title: Breast Cancer Stem Cells Regulated Metabolism Remodeling of Bulk Tumor Drives Immune Evasion

city: hefei

first: linlin

last: yan

state: anhui

address: No. 96, Jinzhai Road

institute: University of Science and Technology of China

summary: Reprogramming of energy metabolism plays pivotal roles in cancer progression and immune surveillance. Here, we demonstrated that hypoxic cancer cell secretome and breast cancer stem cell (BCSC) secretome similarly promoted the global reprogramming of metabolic pathways, particularly glycolysis, in normoxic cancer cells. Screening of BCSC secretome identified MIF as a pivotal factor potentiating glycolysis via transcriptional activation of ALDOC expression by MYC. Targeting MIF attenuated glycolysis as well as impaired tumor growth and metastasis. Cell-intrinsic MIF depletion in breast cancer augmented intratumoral cytolytic CD8+ T cells and pro-inflammatory macrophages while decreased tumor-associated neutrophils. Targeting MIF improved anti-PD-L1 immune checkpoint therapy of triple-negative breast cancer cells. Hence, this study presented a paradigm of heterocellular metabolic communication in modulating tumor energy metabolism and immune surveillance, and thereby proposes strategies to ameliorate immunotherapy in breast cancer.

overall_design: Samples consisting of three dissociated tumors were sorted for the CD45+ immune cell population to scRNA-seq data analysis, and each group was represented by two biologically independent samples from wild-type and  Mif-depleted 4T1 tumors derived BALB/c mice.

first: Linlin

last: Yan

first: Mingming

last: Wu

first: Tao

last: Zhu


title: Dynamics of the UV response of the foliar pathogen Fusarium mangiferae

status: Public on Jan 30 2024

city: Rehovot

first: Shay

last: Covo

state: Central District

address: POB 12

department: Plant Pathology and Microbiology

country: Israel

institute: Hebrew University of Jerusalem

type: Expression profiling by high throughput sequencing

summary: We previously revealed developmental regulation over UV repair capacity in the soilborne pathogen Fusarium oxysporum (F. oxysporum). We demonstrated that photoreactivation assisted survival and repair were high during early stages of germination and low at later stages. In agreement, the expression of the UV specific repair genes photolyase and uvde showed opposite trends. While early on photolyase is induced and uvde is reduced, the trend is reversed later. Here, we tested the dynamic of transcription of photolyase, UV survival, repair capacity, and UV induced mutagenesis in the foliar pathogen Fusarium mangiferae. Unlike F. oxysporum, neither did we observe developmental control over photoreactivation dependent repair nor the changes in gene expression of phr1 and uvde throughout the experiment. Similarly, photo-reactivation assisted reduction in UV induced mutagenesis was similar throughout the development of F. mangiferae but fluctuated during the development of F. oxysporum. To generate hypotheses regarding the recovery of F. mangiferae after UV exposure, an RNAseq analysis was performed after irradiation at different timepoints. The most striking effect of UV on F. mangiferae was developmental-dependent induction of translation related genes. We further report a complex dynamic response that involves translation, cell cycle and lipid biology related genes.

overall_design: Germinating condia of Fusarium mangiferae strain MRC7560 at either 8 or 14 h post innoculation. 8 h post innoculation spores were irradiated with either 50 or 200J/m2 and allowed to recover for additional 30 m. 14 h post innoculation spores were irradiated with either 50 or 200J/m2 and allowed to recover for additional 0, 30, or 60 m. 8 h or 14 h untreated populations were used as a control. Each treatment includes three biological replicates.

first: Shira

last: Milo


title: Enhancer activation by the oncogenic herpesvirus KSHV tethering protein LANA

status: Public on Jan 31 2024

city: Nashville

first: Jeff

last: Rathmell

state: TN

address: 1211 Medical Center Drive

department: PMI

country: USA

institute: Vanderbilt University Medical Center

summary: Higher order genome structure influences the transcriptional regulation of cellular genes through the juxtaposition of regulatory elements, such as enhancers, close to promoters of target genes. While enhancer activation has emerged as an important facet of Kaposi’s sarcoma-associated herpesvirus (KSHV) biology, mechanisms controlling enhancer-target gene expression remain obscure. Here, we discover that the KSHV genome tethering protein latency-associated nuclear antigen (LANA) potentiates enhancer-target gene expression in primary effusion lymphoma (PEL), a highly aggressive B cell lymphoma causally associated with KSHV. Genome-wide analyses demonstrate increased levels of enhancer RNA transcription as well as activating chromatin marks at LANA bound enhancers. 3D-genome conformation analyses identified genes critical for latency and tumorigenesis as targets of LANA occupied enhancers, and LANA depletion results in their downregulation. These findings reveal a previously unknown mechanism in enhancer-gene coordination and describe a role through which the main KSHV tethering protein regulates essential gene expression in PEL.

overall_design: This experiment employ HiChIP, CUT&RUN, ATACme and RNAseq profile systematically how KSHV encoded protein LANA regulate enhancer activity and gene expression in latent TREx-BCBL1-RTA and BC3 cells.

first: Xiang

last: Ye

first: Tony

middle: Z

last: Chen

first: Lindsey

middle: N

last: Guerin

first: John

last: Karijolich


title: Xrp1 governs the stress response program to spliceosome dysfunction

city: Cologne

first: Mirka

last: Uhlirova

state: NRW

address: Joseph-Stelzmann Str. 26

department: Institute for Genetics

country: Germany

institute: CECAD Research Center  Cologne

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Unperturbed pre-mRNA splicing is crucial for proper expression of eukaryotic genes, but also maintenance of genome integrity. Loss of splicing factors is associated to DNA damage arising through different mechanisms, among which is the accumulation of mutagenic DNA/RNA hybrids (R-loops). RNase H1 recognizes and degrades the RNA moiety within R-loops, and is a commonly used marker for R-loops. Ecdysoneless protein was previously shown to participate in pre-mRNA splicing through its association with the U5 snRNP. In this study we have performed in vivo Targeted DamID using the Dam-RNase H1 construct to map the localization of R-loops throughout the Drosophila genome in control and Ecd-deficient Drosophila wing imaginal discs.

overall_design: Low-level expression of UAS-LT3-Dam (noise control) and UAS-LT3-Dam-RNase H1 (binding signals) was performed in wing imaginal discs (WDs) of Drosophila third instar larvae in control animals and temperature-sensitive ecd1 mutants. Heat-sensitive tub-Gal80 prevented the expression of the Dam constructs at 25 °C at which the larval tissues were developing. The larvae were heat shocked at 29 °C for 24 hours prior to dissection on day 7 after egg laying. The heat shock simultaneously enables the transcription of the Dam constructs, and abolishes the function of the mutant ecd1 protein. The WDs were dissected, processed according to established Targeted DamID protocols and the derived material sequenced in three biological replicates.

first: Dimitrije

last: Stankovic

first: Luke

middle: S

last: Tain


summary: We investigated the changes in gene expression in response to reduced levels of Ecdysoneless (Ecd) in the developing wing disc of Drosophila. To do this we utilized the GAL4/UAS system to express UAS-ecd-RNAi in the nubbin-domain (nub-Gal4, UAS-mRFP, UAS-ecd-RNAi) of the wing disc and used Nub-Gal4/+ wing discs as controls.  We quantified transcriptomic expression analysis from four biological replicates for both experimental groups

overall_design: mRNA profiles were generated by single-end, total RNA Illumina sequencing (HiSeq2500) from wandering L3 stage wing discs of nub-Gal4, UAS-mRFP, UAS-ecd-RNAi and nub-Gal4, UAS-mRFP Drosophila larvae.


title: RNA-sequencing of PDX from small round cell sarcomas

city: Houston

first: Danh

last: Truong

address: 1515 Holcombe Blvd

institute: MD Anderson Cancer Center

summary: The goal of this study was to profile the transcriptome of different small round cell sarcomas from patient-derived xenografts. We derive unique gene expression associated with each of the fusion proteins driving these sarcomas.

overall_design: We obtained fresh-frozen tissue from the PDXs from small round cell sarcomas including Ewing sarcoma, CIC-DUX4 sarcoma and desmoplastic small round cell tumors.  After removal, RNA was extracted and processed for RNA-sequencing.

first: Joseph

last: Ludwig


title: Sorghum SbGhd7 is a major regulator of floral transition and directly represses genes crucial for flowering activation

city: Upton

first: Meng

last: Xie

state: NY

address: 50 Bell Ave

institute: Brookhaven National Laboratory

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

overall_design: Refer to individual Series


summary: Sorghum is multipurpose crop worldwide serving as food, feed, and feedstock for biofuels, whose floral transition and vegetative growth heavily depend on photoperiod. Although multiple sorghum maturity loci (Ma1-Ma6) have been associated with photoperiod sensitivity in previous QTL studies, the underlying molecular mechanisms remain poorly understood. By functional characterizing sorghum SbGhd7 (Ma6) and integrating RNA-seq analysis of Ghd7 overexpression sorghum, ChIP-seq analysis of SbGhd7 binding sites in protoplasts and molecular studies, we discovered that SbEhd1 and SbFT10 are the direct targets of SbGhd7. SbGhd7 is a transcriptional repressor and inhibits florigen-induced floral transition by repressing SbEhd1 and SbFT10 expression.

overall_design: Transient chromatin immunoprecipitation DNA-sequencing (transient ChIP-seq) for SbGhd7 in sorghum mesophyll protoplasts.


title: BCLXL PROTAC degrader DT2216 targets secondary plasma cell leukemia addicted to BCLXL for survival

city: Nantes

first: David

last: Chiron

address: 8 quai Moncousu

country: France

institute: CRCI2NA, INSERM_U1307, CNRS_6075, Nantes_Université

summary: We demonstrated that secondary Plasma Cell Leukemia, addicted to BCLXL, a small but a very challenging group of patients, could benefit from DT2216, underpinning its clinical evaluation.

overall_design: RNA CD138 positive human plasma cells  were extracted with RNeasy Mini Kit (Qiagen) and sequenced on 3’ends.

first: Ophélie

last: Champion

first: Alana

last: Soler

first: Sophie

last: Maïga

first: Celine

last: Bellanger

first: Catherine

last: Pellat-Deceunynck

first: Alexis

last: Talbot

first: Cyrille

last: Touzeau

first: Martine

last: Amiot

first: Patricia

last: Gomez-Bougie


title: Protein-truncating variants in BSN are associated with severe adult-onset obesity, type 2 diabetes and fatty liver disease

status: Public on Jan 29 2024

city: Cambridge

first: Brian

middle: Yee Hong

last: Lam

address: Box 289 Addenbookes Hospital, Hills Road

department: Metabolic Research Laboratories

country: United Kingdom

institute: University of Cambridge

summary: Obesity is a major risk factor for many common diseases and has a significant heritable component. While clinical and large-scale population studies have identified several genes harbouring rare alleles with large effects on obesity risk, there are likely many unknown genes with highly penetrant effects remaining. To this end, we performed whole exome-sequence analyses for adult body mass index (BMI) in up to 587,027 individuals. We identified rare, loss of function variants in two genes – BSN and APBA1 – with effects on BMI substantially larger than well-established obesity genes such as MC4R. One in ~6500 individuals carry a heterozygous protein truncating variant (PTV) in BSN, which confers a 6.6, 3.7 and 3-fold higher risk of severe obesity (BMI >40kg/m2), non-alcoholic fatty liver disease and type 2 diabetes, respectively. Rare PTVs in BSN were found in three patients with severe early onset obesity, but in contrast to most other obesity-related genes, rare variants in BSN and APBA1 were not associated with normal variation in childhood adiposity. Furthermore, BSN PTVs magnified the influence of common genetic variants associated with BMI, with a common polygenic score exhibiting an effect on BMI twice as large in BSN PTV carriers than non-carriers. Finally, we explored the plasma proteomic signatures of BSN PTV carriers as well as the functional consequences of BSN deletion in human iPSC-derived hypothalamic neurons. These approaches highlighted a network of differentially expressed genes that were collectively enriched for genomic regions associated with BMI, and suggest emerging roles for neurodevelopment, neurogenesis, and altered neuronal oxidative phosphorylation in the etiology of obesity.

overall_design: To investigate the difference in iPSC derived hypothalamic neurons between BSN+/- and WT

first: Maria

last: Chukanova

first: Yajie

last: Zhou

first: Katherine

middle: A

last: Kentistou

first: Zammy

last: Fairhurst-Hunter

first: Anna

middle: M

last: Sieger

first: Raina

last: Jia

first: Georgina

middle: K

last: Dowsett

first: Eugene

middle: J

last: Gardner

last: Lawler

first: Felix

middle: R

last: Day

first: Lena

last: Kaisinger

first: Yi-Chun

middle: L

last: Tung

middle: Y

first: Hsiao-Jou

middle: C

last: Chen

first: Quanli

last: Wang

first: Jaime

last: Berumen-Campos

first: Pablo

last: Kuri-Morales

first: Roberto

last: Tapia-Conyer

first: Jesus

last: Alegre-Diaz

first: Inês

last: Barroso

first: Jonathan

last: Emberson

first: Jason

last: Torres

first: Rory

last: Collins

first: Danish

last: Saleheen

last: Smith

first: Dirk

last: Paul

first: Florian

last: Merkle

first: I

last: Farooqi

first: Nick

last: Wareham

first: Slavé

last: Petrovski

first: Steve

last: O'Rahilly

first: Ken

last: Ong

first: Giles

last: Yeo

first: John

last: Perry


title: Near-cognate tRNAs increase the efficiency and precision of pseudouridine-mediated readthrough of premature termination codons [PRAISE]

city: Beijing

first: Wenqing

last: Liu

email: puyulwq@163.com, liuwq21@gamils.com

address: No. 5 Summer Palace Road, Haidian District, Beijing

country: China

institute: Peking University

summary: Nonsense mutations, responsible for ~11% of genetic diseases, create premature stop codons (PTCs) and lead to truncated and often non-functional proteins. Recently, programmable RNA pseudouridylation has emerged as a new type of RNA base editor to suppress PTCs. However, current methods suffer from low efficiency and limited precision. Here, we develop RESTART v3, an updated version of RESTART, which utilizes near-cognate tRNAs to improve the readthrough efficiency of pseudouridine-modified PTCs. We show an average of ~5-fold higher editing efficiency than RESTART v2 in cultured cells, currently the most active RNA pseudouridylation tool to mediate PTC readthrough. Moreover, RESTART v3 achieves functional PTC readthrough in disease cell models of cystic fibrosis and Hurler syndrome. Furthermore, RESTART v3 enables accurate incorporation of the original amino acid for nearly half of the PTC sites, considering the naturally occurring frequencies of sense to nonsense codons. In line with the benign off-target effect of RESTART, RESTART v3 does not change the coding information nor the expression level of transcripts with off-target editing; except for the specifically overexpressed tRNA molecule, it does not alter the expression level of endogenous tRNA pool. Overall, RESTART v3 represents an enhanced RNA base editor with increased efficiency and accuracy.

overall_design: we performed transcriptome-wide quantitative Ψ detection in HEK293T treated or untreated with RESTART v3 to evaluate the off-target edits


title: Near-cognate tRNAs increase the efficiency and precision of pseudouridine-mediated readthrough of premature termination codons

type: Non-coding RNA profiling by high throughput sequencing

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: ELAPOR1 induces the classical/progenitor subtype and contributes to reduced disease aggressiveness through metabolic reprogramming in pancreatic cancer

city: Bethesda

first: Stefan

last: Ambs

state: MD

address: Building 37, Room 3050B

department: CCR

institute: NCI

summary: Pancreatic cancer is a heterogenous disease and consists of distinct subtypes. Here, we searched for candidate suppressor genes of the basal-like/squamous subtype, a more aggressive molecular subtype of pancreatic ductal adenocarcinoma (PDAC). Through an integrated transcriptome analysis, we identified the ELAPOR1/KIAA1324, as being downregulated in the basal-like/squamous PDAC using a discovery and validation approach. ELAPOR1 downregulation associated with decreased patient survival. The goal of this study was aimed to identify ELAPOR1 induced molecular signatures in PDAC.

overall_design: RNA was isolated from PDAC cell lines (n = 8 Panc 10.05_Ctrl vs ELAPOR1) using TRIzol method. The mRNA profiles of the above human PDAC cell lines were generated by deep sequencing using Illumina NextSeq.

first: Yuuki

last: Ohara

first: Huaitian

first: Amanda

last: Craig

first: Shouhui

last: Yang

first: Paloma

last: Moreno

first: Tiffany

middle: H

last: Dorsey

first: Helen

last: Cawley

first: Azadeh

last: Azizian

first: Jochen

last: Gaedcke

first: Michael

last: Ghadimi

first: Nader

last: Hanna

first: S

middle: P

last: Hussain


title: Decoding the Universal Human Chromatin Landscape through Teratoma-Based Profiling [ChIP-seq]

city: Detroit

first: Benjamin

last: Kidder

state: MI

address: 4100 John R St

department: Oncology

institute: Wayne State University

summary: Teratoma formation is key for evaluating differentiation of human pluripotent stem cells into embryonic germ layers and serves as a model for understanding stem cell differentiation and developmental processes. Its potential for insights into epigenome and transcriptome profiling is significant. This study integrates the analysis of the epigenome and transcriptome of hESC-generated teratomas, comparing transcriptomes between hESCs and teratomas. It employs cell type-specific expression patterns from single-cell data to deconvolve RNA-Seq data and identify cell types within teratomas. Our results provide a catalog of activating and repressive histone modifications, while also elucidating distinctive features of chromatin states. Construction of an epigenetic signature matrix enabled the quantification of diverse cell populations in teratomas and enhanced the ability to unravel the epigenetic landscape in heterogeneous tissue contexts. This study also includes a single cell multiome atlas of expression (scRNA-Seq) and chromatin accessibility (scATAC-Seq) of human teratomas, further revealing the complexity of these tissues. A histology-based digital staining tool further complemented the annotation of cell types in teratomas, enhancing our understanding of their cellular composition. This research is a valuable resource for examining teratoma epigenomic and transcriptomic landscapes and serves as a model for epigenetic data comparison.

overall_design: ChIP-Seq of human teratomas generated from H1 hESCs


title: Centromere Pairing Enables Correct Segregation of Meiotic Chromosomes

status: Public on Jan 28 2024

city: Oklahoma City

first: DEAN

last: DAWSON

state: OK

address: 825 NE 13th St.

department: Cell Cycle and Cancer Biology Research Program

institute: Oklahoma Medical Research Foundation

summary: Comparison of genome binding/occupancy profiling of the cohesin subunit Rec8 by high throughput sequencing in WT and ctf19-9A strains in meiotic prophase I.

overall_design: ChEC-seq of Rec8 in WT and ctf19-9A strains is made in 3 biological replicates that are arrested in meiotic prophase I with ndt80 deletion.

first: Asli

last: Sadli

first: Bartosz

last: Rapacz

first: Dean

last: Dawson


title: Sister chromatid cohesion halts DNA loop expansion

city: Toulouse

first: christophe

last: chapard

address: 118 route de Narbonne

department: CBI - MCD

institute: CNRS

summary: Eukaryotic genomes are folded into DNA loops mediated by SMC complexes, like cohesin, condensin and Smc5/6. This organization regulates different DNA related processes along the cell cycle such as transcription, recombination, segregation and DNA repair. During G2/M phases, SMC complexes mediated DNA loops coexist with cohesin complexes involved in sister chromatid cohesion (SCC).  It remains unknown whether SCC and DNA loop expansion influence each other and if they cooperate to regulate DNA-related processes. Here we show that SCC is indeed a barrier to DNA loop expansion mediated by cohesin in G2.

overall_design: Examination of the interplay between DNA loop extrusion mediated by cohesin and sister chromatid cohesion in yeast using HiC and ChIP-seq methods, yeast mutants, cell synchonisation and protein depletion and or expression

first: Bastié

last: Nathalie

first: Chapard

last: Christophe

first: Henri

last: Mboumba

first: Koszul

last: Romain

first: Beckouët

last: Frédéric


first: Meng

last: Xie

overall_design: Comparative gene expression profiling analysis of RNA-seq data for leaf tissues from 4-week-old empty vector control and SbGhd7 overexpression (SbGhd7 OE) sorghum plants.


title: Response to Bruton's tyrosine kinase inhibitors in aggressive lymphomas linked to chronic selective autophagy

first: David

last: Huang

state: Maryland

address: 9000 Rockville Pike

institute: National Cancer Institute

summary: We uncovered an autophagic pathway regulating survival in ABC DLBCL upon BTK inhibition using genome wide CRISPR screening.  To investigate the mechanism of action of this unique form of autophagy, we performed RNA-seq on TMD8 cells knocked out for various ATG genes that either showed resistance to BTK inhibitors (ATG9A, ATG101, ATG14, RB1CC1, WIPI2),  those that did not (ATG5, ATG7, ULK1 and 2 DKO, or non-targetingcontrol), or TMD8 cells knocked out for the known NF-kB negative regulator TNFAIP3.  Cells were treated for 24 hours with the BTK inhibitor, acalabrutinib, and gene expression signatures were calculated from normalized RNA-seq reads.  We found NF-kB gene expression signatures to be downregulated in control cells upon BTK inhibition, but not in ATG KO cells conferring BTK inhibitor resistance.

overall_design: RNA sequencing was performed on TMD8 cells treated with vehicle (DMSO) or 25 nM acalabrutinib for 24 hours.

first: James

middle: D

last: Phelan

first: Sebastian

last: Scheich

first: Louis

last: Staudt

first: Thomas

last: Oellerich


title: The effects on the deficient of PRMT1 on gene expression for the acquisition of chemoresistance in triple negative breast cancer cells.

city: Tokyo

first: Takehiro

last: Yamamoto

address: Shinanomachi 35, Shinjuku-ku, tokyo, Japan

department: Biochemistry

country: Japan

institute: Keio University

summary: In cancer cells, metabolic programming is regulated by gene expression and/or by post-translational modifications of metabolic enzymes. The correlation between metabolic characteristics of breast cancer and their drug sensitivity has attracted much attention. Here, we investigated whether PRMT1, a major arginine methyltransferase, affects the expression levels of metabolic enzymes by mRNA expression profiling of wild-type and PRMT1 knockout cells of the breast cancer cell line MDA-MB-468 cells. The results strongly suggest that PRMT1-mediated breast cancer drug sensitivity is regulated by post-transcriptional rather than transcriptional regulation.

overall_design: To investigate the function of PRMT1 for metabolic remodeling, we established PRMT1-knocking out cells in breast cancer cell line, MDA-MB-468.

first: Makoto

last: Suematsu


title: Mediator Complex Subunit 1 Promotes Differentiation and Function of a Tumor-Specific T Regulatory Cell Population

city: Chicago

first: Deyu

last: Fang

address: 303 E Chicago Avenue

institute: Northwestern University

summary: T regulatory (Treg) cells engage specific transcriptional programs depending on the environmental and immunologic context. The Tregcell-intrinsic mechanisms differentially regulating these programs between tumors and other inflammatory settings remain to be determined. Here we show that Mediator complex subunit 1 (MED1) is required for Treg cell function specifically in the context of the tumor microenvironment. Treg cell-specific deletion of MED1 did not predispose mice to autoimmunity or affect Treg cell function in models of colitis, experimental autoimmune encephalomyelitis, or influenza infection. Surprisingly, mice with Treg cell-specific deletion of MED1 showed reduced growth in a wide range of tumors using both syngeneic and spontaneous cancer models. Single-cell RNA sequencing revealed that MED1 is required for the terminal differentiation of effector Treg cells in the tumor. Suppression of these terminally differentiated Treg cells, high in Tigit, Tnfrsf18, Tnfrsf9, Icos, and Ccr8, was sufficient for eliciting antitumor immunity. MED1 did not regulate chromatin accessibility or expression of effector transcription factors in intratumoral Treg cells but did promote transcription of genes involved in effector differentiation. Lastly, we found that both human and murine Treg cells experienced divergent paths of differentiation by comparing cells in tumors and matched tissues with non-malignant inflammation to demonstrate this subset's specificity to tumors. Collectively, we identified a novel pathway regulating the differentiation of a Treg cell effector subset specific to tumors and demonstrated that suppression of a subset of Treg cells is sufficient for promoting antitumor immunity in the absence of autoimmune consequences.

overall_design: Treg cells were isolated from different tissues and subjected to sequencing.

first: Shuvam

last: Chaudhuri

first: Dongmei

last: Wang

first: Samuel

middle: E

last: Weinberg


title: Eph-ephrin signaling couples endothelial cell sorting and arterial specification

city: Münster

first: Kai

last: Kruse

state: North Rhine-Westphalia

address: Roentgenstr. 20

department: Bioinformatics Core Facility

institute: Max Planck Institute for Molecular Biomedicine

summary: Cell segregation allows the compartmentalization of cells with similar fates during morphogenesis, which can be enhanced by cell fate plasticity in response to local molecular and biomechanical cues. Endothelial tip cells in the growing retina, which lead vessel sprouts, give rise to arterial endothelial cells and thereby mediate arterial growth. Here, we have combined cell type-specific and inducible mouse genetics, flow experiments in vitro, single cell RNA sequencing and biochemistry to show that the balance between ephrin-B2 and its receptor EphB4 is critical for arterial specification, cell sorting and arteriovenous patterning. At the molecular level, elevated ephrin-B2 function after loss of EphB4 enhances signaling responses by the Notch pathway, VEGF and the transcription factor Dach1, which is influenced by endothelial shear stress. Our findings reveal how Eph-ephrin interactions integrate cell segregation and arteriovenous specification in the vasculature, which has potential relevance for human vascular malformations caused by EPHB4 mutations.

overall_design: HUVEC cells treated with control, EPHB4 and EFNB2 siRNAs. Two 6-well replicates were pooled for each sample.

first: Jonas

last: Stewen

first: Anca

middle: T

last: Godoi-Filip

first: Zenia

last: Zenia

first: Hyun-Woo

last: Jeong

first: Susanne

last: Adams

first: Frank

last: Berkenfeld

first: Martin

last: Stehling

first: Kristy

last: Red-Horse

first: Ralf

first: Mara

last: Pitulescu


title: Prolonged T–DC macro-clustering in lymph nodes initiates site-specific Th2 differentiation

city: Seattle

first: Michael

last: Gerner

state: WA

address: 750 Republican St

department: Immunology

institute: University of Washington

summary: T helper 2 (Th2) responses protect against pathogens while also driving allergic inflammation, yet how large-scale Th2 responses are generated in tissue context remains unclear. Here, we used quantitative imaging to investigate early Th2 differentiation within lymph nodes (LNs) following cutaneous allergen administration. Contrary to current models, we observed extensive activation and ‘macro-clustering’ of early Th2 cells with migratory type-2 dendritic cells (cDC2s) generating specialized Th2-promoting microenvironments. Macro-clustering was integrin mediated and promoted localized cytokine exchange among T cells to reinforce differentiation, which contrasted behavior during Th1 responses. Unexpectedly, formation of Th2 macro-clusters was dependent on the site of skin sensitization. Differences between sites were driven by divergent activation states of migratory cDC2 from different dermal tissues, with enhanced costimulatory molecule expression by cDC2 in Th2-generating LNs promoting prolonged T cell activation, macro-clustering, and cytokine sensing. Thus, generation of dedicated Th2 priming microenvironments through enhanced costimulatory molecule signaling initiates Th2 responses in vivo and occurs in a skin site-specific manner.

overall_design: To investigate transcriptional differences in dendritic cells between auricular (Au) and brachial (Br) draining lymph nodes (LNs), we immunized mice in the ear or forepaw with papain and EaGFP. LN tissues were harvested 48 hours later and mechanically disrupted and subject to digestion in PBS with 10% fetal bovine serum (FBS) with DNase I (100µg/mL; Sigma), Dispase II (800µg/mL; Sigma), and Collagenase P (200µg/mL; Sigma) at 37°C shaking at 150rpm for 30 minutes with periodic manual disruption. We then stained and FACS sorted cells from three populations of EaGFP+ MHC-II-hi CD11c-int antigen bearing cDC2 (CD301b+CD11b+, CD301b-CD11b+, and CD301b-CD11b-) from Au or Br draining LNs 48 hours post-immunization. We also sorted non-antigen bearing DCs of the same populations from naive mice. Cells were sorted from pooled dLNs from 3 individual mice for each group. 500 of each cDC2 cell type was sorted on an Aria III (BD Biosciences) directly into reaction buffer from the SMART-Seq v4 Ultra Low Input RNA Kit for Sequencing (Takara), and reverse transcription was performed followed by PCR amplification to generate full length amplified cDNA.

first: Miranda

last: Lyons-Cohen

first: Elya

last: Shamskhou


title: Decoding the Universal Human Chromatin Landscape through Teratoma-Based Profiling [Multiome]

first: Benjamin

last: Kidder

overall_design: Single-Nuclei Multiome (ATAC + Gene Expression) Sequencing of teratomas generated from H1 hESCs


title: Whole genome bisulfite sequencing identifies stage- and subtype-specific DNA methylation signatures in pancreatic cancer

city: Davis

first: Sarah

state: CA

address: 1 Shields Ave

institute: University of California, Davis

type: Methylation profiling by high throughput sequencing

summary: In pancreatic ductal adenocarcinoma (PDAC), no recurrent metastasis-specific mutation has been found, suggesting that epigenetic mechanisms, such as DNA methylation, are the major contributors of late-stage disease progression. Here, we performed the first whole genome bisulfite sequencing (WGBS) on mouse and human PDAC organoid models to identify stage-specific and subtype-specific DNA methylation signatures in PDAC. With this approach, we identified thousands of DMRs that can distinguish between the stages and subtypes of PDAC. Stage-specific DMRs are associated with genes related to nervous system development and cell-cell adhesions and are enriched in promoters and bivalent enhancers. Subtype-specific DMRs showed hypermethylation of GATA6 transcriptional networks in the squamous subtype and hypermethylation of EMT transcriptional networks in the progenitor subtype. These results indicate that aberrant DNA methylation contributes to both PDAC progression and subtype differentiation, resulting in significant and reoccurring DNA methylation patterns with diagnostic and prognostic potential.

overall_design: WGBS of PDAC organoids. 12 mouse organoids (2 normal-derived, 2 PanIN-derived, 4 tumor-derived, and 4 metastasis-derived) and 35 human organoids (4 normal-derived and 31 primary tumor-derived)

first: Chang-il

last: Hwang


title: Single-nucleus RNA-seq (snRNA-seq) of two human primary prostate cancer tissues with different histology

status: Public on Jan 27 2024

city: Changsha

first: Dingxiao

last: Zhang

state: Hunan

address: Yuelushan

department: School of BioMedical Sciences

institute: Hunan University

summary: Heterogeneity at different dimensions is a hallmark of prostate cancer (PCa). To carefully dissect the cellular and molecular landscape of PCa with distinct phenotypes, we conducted spatial transcriptomics (ST) and parallel single-nucleus RNA sequencing (snRNA-seq) technologies to identify areas of vulnerability amenable to therapeutic intervention. We here report the identification of an androgen receptor positive (AR+) but neuroendocrine-null primary PCa subtype with morphologic and molecular characteristics of small-cell carcinoma (SCLPC). snRNA-seq was performed on two samples representative of adenocarcinoma and SCLPC phenotypes, respectively. Further molecular characterization and wet experiments prioritized protein translation, represented by upregulation of many ribosomal proteins (RPs), and SP1, a transcriptional factor that drive SCLPC phenotype, as two potential therapeutic targets for treating AR-independent CRPC. Altogether, our studies reveal the non-rare existence of SCLPC in admixed PCa pathology, which may serve as a cell-of-origin for CRPC, and establish SP1 and translation elongation as actionable therapeutic targets for CRPC.

overall_design: Two frozen PCa tissues were enzymatically digested and single nucleus were counted and collected. Single-cell RNA-seq libraries were constructed using Single Cell 3’ Library and Gel Bead Kit V3.1 according to the manufacture’s introduction, and finally sequenced by Illumina Nova-seq 6000 platform.

first: Cheng

last: Zou

first: Bin

last: Xu


title: Transcriptomic landscape of the anterior thalamic nuclei of the mouse

city: Vancouver

first: Mark

middle: Steven

last: Cembrowski

state: BC

address: 2350 Health Sciences Mall

department: Cellular and Physiological Sciences

country: Canada

institute: University of British Columbia

summary: Here, we examined diversity in the cell-type composition of the mouse anterior thalamic nuclei (ATN) using single-cell RNA-seq.

overall_design: Examination of anterior thalamic nuclei neuron transcriptomes.


title: TMEM65 promotes gastric tumorigenesis by targeting YWHAZ to activate PI3K-Akt-mTOR pathway and is a therapeutic target

status: Public on Jan 26 2024

city: Shijiazhuang

first: Lingxue

last: Shi

address: No.89 Donggang Road

institute: The First Hospital of Hebei Medical University

summary: Copy number alterations are crucial for the development of gastric cancer (GC). Here, we identified Transmembrane Protein 65 (TMEM65) amplification by genomic hybridization microarray to profile copy-number variations in GC. TMEM65 mRNA level was significantly up-regulated in GC compared to adjacent normal tissues, and was positively associated with TMEM65 amplification. High TMEM65 expression or DNA copy number predicts poor prognosis in GC. Furtherly, GC patients with TMEM65 amplification or overexpression significantly associated with shortened survival. Ectopic expression of TMEM65 significantly promoted cell proliferation, cell cycle progression and cell migration/invasion ability, but inhibited apoptosis. Conversely, silencing of TMEM65 in GC cells showed opposite abilities on cell function in vitro and suppressed tumor growth and lung metastasis in vitro. Moreover, TMEM65 depletion by VNP-encapsulated TMEM65-siRNA significantly suppressed tumor growth in subcutaneous xenograft model. Mechanistically, TMEM65 exerted oncogenic effects through activating PI3K-Akt-mTOR signaling pathway, as evidenced of increased expression of key regulators (p-Akt, p-GSK-3β, p-mTOR) by Western blot. YWHAZ (Tyrosine 3-Monooxygenase/Tryptophan 5-Monooxygenase) was identified as a direct downstream effector of TMEM65. Direct binding of TMEM65 with YWHAZ in the cytoplasm inhibited ubiquitin-mediated degradation of YWHAZ. Moreover, oncogenic effect of TMEM65 was partly dependent on YWHAZ. In conclusion, TMEM65 promotes gastric tumorigenesis by activating PI3K-Akt-mTOR signaling via cooperating with YWHAZ. TMEM65 overexpression may serve as an independent new biomarker and is a therapeutic target in GC.

overall_design:
To investigate the effect of TMEM65 on gastric cancer cells, we used a TMEM65-expressing lentivirus to construct AGS stable expressing TMEM65 cells.
We then performed gene expression peofiling analysis of RNA-seq data for AGS overexpressed or vector.


title: Near-cognate tRNAs increase the efficiency and precision of pseudouridine-mediated readthrough of premature termination codons [RNA-seq]

first: Wenqing

last: Liu

overall_design: we perform Trace-seq to examine the global RNA expression level of RESTART v3 treated and untreated HEK293T cells, evaluating the possible off-target effect of RESTART v3.


title: Near-cognate tRNAs increase the efficiency and precision of pseudouridine-mediated readthrough of premature termination codons [Target-seq]

overall_design:
we examined the modification level of the PTC sites in HEK293T under the three versions of the RESTART system to evaluate the modification influence of near-cognate tRNAs

we examined the modification level of the PTC sites in 16HBE and MEF under the action of RESTART v3 system to evaluate the modification efficiency.


title: Near-cognate tRNAs increase the efficiency and precision of pseudouridine-mediated readthrough of premature termination codons [tRNA-seq]

overall_design:
we examined the natural tRNA abundance in the HEK293T treated or untreated with RESTART v3 system, to evaluate the how the  overexpressed tRNAs influence the global tRNA pools

we examined the target tRNA abundance in the HEK293T treated or untreated with RESTART v3 system, to evaluate the level of natural tRNAs that were overexpressed.


title: Co-targeting CDK4/6 and BRD4 promotes senescence and ferroptosis sensitivity in cancer

city: Montreal

first: Sidong

last: Huang

address: McIntyre Medical Sciences Building 3655 promenade Sir-William-Osler

institute: McGill University

summary: Cyclin-dependent kinase 4/6 (CDK4/6) inhibitors are approved for breast cancer treatment and show activity against other malignancies, including KRAS-mutant non-small cell lung cancer (NSCLC). However, the clinical efficacy of CDK4/6 inhibitors is limited due to frequent drug resistance and their largely cytostatic effects. Through a genome-wide cDNA screen, we identified that bromodomain-containing protein 4 (BRD4) overexpression conferred resistance to the CDK4/6 inhibitor palbociclib in KRAS-mutant NSCLC cells. Inhibition of BRD4, either by RNA interference or small-molecule inhibitors, synergized with palbociclib to induce senescence in NSCLC cells and tumors, and the combination prolonged survival in a KRAS-mutant NSCLC mouse model. Mechanistically, BRD4-inhibition enhanced cell cycle arrest and reactive oxygen species (ROS) accumulation, both of which are necessary for senescence induction; this in turn elevated GPX4, a peroxidase that suppresses ROS-triggered ferroptosis. Consequently, GPX4 inhibitor treatment selectively induced ferroptotic cell death in the senescent cancer cells, resulting in tumor regression. Co-targeting CDK4/6 and BRD4 also promoted senescence and ferroptosis vulnerability in pancreatic and breast cancer cells. Together, these findings reveal therapeutic vulnerabilities and effective combinations to enhance the clinical utility of CDK4/6 inhibitors.

overall_design: To invertigate the gene expression program that can be affected by inhibition of CDK4/6 or BRD4, we treated KRAS mutant cell line H358 with CDK4/6 inhibitior (palbociclib) or BRD4 inhibitor (IBET726), or both for 9 days.  We then performed gene expression profiling analysis using the RNA-Seq data generated in these cells.

first: David

last: Dankort

first: Xianbing

last: Zhu

first: Zheng

last: Fu


title: Upregulation of developmentally-downregulated miR-1247-5p promotes neuroprotection and axon regeneration in vivo [Small-RNA-seq]

city: Farmington

first: Ephraim

middle: F

last: Trakhtenberg

state: CT

address: 263 Farmington Ave. RM L4005

department: Neuroscience

institute: University of Connecticut School of Medicine

summary: Numerous micro-RNAs (miRNAs) affect neurodevelopment and neuroprotection, but potential roles of many miRNAs in regulating these processes are still unknown. Here, we used the retinal ganglion cell (RGC) central nervous system (CNS) projection neuron and optic nerve crush (ONC) injury model, to optimize a mature miRNA arm-specific quantification method for characterizing the developmental regulation of miR-1247-5p in RGCs, investigated whether injury affects its expression, and tested whether upregulating miR-1247-5p-mimic in RGCs promotes neuroprotection and axon regeneration. We found that, miR-1247-5p is developmentally-downregulated in RGCs, and is also further downregulated after ONC. Importantly, RGC-specific upregulation of miR-1247-5p promoted neuroprotection and axon regeneration after injury in vivo. To gain insight into the underlying mechanisms, we analyzed by bulk-mRNA-seq embryonic and adult RGCs, along with adult RGCs transduced by miR-1247-5p-expressing viral vector, and identified developmentally-regulated cilial and mitochondrial biological processes, which were reinstated to their embryonic levels in adult RGCs by upregulation of miR-1247-5p. Because axon growth is also a developmentally-regulated process, in which mitochondrial dynamics play important roles, it is possible that miR-1247-5p promoted neuroprotection and axon regeneration through regulating mitochondrial functions.

overall_design: Approximately 200,000 of Thy1-immunopanned RGCs were collected (from each condition) and RNA was isolated immediately for preparing cDNA libraries using adaptors that ligate selectively to the endogenously modified 5’ and 3' ends of processed small RNAs, thereby enriching specifically for mature small RNAs, including miRNAs, and excluding degraded RNA fragments, precursors, and other RNA species (TruSeq Small RNA Kit, Illumina). Small RNA cDNA library products in the range of 140-160 base pairs (including 5’ and 3’ adapters and an insert corresponding to a small RNA) were sequenced as paired reads 100 bp from each end on HiSeq 2000 Sequencer (Illumina), passed QC filters, mapped to the mm39 mouse genome and transcriptome by Bowtie, and quantified/analyzed using featureCounts from the subread package and DESeq2.

first: Agnieszka

last: Lukomska

first: Jian

last: Xing


title: Decoding the Universal Human Chromatin Landscape through Teratoma-Based Profiling [RNA-seq]

overall_design: RNA-Seq of teratomas generated from H1 hESCs


title: Next Generation Sequencing of T regulatory cells (GFP+, dTomato+) and ex-T regulatory cells (GFP-, dTomato+) harvested from lymph nodes of bone marrow chimeras (50% Foxp3GFP-tdT  or Foxp3GFP-Fth/-tdT and 50% C57BL/6 bone marrow into Rag2-/- recipients)

city: Oeiras

first: Rui

last: Martins

address: Rua da Quinta Grande 6

department: Inflammation

country: Portugal

institute: Gulbenkian Institute for Molecular Medicine

summary:
Purpose: To address whether FTH regulates gene expression in TREG cells, irrespective of bystander inflammation. For this, we compared the gene expression profile of TREG and Ex-TREG cells sorted from the lymph nodes of mixed BM chimeras
Results: Using an optimized data analysis workflow, we mapped an average 30.14 million sequence reads per sample to the mouse genome (build mm39) and identified 34,321 transcripts in the mRNA of Treg and Ex-Treg cells from mixed bone marrow chimeras. Differential gene expression was performed using the DESeq2 R package (v.1.32) and gene expression was modeled by genotype and condition. Differentially expressed genes were considered for genes with an adjusted p-value<0.05 and an absolute log2 fold change>0.
Conclusions: Analysis of RNAseq data showed that Fth deletion in TREG cells (CD45.2+GFP+tdT+) developing from the BM of Foxp3GFP-Fth∆/∆-tdT mice, up-regulated 149 genes and down-regulation of 96 genes, compared to TREG cells from control Foxp3GFP-tdT mice. Ex-TREG cells (CD45.2+GFP-tdT+) up-regulated 90 genes and down-regulated 36 genes. The genes regulated in TREG and ex-TREG cells originating from the BM of Foxp3GFP-Fth∆/∆-tdT vs. Foxp3GFP-tdT mice were associated with the oxidative stress response regulated by NRF2 . This suggests that FTH exerts cell-autonomous control of TREG cell redox homeostasis, irrespective of bystander inflammation.

overall_design: Methods: T-reg and Ex-T-reg cells were collected from the lymph nodes of mixed bone marrow chimeras, 6 weeks after reconstitution. Mixed bone chimeras were generated by non-lethally irradiating Rag2-/- mice and reconstituting with a mix of 50% Foxp3GFP-tdT  or Foxp3GFP-Fth/-tdT and 50% C57BL/6 bone marrow (n=3-4 per combination). RNA was extracted and libraries prepared. mRNA expression profiles were generated by next generation sequencing,  using Illumina High output kit v2.5 and NexSeq500 sequencer.

first: Qian

last: Wu

first: Ana

last: Carlos

first: Faouzi

last: Braza

first: Marie-Louise

last: Bergman

first: Patrícia

last: Bastos-Amador

first: Jamil

last: Kitoko

first: Eloy

last: Cuadrado

first: Brendon

last: Sicluna

first: Jonathan

last: Landry

first: Ferris

last: Jung

first: Temitope

middle: W

last: Ademolue

first: Mirko

last: Peitzsch

first: José

last: Almeida-Santos

first: Jessica

last: Thompson

first: Sílvia

last: Cardoso

first: Pedro

last: Ventura

first: Vanessa

last: Ribeiro

first: Vital

last: Domingues

first: Inês

last: Cabral

first: Sebastian

last: Weis

first: Marco

last: Groth

first: Cristina

last: Ameneiro

first: Miguel

last: Fidalgo

first: Jocelyne

last: Demengeot

first: Derk

last: Amsen

last: Soares


title: Atopic dermatitis-like epicutaneous infection with S. aureus in WT and eosinophil KO mice

city: Baltimore

first: Alex

middle: Jea-Eun

last: Lee

address: 1650 Orleans Street

institute: Johns Hopkins University

summary: Investigation of the pathways involved in eosinophil-mediated inflammation in S. aureus epicutaneous infection

overall_design: Dorsal fur of mice was shaved and depilated, followed by epicutenous infection using 1e8 S. aureus CFU for 7 days. A 10mm punch was then excised and sent for RNA purification and sequencing

first: Sabrina

last: Nolan

first: Nicholas

last: Orlando

first: Nathan

last: Archer


title: Drp1 Controls Complex II assembly and Fatty Acid Oxidation by Sdhaf2 Action on Mitochondria

city: Los Angeles

first: Peter

middle: Huy

last: Tran

address: 650 Charles E Young Dr. S

department: Medicine

institute: UCLA

summary: The Dynamin-related GTPase, Drp1 (encoded by Dnm1l) plays a central role in mitochondrial fission and is requisite for numerous cellular processes however its role in oxidative metabolism remains unclear. Herein, we show that among human tissues, skeletal muscle exhibits the strongest correlations with the DNM1L gene. Knockdown of Drp1 (Drp1-KD) promoted mitochondrial hyperfusion in the muscle of male mice. Reduced fatty acid oxidation and accumulation of intramyocellular lipids along with increased muscle succinate was observed in Drp1-KD muscle. Muscle Drp1-KD reduced Complex II assembly and activity as a consequence of diminished mitochondrial translocation of succinate dehydrogenase assembly factor 2 (Sdhaf2). Restoration of Sdhaf2, normalized Complex II activity and lipid oxidation in Drp1-KD myocytes. Drp1 appears critical in maintaining mitochondrial Complex II assembly and fatty acid oxidation, suggesting a mechanistic link between mitochondrial morphology and skeletal muscle lipid metabolism which is of clinical significance in combatting metabolic diseases.

overall_design: We generated a conventional muscle-specific Drp1 knockout heterzygous mice (Drp1-KD) and performed RNAseq in the quadricep muscle of the Control f/f and mDrp1HET mice.

first: Zhenqi

last: Zhou

first: Alice

last: Ma

first: Timothy

last: Moore

first: Dane

last: Wolf

first: Nicole

last: Yang

first: Mayuko

last: Segawa

first: Alexander

last: Strumwasser

first: Wenjuan

last: Ren

first: Kai

last: Fu

first: Jonathan

last: Wanagat

last: van der Bliek

first: Rachelle

last: Crosbie-Watson

first: Marc

last: Liesa

first: Linsey

last: Stiles

first: Sushil

last: Mahata

first: Orian

last: Shirihai

first: Mark

middle: O

last: Goodarzi

first: Michal

last: Handzlik

first: Christian

last: Metallo

first: David

last: Walker

first: Andrea

last: Hevener


title: Expression of ALS-PFN1 impairs vesicular degradation in iPSC-derived microglia.

city: Worcester

first: Elaine

last: Lim

state: MA

address: 366 Plantation Street, Paul J DiMare 5-1025

institute: UMass Chan Medical School

summary: Microglia play a pivotal role in neurodegenerative disease pathogenesis, but the mechanisms underlying microglia dysfunction and toxicity remain to be elucidated. To investigate the effect of neurodegenerative disease-linked genes on the intrinsic properties of microglia, we studied microglia-like cells derived from human induced pluripotent stem cells (iPSCs), termed iMGs, harboring mutations in profilin-1 (PFN1) that are causative for amyotrophic lateral sclerosis (ALS). ALS-PFN1 iMGs exhibited elevated lipid droplets, upregulation of autophagic factors and deficient phagocytosis, a canonical microglia function. Mutant PFN1 also displayed enhanced binding affinity for PI3P, a critical signaling molecule involved in autophagic and endocytic processing. Our cumulative data implicate a gain-of-toxic function for mutant PFN1 within the autophagic and endo-lysosomal pathways, as administration of rapamycin rescued phagocytic dysfunction in ALS-PFN1 iMGs. These outcomes demonstrate the utility of iMGs for neurodegenerative disease research and implicate microglial vesicular degradation pathways in the pathogenesis of these disorders.

overall_design: We performed RNA sequencing on wildtype (WT) and mutant (C71G het) iMGs with 4 replicates each, and wildtype (WT) and mutant (M114T het) with 3 replicates each.

first: Daryl

last: Bosco


title: Identification of endothelial gene regulatory elements during cardiopharyngeal lineage differentiation [ATAC-seq]

city: Naples

first: Antonio

last: Baldini

state: Naples

address: Via Sergio Pansini 5

department: Department of Molecular Medicine and Medical Biotechnology

country: Italy

institute: University of Naples Federico II

summary: Endothelial cells (EC) differentiate from multiple sources during embryonic development, including the cardiopharyngeal mesoderm, which gives rise to multiple tissues including the cardiac and branchiomeric muscle. Here, we have used a cardiogenic mesoderm cell differentiation model that also activates an endothelial transcription program. Comparing our chromatin remodeling data with publicly available single-cell RNA-seq data from mouse embryos, we derived a list of over 100 putative regulatory elements activated in EC marker genes. We applied a machine-learning strategy trained with validate enhancers to predict the probability of these sequences to function as enhancers. The computational assay returned over 50% of these sequences to be likely enhancers, some of which previously reported. In addition, using genetic and epigenetic perturbations of some of these elements, we established their requirement for gene expression during endothelial differentiation. Finally, we used the integration of multiple data sources and computational tools to search for transcriptional factor binding motifs related to endothelial cell fate specification. We found motifs of GATA, ETS, and FOS/AP-1 factors as the most significantly enriched in these sequences. Overall, our approach has been efficient in the identification of novel EC regulatory sequences with high likelihood to be enhancers, and validated a subset of them. Motif analyses revealed that the core EC transcription factors GATA/ETS/FOS is a likely driver of EC differentiation in cardiopharyngeal mesoderm.

overall_design: In this study we investigated the gene expression and the chromatin accessibility profile in the early phases of multipotent cardiac progenitors' differentiation into mature endothelial cells (ECs), starting from mouse embryonic stem cells (mESCs). Differentiating mESCs were collected at day2 and day4 and two biological replicates for each condition were  used for ATAC-seq and RNA-seq assay.

first: Ilaria

last: Aurigemma

first: Olga

last: Lanzetta

first: Varsha

last: Krishnan

first: Claudia

last: Angelini

first: Elizabeth

last: A Illingworth


title: Histological heterogeneity in primary prostate

first: Dingxiao

last: Zhang

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

overall_design: Refer to individual Series


title: Single-mitosis dissection of acute and chronic DNA mutagenesis and repair

city: Heidelberg

first: Paul

middle: Adrian

last: Ginno

state: Baden-Württemberg

address: Im Neunheimer Feld 280

institute: DKFZ

summary: The mammalian genome is constantly exposed to genotoxins that can lead to mutations and ultimately tumorigenesis. Despite cancer being recognized as a genetic disease, much is still unknown regarding tumor induction and evolution of the cancer genome. To study gradual (reactive oxygen species, ROS) and acute (ultraviolet radiation, UV) mutagenic processes both in vitro and then in vivo, we first use microfluidic separation of cells after a single mitosis following pulse mutagenesis. While UV mutations tend to be fixed after one cell cycle and show mirror image mutation patterns between sister cells from a single mitosis, ROS mutations do not. We next show that while both ROS and UV mutation rates are reduced as a function of transcription, only UV has a strand-specific repair signature. Lastly, we resolve pulse induced lesion segregation to single haplotypes by analysing liver tumour genomes from F1 mice treated with N-nitroso-diethylamine (DEN). In summary, we have established highly controlled systems to study 3 distinct forms of mutagenesis at their inception in mammalian cells, providing insights into how these mutations are fixed at the genome scale.

sample_organism:
Mus musculus
Mus musculus castaneus

platform_organism:
Mus musculus
Mus musculus castaneus

overall_design: Whole Genome sequencing of the PF1 cell line following UV treatment and seperation of mitotic sisters. Whole genome sequencing of liver tumors from male F1 mice of a Mus castaneus/C3H cross. Ribo-zero bulk RNA and Omni-ATAC samples of the PF1 cell line

middle: A

first: Duncan

last: Odom

first: Martin

middle: S

last: Taylor


title: Revealing the Grammar of Small RNA Secretion Using Interpretable Machine Learning IV

city: San Francisco

first: Hani

last: Goodarzi

department: Biochemistry and Biophysics

institute: UCSF

summary: Small non-coding RNAs can be secreted through a variety of mechanisms, including exosomal sorting, in small extracellular vesicles, and within lipoprotein complexes. However, the mechanisms that govern their sorting and secretion are still not well understood. In this study, we present ExoGRU, a machine learning model that predicts small RNA secretion probabilities from primary RNA sequence. We experimentally validated the performance of this model through ExoGRU-guided mutagenesis and synthetic RNA sequence analysis, and confirmed that primary RNA sequence is a major determinant in small RNA secretion. Additionally, we used ExoGRU to reveal cis and trans factors that underlie small RNA secretion, including known and novel RNA-binding proteins, e.g., YBX1, HNRNPA2B1, and RBM24. We also developed a novel technique called exoCLIP, which reveals the RNA interactome of RBPs within the cell-free space. We used exoCLIP to reveal the RNA interactome of HNRNPA2B1 and RBM24 in extracellular vesicles. Together, our results demonstrate the power of machine learning in revealing novel biological mechanisms. In addition to providing deeper insight into complex processes such as small RNA secretion, this knowledge can be leveraged in therapeutic and synthetic biology applications.

overall_design: Using a machine learning model to dissect the primary sequences and structural features of small RNAs in the extracellular and intracellular compartments.

first: Bahar

last: Zirak

first: Mohsen

last: Naghipourfar

first: Ali

last: Saberi

first: Delaram

last: Pouyabahar

first: Amirhossein

last: Zarezadeh

first: Lixi

last: Luo

first: Lisa

last: Fish

first: Doowon

last: Huh

last: Sharifi-Zarchi


title: Revealing the Grammar of Small RNA Secretion Using Interpretable Machine Learning III


title: Revealing the Grammar of Small RNA Secretion Using Interpretable Machine Learning I


title: RNA-Seq analysis in human trophoblast stem cells (CT27) upon depletion of transcription factor GATA2 or GATA3

city: Kansas City

first: Soumen

last: Paul

state: KS

address: Hemenway Building 3901 Rainbow Boulevard

department: Department of Pathology & Laboratory Medicine

institute: University of Kansas Medical Center

summary: The GATA transcription factors, particularly GATA2 and GATA3 are responsible for trophoblast stem cells self renewal and differentiation in mice. However, we have a poor understanding about the contribution of GATA2 and GATA3, in the trophoblast cells in humans. Thus, the goal of this study is to define the importance of GATA factors in orchestrating global gene expression program in human trophoblast stem cells. Using a RNA interference strategy (RNAi), we depleted GATA2 or GATA3 expression in human trophoblast stem cells (GATA2 KD or GATA3 KD) . We then differentiated GATA2 KD and GATA3 KD human trophoblast stem cells into syncytiotrophoblasts(SynT) and compared global gene expression patterns via RNA sequencing (RNA-seq). Three individual experiments were performed for the differentiated SynT state. The submitted files contain the data (raw fastq files, count data and deseq2 output) from  those experiments.

overall_design: RNA-Seq

first: Rajnish

last: Kumar

first: Ananya

last: Ghosh


title: Spatial transcriptomics reveals unexpected histological heterogeneity in primary prostate

last: Zhang

overall_design: Prostate biopsies from eight Chinese patients with histologically diagnosed PCa were collected. According to the manufacture’s introduction, Visum spatial libraries were constructed using Visum spatial library construction kit (10x Genomics, PN-1000184). The libraries were finally sequenced using an Illumina Nova-seq 6000 platform.

first: Cheng

last: Zou


title: Acute STING signaling from microglia and peripheral white blood cells drive neutrophil infiltration influencing brain injury

city: Blacksburg

first: Alicia

last: Pickrell

state: VA

address: 970 Washington Street SW

institute: Virginia Tech

summary:
To investigate transcriptomic changes 2 hours after injury in mice ablating STING in microglia after brain injury
We then performed gene expression profiling analysis using data obtained from RNA-seq of ipsilateral and contralateral cortices.

overall_design: CX3CR1cre-er/+ and CX3CR1cre/+ STING fl/fl animals

middle: M


title: Upregulation of developmentally-downregulated miR-1247-5p promotes neuroprotection and axon regeneration in vivo [RNA-seq]

first: Ephraim

last: Trakhtenberg

overall_design: Adult RGCs transduced with AAV2 expressing miR-1247-5p mimic or scramble shRNA mimic were Thy1-immunopanned from single cell retinal suspension (from 10 retinas per condition) and FACS’ed for mCherry+ cells. Approximately, 5,000 RGCs (per condition) were collected by FACS. Approximately, 50,000 of embryonic day 18 (E18) RGCs were Thy1-immunopanned from single cell retinal suspension. RNA was isolated immediately using the Zymo QuickRNA microprep kit. Total RNA with RNA Integrity Number (RIN) ≥ 9 (by Bioanalyzer 2100 using the Nano 6000 kit, Agilent) was extracted using Direct-zol RNA MiniPrep kit (R2050, Zymo Research). cDNA libraries were prepared using polyA-selected RNA (TruSeq RNA Library Prep Kit, Illumina), paired reads sequenced 100 bp from each end on HiSeq 2000 Sequencer (Illumina), passed QC filters, mapped to the mm39 genome and transcriptome by Hisat2, gene expression was normalized as fragments per kilobase of transcript per million mapped reads (FPKM), and analyzed by Cufflinks/CuffDiff.

first: Agnieszka

last: Lukomska

first: Jian

last: Xing


title: Identification of PRMT1 as a suppressor of MHC-I and anti-tumour immunity

city: Clayton

first: Wei

last: Shi

state: VIC

address: 15 Innovation Walk

department: Department of Biochemistry and Molecular Biology

country: Australia

institute: Monash University

summary: To understand the mechanisms by which Prmt1 knockout or inhibition sensitizes B16F10 to T cell mediated killing.

overall_design: B16F10 cells were pre-treated with Prmt1-inhibitor GSK3368715 (2.5 µM) for 2 days. Afterwards, untreated wildtype (controls), Prmt1-inhibitor treated or Prmt1-/- B16F10 were cultured for one more day in the presence or absence of mouse IFNγ (1 ng/ml). There are three independent replicates for each condition. Total RNA was then extracted, quantitated with 4200 Tapestation and finally, 1 µg of total RNA per sample was subjected to NGS library preparation (TruSeq RNA). NGS libraries were pooled and sequenced using a P2 100-cycle Illumina sequencing kit on the NextSeq 2000 sequencing system with paired-end 66bp reads.

first: Tirta

last: Djajawi

first: Liam

last: Neil

first: Bhupinder

last: Pal

first: Yang

last: Liao

first: Conor

middle: J

last: Kearney


title: Identification of endothelial gene regulatory elements during cardiopharyngeal lineage differentiation

first: Antonio

last: Baldini


title: Bulk RNA-seq reveals effects of tamoxifen on endothelial gene expression

status: Public on Jan 25 2024

city: Jamaica Plain

first: Xiaoran

address: 14 Perkins Square, Apt 11

department: Cardiology

institute: Boston Children's Hospital

summary: Transgenic line combination with tamoxifen induction is widely used for biological study. Considering the possibility that tamoxifen induction may alter gene expression, we performed this study to elucidate the potential effect of tamoxifen gavage on the gene expression profile of endothelial cells.

overall_design: Cdh5-CreERT2(PAC);Rosa26fsTRAP transgenic mice were divided into 2 groups: with tamoxifen gavage group and withou tamoxifen gavage group.Tamoxifen gavage was conducted at E12.5 for pregnant mice, then E15.5 embryos were harvest to perform endothelial cells (ECs) sorting with FACS. ECs purified from different samples were stored at -80℃ and then processed at the same time to produce sequencing libraries. rRNAs were deleted with NEBNext rRNA Depletion Kit (V2) and libraries were made with NEBNext Ultra II RNA Library Prep Kit.

last: Chen

first: William

last: Pu


title: Identification of endothelial gene regulatory elements during cardiopharyngeal lineage differentiation [RNA-seq]

first: Ilaria

last: Aurigemma

first: Olga

last: Lanzetta

first: Varsha

last: Krishnan

first: Claudia

last: Angelini

first: Elizabeth

last: A Illingworth


title: Effect of Dyrk1a inhibition on the transcriptome of embryonic hearts from the Dp1Tyb mouse model of Down syndrome using bulk RNAseq

city: London

first: Victor

last: Tybulewicz

address: 1 Midland Road

institute: Francis Crick Institute

summary: Embryos from the Dp1Tyb mouse model for Down syndrome (DS) present with congenital heart defects similar to the heart defects seen in humans with DS. We found that genetically reducing the copy number of the Dyrk1a gene (one of the genes in 3 copies in DS) from 3 to 2, normalised some of the transcriptomic changes in Dp1Tyb embryonic hearts and rescued congenital heart defects. Here we treated pregnant mice carrying Dp1Tyb and wild-type (WT) embryos with a Dyrk1a pharmacological inhibitor (Leucettinib-21 or L21) or an inactive isomer (Iso-L21) to study the effect of L21 on the transcriptome of Dp1Tyb and WT embryonic hearts.

overall_design: C56BL/6J females were set up in timed matings  with Dp1Tyb males and were treated daily by oral gavage with 30mg/kg of L21 or Iso-L21 from embryonic day 5.5 to E13.5. 20 embryonic hearts were used for this study; five biological repeats per condition (4 conditions: 2 genotypes treated with L21 or iso-L21) were used for bulk RNAseq.

first: Eva

last: Lana-Elola

first: Tegan

last: Gilmore


title: Tumor circadian clock strength influences metastatic potential and predicts patient prognosis in Luminal A breast cancer

city: Manchester

first: Qing-Jun

last: Meng

state: Manchester

address: A.V.Hill Building, The University of Manchester, Oxford Road

department: Division of Cell Matrix Biology & Regenerative Medicine

institute: University of Manchester

summary: To investigate circadian clocks in human breast tumors, 43 pairs of human samples (tumor and matched normal breast tissues from the same patient) were collected to run bulk-RNAseq, including 29 pairs of Luminal A, 3 pairs of Luminal B, 2 pairs of HER2 and 9 pairs of Triple-negative breast cancer

overall_design: Differential genes expression levels and circadian clock time in human normal and paired tumor samples were analysed based on our RNAseq data

first: Shi-Yang

last: Li


title: Snowball-like mushroom morphologies and a novel gene family required for fruiting body development

city: Szeged

first: Zsolt

last: Merényi

state: Csongrad-Csanad

address: Temesvári krt. 62, 5. emelet 519. szoba (vagy az épület főportája)

country: Hungary

institute: Biological Research Centre

summary: The morphogenesis of sexual fruiting bodies of fungi is a complex process determined by a genetically encoded programme. Fruiting bodies reached highest complexity levels in the Agaricomycetes, yet, the underlying genetics is currently poorly known. In this work, we functionally characterised an unannotated gene we term snb1, whose expression level increases rapidly during fruiting body initiation. According to phylogenetic analyses, orthologues of snb1 are present in almost all Agaricomycetes and may represent a novel conserved gene family that plays a substantial role in fruiting body development. We disrupted snb1 using CRISPR/Cas9 in the agaricomycete model organism Coprinopsis cinerea. Snb1 mutants formed unique, snowball-shaped, rudimentary fruiting bodies that could not differentiate caps, stipes and lamellae. We took advantage of this phenotype to study fruiting body differentiation using RNA-Seq analyses. This revealed a multitude of differentially regulated genes and gene families that may be related to tissue differentiation and the formation of structures. Taken together, the novel gene family of snb1 and the differentially expressed genes in the snb1 mutants provide valuable insights into the complex mechanisms underlying tissue differentiation in the Agaricomycetes.

overall_design: Wild type Coprinopsis cinerea (AmutBmut1) and snb1 knock out mutant strains were sampled for RNA sequencing. From both strains, samples were collected after 24 h (secondary hyphal knots) and 48 h (stage 1 primordia) post-light induction. Each sample group contains three biological replicates.

first: Csenge

last: Földi

last: Merenyi

first: Bálint

last: Balázs

first: Árpád

last: Csernetics

first: Nikolett

last: Miklovics

first: Hongli

last: Wu

first: Botond

last: Hegedüs

first: Máté

last: Virágh

first: Zhihao

last: Hou

first: Xiao-Bin

last: Liu

first: László

last: Galgóczy

middle: G

last: Nagy


title: Differential gene expression between porcine plantar, trunk, dorsal foot, and snout skin

city: Boston

first: Christiane

last: Fuchs

address: 40 Blossom Street

department: Wellman Center for Photomedicine

institute: Massachusetts General Hospital / Harvard Medical School

summary: Porcine plantar skin was compared to 3 other skin types, namely trunk, snout and dorsal foot skin, to identify potential differences in gene expression profiles, which could lead to insights into the mechanisms underlying the many specializations of plantar skin tissue.

overall_design: We performed Differential Gene Expression profiling from the obtained RNA-Seq data of 4 different skin sites (plantar, trunk, snout, and dorsal foot skin) from 5 different animals (all female adults).

first: Katherine

last: Stalnaker

first: Ying

last: Wang

first: Linh

last: Pham

first: Clifton

last: Delgard

first: Gauthaman

last: Sukumar

first: Daniel

last: Hupalo

first: Sunhhun

last: Cho

first: Xunwei

first: Ivan

last: Jozic

first: Richard

middle: R

last: Anderson

first: Jon

last: Meyerle

first: Joshua

last: Tam


title: Cut and run analysis in human trophoblast stem cells (CT27) upon pulling down with GATA2 and GATA3 antibody

first: Soumen

last: Paul

summary: GATA transcription factors, particularly GATA2 and GATA3 are selectively expressed in the extraembryonic trophoblast lineage and regulates gene expression to promote trophoblast self-renewal during mammalian development. However, we have a poor understanding about the contribution of these GATA factors, in the process of syncytiotrophoblast (SynT) development. Thus, the goal of this study is to define the importance of GATA2 and GATA3 in orchestrating a global gene expression program that poises and commits mammalian trophoblast stem cells to the SynT lineage. Using the cut and run technique for GATA2 and GATA3 antibodies in the human trophoblast stem cells we are aiming to identify the direct targets of the GATA transcription factors that are essential for determining the SynT lineage. Three individual experiments were performed.

overall_design: Cut&Run experiments using GATA2 and GATA3 antibodies in human trophoblast stem cells.

first: Rajnish

last: Kumar

first: Ananya

last: Ghosh

first: Ram


title: miRNA levels in F1-F3 male germ cells and female F2 PFC  following prenatal exposure to sGC in guinea pigs

city: Toronto

first: Christopher

last: Casciaro

state: ON

address: 1 King's College Circle

department: Department of Physiology

institute: University of Toronto

summary:
Stress and glucocorticoid exposure during pregnancy alters neurodevelopment and behavior in offspring, and these effects extend multiple generations through a paternal lineage. The epigenetic mechanisms that govern this transgenerational transmission are unclear. We hypothesized that maternal exposure to multiple courses of sGC in late pregnancy would result in altered miRNA levels in germs cells of male guinea pigs across three generations. Further, our behavioral data indicate that F2 females exhibit altered behaviors following sGC exposure. Thus, we evaluated the miRNA profile of F2 female prefrontal cortex to determine the mechanisms responsible for these behavioral changes and to compare these data to our germ cell miRNA analysis.
We used miRNA microarray to evaluate the miRNA levels in F1-F3 germ cells and F2 female PFC in guinea pigs that were exposed to control and F0 prenatal sGC exposure. We identified no significant changes to miRNA levels in both F1-F3 germ cells and F2 PFC from guinea pigs in the sGC group.

overall_design: Germ cells were isolated from male guinea pigs that either were in the control or sGC group F1_Ctrl (n=7), F1_sGC (n=6), F2 (n=6/gp), and F3 (n=6/gp). PFC was isolated from F2 female guinea pigs (n=6/group). Total RNA was extracted from all samples and microarray analysis was performed using miRNA 4.0 microarray (Affymetrix).

first: Hirotaka

last: Hamada

first: Vasilis

last: Moisiadis

first: Andrea

last: Constantinof

first: Alisa

last: Kostaki

first: Stephen

last: Matthews


title: Single cell RNA sequencing analysis using embryonic day 9.5 placenta from control and GATA DKO mice

summary: The placenta establishes a maternal–fetal exchange interface which ensures transportation of nutrients and gases between the mother and the fetus. A critically conserved step in the establishment of this exchange surface in mammals is the fusion of trophoblast cells into a multinucleated syncytiotrophoblasts (SynT).  In mice, SynTs develop via differentiation of the labyrinth trophoblast progenitors (LaTP) of the developing placenta, and in humans, SynTs develop via differentiation of villous cytotrophoblast (CTB) progenitors. Despite being an indispensable step for mammalian development in utero, conserved molecular mechanisms that ensure SynT development are poorly understood. Herein, we show that GATA2 and GATA3 (GATA factors) plays an essential role in labyrinth trophoblast progenitors (LaTPs) thereby committing these cells towards the SynT differentiation program. Mature syncytial layer specific immunostaining with established markers, and ultrastructure analysis of the labyrinth revealed that the loss of GATA factors was impairing the formation of mature SynT cells. We therefore focused our attention on the status of the labyrinth progenitor trophoblast cells in the GATA DKO vs the control placentae. To do that we performed single cell RNA Sequencing (scRNA-Seq) with E9.5 control and Gcm1Cre GATA KO placentae.

overall_design: Single cell RNA sequencing of mouse placenta were generated by deep sequencing using the 10x Genomics Chromium Single Cell Gene Expression Solution (10xgenomics.com).


title: MERFISH reveals embryonic endothelial heterogeneity

first: Xiaoran

last: Zhang

summary: The heterogeneity of embryonic endothelial cells (ECs) especially the distinction of arteriovenous ECs remains incompletely characterized. We established a mouse single-EC transcriptomic landscape at mid-to-late gestation stage and identified 19 subclusters, including Etv2+Bnip3+ early ECs and 2 specialized ECs. Most of these subtypes were grouped by their vascular-bed types, while ECs from brain, heart and liver were grouped by their tissue origins. Unlike arterial ECs (aECs), embryonic venous (vECs) and capillary ECs (cECs) shared less markers with their adult counterparts. Notably, capillary clusters showed some venous characteristics and one of them served as an intermediate state of arteriovenous specification. Compared to the more early stage, a clear arteriovenous branch which also going through a venous plexus was identified.  aECs and vECs showed distinct transcriptional modules including specific regulatory networks of transcription factors. Especially, USF1 and MECOM were verified functioning in arteriovenous differentiation through human induced pluripotent stem cells (hiPSC) differentiation models. We therefore provide a new map of endothelial heterogeneity highlighting regulation of arteriovenous specification.

overall_design: Vizgen MERSCOPE platform was employeed to perform the spatial transcriptome study. 2 sagittal sections were obtained from 2 different E9.5 wild-type embryos, and 1 sagittal section was obtained from wild-type E15.5 embryo. A 300-genes gene panel was used for probe hybridization.

first: Jian

last: Chen

first: Michael

last: Trembley

first: William

last: Pu


title: Dyrk1a dosage effects on the transcriptome of embryonic hearts from the Dp1Tyb mouse model of Down syndrome using scRNAseq

first: Victor

last: Tybulewicz

summary: We performed single cell RNAseq on whole hearts from E13.5 Dp1Tyb embryos and from wild-type littermates, as well as hearts from E13.5 Dp1TybDyrk1a+/+/- embryos and wild-type littermates. We identified 14 clusters which were assigned to individual cell types based on expression of marker genes.

overall_design: Single cell RNAseq analysis of 1 E13.5 Dp1Tyb heart, 3 Dp1TybDyrk1a+/+/- embryonic hearts and 3 wild-type embryonic hearts.

first: Eva

last: Lana-Elola

first: Miriam

last: Llorian


title: Dyrk1a dosage effects on the transcriptome of embryonic hearts from the Dp1Tyb mouse model of Down syndrome using bulk RNAseq

summary: We performed bulk RNAseq on whole hearts from E13.5 Dp1Tyb embryos and from wild-type littermates, as well as hearts from E13.5 Dp1TybDyrk1a+/+/- embryos and wild-type littermates. Analysis showed the expected increased expression of the Hsa21-orthologous genes on Mmu16 that are present in 3 copies. Gene set enrichment analysis identified pathways that are altered in Dp1Tyb hearts. Some of these changes were dependent on 3 copies of Dyrk1a.

overall_design: RNAseq on 5 E13.5 Dp1Tyb embryonic hearts and 5 wild-type littermate embryonic hearts, and on 5 E13.5 Dp1TybDyrk1a+/+/- embryonic hearts and 5 wild-type littermate embryonic hearts.


title: Physical interaction with Spo11 mediates the localisation of Mre11 to chromatin in meiosis and promotes its nuclease activity

city: Vienna

first: Franz

last: Klein

address: Dr. Bohr-Gasse 9

department: Chromosome Biology

country: Austria

institute: Max Perutz Labs, University of Vienna

summary: Meiotic recombination is of central importance for the proper segregation of homologous chromosomes and is initiated by DNA double-strand breaks (DSB) that depend on Spo11 and Mre11. Calibrated chromatin immunoprecipitation (ChIP) for Mre11 shows that Spo11 promotes Mre11 recruitment to chromatin, independent of DSB formation. A C-terminal deletion mutant deficient in Spo11 interaction severely reduces the association of Mre11 with meiotic chromatin. Consistent with the reduction of Mre11 foci in this mutant, it strongly impedes DSB formation, leading to spore death.

sample_organism:
Saccharomyces cerevisiae
Nakaseomyces glabratus

overall_design: Six ChIP-seq samples and six corresponding whole cell extract (WCE) samples were derived from Saccharomyces cerevisiae (S. cer.) SK1 cells with HA-tagged (or untagged) Mre11 grown in meiotic culture for 5 hrs ("t5") and mixed with Candida glabrata (C. gla., with Scc1-HA6) CBS138 cells for calibration and downstream quantiative comparison. Paired-end (75bp) fastq.gz files are provided, as well as calibrated processed files ("filled dpp" with chromosome, position, and depth information).

first: Lingzhi

last: Huang

first: Kuldeep

last: Nangalia

first: Doris


title: NRF2-independent iron-triggered signaling via MAPK p38-ETS1 regulates LSEC Bmp6 mRNA expression [I]

city: Warsaw

first: Michal

last: Mikula

address: Roentgena 5

department: Experimental Oncology

country: Poland

institute: Maria Skłodowska-Curie National Research Institute of Oncology

summary: BMP6 is an iron-sensing cytokine produced by liver sinusoidal endothelial cells (LSECs). The transcription factor NRF2 was proposed to induce Bmp6 expression in response to iron-triggered oxidative stress, and NRF2 knock-out mice were shown not capable of regulating Bmp6 in response to massive diet-imposed iron overload. However, LSEC Bmp6 expression levels in the hemochromatosis mouse model reflect hepatocytic, not endothelial iron content. Hence, it is not fully resolved how iron signals translate into alterations of Bmp6 mRNA levels. To further explore the mechanisms of Bmp6 regulation, we employed female mice aged 10-11 months that are hallmarked by hepatocytic, but not LSEC iron accumulation and decreased transferrin saturation, hinting at the absence of systemic iron overload. We found that LSECs of aged mice exhibit increased Bmp6 levels as compared to young controls, but do not show oxidative stress or a transcriptional signature characteristic of activated NFR2-mediated signaling in FACS-sorted LSECs. We further observed that primary murine LSECs derived from both wild-type and NRF2 knock-out mice induce Bmp6 in response to acute iron exposure. By analyzing RNA sequencing data of FACS-sorted LSECs from aged versus young mice, as well as upon acute injections of iron citrate in young mice, we identified ETS1 as a candidate transcription factor involved in Bmp6 regulation. By conducting siRNA-mediated knock-down and small-molecule treatments in primary LSECs, we show that Bmp6 transcription is regulated in an NRF2-independent manner via ETS1, which is activated downstream of  p38 MAP kinase-mediated signaling. This knowledge expands the understanding of Bmp6 transcriptional control in both iron-triggered oxidative stress and under the conditions uncoupled to LSECs reactive oxygen species levels.

overall_design: We analyzed in total 5 versus 5 RNA samples derived from FAC-sorted LSECs obtained from young female Balb/c mouse mice, intravenously injected with citrate buffer or iron-citrate solution (150 ug), respectively.

first: Gabriela

last: Zurawska

first: Aneta

last: Jończy

first: Zuzanna

last: Sas

first: Izabela

last: Rumieńczyk

first: Maria

last: Kulecka

first: Katarzyna

last: Piwocka

first: Tomasz

last: Rygiel

first: Michał

last: Mleczko-Sanecka


title: An expanded transcriptome atlas for Bacteroides thetaiotaomicron reveals a small RNA that modulates tetracycline sensitivity

city: Würzburg

first: Alexander

middle: J.

last: Westermann

address: Josef-Schneider-Straße 2/D15

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

summary: Gene expression plasticity allows bacteria to adapt to diverse environments, tie their metabolism to available nutrients, and cope with stress. This is particularly relevant in a niche as dynamic and hostile as the human intestinal tract, yet transcriptional networks remain largely unknown in gut Bacteroides spp. Here, we map transcriptional units and profile their expression levels in Bacteroides thetaiotaomicron over a suite of 15 defined experimental conditions that are relevant in vivo, such as variation of temperature, pH, and oxygen tension, exposure to antibiotic stress, and growth on simple carbohydrates or on host mucin-derived glycans. Thereby, we infer stress- and carbon source-specific transcriptional regulons, including conditional expression of capsular polysaccharides and polysaccharide utilization loci, and expand the annotation of small regulatory RNAs (sRNAs) in this organism. Integrating this comprehensive expression atlas with transposon mutant fitness data, we identify conditionally important sRNAs. One example is MasB, whose inactivation led to increased bacterial tolerance of tetracyclines. Using MS2 affinity purification coupled with RNA sequencing, we predict targets of this sRNA and discuss their potential role in the context of the MasB-associated phenotype. Together, this transcriptomic compendium in combination with functional sRNA genomics—publicly available through a new iteration of the ‘Theta-Base’ web browser (www.helmholtz-hiri.de/en/datasets/bacteroides-v2)—constitutes a valuable resource for the microbiome and sRNA research communities alike.

overall_design: Samples 1-32: RNA-seq of Bacteroides thetaiotaomicron VPI-5482 (wild type) grown in TYG-medium and exposed to different stress conditions (Cold, Heat, Acid, Aerobe, H2O2, Deoxycholate, Bile salts and Gentamicin) or in minimal medium supplemented with different carbon sources (Glucose, Arabinose, Xylose, Maltose, GlcNAc, Mucin and Starvation). Two biological replicates per strain and condition. Samples 33, 34: Genome-wide mapping of transcription start sites via differential RNA-seq (dRNA-seq) of pooled RNA from the above mentioned samples 1-32. Two biological replicates. Samples 35-38: Identification of RNAs (RNA-seq) co-purified with MS2 aptamer tagged MasB sRNA in a masB deletion background and an untagged MasB control strain. Two biological replicates.

first: Daniel

last: Ryan

first: Elise

last: Bornet

first: Gianluca

last: Prezza

first: Shuba

middle: V

last: Alampalli

first: Taís

last: de Carvalho

first: Hannah

last: Felchle

first: Titus

last: Ebbecke

first: Regan

last: Hayward

first: Adam

last: Deutschbauer

first: Lars

last: Barquist


title: Orai-mediated calcium entry regulates the excitability of central dopaminergic neurons

city: Cambridge

first: Rishav

last: Mitra

state: Massachusetts

address: 16 Divinity Avenue

institute: Harvard University

summary: Transcriptomic screen from Drosophila pupal CNS to identify genes regulated by loss of Orai-mediated Ca2+ entry in a subset of dopaminergic neurons  by means of expression of the OraiE180A dominan negative allele

overall_design: Comparative gene expression profiling  for cell sorted 72 hrs APF THD' neurons with or without expression of OraiE180A

first: Shlesha

last: Richhariya

first: Gaiti

last: Hasan


title: Down Syndrome

first: Victor

last: Tybulewicz

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

overall_design: Refer to individual Series


title: Gene expression data of c-kit+ BM cells from Gadd45g-deficient mice with MPN and those from controls

city: TianJin

first: Xiao Tong

last: Ma

state: TianJin

address: Blood hospital, 288 Nanjing Road

department: Institute of Hematology & Blood Diseases Hospital

institute: Chinese Academy of Medical Sciences

summary: To elucidate the mechanisms underlying the pathogenic role of Gadd45g deficiency in myeloproliferative neoplasms, we performed RNA sequencing on c-kit+ BM cells from heterozygous Gadd45g knockout mice  with MPN and those from Ctrl mice.

overall_design: RNA-seq analyses on c-kit+ BM cells from  Gadd45g-deficient mice with MPN in comparison with those from Ctrl.

first: Xiaotong

last: You

first: Peiwen

last: Zhang


title: Transcriptomic changes in embryonic hearts from Dp3Tyb and Ts1Rhr mouse models of Down Syndrome

summary: We performed bulk RNAseq on whole hearts from E13.5 Dp3Tyb embryos and from wild-type littermates, as well as hearts from E13.5 Ts1Rhr embryos and wild-type littermates. Analysis showed the expected increased expression of the Hsa21-orthologous genes on Mmu16 that are present in 3 copies. Gene set enrichment analysis identified pathways that are altered in Dp3Tyb and Ts1Rhr hearts.

overall_design: RNAseq on 5 E13.5 Dp3Tyb embryonic hearts and 5 wild-type littermate embryonic hearts, and on 5 E13.5 Ts1Rhr embryonic hearts and 5 wild-type littermate embryonic hearts.

first: Eva

last: Lana-Elola

first: Miriam

last: Llorian


title: Global chromatin accessibility using an Assay for Transposase-Accessible Chromatin sequencing (ATAC-seq) reveals epigenomic reprogramming of Adipose tissue macrophages toward proinflammatory and proangiogenic phenotypes in formerly obese mice.

city: Montreal

first: Mike

last: Sapieha

state: Quebec

address: 5415 Boul. L'Assomption

institute: Hôpital Maisonneuve-Rosemont

summary: We report sustained changes in the chromatin accessibility landscape of adipose tissue macrophages (ATMs) from high fat diet (HFD)-fed mice persist long after return to regular diet (RD). We compared the data of ATAC-seq performed on nuclei extracted from FACS-sorted ATMs isolated from HFD, RD-RD and HFD-RD-fed mice. Inter-group comparisons revealed the highest diversity and hence the greatest number of differentially accessible regions (DARs) to occur between ATMs from RD-RD and HFD-RD-fed mice, and considerably fewer DARs were identified between ATMs from HFD-RD vs HFD-fed mice. Association of DARs with the nearest gene and gene ontology (GO) enrichment analysis revealed considerable pathway enrichment, especially pathways coding for angiogenesis and inflammatory response, in HFD-RD group when compared to  RD-RD group. The results altogether indicated that most changes in chromatin landscape induced by HFD-feeding are maintained as open chromatin positions for a long time, suggesting that HFD-feeding leads to long-term reprograming of ATMs and renders them prone to pro-angiogenic and pro-inflammatory responses.

overall_design: We performed ATAC-seq on nuclei extracted from FACS-sorted ATMs isolated from HFD, RD-RD and HFD-RD-fed mice.

first: Masayuki

last: Hata


title: Transcriptomic analysis of human foetal hearts with or without Down Syndrome

summary: We performed bulk RNAseq on whole hearts from foetuses with Down syndrome and age-matched, sex-matched euploid controls. Analysis showed the expected increased expression of the genes on Hsa21 that are present in 3 copies. Gene set enrichment analysis identified pathways that are altered in DS hearts.

overall_design: RNAseq on 5 human embryonic hearts with Down syndrome and on 5 euploid age- and sex-matched control hearts.


title: Extracellular niche and tumor microenvironment enhance the impact of KRAS inhibitors in pancreatic cancer models [RNA-seq]

status: Public on Jan 24 2024

city: Buffalo

first: Jianxin

last: Wang

state: NY

address: 665 Elm Street

department: Molecular and Cellular Biology

institute: Roswell Park Comprehensive Cancer Center

summary: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease for which new therapeutic interventions are needed. Here, we assessed the cellular response to pharmacological KRAS inhibition, targeting the central oncogenic factor in PDAC. In a panel of PDAC cell lines, pharmaceutical inhibition of the KRASG12D allele with MRTX1133 yields variable efficacy in the suppression of cell growth and downstream gene expression programs in 2D culture. CRISPR-Cas9 loss-of-function screens identified ITGB1 as a novel driver for enhanced therapeutic response that regulates mechanotransdcution signaling and YAP/TAZ expression, which is further confirmed by gene specific knockdowns and combinatorial drug synergy. Interestingly, MRTX1133 is considerably more efficacious in the context of 3D cell cultures. Moreover, MRTX1133 elicits a more pronounced cytostatic effect in controlling the in vivo tumor growth in PDAC patient-derived xenografts. In syngeneic models, KRASG12D inhibition leads to potent tumor regression that did not occur in immune-deficient hosts. Digital spatial profiling on tumor tissues indicates that MRTX1133-mediated KRAS inhibition enhances interferon-γ signaling and induces antigen presentation that modulates the tumor microenvironment. Further investigation on the immunological response using single-cell sequencing and multispectral imaging reveals that tumor regression is associated with suppression of neutrophils and influx of effector CD8+ T-cells. Thus, both tumor cell-intrinsic and extrinsic events contribute to response and credential KRASG12D inhibition as a promising strategy for a large percentage of PDAC tumors.

overall_design: Patient derived PDAC cell lines, 1222 and 3226 were seeded in 6 well dish at a denisty of 100,000 cells/well. Following 24 hours, the cells were treated with 0.1% DMSO and MRTX1133 (500 nM). The tretments were done in triplicates. The cells were exposed to the MRTX1133 for 48 hours and RNA was extracted using Quiagen RNeasy plus kit.

first: Vishnu

last: Kumarasamy

first: Costakis

last: Frangou

first: Yin

last: Wan

first: Andrew

last: Dynka

first: Hanna

last: Rosenheck

first: Prasenjit

last: Dey

first: Ethan

last: Abel

first: Erik

last: Knudsen

first: Agnieszka

middle: K

last: Witkiewicz


title: Extracellular niche and tumor microenvironment enhance the impact of KRAS inhibitors in pancreatic cancer models.


title: Extracellular niche and tumor microenvironment enhance the impact of KRAS inhibitors in pancreatic cancer models [scRNA-seq]

overall_design: Single cell sequencing was performed on tumor tissues that were derived from AKB6 xenografts developed in  C57BL/6 strain. Mice were treated with vehicle and MRTX1133 (30 mg/kg) once a day via IP administration.Single cells isolated from vehicle treated mice (n=2) were pooled together. SImilarly, single cells isolated from MRTX1133 treated tissues (n=2) were pooled together.


title: Extracellular niche and tumor microenvironment enhance the impact of KRAS inhibitors in pancreatic cancer models [DSP]

overall_design: Digital spatial profiling (DSP) was performed on tumor tissues that were derived from AKB6 xenografts developed in  C57BL/6 strain. Mice were treated with vehicle and MRTX1133 (30 mg/kg) once a day via IP administration. The DSP analysis was performed on vehiclle (n=2) and MRTX1133 treated (n=4) tissues. A total of 6 ROIs  were selected from vehicle treted tissues and 23 ROIs were selected from MRTX1133 treated tissues.


title: Unfolded protein Response is a major hurdle in direct neuronal reprogramming of human astrocytes [scRNA-seq day 20]

city: Planegg-Martinstied

first: Giacomo

last: Masserdotti

address: Groshaderner Strasse 9

department: ISF

institute: Helmholtz Zentrum

summary: Mitochondria account for essential cellular pathways, from ATP production to nucleotide metabolism, and their deficits lead to neurological disorders and contribute to the onset of age-related diseases. Direct neuronal reprogramming aims at replacing neurons lost in such conditions, but almost nothing is known about the impact of mitochondrial dysfunction in human cell direct reprogramming. Here we explore the defects and how to improve the neuronal reprogramming of iPSC-derived astrocytes carrying mutations in NDUFS4 gene, important for Complex I and associated to Leigh Sydrome. This led to the identification of the unfolded protein response as a major hurdle in the direct neuronal conversion of not only patient, but also control astrocytes and fibroblasts. Our data highlight mitochondria-ER stress-mediated inhibition of translation as a key hurdle in direct neuronal reprogramming, emphasizing how disease modelling using patient cells can contribute to unravel novel pathways in human glia cell reprogramming.

overall_design:
pAstros were transduced with control or Ngn2-expressing retrovirus. 2DPT, media was replaced with differentiation media in absence of presence of AMG; 20DPT, DsRed-positive cells were sorted, barcoded with CMOs and analyzed by scRNAseq via 10XGenoics platform

***************************************************************
Raw data will be deposited in GHGA when the database is available
***************************************************************

first: Giovanna

last: Sonsalla

first: Holger

last: Prokisch


title: Autophagy-Mediated Degradation of Integumentary Tapetum Is Critical for Embryo Pattern Formation

status: Public on Feb 20 2024

city: Wuhan

first: Peng

last: Zhao

address: Luojia Hill

department: College of Life Sciences

institute: Wuhan University

summary: To investigate molecular links and underlying mechanisms between autophagy in the integumentary tapetum and embryo pattern formation, we isolated embryos from wild-type plants, atg5 mutants, and atg5 mutants expressing proTPE8:ATG5-GFP for RNA-sequencing.

overall_design: 8-cell embryos from wild-type, atg5 mutants, and atg5 mutants expressing proTPE8:ATG5-GFP were collected for RNA-sequencing. Three independent biological replicates were performed for each genotype.

first: Lin-lin

first: Yicheng

last: Zhong


title: Unfolded protein Response is a major hurdle in direct neuronal reprogramming of human astrocytes


title: The level of protein in the maternal diet modulates facial appearance via mTORC1 signaling

status: Public on Jan 23 2024

city: Tokyo

first: Ruslan

last: Deviatiiarov

address: Hongo 2-1-1, Bunkyo-ku

department: Intractable Disease Research Center

institute: Juntendo University

summary: To elucidate enhancers participating in sculpting human face, we performed CAGE-sequencing of facial mesenchyme of human embryos and cross-referenced identified active enhancers against GWAS datasets for human normal-range facial appearance. Among identified single nucleotide polymorphisms (SNPs) in the active cis-enhancers, several belonged to the components of mTORC1 (Mechanistic Target of Rapamycin Complex 1) pathway. To assess the role of this pathway functionally, genetic and pharmacological manipulations of mTORC1 pathway in mice and zebrafish were performed. They revealed the modulatory role of mTORC1 signaling in fine craniofacial shaping.

overall_design: CAGE-sequencing performed on human facial embryonic material (3-12 weeks).

first: Meng

last: Xie

first: Markéta

last: Tesařová

first: Yakov

last: Gerstein

first: Daniela

last: Schnyder

first: Guzel

last: Gazizova

first: Elena

last: Shagimardanova

first: Tomáš

last: Zikmund

first: Greet

last: Kerckhofs

first: Evgeny

last: Ivashkin

first: Dominyka

last: Batkovskyte

first: Phillip

middle: T

last: Newton

first: Olov

last: Andersson

first: Kaj

last: Fried

first: Oleg

last: Gusev

first: Hugo

last: Zeberg

first: Jozef

last: Kaiser

first: Igor

last: Adameyko

first: Andrei

last: Chagin


title: Unfolded protein Response is a major hurdle in direct neuronal reprogramming of human astrocytes [scRNA-seq 5 day]

first: Giacomo

last: Masserdotti

overall_design:
pAstros were transduced with control or Ngn2-expressing retrovirus. 2DPT, media was replaced with differentiation media in absence of presence of AMG; 5DPT, DsRed-positive cells were sorted, barcoded with CMOs and analyzed by scRNAseq via 10XGenoics platform

***************************************************************
Raw data will be deposited in GHGA when the database is available
***************************************************************

first: Giovanna

last: Sonsalla

first: Holger

last: Prokisch


title: Extracellular niche and tumor microenvironment enhance the impact of KRAS inhibitors in pancreatic cancer models [CRISPR]

first: Jianxin

last: Wang

overall_design: ASPC1 and UM53 cells were infected with the TKOv3 lentiviral particles at an MOI ~0.3 and the positive clones were selected using puromycin to achieve a mutant pool comprising at least 200-fold coverage of the library. The resulting mutant pool was grown in the absence and presence of KRAS inhibitors, which include MRTX1133 (6.25 nM) and MRTX849 ( 500 nM)  for at least 5 passages. Genomic DNA was extracted and subjected to sequencing library construction by amplifying the gRNA inserts

first: Vishnu

last: Kumarasamy

first: Costakis

last: Frangou

first: Yin

last: Wan

first: Andrew

last: Dynka

first: Hanna

last: Rosenheck

first: Prasenjit

last: Dey

first: Ethan

last: Abel

first: Erik

last: Knudsen

first: Agnieszka

last: Witkiewicz