025

city: Gainesville

first: Zongliang

last: Jiang

state: FL

address: 2033 Mowry Rd

department: Animal Sciences

institute: University of Florida

summary: Extraembryonic endoderm stem (XEN) cells resemble the primitive endoderm of blastocyst and provide valuable alternatives for understanding hypoblast development and building stem cell-based embryo models. Here we report that a chemical cocktail (FGF4, BMP4, IL-6, XAV939, and A83-01) enables the de novo derivation and long-term culture of bovine XENs (bXENs) from blastocysts. Transcriptomic and epigenomic analyses confirm the identity of bXENs and reveal that bXENs resemble the hypoblast lineages of early bovine peri-implantation embryos. Furthermore, we show that bXENs maintain the stemness of expanded embryonic stem cells (EPSCs) and prevent them from differentiation. The chemical cocktail sustaining bXENs also facilitates the growth of epiblasts in developing pre-implantation embryos. Finally, 3D assembly of bXENs with bovine EPSCs and trophoblast stem cells (TSCs) enables efficient generation of blastoids that resemble blastocysts. Bovine XENs and blastoids established here represent accessible in vitro models for understanding embryogenesis and improving reproductive efficiency in domestic livestock.

overall_design: Cultured bovine XEN cells were used for RNA-seq and ATAC-seq

first: Zongliagn

first: Hao

last: Ming


title: The response to individual deletion of two transptional repressor genes in Hyphomicrobium denitrificans

city: Bonn

first: Christiane

last: Dahl

address: Meckenheimer Allee 168

department: Institut für Mikrobiologie & Biotechnologie

country: Germany

summary: Bacteria use different strategies to sense and respond to reduced sulfur compounds such as sulfide or thiosulfate. In the methylotrophic Alphaproteobacterium Hyphomicrobium denitrificans, which uses thiosulfate as an accessory electron donor, this involves two distinct but related ArsR-type transcriptional repressors, sHdrR and SoxR. Here, we focused on identifying target genes regulated by these repressors. This was achieved by generating individual deletions of each regulator gene, and performing comparative RNA-seq analysis.

overall_design: To identify SoxR and sHdrR-regulated genes, we performed an RNA-sequencing transcriptomic analysis of strains H. denitrificans ΔtsdA ΔsoxR and H. denitrificans ΔtsdA ΔshdrR. In a previous project (accession number GSE278992), we have identified thiosulfate-inducible genes in the reference strain H. denitrificans ΔtsdA. This strain lacks the tetrathionate-forming enzyme thiosulfate dehydrogenase. Here, we identified SoxR and sHdrR regulated genes by comparing expression levels in the reference strain vs. the repressor-deficient strains grown on methanol-containing medium under aerobic conditions in the absence of thiosulfate.

first: Jingjing


title: Self-organization of bovine blatocyst-like structure from three pluripotent stem cells

summary: Here we report the derivation and long-term culture of bovine extraembryonic endoderm cells (bXENs). We showed that bXENs enhance maintain the stemness of bovine ESCs and prevent them from differentiation. In the present of signaling cocktail sustaining bXENs, the growth and progression of epiblasts are also facilitated in the developing pre-implantation embryo. Through 3D assembly of bXENs with bovine ESCs and TSCs, we developed an improved bovine blastocyst like structure (bovine blastoid). The bovine blastoids established in this study represents accessible in vitro models for understanding early embryogeneisis and improving reproductive efficiency in livestock species.

overall_design: Bovine blastocyst-like structure generated from three cell lineages (bEPSC, bTSC, bXEN) and day8 bovine blasytocyst were collected for single nuclei isolation followed by snRNA-seq.


title: Defective Processing of Cytoplasmic and Chloroplast Ribosomal RNA in the Absence of Arabidopsis DXO1 [3'RNA-seq]

status: Public on Feb 11 2025

city: Warsaw

first: Michal

last: Krzyszton

address: Pawinskiego 5a

country: Poland

institute: Institute of Biochemistry and Biophysics, Polish Academy of Sciences

summary:
Decapping 5'-3' exoribonucleases from the DXO/Rai1 family are highly conserved among eukaryotes and exhibit diverse enzymatic activities depending on the organism. The biochemical and structural properties of the plant DXO1 differ from the yeast and animal counterparts, which is reflected in the in vivo functions of this enzyme. Here we show that Arabidopsis DXO1 contributes to the efficient processing of rRNA precursors in both nucleolar/cytosolic and chloroplast maturation pathways. However, the processing defects in DXO1-deficient plants do not depend on the catalytic activity of the enzyme but rely on its plant-specific N-terminal extension, which is responsible for the interaction with the mRNA cap methyltransferase RNMT1. Our RNA sequencing analyses show that the dxo1 mutation deregulates the expression of many ribosomal protein genes, most likely leading to inefficient or delayed pre-rRNA maturation. These phenotypes are partially suppressed by RNMT1 overexpression, suggesting that defective cap synthesis may be responsible, at least to some extent, for the observed effects.

Founded by National Science Centre Grant, Poland (UMO‐2018/29/B/NZ3/01980, UMO‐2021/40/Q/NZ1/00014, UMO‐2014/13/B/NZ3/00405, MINIATURA 2017/01/X/NZ1/00332, UMO‐2021/41/B/NZ3/02605).

overall_design: Using 3’RNA-seq, we analyzed transcriptomes (4 replicas) of Col-0, dxo1-2 mutant and a set of dxo1-2 complementation lines. Constructs encoding different versions of DXO1 protein were introduced to the dxo1-2 mutant (Kwasnik et al. 2019). These were wild type protein - wt (35S::DXO1), catalytic mutant - mut (35S::DXO1(E394A/D396A)), N-truncated protein - wt_dN (35S::DXO1(ΔN194)) and catalytically inactive N-truncated protein - mut_dN (35S::DXO1(ΔN194/E394A/D396A)).

first: Monika

last: Zakrzewska-Placzek

first: Aleksandra

last: Kwasnik

first: Anna

last: Golisz

first: Joanna

last: Kufel


title: Defective Processing of Cytoplasmic and Chloroplast Ribosomal RNA in the Absence of Arabidopsis DXO1 [RNA-seq]

overall_design: Transcriptome profiles of Col-0 wild type, and dxo1-2, xrn3-8 and double dxo1-2 xrn3-8 mutant plants using RNA-seq (three biological replicates). Plants growth, RNA-isolation, libraries preparation and Illumina sequencing were performed in parallel for Col-0 and all mutants. However, data for different mutants were submitted as separate GEO accession along with published manuscripts describing these mutants. Data for Col-0 and xrn3-8 mutant are part of the GSE95473. Data for dxo1-2 mutant are available upon GSE99600. Here, we provide data for double dxo1-2 xrn3-8 mutant. The supplementary file contains gene counts combined for all libraries sequenced in this experiment.


title: Circular RNA research in AML

status: Public on Feb 08 2025

city: Wuhan

first: Chunjiang

last: He

state: Hubei

address: Donghu Road 185

institute: Wuhan University

type: Non-coding RNA profiling by high throughput sequencing

summary: To investigate the effects of circRNA in AML, we collected the PBMC of AML patients and healthy volunteers and performed high-throughput sequencing, the results showed that circRNA_01 was differentially expressed in AML and normal samples. This suggests that circRNA_01 might play an important role in regulating AML. In order to find out the cohort of genes whose expression was regulated by circRNA_01, we constructed THP-1/U937 cell lines stably infected with lentiviral particles carrying control shRNA (sh_NC) or circRNA_01 shRNA. Subsequently, total RNA was isolated and then performed transcriptome RNA sequence

overall_design: we performed circular RNAs profiling of AML patients and healthy volunteers and mRNA profiling of THP-1/U937 cell lines infected with lentiviral particles carrying control shRNA (sh_NC) or circAFF2 shRNA

first: Xin

last: Dong


title: Ribo-seq analysis elucidates the dynamic translation landscape of yak ovarian tissue across different reproductive stages

status: Public on Feb 05 2025

city: Lanzhou

first: Liyan

last: hu

address: No. 335, Mogou Lane, Qilihe District

department: Key Laboratory of Yak Breeding in Gansu Province

institute: Lanzhou Institute of Animal Husbandry and Veterinary Pharmaceutics, Chinese Academy of Agricultural Sciences

summary: Translation, as a crucial link in the transmission of genetic information, plays an indispensable regulatory role in follicular development, ovulation, corpus luteum formation, and degeneration. In order to deeply analyze the molecular mechanisms of reproductive regulation at the translation level, our study focused on the ovarian tissues of yaks at different reproductive stages to analyze gene expression changes using Ribosome profiling and integrating RNA-seq data. Meanwhile, the small open reading frames (sORFs) of ovarian tissue was characterized, and the impact of the translation efficiency of focused genes on their sequence feature was analyzed. The results showed that over 80% of genes in the two groups exhibited inconsistent changes at the transcription and translation levels, indicated that the changes in genes at both levels were complex and not simply synergistic. Upon the pathway enrichment analysis, the signaling pathways involved in these differentially expressed genes were further clarified, such as the estrogen signaling pathway PI3K-Akt, MAPK, calcium signaling pathway and ovarian steroidogenesis. Further research has found that some genes related to ovarian function have inconsistent expression at two levels and exhibit dynamic changes in translation activity, including PALB2, EGR2, BMP7, SOX9, PIK3R2, and WNT2B. Besides, we identified 66 predicted translatable small open reading frames (sORFs) and also explored the impact of upstream ORFs (uORFs) on the translation efficiency of downstream major ORFs (mORFs). This study systematically revealed the gene expression characteristics of ovarian tissues in yaks at different reproductive stages from the perspective of translational level for the first time, providing a new perspective for a deeper understanding of the physiological mechanisms of yak reproduction.

overall_design: Construction of ribosomal footprint map of yak ovaries

last: Hu

first: Shaoke

last: Guo

first: Mengli

last: Cao

first: Xian


title: H3K36me3 Modification by SETD2 is Essential for Col11a2 and Sema3e Transcription to Maintain Dentinogenesis in Mice

status: Public on Feb 03 2025

first: 佳欣

last: 牛

state: 洪山区

address: State Key Laboratory of Oral & Maxillofacial Reconstruction and Regeneration, Key Laboratory of Oral Biomedicine Ministry of Education, Hubei Key Laboratory of Stomatology, School & Hospital of Stomatology

summary: Dentin is a major mineralized component of teeth generated by odontoblasts. Several types of histone methylation have been reported to play important roles in odontoblast differentiation and dentinogenesis. However, the role of methylation on histone 3 at lysine 36 (H3K36) remains enigmatic. Here we find a high expression of SETD2, a methyltransferase catalyzing the trimethylation of H3K36 (H3K36me3), in the odontoblast layer. In vitro knockdown experiments and in vivo observations of two conditional knockout mouse models reveal SETD2 is essential for odontoblast differentiation and dentinogenesis. Integrated analyses of RNA sequencing and Spike-in Cleavage Under Targets and Tagmentation sequencing data show SETD2 is crucial for both H3K36me3 occupancy on the loci of Col11a2 and Sema3e and their transcription. Further experiments verify COL11A2 and SEMA3E act upstream of AKT1 signaling, promoting odontoblastic differentiation. In vitro and in vivo activation of AKT1 using SC79 (an AKT activator) partially rescues the impaired odontoblast differentiation caused by Setd2 knockdown or deficiency. Therefore, our findings indicate that H3K36me3 mediated by SETD2 is essential for dentinogenesis through regulating the expression of Col11a2 and Sema3e and AKT1 signaling.

overall_design: Mouse dental papilla cells were transfected with Setd2 or scramble siRNAs, induced for odontoblastic differentiation, and then used for H3K36me3 CUT&Tag-seq experiments.

first: Jiaxin

last: Niu

first: Jing

last: Fu

last: Feng

first: Jiahao

last: Han

first: Zhi

last: Chen

first: Guobin

last: Yang

first: Guohua

last: Yuan


overall_design: Gene expression profiling analysis of RNA-seq data from the mouse dental papilla cells of the first mandibular molars in control mice and Dmp1-Cre; Setd2fl/fl mice at P6.

first: Hao


title: Microglial reprogramming enhances antitumor immunity and immunotherapy response in melanoma brain metastases

status: Public on Feb 06 2025

city: San Juan de Alicante

first: Francisco Javier

last: Rodríguez Baena

state: Alicante

address: Avenida Ramón y Cajal S/N Campus de San Juan

country: Spain

institute: Instituto de Neurociencias CSIC-UMH

summary: Melanoma is one of the solid tumor types with the highest risk of brain metastasis. However, the biology of melanoma brain metastasis and the contribution of the brain immune microenvironment to the responses to therapies remain insufficiently characterized. By using preclinical models and single-cell transcriptomics, we identify microglia, as critical regulators of melanoma biology in the brain. We show that activation of the Rela/NF-kB pathway in microglia promotes melanoma brain metastasis and that targeting this pathway elicits microglia reprogramming towards a pro-inflammatory phenotype that enhances anti-tumor immunity and reduces brain metastatic burden. Additionally, canonical and pro-inflammatory microglial markers in melanoma brain metastasis correlate with better responses to immune checkpoint inhibitors in patients and we show that Rela/NF-kB targeting improves responses to these therapies in the brain. Thus, we propose targeting Rela/NF-kB in activated microglia as a strategy to promote anti-tumor immunity and to improve the response to immune checkpoint inhibitors in melanoma brain metastasis.

overall_design:
Refer to individual Series. This SuperSeries is composed of the SubSeries listed below.

Figure 1: scRNAseq of a bulk sample 1014 NRAS*. Sample GSM8195555.

Figure 2: RNAseq. Samples GSM8195543 to GSM8195554.

Figures 4 - 6: scRNAseq of CX3CR1+/CD45+ samples from NRAS* MBM. Samples from GSM8706608 to GSM8706613.

first: Berta

last: Sánchez-Laorden

first: Ángel

last: Márquez-Galera


title: Low input ribosome profiling reveals stage-specific translational regulation during early C. elegans embryogenesis

city: Austin

first: Can

last: Cenik

state: Texas

address: Moffett Molecular Biology Building 2.122 2500 Speedway

department: Molecular Biosciences

institute: The University of Texas at Austin

summary: Translation of maternal mRNAs guides early embryonic development. C. elegans embryos exhibit an invariant cell lineage from the first division, offering a unique opportunity to trace the impact of post-transcriptional gene regulation on subsequent embryonic development. Although prior work has explored the translation of a select few transcripts, we lack a transcriptome-wide view of translation control during early embryogenesis. We developed a method for low-input ribosome profiling and combined it with low-input RNA sequencing to measure new protein translation during the first four cell cycles of C. elegans embryogenesis. Strikingly, our results show a poor correlation between RNA abundance and ribosome occupancy in this system compared to other model systems, emphasizing the predominant role of variation in translation efficiency in this system. We found numerous translationally regulated transcripts that had not been previously identified. Housekeeping genes involved in ribosome function and biogenesis were stably translated throughout the studied stages, while genes involved in mRNA processing and mRNA binding are dynamically regulated, consistent with roles in cell fate specification. A gain-of-function mutation in a known translational repressor protein, OMA-1, stalls the developmental program by preventing the translational remodeling that should occur at the 2-cell stage. These insights advance our understanding of the post-transcriptional regulatory mechanisms that shape the embryonic proteome and direct cell differentiation, and provide a valuable resource studies of C. elegans development.

overall_design: Ribosome profiling via isotachophoresis (RiboITP) and RNA-seq were performed on pooled Caenorhabditis elegans embryos at distinct early developmental stages (1-cell, 2-cell, 4-cell, and 8-cell) in both wild-type (N2) and OMA-1 mutant (TX20 strain, zu405 allele) backgrounds. For wild-type, samples were collected at all four stages, while the OMA-1 mutant was analyzed at 1-cell, 2-cell, and 4-cell stages. Each stage-specific sample consisted of pooled embryos to ensure adequate material for both RiboITP and RNA-seq analysis. The RiboITP and RNA-seq measurements enabled calculation of translational efficiency at each developmental stage, allowing comparison of translational regulation between wild-type and the OMA-1 mutant during early embryogenesis.

first: Yash

last: Shukla

first: Vighnesh

last: Ghatpande

first: Cindy

first: Daniel

last: Dickinson


title: Dynamic changes of immune cells in systemic lupus erythematosus with nephritis using longitudinal peripheral blood single-cell RNA-seq

status: Public on Feb 01 2025

city: Suwon

first: Seung-Jun

last: Paek

address: Worldcup-ro

department: Department of Physiology

institute: Ajou University School of Medicine

summary: Disease activity in patients with systemic lupus erythematosus (SLE) may fluctuate between flares and remissions, complicating effective disease management. The aim of this study is to detect early signs of cellular attributes responsible for flares and to understand dynamic changes in the immune system. Peripheral blood mononuclear cells were collected at different time points throughout the disease course from 6 patients with SLE (n = 19) and 32 healthy donors fromthe Asian Immune Diversity Atlas (AIDA), and subsequently underwent to generate single cell gene expression combined with T cell receptor (TCR) and B cell receptor (BCR) clonotypes data (10X Chromium 5’, TCR sequencing).

overall_design: Blood samples were collected from 6 patients before and after six instances of flare, and peripheral blood mononuclear cells (PBMCs) were isolated and cryopreserved.

first: Hye-Soon

last: Lee

first: Kyu-Tae

last: Kim

first: Sang-Cheol

last: Bae


title: The unique expression of non-coding microRNAs in radioresistant fraction of acute promyelocytic leukemia, HL60 cell

status: Public on Jan 31 2025

first: Satoru

last: Monzen

summary: A one of general treatments of patient with acute promyelocytic leukemia (APL) is performed a high dose rate of total-body irradiation therapy. However, the appearance of radioresistant cell that is rarely expressed induces a harmful outcome even performing radiotherapy. We have been established a radioresistant APL cell model (Resistant type), and some characteristics was clarified. But little is known as to the micro RNAs (miRs) expression profile in this cell. In the present study, we analyzed on the expression profile of miRs included in small RNAs using miRNA microarray, The focused miRs that is enabled to investigate by normalization was 1187 molecules. Among these miRs, 27 miRNAs (10 up-regulated miRs and 17 down-regulated miRs) in Res-HL60 showed a p-value < 0.05, Fold change > 1.5 or Fold change < 0.66 in comparison to Wt-HL60. In addition, five miRs were reproducible in RT-qPCR (miR-146a-5p, miR-30c-1-3p, miR-671-5p, miR-610, miR-3675-5p). miRNA-target gene regulation networks were looked for the five miRNAs using OmicsNet 2.0. Interestingly, the five miRNAs were Superoxide Dismutase 2 (SOD2) mRNA. These results were suggested that radioresistant leukemia cell acquires antioxidant system through from specific miRs endogenously.

overall_design: Human acute promyelocytic leukemia cell line HL60 was prepared after frequent irradiation (4Gy X-irradiation ×4 fractions per 4 weeks) with confirmed radiation-resistance characteristics.  The extracted total RNA in wild type cell and radiation-resistant cell were performed micro RNA microarray and calculate the expression of micro RNAs.

first: Hikoto

last: Sugiyama

first: Mitsuru

last: Chiba


title: LACTIPLANTIBACILLUS PLANTARUM STRENGTHENS THE INTESTINAL BARRIER: INVOLVEMENT OF THE ENDOCANNABINOIDOME

city: Quebec City

first: Frederic

last: Raymond

state: Quebec

address: 2440 Bd Hochelaga Suite 1710

department: École de nutrition

country: Canada

institute: Université Laval

summary: Probiotics have been suggested to ameliorate the function of the intestinal epithelial barrier and so have several mediators and receptors of the expanded endocannabinoid system, the endocannabinoidome (eCBome). Here we cocultured three live strains of Lactiplantibacillus plantarum with intestinal epithelial organoids to study their effects on the gut barrier function and the possible involvement of the eCBome in this effect. All three L.plantarum strains variously reduced the trans-epithelial permeability of intestinal organoids and promoted increased mRNA expression of several tight junction proteins and intestinal barrier proteins. Concomitantly, the three strains upregulated the expression of genes encoding biosynthetic enzymes (i.e., NapePLD, Abdh4, Gde1, Daglb) and receptors (i.e., Cnr1, Cnr2, Gpr55, and Ppara), while concurrently downregulating the expression of two essential catabolic enzymes (i.e. Faah and Naaa), involved in the signaling of several eCBome mediators known for their role in regulating the intestinal epithelial barrier. Selective inhibitors of eCBome mediator degrading enzymes FAAH and MAGL, i.e., URB597 and JZL184, increased N-acyl-ethanolamine (NAE) and 2-monoacylglycerol (2-MAG) levels, respectively, enhanced the expression of intestinal epithelial barrier genes and reduced the trans-epithelial permeability of organoids, as for L. plantarum strains. Interestingly, inflammation-induced trans-epithelial permeability in organoids was also reversed by both FAAH and MAGL inhibitors. We surmise that elevated endogenous levels of either NAEs or 2-MAGs promote improvement in small intestine trans-epithelial permeability and that L. plantarum strains may exploit this mechanism to promote these beneficial effects.

overall_design: Intestinal organoids exposed to inhibitors of enzymes part of the endocannabinoid system

first: Bénédicte

last: mAllam-Ndoul

first: Elena

last: Pulido Mateos

first: Frédéric

last: Bégin

first: Thomas

last: Mayer

first: Denis

last: Roy

first: Yves

last: Desjardins

first: Vincenzo

last: Di Marzo

first: Alain

last: Veilleux


title: Role of different epigenetic drugs in reshaping murine melanoma tumor microenvironment

city: Genova

first: Adriana

middle: Agnese

last: Amaro

state: Italy

address: Largo Rosanna Benzi 10

country: Italy

institute: IRCCS Ospedale policlinico San Martino

summary: To highlight the role of epigenetics in immune responses, we exploited a model of melanoma using B16F10 injected subcutaneously in syngeneic C57black6j mice and studied the effect of inhibitors of (i) BET bromodomain (OTX-015) (ii) DNA methyltransferase (DNMT) (guadecitabine) on in vivo tumour growth. OTX-015 showed significant in vivo effects in reducing tumour growth in our setting. Guadecitabine induced MHC class I expression on tumour cells, but the latter also increased immune checkpoint expression on cancer cells. Each epigenetic agent altered the tumour microenvironment (TME), mostly by reducing immune suppression, particularly of Tregs or MDSC, or both. OTX-015 and guadecitabine had strong effects, but in opposite ways. OTX-015 downmodulated antitumour responses, whereas guadecitabine remodeled the TME to be more immune-responsive. Epigenetic therapy may enhance immunotherapy, but specific schedules for treatments need to be studied for each drug. Among the drugs tested, guadecitabine seems to be the most promising drug for successful combination immunotherapy.

overall_design:
RNAseq gene expression analysis on B16F10 cell line tumor samples derived from mice treated or not with VORINOSTAT (3 treated 3 untreated)
Contributor:  EPigenetic Immune-oncology Consortium Airc (EPICA)

first: Valentina

last: Rigo

first: Rosaria

last: Gangemi

first: Francesco

last: Reggiani

first: Alessia

last: Parodi

first: Daniela

last: Fenoglio

first: Barbara

last: Banelli

first: Simonetta

last: Astigiano

first: Laura

last: Emionite

first: Stefania

last: Martini

first: Giuseppina

last: Conteduca

first: Tiziana

last: Altosole

first: Mariangela

last: Petito

first: Cecilia

last: Profumo

first: Guido

last: Ferlazzo

first: Gilberto

last: Filaci

first: Michele

last: Maio

first: Ulrich

last: Pfeffer

first: Michela

last: Croce


title: Dysregulation of adipocytokines regulated by hsa-miR-548ay-3p/PPARγ signaling pathway leaded to cardiac fibrosis

status: Public on Jan 30 2025

city: Changsha

first: Yubin

last: Chen

state: Hunan

address: Furong Road

department: Department of Cardiovascular Surgery

institute: Xiangya Hospital, Central South University

summary: Cardiac fibrosis is the key to the onset of atrial fibrillation (AF), but the mechanism is not fully understood. Our previous study found that fibrosis-related adipocytokines were dysregulated in epicardial adipose tissue (EAT) from patients with AF. In this study, we explored the factor that induce adipocytokines dysregulation and the mechanism underlying adipocytokines dysregulation-induced cardiac fibrosis.

overall_design: In part 1, cardiac fibroblasts(CFs) were treated with conditioned medium (CM) of adipocyte infected with hsa-miR-548ay overexpression lentivirus or vector lentivirus. In part 2, CFs were treated with TGF-beta1 and CM of adipocyte infected with hsa-miR-548ay inhibition lentivirus or vector lentivirus.

first: Fanyan

last: Luo


title: Vaccines combining slow release and follicle targeting of antigens increase germinal center B cell clonal diversity and clonal expansion

status: Public on Feb 04 2025

city: Cambridge

first: Darrell

last: Irvine

state: MA

address: 77 Massachusetts Ave

institute: Massachusetts Institute of Technology

summary: Vaccines incorporating slow release, multivalent antigen display, and/or immunomodulation through adjuvants have important roles in shaping the humoral immune response. Here, we analyzed mechanisms of action of a clinically relevant combination adjuvant strategy, where phosphoserine (pSer)-tagged immunogens bound to aluminum hydroxide (alum) adjuvant (promoting prolonged antigen release to draining lymph nodes) are combined with a potent saponin nanoparticle adjuvant termed SMNP (which alters lymph flow and antigen entry into lymph nodes). When employed with a stabilized HIV Env trimer antigen in mice, this combined adjuvant approach promoted substantial enhancements in germinal center (GC) and antibody responses relative to either adjuvant alone. Using scRNA-seq and scBCR-seq, we found that the alum-pSer/SMNP combination augmented the clonal expansion and diversity of GC B cell repertoire, coincident with increased expression of positive selection markers and proportion of S-phase GC B cells. To gain insight into the source of these changes in the GC response, we analyzed antigen biodistribution and structural integrity in draining lymph nodes and found that the combination adjuvant approach, but not alum-pSer delivery or SMNP alone, promoted accumulation of intact antigen on follicular dendritic cells (FDCs), reflecting an integration of the effects by slowing antigen release and altering lymph node uptake. Ablating the Cr1/2 complex on FDCs significantly hampered antigen-specific GC response, corroborating enhanced FDC decoration as the key mechanism of improved vaccine response. These results demonstrate how adjuvants with complementary mechanisms of action impacting vaccine biodistribution and kinetics can synergize to enhance humoral immunity.

overall_design: On day 14 post-immunization, antigen-specific germinal center B cells were isolated from inguinal lymph nodes using fluorescent-activated cell sorting. Sorted cells were immediated loaded onto SeqWell arrays for scRNA-seq.

first: Yiming J.

last: Zhang

first: Duncan M.

last: Morgan

first: J. Christopher

last: Love

first: Darrell J.