title: Plasticity of intragraft alloreactive T cell clones in human gut correlates with transplant outcomes

city: New York

first: Jianing

last: Fu

state: NY

institute: Columbia University Medical Center

summary: The site of transition between tissue resident memory (TRM) and circulating phenotypes of T cells is unknown. We integrated clonotype, alloreactivity and gene expression profiles of graft-repopulating recipient T cells in the intestinal mucosa at the single cell level after human intestinal transplantation. Host-versus-graft (HvG)-reactive T cells mainly distributed to TRM, Teff/TRM and T follicular helper compartments. RNA velocity analysis demonstrated a trajectory from TRM to effector T (Teff)/TRM clusters in association with rejection. By integrating pre- and post-Tx mixed lymphocyte reaction-determined alloreactive repertoires, we observed that pre-existing HvG-reactive T cells that demonstrated tolerance in the circulation were dominated by TRM profiles in quiescent allografts. Putative de novo HvG-reactive clones showed a transcriptional profile skewed to cytotoxic effectors in rejecting grafts. Inferred protein regulon network analysis revealed upstream regulators that accounted for the effector and tolerant T cell states. We demonstrate Teff/TRM interchangeability for individual T cell clones with known (allo)recognition in human gut, providing novel insight into TRM biology.

overall_design: ScRNA-seq data were generated from FACS-sorted recipient HLA+ CD45+ CD3+ T cells from fresh or frozen/thawed suspensions of intraepithelial lymphocytes (IEL) and/or lamina propria lymphocytes (LPL), isolated from ileal biopsies or tissue resections collected during the late post-Tx (POD626-1764) period from a total of six pediatric ITx patients, including 6 quiescent and 5 rejecting allograft specimens. Samples from Pt15 MVTx POD1194 (IEL + LPL mixed), Pt13 iITx POD1032 IEL and LPL, Pt16” (secondary Tx of Pt16) MVTx POD1004 IEL and LPL, and Pt21 MVTx POD626 (IEL+LPL mixed) were quiescent and free of rejection. All rejecting allograft specimens used in the scRNA-seq study were collected during ileal graft explantation, providing a snapshot of immunological events during graft loss.


title: RNAseq from endothelial cells (ECs) isolated from Li-TSC1KO RagAGTP and wild-type livers at p14 stage of postnatal development

status: Public on Jan 15 2024

city: Madrid

first: Elena

last: Piñeiro-Yáñez

address: Calle de Melchor Fernández Almagro, 3

country: Spain

institute: Spanish National Cancer Research Centre (CNIO)

type: Expression profiling by high throughput sequencing

summary: RNAseq was performed from ECs from four Li-TSC1KO RagAGTP and five wild-type mice at p14.

overall_design: Liver was collected from 4 Li-TSC1KO RagAGTP and 5 wild-type mice at p14. ECs were isolated following a protocol of digestion and immediately collected on Trizol to perform RNA extraction

first: Ana

last: Belén Plata

last: Piñeiro

first: Alejo

last: Efeyan


title: IL-10

status: Public on Jan 11 2024

city: New Haven

first: Rihao

last: Qu

state: CT

address: 300 George Street

country: USA

institute: Yale University

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

overall_design: Refer to individual Series


title: Clinical and molecular features of acquired resistance to immunotherapy [ATAC-seq]

city: Cambridge

first: Danish

last: Memon

address: Robinson Way

department: Cancer Research UK Cambridge Institute

country: United Kingdom

institute: University of Cambridge

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Although immunotherapy with PD-(L)1 blockade is routine for lung cancer, little is known about acquired resistance. Among 1,201 patients with non-small cell lung cancer (NSCLC) treated with PD-(L)1 blockade, acquired resistance is common, occurring in >60% of initial responders. Acquired resistance shows differential expression of inflammation and interferon (IFN) signaling. Relapsed tumors can be separated by upregulated or stable expression of IFNγ response genes. Upregulation of IFNγ response genes is associated with putative routes of resistance characterized by signatures of persistent IFN signaling, immune dysfunction, and mutations in antigen presentation genes which can be recapitulated in multiple murine models of acquired resistance to PD-(L)1 blockade after in vitro IFNγ treatment. Acquired resistance to PD-(L)1 blockade in NSCLC is associated with an ongoing, but altered IFN response. The persistently inflamed, rather than excluded or deserted, tumor microenvironment of acquired resistance informs therapeutic strategies to effectively reprogram and reverse acquired resistance.

overall_design: The molecular changes on acquired resistance (AR) to anti-PD1 were examined using CT26 murine model system. To model acquired resistance, persistent viable CT26 cells following anti-PD-1 treatment were excised, cultured in vitro, and reimplanted in mice. This process was repeated for several passages until CT26 tumors were no longer responsive to anti-PD-1 antibody therapy. ATAC-Seq was performed on the ICB-resistant cancer cell lines derived from tumors from 4th round (n=5) of in vivo passage and compared against the ICB-sensitive in vivo passaged tumor cell line (n=5) or  parental cell line (n=2).

first: George

last: Fromm

first: Taylor

middle: H

last: Schreiber

first: Jayalaksmi

last: Miriyala

first: Kyung

middle: J

last: Yoo


title: Single-cell RNA sequencing analysis of colonic macrophage

summary: Single cell RNA sequencing of sorted Cd45.2 colonic macrophage from bone marrow chimeras from IL-10R KO or IL-10R/CerS2 DKO donors

overall_design: Single cell RNA sequencing of sorted CD45.2/CD11B/CD11c/CD64+ cells isolated from the colons of chimericic mice with IL-10R KO or IL-10R/CerS2 DKO bone marrow donors

first: Richard

middle: A

last: Flavell

first: Autumn

last: York


title: Bulk RNA sequencing analysis of WT and IL-10 knockout bone marrow derived macrophage (BMDMs)

summary: The goal of this experiment was to identify differentially expressed genes in response to 24h TLR2 activation from BMDMs from WT (JAX 000664) or IL-10 KO ( JAX 002251)

overall_design: Two biological replicates per group. Bone marrow was differentiated for 8 days with MCSF before 24h TLR2 activation with PamCSK4. Samples were harvested eith trizol and extracted with combination Trizol/RNAeasy kit for RNA isolation.


title: Histone butyrylation in the mouse intestine is mediated by the microbiota and associated with regulation of gene expression

status: Public on Jan 13 2024

city: Cleveland

first: Leah

last: Gates

state: Ohio

address: Case Western Reserve University

institute: Case Western Reserve University - Cleveland, OH

summary: We characterized the genomic localization of select histone modifications  in mouse cecal intestinal epithelial cells (IECs), including butyrulation on histone H3 lysine 27 (H3K27bu), as well as H3K27ac, H3K4me3, and H3K27me3. We also analyzed the response of IECs to different treatments of ampicillin and tributyrin (an analog of butyrate) by RNA-seq.

overall_design: Characterization of histone modifications in cecal intestinal epithelial cells (IECs) and analysis of RNA expression in mice treated with or without ampicillin and tributyrin.

first: C

middle: D

last: Allis

first: Matthew

middle: R

last: Paul

first: Thomas

middle: S

last: Carroll


title: Cell atlas of the Atlantic salmon spleen reveals immune cell heterogeneity and cell-specific responses to bacterial infection

city: Edinburgh

first: Jianxuan

last: Sun

state: Midlothian

address: The University of Edinburgh, Easter Bush Campus

institute: Roslin Institute

summary: The spleen is a conserved secondary lymphoid organ that emerged in parallel to adaptive immunity in early jawed vertebrates. Recent studies have applied single cell transcriptomics to reveal the cellular composition of spleen in several species, cataloguing diverse immune cell types and subpopulations. In this study, 51,119 spleen nuclei transcriptomes were comprehensively investigated in the commercially important teleost Atlantic salmon (Salmo salar L.), contrasting control animals with those challenged with the bacterial pathogen Aeromonas salmonicida. We identified clusters of nuclei representing the expected major cell types, namely T cells, B cells, natural killer-like cells, granulocytes, mononuclear phagocytes, endothelial cells, mesenchymal cells, erythrocytes and thrombocytes. We discovered heterogeneity within several immune lineages, providing evidence for resident macrophages and melanomacrophages, infiltrating monocytes, several candidate dendritic cell subpopulations, and B cells at distinct stages of differentiation, including plasma cells and an igt+ B subset. We provide evidence for twelve candidate T cell subsets, including cd4+ T helper and regulatory T cells, one cd8+ subset, three γδT subsets, and populations double negative for cd4 and cd8. The number of genes showing differential expression during the early stages of infection with Aeromonas was highly variable across cell types, with the largest changes observed in resident macrophages and infiltrating monocytes, followed by resting mature B cells. Our differential expression analysis offers evidence for a strong inflammatory response to bacteria in the spleen driven by macrophages and monocytes, and the recruitment of splenic igt+ B cells to the gut. Overall, this study provides a new cell-resolved perspective of the immune actions of Atlantic salmon spleen, highlighting extensive heterogeneity hidden to bulk transcriptomics. We further provide a large catalogue of cell-specific marker genes that can be leveraged to further explore the function and structural organization of the salmonid immune system.

overall_design: Twenty Atlantic salmon were anaesthetized and given an intraperitoneal injection of either PBS (n=10), or the pathogenic Hooke strain of A. salmonicida (n=10). Sampling was performed 24 h post-injection and control (n=2) and infected (n=2) fish were selected based on immune response validated with qPCR. Single nucleus libraries were constructed according using the 10x Genomics Chromium platform.

first: Rose

last: Daniels

first: Adam

last: Balic

first: Adriana

middle: M

last: Andresen

first: Håvard

last: Bjørgen

first: Ross

last: Dobie

first: Neil

middle: C

last: Henderson

first: Erling

middle: O

last: Koppang

first: Samuel

last: Martin

first: Johanna

last: Hol Fosse

last: Taylor

first: Daniel

last: Macqueen


title: Clinical and molecular features of acquired resistance to immunotherapy [RNA-seq]

first: Danish

last: Memon

overall_design: The molecular changes on acquired resistance (AR) to anti-PD1 were examined using CT26 murine model system. To model acquired resistance, persistent viable CT26 cells following anti-PD-1 treatment were excised, cultured in vitro, and reimplanted in mice. This process was repeated for several passages until CT26 tumors were no longer responsive to anti-PD-1 antibody therapy. Bulk RNA-seq was performed on the ICB-resistant cancer cell lines derived from tumors from the 2nd round (n=2) and 4th round (n=4) of in vivo passage and compared against the ICB-sensitive parental cell line (n=3). Examination of mRNA levels from ICB-sensitive parental cell line (n=3) and anti-PD1 resistant cancer cell lines derived from tumors from the 2nd round (n=2) and 4th round (n=4) of in vivo passage, with or without in vitro IFNγ stimulation.

first: George

last: Fromm

first: Taylor

last: Schreiber

first: Jayalaksmi

last: Miriyala

first: Kyung

last: Yoo


title: A precise balance of TETRASPANIN1/TORNADO2 activity is required for vascular proliferation and ground tissue patterning in Arabidopsis

city: Ghent

first: Bert

last: De Rybel

address: Technologiepark 71

department: Center for Plant Systems Biology

country: Belgium

institute: VIB

summary: The molecular mechanisms guiding oriented cell divisions in the root vascular tissues of Arabidopsis thaliana are still poorly characterized. By overlapping bulk and single-cell transcriptomic datasets, we unveiled TETRASPANIN1 (TET1) as a putative regulator in this process. TET1 is expressed in root vascular cells and loss-of-function mutants contain fewer vascular cells files. We further generated and characterized a CRISPR deletion mutant and show, unlike previously described mutants, that the full knock out is additionally missing endodermal cells in a stochastic way. Finally, we show that HA-tagged versions of TET1 are functional in contrast to fluorescent TET1 translational fusions. Immunostaining using HA-TET1 lines complementing the mutant phenotype suggested a dual plasma membrane and intracellular localization in the root vasculature and a polar membrane localization in young cortex, endodermal and initial cells. Taken together, we show that TET1 is involved in both vascular proliferation and ground tissue patterning. Our initial results pave the way for future work into deciphering its precise mode of action.

overall_design: Arabidopsis seedlings of the pRPS5A::DOF2.1-GR line were grown on control plates and transferred to plates suppemented with 10uM DEX for the indicated times to induce the DOF2.1 transcription factor. Root meristems were harvested and RNA extraction was performed before RNA sequencing.

first: Nataliia

last: Konstantinova

first: Eliana

last: Mor

first: Eline

last: Verhelst

first: Jonah

last: Nolf

first: Kenzo

last: Vereecken

first: Feng

last: Wang

first: Mieke

last: van Lijsebettens

last: Van Damme

first: Matouš

last: Glanc


title: Clinical and molecular features of acquired resistance to immunotherapy in non-small cell lung cancer

overall_design: Tumor tissue samples from pre- and post-immunotherapy treatment timepoints were obtained from patients with NSCLC patients treated with PD-1 blockade (n = 29). All samples were processed as FFPE. 13 samples were obtained prior to initiation of therapy (pre-treatment) and 29 samples were obtained at time of acquired resistance (post-treatment). The RNA samples quantification was performed using the Affymetrix Clariom D Pico assay.

first: Martin

middle: L

last: Miller

first: Matthew

last: Hellmann

first: Adam

last: Schoenfeld


title: Starvation resistant cavefish reveal conserved mechanisms of starvation-induced hepatic lipotoxicity

city: Greater Noida

first: Sumeet

middle: Pal

last: Singh

state: Uttar Pradesh

address: NH91, Tehsil Dadri

department: Life Sciences

country: India

institute: Shiv Nadar Institution of Eminence

summary: Starvation is a severe form of malnutrition that occurs when an individual's intake of food is inadequate to meet their body's energy requirements. Prolonged starvation can cause permanent organ damage, stunted growth in children, and death if left untreated. It is estimated that approximately 45% of deaths among children under the age of 5 years are linked to undernutrition. Notably, the liver’s health is compromised during starvation. More than 60 years ago, it was recognized that chronic severe malnutrition in children, such as kwashiorkor, leads to fatty liver. Subsequently, starvation has been shown to induce accumulation of liver fat (steatosis) in drosophila, zebrafish, and humans. To uncover potential strategies for protecting the liver, we turned to adaptation strategies present in a genetically tractable and naturally occurring model of starvation resistance - the Astyanax mexicanus model system. Cave populations of this species have adapted to survive in conditions of extreme starvation, while the surface populations of the same species display relatively normal vertebrate physiology. We took advantage of this unique system to study the response of the liver to starvation and identified reduced expression of the fatty acid uptake gene slc27a2a/fatp2 allowing cavefish to prevent liver damage under starvation. Targeting this pathway in both fish and flies mitigates starvation-induced hepatic steatosis and protects the liver from lipotoxicity in zebrafish. This demonstrates that the identified pathway is evolutionary conserved for over 400 million years, highlighting its potential as a druggable target.

overall_design: For RNA-Seq, RNA was isolated from livers from fed and fasting animals. For this, livers from 40 larvae / replicate were dissected and collected in the lysis buffer from ReliaPrepTM RNA Miniprep Systems (Promega, Z6110). RNA was isolated from the lysed tissue by following the manufacture’s instruction. cDNA synthesis, library preparation and Illumina sequencing was performed according to the manufacturer’s instructions using the TruSeq Stranded mRNA Preparation Kit for Illumina (Ilumina #20020594). Purified libraries were quantified using a Quibit fluorometer (Life Technologies) and the quality assessed on the Agilent Bioanalyzer (Agilent Technologies). Sequencing was performed on the Illumina NextSeq 500 instrument as High Output with single-end 75bp reads. Following sequencing, raw reads were demultiplexed into Fastq format using Ilumina bclfastq2 v2.18. Raw reads were mapped using STAR aligner v2.7.3a against the UCSC genome astMex_2.0 genome, using Ensembl 102 gene models for annotations. Transcript abundance was quantified using RSEM v1.3.0.

first: Rohner

last: Nicolas


title: Compact zinc finger architecture utilizing toxin-derived cytidine deaminases for highly efficient base editing in human cells-K562 cells

status: Public on Jan 12 2024

city: Richmond

first: Jessica

last: Davis

state: CA

address: 501 Canal Blvd

institute: Sangamo Therapeutics

summary: Nucleobase editors represent an emerging technology that enables precise single-base edits to the genomes of eukaryotic cells. Most nucleobase editors use deaminase domains that act upon single-stranded DNA and require RNA-guided proteins such as Cas9 to unwind the DNA prior to editing. However, the most recent class of base editors utilizes a deaminase domain, DddAtox, that can act upon double-stranded DNA. Here, we target DddAtox fragments and a FokI-based nickase to the human CIITA gene by fusing these domains to arrays of engineered zinc fingers (ZFs). We also identify a broad variety of Toxin-Derived Deaminases (TDDs) orthologous to DddAtox that allow us to fine-tune properties such as targeting density and specificity. TDD-derived ZF base editors enable up to 73% base editing in T cells with good cell viability and favorable specificity.

overall_design: Primer pools targeting potential base edited sites identified by the ZF-CBE genome-wide specificity assay were designed and synthesized. Potential base edited sites were amplified from genomic material isolated from transfected K562 cells and analyzed for base editing levels.

first: Friedrich

last: Fauser

first: Bhakti

middle: N

last: Kadam

first: Sebastian

last: Arangundy-Franklin

middle: E

first: Vishvesha

last: Vaidya

first: Nicola

last: Schmidt

first: Garrett

last: Lew

first: Danny

middle: F

last: Xia

first: Rakshaa

last: Mureli

first: Colman

last: Ng

first: Yuanyue

last: Zhou

first: Nicholas

last: Scarlott

first: Jason

last: Eshleman

first: Yuri

last: Bendaña

first: David

last: Shivak

first: Andreas

last: Reik

first: Patrick

last: Li

first: Gregory

first: Jeffrey


title: Spatial Transcriptomics of Pancreatic Tumors Reveal Distinct Compartmentalisation of Neutrophil Subsets [Spatial Transcriptomics]

city: Singapore

first: Melissa

address: 8A Biomedical Grove, #03-06 Immunos Building, Singapore

country: Singapore

institute: Singapore Immunology Network

summary: Neutrophils are increasingly recognized as key players in the tumour immune response, and are associated with poor clinical outcomes. Multiple studies have characterised various neutrophil states within the tumor, with associated pro- and anti- tumoral functions. However, it is unknown if this heterogeneity of neutrophils is contributed by their surrounding tumor microenivironment. To answer this, we performed 10X Visium spatial transcriptomics on a murine model of pancreatic cancer. We discovered that terminally differentiated T3 neutrophils localise primarily in regions near hypoxic and glycolytic areas, while transitional immature T1 and mature T2 neutrophils localise towards the peripheral regions of the tumors. This results points to a spatially dependent role of T3 neutrophils, which may be critical to carry out pro-tumorigenic functions.

overall_design: Frozen samples were quartered before embedding in optimal cutting temperature‒filled molds and sectioned to a thickness of 10 μm at −20 °C with cryotome (CM3050S, Leica). To ensure capture of tumor-infiltrating neutrophils, cryosections were first screened for Ly6G immunofluorescence before mounting on chilled Visium Tissue Optimization Slides (1000193, 10X Genomics) and Visium Spatial Gene Expression Slides (1000184, 10X Genomics). For processing, we fixed tissue sections onto the slide in chilled methanol and prepared them for immunostaining according to manufacturer’s protocol. Sections were stained with Rabbit anti-wide spectrum cytokeratin (Abcam) and Rat Ly6G (1A8, Biolegend) for 30 min. Subsequently, sections were washed with wash buffer for 5 times before secondary staining with Anti-Rabbit Alexa Fluor 488 (Invitrogen) and Anti-Rat Alexa Fluor 555 (Invitrogen) and DAPI for 30 minutes. Upon completion of immunostaining, slides were scanned with the slide scanner (EVOS Auto FL2, Thermo Scientific). After immunofluorescence imaging, Visium Spatial Gene Expression libraries were prepared according to the manufacturer’s protocol. The libraries were sequenced on either HiSeq X or NovaSeq 6000 S4 (Illumina) at PE150 and a read depth of 50,000 read-pairs per tissue-covered spot.

first: Immanuel

last: Kwok

first: Leonard

last: Tan

first: Yingrou

first: Shih

last: Changming

first: Mengwei

last: Ang

first: Jinmiao

last: Chen

first: Laiguan


title: NIC/VC and NIC/p66 mammary tumors

city: Montréal

first: Claudia

last: Kleinman

state: Québec

address: 3999 Côte Ste-Catherine Road

department: Human Genetics

country: Canada

institute: Lady Davis Institute for Medical Research

summary: Mammary tumors of parental and aggressive variants of an ErbB2+ breast cancer mouse model that differs in the degree to which is is experiencing oxidative stress as a result of forced expression of an oxidative stress inducer called p66ShcA.

overall_design: NIC are ErbB2+ murine breast cancer cells, which lack p66ShcA.  We engineered the parental cells to constitutively overexpress p66ShcA.  These NIC/p66Low and NIC/p66High breast cancer cells were serially passaged in the mammary fat pads of mice and after two rounds of in vivo selection, the cell lines were explanted back into culture.  Both explanted populations were more aggressive (higher tumor growth rate and increased metastasis) compared to their parental counterparts.  RNAseq was performed on bulk tumors (3 tumors each) from the following groups: NIC/p66Low-Parental; NIC/p66Low-Aggressive; NIC/p66High-Parental; NIC/p66High-Aggressive.

first: Rachel

last: La Selva

first: Steven

last: Hebert

first: Josie

last: Ursini-Siegel


title: Profiling chromatin accessibility of neutrophil subsets in a mouse model of pancreatic cancer [ATAC-Seq]

last: Ng

summary: Neutrophils are increasingly recognized as key players in the tumour immune response, and are associated with poor clinical outcomes. Despite recent advances characterizing the diversity of neutrophil states in cancer, common trajectories and mechanisms governing the ontogeny and relationship between these neutrophil states remain undefined.Using Assay for Transposase Accessible Chromatin (ATAC) sequencing, we show that immature and mature neutrophils infiltrating the tumor share a similar chromatin accessibility profile that was not observed in non-tumor populations (in both tumor-bearing and wild type mice). Our results thus indicate that tumor-infiltrating neutrophils can be reprogrammed by the tumor microenvironment regardless of their stage of maturity by converging chromatin trajectories towards a distinct T3 state.

overall_design: CD11b+CD115-Ly6G+CD101+ mature and CD101- immature neutrophils were sorted by fluorescence activated cell sorting (FACS) from the bone marrow (BM), spleen (SPL), blood (BLD) and tumor (TUM) from mice bearing orthotopic tumors previously established from the Pdx1Cre; KrasG12D/+; Trp53R172H/+ (KPC) genetically modified mouse model (n=3 for all except mature blood neutrophils, n=3). Mature and immature neutrophils from the bone marrow of wild type mice were also further sorted as controls (n=3).

first: Changming

last: Shih

first: Nicholas


title: Deterministic reprogramming of neutrophils in tumors


title: Single cell RNA profiling of neutrophils in a model of pancreatic orthotopic cancer

first: Li

last: Zeng

address: 11 Jln Tan Tock Seng

institute: National Neuroscience Institute

summary: Neutrophils are increasingly recognized as key players in the tumour immune response, and are associated with poor clinical outcomes. Despite recent advances characterizing the diversity of neutrophil states in cancer, common trajectories and mechanisms governing the ontogeny and relationship between these neutrophil states remain undefined. Single-cell RNA sequencing (scRNAseq) of neutrophils from the bone marrow, spleen, and blood in addtion to the pancreatic tumor thus uncovers an unique tumor differentiation trajectory, where immature and mature neutrophils converge into a terminally differentiated tumor neutrophil state.

overall_design: CD11b+CD115-Ly6G+ neutrophils were sorted by fluorescence activated cell sorting (FACS) from the bone marrow, spleen, blood and tumor from mice bearing orthotopic tumors previously established from the Pdx1Cre; KrasG12D/+; Trp53R172H/+ (KPC) genetically modified mouse model (n=2). Each sample was independently labelled with TotalSeq-A hashtag antibodies (A0301-A0308), before pooling for 10X v3 3' scRNAseq.


title: Effect of GNAS mutation on the gene expression of HCT116 colon cancer cell line

city: Helsinki

first: Pirjo

last: Nummela

address: Haartmaninkatu 8

department: Research Programs Unit

country: Finland

institute: University of Helsinki

summary:
To investigate the effect of activating GNAS mutation on gene expression of colorectal cancer cells, we utilized the GNAS R201C knockin cell line produced by Horizon Discovery Ltd.
We compared the gene expression of parental and GNAS R201C/+ HCT116 cells with RNA-Seq, using four biological replicates.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for parental HCT116 cell line and its GNAS mutant derivative.

first: Ari

last: Ristimäki


title: SARS-CoV-2-induced disruption of a vascular bed in a microphysiological system caused by type-I interferon from bronchial organoids

city: Kyoto

first: Kazuo

last: Takayama

state: Kyoto

address: Shogoin Kawaharacho 53, Sakyo-ku

department: Center for iPS Cell Research and Application

country: Japan

institute: Kyoto University

summary: Blood vessels show various COVID-19-related conditions including thrombosis and cytokine propagation. Existing in vitro blood vessel models cannot represent the consequent changes in the vascular structure or determine the initial infection site, making it difficult to evaluate how epithelial and endothelial tissues are damaged. Here, we developed a microphysiological system (MPS) that co-culture the bronchial organoids and the vascular bed to analyze infection site and interactions. In this system, virus-infected organoids caused damage in vascular structure. However, vasculature was not damaged or infected when the virus was directly introduced to vascular bed. The knockout of interferon-related genes and inhibition of the JAK/STAT pathway reduced the vascular damage, indicating the protective effect of interferon response suppression. The results demonstrate selective infection of bronchial epithelial cells and vascular damage by cytokines and also indicate the applicability of MPS to investigate how the infection influences vascular structure and functions.

overall_design: RNA-seq in the uninfected and infected BO : Total RNA was collected, and then RNA-seq was performed.


title: WDR5 activity regulates nuclear changes and migration of acute lymphoblastic leukemia cells across confined spaces [Jurkat]

first: Javier

last: Redondo-Muñoz

state: Madrid

address: Ramiro de Maeztu,9

department: Molecular Biomedicine

institute: CIB-CSIC

summary: Purpose: Acute lymphoblastic leukemia (ALL) is the most common pediatric cancer and  infiltration of leukemic cells is critical for disease progression and relapse. In spite of the  canonical functions of epigenetics in transcription and DNA homeostasis, its contribution to  the nuclear deformability of migrating leukemic cells remains unclear. The objective was to  evaluate the role of WDR5 (a core subunit of the histone H3K4 methyltransferases) during  leukemia cell migration.  Experimental design: We used ALL cell lines and primary samples from patients to study  their response and migration in 3D environments and how they extravasate in a mouse  xenograft model. We also performed, biophysical, transcriptional and ChIP-sequencing  analyses to determine the mechanism by which WDR5 controls ALL progression.  Results: We observed that WDR5 activity is critical for ALL cell invasion into 3D collagen  matrices. Notably, blocking WDR5 activity also decreased the extravasation of ALL cells in  an in vivo model. We identified that 3D constrained conditions promoted H3K4 methylation  in ALL cells that altered the global chromatin configuration and transcriptional changes  related to cell cycle and DNA replication. WDR5 inhibition did not impair cell adhesion or  cell response to chemokines; but we demonstrated that 3D conditions affected the  deformation and biophysical behavior of the nucleus in ALL cells.  Conclusions: We conclude that confined conditions have a fundamental role in ALL cell  biology and provide novel molecular and biophysical mechanisms used by leukemia cells to  disseminate. Targeting WDR5 might be a promising therapeutic strategy against ALL  infiltration and dissemination.

overall_design: 3 replicates of Jurkat control and cultured in 3D conditions for 3 hours were used for chromatin immunoprecipitation of H3K4me3 and their DNA was extracted and sequenced. DNA from a control sample of the ChIP and from cells before de ChIP (input) was also sequenced.

first: Raquel

last: González-Novo

first: África

last: González-Murillo

first: Elena

last: Madrazo

first: David

last: Acitores

first: Mario

last: García de Lacoba

first: Manuel

last: Ramírez


title: scRNAseq of bone marrow (BM) T cells and hematopoietic stem and progenitor cells (HSPCs) of acute myeloid leukemia patients post allogeneic stem cell transplantation (alloSCT).

city: Heidelberg

first: Anna

last: Mathioudaki

state: None / N/A

address: Meyerhofstraße 1, 1

country: Germany

institute: EMBL Heidelberg

summary: Gene expression profile at single cell level of bone marrow (BM) T cells and hematopoietic stem and progenitor cells (HSPCs) of acute myeloid leukemia patients post allogeneic stem cell transplantation (alloSCT) of patients in complete remission (CR) and relapse (REL).

overall_design: HSPCs and T cells were isolated using Fluorescence-activated cell sorting (FACS) according to the presence of CD34 and CD3 respectively, and were then analyzed using scRNAseq. Cells from all samples per group were pooled in one 10x reaction.

first: Xizhe

last: Wang

first: Judith

last: Zaugg

first: Caroline

last: Pabst


title: YAP promotes cardiomyocyte cell cycle entry

city: Houston

first: Jong

last: Kim

state: TX

address: One Baylor Plaza

institute: Baylor College of Medicine

summary: Specialized adult somatic cells, such as cardiomyocytes (CMs), are highly differentiated with poor renewal capacity, an integral reason underlying organ failure in disease and aging. Among the least renewable cells in the human body, CMs renew approximately 1% annually. Consistent with poor CM turnover, heart failure is the leading cause of death. Here, we show that an active version of the Hippo pathway effector YAP, termed YAP5SA, partially reprograms adult mouse CMs to a more fetal and proliferative state. One week after induction, 19% of CMs that enter S-phase do so twice, CM number increases by 40%, and YAP5SA lineage CMs couple to pre-existing CMs. Genomic studies showed that YAP5SA increases chromatin accessibility and expression of fetal genes, partially reprogramming long-lived somatic cells in vivo to a primitive, fetal-like, and proliferative state.

overall_design: We conditionally expressed a Yap isoform incapable of being phosphorylated by Lats1/2, YAP5SA, in CMs using a mouse model that we described previously (Monroe et al., 2019). We then performed cardiac Drop-seq on Myh6-MerCreMer/+; YAP5SA-tg/+ (hereafter referred to as YAP5SA) mice, in addition to control animals.

first: Matthew

last: Hill


title: Mitochondrial complex I activity in microglia sustains neuroinflammation [scRNA-seq II]

status: Public on Jan 10 2024

first: Irina

last: Mohorianu

address: Puddicombe Way

department: Wellcome-MRC Cambridge Stem Cell Institute

summary: Sustained smouldering, or low grade, activation of myeloid cells is a common hallmark of several chronic neurological diseases, including multiple sclerosis (MS). Distinct metabolic and mitochondrial features guide the activation and the diverse functional states of myeloid cells. However, how these metabolic features act to perpetuate neuroinflammation is currently unknown. Using a multiomics approach, we identified a new molecular signature that perpetuates the activation of myeloid cells through mitochondrial complex I (CI) activity driving reverse electron transport (RET) and the production of reactive oxygen species (ROS). Blocking CI activity in pro-inflammatory microglia protected the central nervous system (CNS) against neurotoxic damage and improved functional outcomes in animal disease models in vivo. Our data show that mitochondrial CI in microglia is a potential new therapeutic target to foster neuroprotection in smouldering inflammatory CNS disorders.

overall_design: MOG-EAE induction in WT and Nd6 mice

first: Luca

last: Peruzzotti-Jametti

first: Cory

last: Willis

first: G

last: Krzak

first: Liviu

last: Pirvan

first: Stefano

last: Pluchino


title: AKT1 interacts with DHX9 to mitigate R-loop-induced replication stress in ovarian cancer

city: Bethesda

first: Jayakumar

last: Nair

state: MD

address: 10 center drive

department: Women malignancies Branch

institute: National Cancer Institute

summary: PARP inhibitor (PARPi)-resistant BRCA mutant (BRCAm) high-grade serous ovarian cancer (HGSOC) represents a new clinical challenge with unmet therapeutic needs. Quantitative high-throughput drug combination screen identifies that ATR inhibitor (ATRi) and AKT inhibitor (AKTi) combination is a rational treatment strategy for both PARPi-sensitive and PARPi-resistant BRCAm HGSOC by inducing DNA damage and R-loop-mediated replication stress (RS). Mechanistically, the kinase domain of AKT1 directly interacts with DHX9, thus facilitating recruitment of DHX9 to R-loops. AKTi increases ATRi-induced R-loop-mediated RS by mitigating recruitment of DHX9 to R-loops. Moreover, DHX9 is upregulated in tumors from PARPi-resistant BRCAm HGSOC patients and high co-expression of DHX9 and AKT1 correlates with worse survival. Our study reveals a previously unknown interaction between AKT1 and DHX9 in R-loop resolution and novel mechanisms of action of AKTi and ATRi combination. Our data also provide a rationale for the clinical development of ATRi and AKTi combination for BRCAm HGSOC irrespective of PARPi resistance status and a potential biomarker to predict the response of combination therapy.

overall_design: RNA sequencing (RNAseq) data were obtained from total RNA from fresh frozen tumor biopsy samples from 14 BRCAm PARPi-resistant HGSOC patients enrolled in the clinical trial at the National Cancer Institute (ClinicalTrials.gov, NCT02203513). RNAseq was performed using a HiSeq4000 sequencing system (Illumina) at the CCR Sequencing Facility/NCI on pre-treatment biopsy samples from 14 patients.  Paired-end sequencing was done.  Quartile normalization and log-transformation prior to analysis were performed on datasets.

first: Tzu-Ting

last: Huang

first: Chih-Yuan

last: Chiang

first: Kelli

last: Wilson

first: Ken

last: Cheng

first: Jung-Min

last: Lee


title: Synthetic dual-costimulation for TCR and TCR-like targeted cell therapies

first: Michel

last: Sadelain

address: 1275 York Avenue, Box 182

institute: Memorial Sloan Kettering Cancer Cener

summary: To characterize the phenotype of human HIT T-cells endowed with a novel synthetic co-stimulatory molecule isolated from a mouse xenograft model compared to a control.

overall_design: human CD45+ cells were isolated at day 9 post injection from bone marrows of tumour-bearing mice treated with HIT T-cells.  Experimental and control samples were hash-tagged and pooled for single cell RNAseq.

first: Anton

last: Dobrin

first: Hongyao

last: Xie

first: Karlo

last: Perica

first: Nayan

last: Jain

first: Andrew

last: Chow

first: Jedd

last: Wolchok

first: Taha

last: Merghoub

first: Mohamad

last: Hamieh


title: A phenotypic screening platform for identifying chemical modulators of astrocyte reactivity [BulkRNAseq.ONC]

first: Benjamin

last: Clayton

state: OH

address: 2109 Adelbert Rd BRB739A

department: Genetics and Genome Sciences

institute: Case Western Reserve University

summary: Disease, injury, and aging induce pathological reactive astrocyte states that contribute to neurodegeneration. Modulating reactive astrocytes therefore represents an attractive therapeutic strategy. Here, we describe the development of an astrocyte phenotypic screening platform for identifying chemical modulators of astrocyte reactivity. Leveraging this platform for chemical screening, we identify HDAC3 inhibitors as effective suppressors of pathological astrocyte reactivity. We demonstrate that HDAC3 inhibition reduces molecular and functional characteristics of reactive astrocytes in vitro. Transcriptional and chromatin mapping studies show that HDAC3 inhibition disarms pathological astrocyte gene expression and function while promoting the expression of genes associated with beneficial astrocytes. Administration of RGFP966, a small molecule HDAC3 inhibitor, blocks reactive astrocyte formation and promotes neuroprotection in vivo in mice. Collectively, these results establish a platform for discovering modulators of reactive astrocyte states, inform the mechanisms that control astrocyte reactivity, and demonstrate the therapeutic benefits of modulating astrocyte reactivity for neurodegenerative diseases.

overall_design: Transcriptional profiling of the left and right retina from mice that had their right optic nerve crushed (ONC) and left optic nerve left intact (CTRL) followed by treatment for 7d with daily i.p. injections of either vehicle or 10mg/kg RGFP966. Retina were collected at 7 days post crush.

first: Paul

last: Tesar


title: A phenotypic screening platform for identifying chemical modulators of astrocyte reactivity [ATAC-seq]

overall_design: Assay for transposase-accessible chromatin with high-throughput sequencing (ATACseq) was performed on resting astrocytes, reactive astrocytes (treated with TNFa, IL1a, and C1q), and reactive  astrocytes treated with either RGFP966 or JSH-23 to identify regions of open chromatin and how they change as astrocytes go from a resting to reactive state.


title: Mitochondrial complex I activity in microglia sustains neuroinflammation [scRNA-seq III]

first: Irina

last: Mohorianu

institute: University of Cambridge

overall_design: MOG-EAE induction in NDUFS4_WT and NDUFS4_KO mice

first: Luca

last: Peruzzotti-Jametti

first: Cory

last: Willis

first: G

last: Krzak

first: Liviu

last: Pirvan

first: Stefano

last: Pluchino


title: High/low MNase-seq in foregut, Control and FOXA-TKD

city: Cincinnati

first: Hee-Woong

last: Lim

address: 3333 Burnet Ave. MLC 7024

department: Division of Biomedical Informatics

institute: Cincinnati Children's Hospital Medical Center

summary: Nucleosome occupancy and positioning were examined in foregut stage to compare control and FOXA-TKD using high/low MNase-seq

overall_design: low-MNase-seq for identify accessible/fragile nucleosomes and high-Mnase-seq for identify stable nucleosomes

first: Satoshi

last: Matsui

first: Makiko

last: Iwafuchi


title: Pioneer transcription factors coordinate active and repressive gene expression states to regulate cell fate

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

overall_design: Refer to individual Series


title: Molecular profiling of osteoprogenitor cells reveals FOS as a master regulator of bone non-union [tissue]

city: Bern

first: Daniel

middle: Walter

last: Hanhart

state: Bern

address: Murtenstrasse 24

department: Department for BioMedical Research (DBMR)

country: Switzerland

institute: UroGenus

summary: Despite the advances in bone fracture treatment, a significant fraction of fracture patients will develop non-union. Most non-unions are treated with surgery since identifying the molecular causes of these defects is exceptionally challenging. In this study, compared with marrow bone, we generated a transcriptional atlas of human osteoprogenitor cells derived from healing callus and non-union fractures. Detailed comparison among the three conditions revealed a substantial similarity of callus and nonunion at the gene expression level. Nevertheless, when assayed functionally, they showed different osteogenic potential. Utilizing longitudinal transcriptional profiling of the osteoprogenitor cells, we identified FOS as a putative master regulator of non-union fractures. We validated FOS activity by profiling a validation cohort of 31 tissue samples. Our work identified new molecular targets for non-union classification and treatment while providing a valuable resource to better understand human bone healing biology.

overall_design: Validated FOS activity by profiling a validation cohort of 31 tissue samples.

first: Federico

last: La Manna

first: Peter

last: Kloen

first: Andre

last: van Wijnen

first: George

last: Thalmann

first: Marianna

last: Kruithof-de Julio

first: Panagiotis

last: Chouvardas


title: Type 2 inflammation reduces SARS-CoV-2 replication in the airway epithelium in allergic asthma through functional alteration of ciliated epithelial cells

city: Seattle,

first: Stephanie

last: Osmond

state: WA

address: 1201 9th Ave

institute: Benaroya Research Institute


title: Mitochondrial complex I activity in microglia sustains neuroinflammation [bulk]

first: Irina

last: Mohorianu

overall_design: Derivation of human induced microglia (hiMG) from human induced pluripotent cells (hiPSCs)


title: Molecular profiling of osteoprogenitor cells reveals FOS as a master regulator of bone non-union [timecourse]

overall_design: OPC were cultured in basal medium (DMEM with 1 g/L glucose, supplemented with 1x GlutaMax, 1% penicillin/streptomycin and 10% FBS). Cells were harvested at mac 80% confluence and processed for RNASeq (sampes "day 0") or seeded in 6-well plates to undergo a timecourse of osteogenic differentiation in basal medium supplemented with dexamethasone 10 nM, beta-glycerophosphate 2 mM and L-ascorbic acid-2-phosphate 50 ug/mL. Cells were then harvested after 7, 14, 21 or 28 days of differentiation (samples "day 7", "day 14", "day 21" or "day 28" respectively)


title: A phenotypic screening platform for identifying chemical modulators of astrocyte reactivity [CUT&RUN]

first: Benjamin

last: Clayton

overall_design: CUT&RUN analysis of RelA/p65 and H3K27ac in primary mouse resting astrocytes, reactive astrocytes (treated with TNFa, IL1a, and C1q), reactive astrocytes treated with 5uM RGFP966, and reactive astrocytes treated with 5uM JSH-23.

first: Paul

last: Tesar


title: Divergent transcriptomic signatures from putative mesenchymal stimuli in glioblastoma cells

city: Charlottesville

first: Matthew

last: Lazzara

state: VA

address: 345 Crispell Dr

institute: University of Virginia

summary: In glioblastoma, a mesenchymal phenotype is associated with especially poor patient outcomes. Various glioblastoma microenvironmental factors and therapeutic interventions are purported drivers of the mesenchymal transition, but the degree to which these cues promote the same mesenchymal transitions and the uniformity of those transitions, as defined by molecular subtyping systems, is unknown. Here, we investigate this question by analyzing publicly available patient data, surveying commonly measured transcripts for mesenchymal transitions in glioma-initiating cells (GIC), and performing next-generation RNA sequencing of GICs. Analysis of patient tumor data reveals that TGFβ, TNFα, and hypoxia signaling correlate with the mesenchymal subtype more than the proneural subtype. In cultured GICs, the microenvironment-relevant growth factors TGFβ and TNFα and the chemotherapeutic temozolomide promote expression of commonly measured mesenchymal transcripts. However, next-generation RNA sequencing reveals that growth factors and temozolomide broadly promote expression of both mesenchymal and proneural transcripts, in some cases with equal frequency. These results suggest that glioblastoma mesenchymal transitions do not occur as distinctly as in epithelial-derived cancers, at least as determined using common subtyping ontologies and measuring response to growth factors or chemotherapeutics. Further understanding of these issues may identify improved methods for pharmacologically targeting the mesenchymal phenotype in glioblastoma.

overall_design: Comparative gene expression analysis (RNA-seq) of G816 glioma-initiating control cells or cells treated with TGFβ+TNFα or temozolomide.

first: William

last: Hart


title: A phenotypic screening platform for identifying chemical modulators of astrocyte reactivity

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

Disease, injury, and aging induce pathological reactive astrocyte states that contribute to neurodegeneration. Modulating reactive astrocytes therefore represents an attractive therapeutic strategy. Here, we describe the development of an astrocyte phenotypic screening platform for identifying chemical modulators of astrocyte reactivity. Leveraging this platform for chemical screening, we identify HDAC3 inhibitors as effective suppressors of pathological astrocyte reactivity. We demonstrate that HDAC3 inhibition reduces molecular and functional characteristics of reactive astrocytes in vitro. Transcriptional and chromatin mapping studies show that HDAC3 inhibition disarms pathological astrocyte gene expression and function while promoting the expression of genes associated with beneficial astrocytes. Administration of RGFP966, a small molecule HDAC3 inhibitor, blocks reactive astrocyte formation and promotes neuroprotection in vivo in mice. Collectively, these results establish a platform for discovering modulators of reactive astrocyte states, inform the mechanisms that control astrocyte reactivity, and demonstrate the therapeutic benefits of modulating astrocyte reactivity for neurodegenerative diseases.


title: Bulk liver transcriptomics from Li-TSC1KO RagAGTP mice

city: Madrid

first: Osvaldo

last: Graña

state: Comunidad de Madrid

address: C/ Melchor Fernández Almagro, 3

department: Structural Biology

institute: CNIO

summary: RNA-seq from bulk liver was performed for wild-type, Li-TSC1KO and Li-TSC1KO RagAGTP livers.

overall_design: Mice were fasted for 24h. Some of them were refed during 2 hours or treated with rapamycin. Liver was collected and RNA was extracted for RNA-seq analysis

first: Ana

middle: B

last: Plata-Gómez

last: Graña-Castro

first: Alejo

last: Efeyan


title: Effect of Glucuronic acid on Citrobacter rodentium

status: Public on Jan 09 2024

city: Madison

first: Thibaut

last: Rosay

state: Wisconsin

department: MMI

institute: UW Madison

summary: Previous experiments have shown that hexuronates regulate EHEC virulence, here we look at glucuronic acid effect on citrobacter rodentium

overall_design: 6h culture of Citrobacter rodentium in DMEM low glucose in microaerobic condition in presence or not of Glucuronic acid 100uM

first: Rosay

last: Thibaut

first: Sperandio

last: Vanessa


title: A phenotypic screening platform for identifying chemical modulators of astrocyte reactivity [bulkRNA-seq]

city: Cleveland

overall_design: Transcriptional profiling of primary mouse resting astrocytes, reactive astrocytes (treated with TNFa, IL1a, and C1q), reactive astrocytes treated with 5uM RGFP966, and reactive astrocytes treated with 5uM JSH-23.


title: A phenotypic screening platform for identifying chemical modulators of astrocyte reactivity [scRNAseq]

overall_design: 10x Genomics single cell RNAseq was performed on primary mouse astrocyte cultures in either a reactive (treated with TNFa, IL1a, and C1q) or resting state.


title: USP10-mediated MOF (KAT8) stabilization accelerates the malignant progression of esophageal squamous cell carcinoma via epigenetically activating of ANXA2/Wnt signaling

city: Jinan

first: Peichao

last: Li

state: Shangdong

address: The Second Hospital of Shandong University

department: thoracic surgery department

country: China

institute: Shandong University

summary: Dysregulation of MOF (MYST1, KAT8), a highly conserved H4K16 acetyltransferase, plays important roles in human cancers. However, its expression and function in esophageal squamous cell carcinoma (ESCC) remain unknown. Here, we report that MOF is highly expressed in ESCC tumors and predicts a worse prognosis. Depletion of MOF in ESCC significantly impedes tumor growth and metastasis both in vitro and in vivo, whereas ectopic expression of MOF but not enzyme inactive mutant (MOF E350Q) promotes ESCC progression, suggesting that MOF acetyltransferase activity is crucial for its oncogenic activity. Further analysis reveals that USP10, a deubiquitinase highly expressed in ESCC, binds to and protects MOF from proteosome-dependent protein degradation. MOF stabilization by USP10 promotes H4K16ac enrichment in the ANXA2 promoter to stimulate ANXA2 transcription, which subsequently activates Wnt/ β-Catenin signaling to facilitate ESCC progression. Our findings highlight a novel USP10/MOF/ANXA2 axis as a promising therapeutic target for ESCC.

overall_design: To identify downstream targets mediating the oncogenic effect of MOF on ESCC progression, RNA-seq analysis was performed in Eca-109 cells respectively transfected with siRNAs against MOF (si-MOF#1 or si-MOF#2) or Negative control (si-NC). The amounts and integrity of total exacted RNA were analyzed using Bioanalyzer 2100 system (Agilent Technologies, CA, USA). Library preparation, sequencing and bioinformatic analysis were conducted at Novogene (Beijing, China).

first: Xiaogang

last: Zhao

first: Hong

last: Sun

first: Lingxiao

last: Yang


title: Genome-wide profiling of linker histone variant H1.3 in T47D-MTVL cancer cell line

city: Barcelona

first: Albert

last: Jordan

address: Baldiri Reixac 4

department: Molecular Genomics

institute: IBMB-CSIC

summary: Human somatic cells may contain up to seven members of the histone H1 family contributing to chromatin compaction and regulation of nuclear processes, apparently with certain subtype specificities. Our previous studies of H1.0, H1.2, H1.4, H1.5, and H1X genome-wide distribution determined that H1 variants are distributed in a variant-specific manner throughout the genome. We have been able to extend our study of the genomic distribution of the H1 family from five to six proteins thanks to the acquisition and validation of a new ChIP-grade endogenous antibody against histone H1.3.

overall_design: Genome-wide analysis of linker histone variant H1.3 in T47D breast cancer cells (ChIP-Seq).

first: A

first: M

last: Salinas

first: N

last: Serna


title: Genome-wide profiling of linker histone variant H1X in five different cancer cell lines

summary: Human somatic cells may contain up to seven members of the histone H1 family contributing to chromatin compaction and regulation of nuclear processes, apparently with certain subtype specificities. Previous studies in T47D breast cancer cells determined that H1 variants are distributed in a variant-specific manner throughout the genome. In particular, we observed a clear enrichment of linker histone H1X within SVA and SINE repetitive sequences. To extend our results and study whether H1X is universally enriched within these types of repeats, we have performed ChIP-Seq of endogenous H1X in five additional cancer cell lines (HeLa, MCF-7, SK-N-SH, HCT-116, and SK-MEL-147).

overall_design: Genome-wide analysis of linker histone variant H1X in HeLa, MCF-7, SK-N-SH, HCT-116, and SK-MEL-147 cancer cell lines (ChIP-Seq).


title: IκBζ is a dual-use coactivator of NF-κB and POU transcription factors

city: Cold Spring Harbor

first: Christopher

middle: R.

last: Vakoc

state: NY

address: 1 Bungtown road

institute: Cold Spring Harbor Laboratory

summary: OCA-B, OCA-T1, and OCA-T2 belong to a family of transcriptional coactivators that bind to POU transcription factors (TFs) to regulate gene expression in immune cells. Here, we identify IkBz (encoded by the NFKBIZ gene) as the fourth member of the OCA protein family. While originally discovered as an inducible regulator of NFkB, we show here that IkBz shares a microhomology with OCA proteins and uses this segment to simultaneously bind to POU transcription factors and octamer motif-containing DNA. Our functional reporter assays suggest that IkBz requires its interaction with POU TFs to coactivate immune-related genes. This finding is reinforced by our epigenomic analysis of MYD88 L265P-mutant lymphoma cells, which revealed colocalization of IkBz, the POU transcription factor OCT2, and NFkB:p50 at hundreds of DNA elements harboring octamer and kB motifs. These results suggest that IkBz is a transcriptional coactivator that integrates and amplifies the output of NFkB and POU transcription factors at inducible genes in immune cells.

overall_design: For each ChIP, 20 million cells were used. Cells were cross-linked with 1% formaldehyde for 10 min at room temperature with agitation and quenched with 0.125 M glycine for 5 min at room temperature. After washing twice with PBS, cells were incubated in 1 ml cell lysis buffer (10 mM Tris-HCl (pH 8.0), 10 mM NaCl, 0.2% NP-40 with protease inhibitor) for 15 min on ice. Nuclei were isolated by centrifugation at 600×g for 30 s, resuspended in 1 ml nuclear lysis buffer (50 mM Tris-HCl (pH 8.0), 10 mM EDTA, 1% SDS with protease inhibitor) and sonicated using a Bioruptor Pico (Diagenode) (30s on/off, 10 cycles). In all cases, the chromatin was centrifuged at maximum speed in a tabletop centrifuge for 15 min at 4 °C. The supernatant was mixed with 7 ml IP dilution buffer (20 mM Tris-HCl (pH 8.0), 2 mM EDTA, 150  mM NaCl, 1% Triton X-100), and incubated with the 2.5 µg of the indicated antibody for 2 h. Protein A/G magnetic beads (25 µl, Dynabeads, Thermo Fisher Scientific) were washed twice with PBS and added to the antibody–chromatin mixture at 4 °C overnight. The beads were washed once with IP wash 1 buffer (20 mM Tris-HCl pH 8.0, 2 mM EDTA, 50 mM NaCl, 1% Triton X-100, 0.1% SDS), twice with high salt buffer (20 mM Tris-HCl (pH 8.0), 2 mM EDTA, 500 mM NaCl, 1% Triton X-100, 0.01% SDS), once with IP wash 2 buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 250 mM LiCl, 1% NP-40, 1% sodium deoxycholate) and twice with TE (pH 8.0). Chromatin DNA was eluted, and cross-linking was reversed in 200 µl nuclear extraction buffer with 12 µl of 5 M NaCl and 1 µg /mL RNase A at 65 °C overnight. The beads were then discarded. The DNA-containing supernatant was treated with 4 µg /mL proteinase K at 56 °C for 20 min and purified using the QIAquick PCR purification kit (QIAGEN) in 60 μl water.

first: Aktan

last: Alpsoy

first: Xiaoli

last: Wu

first: Osama

last: Demerdash

first: Benan

last: Nalbant

first: Olaf

last: Klingbeil


title: A Plant Flavonol and Genetic Suppressors Rescue a Pathogenic Mutation Associated with Kinesin in Neurons

status: Public on Jan 08 2024

city: Beijing

first: Yongping

last: Chai

state: Beijing

address: Tsinghua University

institute: Tsinghua University 清华大学

summary: KIF1A, a microtubule-based motor protein responsible for axonal transport, is linked to a group of neurological disorders known as KIF1A-associated neurological disorder (KAND). Current therapeutic options for KAND are limited. Here, we introduced the clinically relevant KIF1A(R11Q) variant into the C. elegans homolog UNC-104, resulting in uncoordinated animal behaviors. Through genetic suppressor screens, we identified intragenic mutations in UNC-104's motor domain that rescued synaptic vesicle localization and coordinated movement. We showed that two suppressor mutations partially recovered motor activity in vitro by counteracting the structural defect caused by R11Q at KIF1A's nucleotide-binding pocket. We found that supplementation with fisetin, a plant flavonol, improved KIF1A(R11Q) worms’ movement and morphology. Notably, our biochemical and single-molecule assays revealed that fisetin directly restored the ATPase activity and processive movement of human KIF1A(R11Q) without affecting wild-type KIF1A. These findings suggest fisetin as a potential intervention for enhancing KIF1A(R11Q) activity and alleviating associated defects in KAND.

overall_design: To examine the unc-104 mRNA expression in the R9Q mutant and/or with the genetic suppressor presence, with the WT animal unc-104 mRNA levels.

first: Jingyi

last: Ke

first: Guangshuo

last: Ou


title: SRSF1 is crucial for male meiosis through alternative splicing during homologous pairing and synapsis in mice

first: Jiali

last: Liu

address: 2 West Yuanmingyuan Road

institute: China Agricultural University


title: Intersection of Immunity, Metabolism, and Muscle Regeneration in an Autoimmune-Prone MRL Mouse Model

first: Kaitlyn

last: Sadtler

state: Maryland

address: 50 S Drive, Room 4341

department: NIBIB

institute: NIH

summary: Due to the limited capacity of mammals to regenerate complex tissues, researchers have worked to understand the mechanisms of tissue regeneration in organisms that maintain that capacity. One example is the MRL/MpJ mouse strain with unique regenerative capacity in ear pinnae that is absent from other strains, such as the common C57BL/6 strain. The MRL/MpJ mouse has also been associated with an autoimmune phenotype even in the absence of the mutant Fas gene described in its parent strain MRL/lpr. Due to these findings, we evaluated the differences between the responses of MRL/MpJ versus C57BL/6 strain in volumetric muscle injury and subsequent material implantation. One salient feature of the MRL/MpJ response to injury was robust adipogenesis within the muscle. This was associated with a decrease in M2-like polarization in response to biologically derived extracellular matrix scaffolds. In pro-fibrotic materials, such as polyethylene, there were fewer foreign body giant cells in the MRL/MpJ mice. As there are reports of both positive and negative influences of adipose tissue and adipogenesis on wound healing, this model could provide an important lens to investigate the interplay between stem cells, adipose tissue, and immune responses in trauma and material implantation.

overall_design: To investigate the metabolic and immune pathways involved in skeletal muscle regeneration, we performed bulk RNA sequencing of mouse quadriceps from MRL/MpJ and C57BL/6 mice 21 days after VML surgeries and biomaterial implantation

first: Tran

last: Ngo

first: Aditya

last: Josyula

first: Sabrina

last: DeStefano

first: Daphna

last: Fertil

first: Mondreakest

last: Faust

first: Ravi

last: Lokwani


title: Illumina sequencing of CRAC data obtained with Ylr419w

city: Paris

first: Gwenael

last: Badis

address: 46 rue d'Ulm

department: Genomique fonctionnelle (Section 2)

country: France

institute: IBENS

summary: 27 years after the yeast genome sequencing, the function of many ORFs remain unknown. Despite the evolutionary distance between human and yeast, homology with the conserved DEAD-box helicase domains allowed us to list DHX29, DHX36 and DHX57 as three putative homologs of the yeast Ylr419w. Functional studies first linked the Ylr419w protein to the translating ribosome, and cross-linking and analysis of cDNA (CRAC) experiments determined the precise region of Ylr419w in contact with the ribosome. It corresponds to the loop of the h16 helix in the 18S rRNA designing the translation initiation factor DHX29, as the functional homolog of Ylr419w.

overall_design: cross-linking and analysis of cDNA (CRAC) experiments of Yhr419w_HTP tagged protein with or without UV treatment

first: Micheline

last: Fromont-Racine

first: Varun

last: Khanna

first: Alain

last: Jacquier


title: Synthetic reversed sequences reveal default genomic states [Yeast_ATACseq]

status: Public on Jan 16 2024

city: New York

first: Brendan

last: Camellato

state: New York

address: 435 E 30th St

department: Institute for Systems Genetics

institute: NYU Langone Health

summary: Understanding default genome states would help interpret whether pervasive transcriptional activity has biological meaning. The genomes of extant organism have undergone billions of years of evolution, making it unclear whether observed genomic activities represent the effects of selection or “noise”. We addressed this question by introducing a novel 101-kb locus into the genomes of S. cerevisiae and M. musculus, and characterizing genomic activity. The locus was designed by reversing but not complementing human HPRT1, including substantial flank-ing regions, retaining basic sequence features but ablating evolved coding or regulatory infor-mation. We observed widespread activity of both reversed and native HPRT1 loci in yeast, de-spite the lack of evolved yeast promoters. In contrast, the reversed locus displayed no activity at all in mouse embryonic stem cells, instead showing repressive chromatin signatures. The re-pressive signature was alleviated in a locus variant lacking CpG dinucleotides; nevertheless this variant too was transcriptionally inactive. These results show that novel genomic sequences lacking coding information are active in yeast, but inactive in mouse embryonic stem cells, con-sistent with a major difference in “default genomic states” between these two divergent eukary-otic cell types, with implications for understanding pervasive transcription, horizontal transfer of genetic information, and new gene birth.

overall_design: Exploratory ATAC-seq was performed to investigate whether synthetic HPRT1 and HPRT1R sequences are accessible in yeast, and to compare to the rest of the respective genome.

first: Ran

last: Brosh

first: Hannah

last: Ashe

first: Matthew

middle: T

last: Maurano

first: Jef

last: Boeke


title: Comparative genomic analysis of Fischer F344 rat livers exposed for 90 days to 3-methylfuran or its parental compound furan.

city: Ottawa

first: Lauren

last: Carter

state: Ontario

address: 251 SIR FREDERICK BANTING

department: 251 Sir Frederick Banting Driveway

institute: Health Canada

summary: Transcriptomic data obtained by RNA-seq from male Fischer 344 rats treated with furan and 3-methylfuran were compared and contrasted. Microarray data from the same rats treated with furan was also used to compare microarray to RNA seq analysis.

overall_design: Compare trascriptomics data between furan and its derivative 3-methylfuran in male Fischer 344 rats using RNA-seq while at the same time comparing microarray data from the same rats treated with furan to that of RNA-seq data from both furan and 3-methylfuran.

first: Steven

last: Bugiel

first: Andrée

last: Nunnikhoven

first: Adrian

last: Verster

first: Nicholas

last: Petronella

first: Santokh

last: Gill

first: Ivan

middle: H

last: Curran


title: Single molecule DNA methylation reveals unique epigenetic identity profiles of T helper cells

city: Bron

first: Hector

last: Hernandez-Vargas

state: Rhone

address: 36 bis Rue de la Batterie

institute: Genomics Consulting

type: Methylation profiling by high throughput sequencing

summary: Identity and plasticity of CD4 T helper (Th) cells are regulated in part by epigenetic mechanisms. Cytosine methylation in CpG context (5mCpG) and cytosine hydroxymethylation (5hmCpG) are DNA modifications that identify stable cell phenotypes. To assess transition states in Th cells, we developed a method based on Cas9-targeted single molecule nanopore sequencing and found that 5mCpG can be used as markers of cellular identity. Targeting as few as 10 mouse selected genomic loci, we were able to distinguish major differentiated T cell subtypes as well as intermediate phenotypes by their native DNA 5mCpG patterns. Moreover, by using off-target sequences we were able to infer transcription factor activities relevant to each cell subtype. Our data highlight the potential to exploit native DNA methylation profiling to study physiological and pathological Th transition states.

overall_design: We polarized mouse naïve Th0 cells towards various Th fates in vitro.  Th0 cells were isolated by FACs-sorting and activated under polarizing conditions towards the Th0, Th1, Th2, Th17 and Treg phenotypes using cytokine cocktails. To test the sensitivity of this approach to detect potential pathogenic Th cell phenotypes, naïve T cells were also polarized under Th17 conditions in the absence of TGFb, referred to as ‘Th17-noTGFb’ cells.

first: Chloe

last: Goldsmith


title: Human fetal brain self-organizes into long-term expanding organoids

city: Utrecht

first: Delilah

last: Hendriks

address: Uppsalalaan 8

country: Netherlands

institute: Hubrecht Institute

overall_design: Refer to individual Series


title: Human fetal brain self-organizes into long-term expanding organoids (scRNA-Seq)

summary: Human brain development involves an orchestrated, massive neural progenitor expansion while a multi-cellular tissue architecture is established. Continuously expanding organoids can be grown directly from multiple somatic tissues, yet to date brain organoids can solely be established from pluripotent stem cells. Here, we show that healthy human fetal brain in vitro self-organizes into organoids (FeBOs), phenocopying aspects of in vivo cellular heterogeneity and complex organization. FeBOs can be expanded over long time periods. FeBO growth requires maintenance of tissue integrity, which ensures production of a tissue-like extracellular matrix (ECM) niche, ultimately endowing FeBO expansion. FeBO lines derived from different areas of the central nervous system (CNS), including dorsal and ventral forebrain, preserve their regional identity and allow to probe aspects of positional identity. Using CRISPR-Cas9, we showcase the generation of syngeneic mutant FeBO lines for the study of brain cancer. Taken together, FeBOs constitute a complementary CNS organoid platform.

overall_design:
Single-cell RNA-sequencing of human fetal brain organoids
-------------------------------------
Authors state "The raw data are not publicly available due to donor privacy concerns."

first: Benedetta

last: Artegiani


title: Autonomous transposons tune their sequences to ensure somatic suppression [totalRNA_HEK293_fractionation]

city: Berlin

first: IBRAHIM

middle: AVSAR

last: ILIK

state: Berlin

address: Ihnestraße 63-73

institute: Max-Planck-Institut für molekulare Genetik

summary: Transposable elements (TEs) are a major constituent of human genes, occupying approximately half of the intronic space. During pre-mRNA synthesis, intronic TEs are transcribed along with their host genes but rarely contribute to the final mRNA product as they are spliced out together with the intron and rapidly degraded. Paradoxically, TEs are an abundant source of RNA-processing signals through which they can create new introns (Huff et al. 2016), and functional (Cosby et al. 2021) or non-functional chimeric transcripts (Clayton et al. 2020). The rarity of these events impliesy the existence of a resilient splicing code that is able to suppress TE-exonization without compromising host pre-mRNA processing. Here we show that SAFB proteins play a unique dual role in defence against TEs by preventing retrotransposition of L1 elements as well as their exonization when they land into genes by coating their adenosine-rich RNA. The suppressive activity of SAFB extends to neuro-, muscle- and testis-specific giant protein-coding cassette exons of ANK3, MAP4 and CLIP1, nested genes as well as Tigger DNA transposons, which are all enriched with adenosine/purine-rich sequences that act as splicing-enhancers in SAFB depleted cells. In mice and flies, SAFB additionally suppresses LTR/ERV elements, which are active in mice and flies, but no longer in humans. Splicing events suppressed by SAFB in somatic cells are activated in the testis, coinciding with low SAFB expression in post-meiotic spermatids. Reminiscent of the division of labour between innate and adaptive immune systems that fight external pathogens, our results uncover SAFB proteins as an RNA-based, pattern-guided, non-adaptive defence system against TEs in the soma, complementing the RNA-based, adaptive Piwi-piRNA pathway of the germline.

overall_design: Gene expression profiling of nuclear and cytoplasmic fractions, and whole cell extracts of HEK293 cells subject to knockdown of all three SAFB proteins (SAFB1, SAFB2 and SLTM) at the same time, and a control treatment. rRNA-depleted RNAs were sequenced in three biological replicates per fraction and treatment (18 libraries in total).

first: Ibrahim

middle: A

last: Ilik

first: Petar

last: Glažar

first: Tugce

last: Aktas


title: Autonomous transposons tune their sequences to ensure somatic suppression [polyA_HCT116]

summary: Transposable elements (TEs) are a major constituent of human genes, occupying approximately half of the intronic space. During pre-mRNA synthesis, intronŁic TEs are transcribed along with their host genes but rarely contribute to the final mRNA product as they are spliced out together with the intron and rapidly degraded. Paradoxically, TEs are an abundant source of RNA-processing signals through which they can create new introns (Huff et al. 2016), and functional (Cosby et al. 2021) or non-functional chimeric transcripts (Clayton et al. 2020). The rarity of these events impliesy the existence of a resilient splicing code that is able to suppress TE-exonization without compromising host pre-mRNA processing. Here we show that SAFB proteins play a unique dual role in defence against TEs by preventing retrotransposition of L1 elements as well as their exonization when they land into genes by coating their adenosine-rich RNA. The suppressive activity of SAFB extends to neuro-, muscle- and testis-specific giant protein-coding cassette exons of ANK3, MAP4 and CLIP1, nested genes as well as Tigger DNA transposons, which are all enriched with adenosine/purine-rich sequences that act as splicing-enhancers in SAFB depleted cells. In mice and flies, SAFB additionally suppresses LTR/ERV elements, which are active in mice and flies, but no longer in humans. Splicing events suppressed by SAFB in somatic cells are activated in the testis, coinciding with low SAFB expression in post-meiotic spermatids. Reminiscent of the division of labour between innate and adaptive immune systems that fight external pathogens, our results uncover SAFB proteins as an RNA-based, pattern-guided, non-adaptive defence system against TEs in the soma, complementing the RNA-based, adaptive Piwi-piRNA pathway of the germline.

overall_design: Gene expression profiling of HCT116 cells subject to knockdown of two SAFB proteins (SAFB1 and SLTM) at the same time, and a control treatment. poly(A)+ RNAs were sequenced in three biological replicates per treatment (6 libraries in total).


title: Autonomous transposons tune their sequences to ensure somatic suppression [SAFKD_1H4_DHX9_FLASH]

overall_design: FLASH libraries of p-SR (mAb 1H4) and DHX9 in HEK293 cells that were either treated with control siRNA or siRNAs against SAFB1, SAFB2 and SLTM.


title: ONTOGENY SHAPES THE ABILITY OF ETV6-RUNX1 TO ENHANCE HEMATOPOIETIC STEM CELL SELF-RENEWAL AND DISRUPT EARLY LYMPHOPOIESIS [in vivo]

city: Lund

first: Mohamed

last: Eldeeb

address: DMH, BMC B12

country: Sweden

institute: Lund University

summary: The ETV6-RUNX1 (E/R) translocation, found in approximately 30% of pediatric B-cell acute lymphoblastic leukemia (ALL) cases, originates prenatally. Although hematopoietic stem cells (HSCs) have been proposed as the cell of origin for E/R-ALL, definitive evidence for this has been lacking. Here, we develop an E/R-inducible transgenic mouse model and observe that E/R rapidly expands the pool of phenotypic HSCs and block early B cell development. By leveraging the reversibility of the model, we demonstrate functional rather than merely phenotypic HSC expansion. At a molecular level, E/R activates immune checkpoint pathways, which is more pronounced in fetal cells. Comparison of fetal and adult HSCs reveals that although E/R disrupts HSC differentiation in both, fetal cells have a competitive advantage, especially in a setting of viral mimicry. These findings provide new insights into the nature of preleukemic clones associated with E/R and suggest potential benefits of immune checkpoint inhibition for their eradication.

overall_design: RNA-seq was performed on HSCs (Lin- Kit+ Sca1+ CD48- CD150+) sorted from control (uninduced) or E/R-induced mice, n = 3 replicates/group

first: Anna

last: Konturek-Ciesla

first: Qinyu

last: Zhang

first: Shabnam

last: Kharazi

first: Johanna

last: Tingvall-Gustafsson

first: Jonas

last: Ungerbäck

first: Mikael

last: Sigvardsson

first: David

last: Bryder


title: Multi-omics analysis of human population variation in immune function and in vivo response to BCG vaccination

city: Vienna

first: Christoph

last: Bock

address: Lazarettgasse 14

country: Austria

institute: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences

summary: Immune responses are tightly regulated, yet highly variable between individuals. To investigate human population variation of trained immunity, we immunized healthy individuals with Bacillus Calmette-Guérin (BCG). This live attenuated vaccine induces not only an adaptive immune response against tuberculosis, but also triggers innate immune activation and memory. We established personal immune profiles and chromatin accessibility maps over a time course of BCG vaccination in 323 individuals. This large resource uncovered genetic and epigenetic predictors of baseline immunity and BCG vaccine response. We found that BCG vaccination enhances the innate immune response only in individuals with dormant immune states at baseline, suggesting that exogeneous induction of trained immunity is not a universal booster of innate immunity, but specifically elevates weak innate immune responses. This study advances our understanding of BCG’s heterologous immune-stimulatory effects and trained immunity in humans. Moreover, our results highlight the value of epigenetic cell states as an “endophenotype” that connects immune function with genotype and the environment.

overall_design: The 300BCG cohort consists of 323 healthy adult individuals (184 females and 139 males) and is part of the Human Functional Genomics Project (HFGP). Study participants were included from April 2017 until June 2018 at the Radboud University Medical Center, the Netherlands. They received a standard dose of 0.1 ml BCG (Bulgaria strain, Intervax) intradermally. Blood was drawn immediately before BCG vaccination (d0), and 14 days (d14) as well as 90 days (d90) after vaccination. Exclusion criteria were any vaccination within three months prior to d0, acute (febrile) illness four weeks before inclusion, a medical history of immunodeficiency or any other chronic disease, use of systemic medication other than oral contraceptives or acetaminophen, use of antibiotics three months before inclusion, ancestry other than Western European, previous BCG vaccination, history of tuberculosis, previous contact with tuberculosis patients, being born in a tuberculosis endemic country, pregnancy, or breastfeeding. During the study, individuals were excluded upon medication use other than oral contraceptives or acetaminophen, or illness at time of study visit. Applying these criteria resulted in 273 individuals with measurements for d0, 253 individuals for d14, and 254 individuals for d90, respectively. >>>Submitter declares that the raw data will be deposited in EGA due to patient privacy concerns<<<

first: Mihai

middle: G

last: Netea

first: Simone

middle: J

last: Moorlag

first: Lukas

last: Folkman

first: Rob

last: ter Horst

first: Thomas

last: Krausgruber

first: Daniele

last: Barreca

first: Linda

last: Schuster

first: Victoria

last: Fife

first: Vasiliki

last: Matzaraki

first: Wenchao

last: Li

first: Stephan

last: Reichl

first: Vera

middle: P

last: Mourits

first: Valerie

last: Koeken

first: L

last: de Bree

first: Helga

last: Dijkstra

first: Heidi

last: Lemmers

first: Bram

last: van Cranenbroek

first: Esther

last: van Rijssen

first: Hans

last: Koenen

first: Irma

last: Joosten

first: Cheng-Jian

last: Xu

first: Yang

first: Leo

first: Reinout

last: van Crevel


title: Autonomous transposons tune their sequences to ensure somatic suppression [directRNA_HEK293]

first: IBRAHIM

last: ILIK

overall_design: Gene expression profiling of mouse 3T3 cells subject to knockdown of three SAFB proteins (Safb1, Safb2 and Sltm) at the same time, and a control treatment. poly(A)+ RNAs were sequenced in three biological replicates per treatment (6 libraries in total).

first: Ibrahim

last: Ilik

first: Petar

last: Glažar

first: Tugce

last: Aktas


title: ONTOGENY SHAPES THE ABILITY OF ETV6-RUNX1 TO ENHANCE HEMATOPOIETIC STEM CELL SELF-RENEWAL AND DISRUPT EARLY LYMPHOPOIESIS

first: Mohamed

last: Eldeeb

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


title: Assessment of wood smoke induced pulmonary toxicity using bronchial, chronic bronchitis-like, and alveolar lung mucosa models at air-liquid interface

city: Neuherberg

first: Johannes

last: Beckers

address: Ingolstaedter Landstr. 1

department: Institute of Experimental Genetics

institute: Helmholtz Zentrum Muenchen

summary:
Chronic obstructive pulmonary disease (COPD) has the highest increased risk due to household air pollution arising from biomass fuel burning. However, knowledge on COPD patho-mechanisms is mainly limited to tobacco smoke exposure. In this study, a repeated direct wood smoke (WS) exposure was performed using normal- (bro-ALI) and chronic bronchitis-like bronchial (bro-ALI-CB), and alveolar (alv-ALI) lung mucosa models at air-liquid interface (ALI) to assess broad toxicological end points.
The bro-ALI and bro-ALI-CB models were developed using human primary bronchial epithelial cells and the alv-ALI model was developed using a representative type-II pneumocyte cell line. The lung models were exposed to WS (10 minutes/exposure; 5-exposures over 3-days; n=6-7 independent experiments). Sham exposed samples served as control. WS composition was analyzed following passive sampling. Cytotoxicity, total cellular reactive oxygen species (ROS) and stress responsive NFkB were assessed by flow cytometry. WS exposure induced changes in gene expression were evaluated by RNA-seq (p≤0.01) followed by pathway enrichment analysis. Secreted levels of proinflammatory cytokines were assessed in the basal media. Non-parametric statistical analysis was performed.
147 unique compounds were annotated in WS of which 42 compounds have inhalation toxicity (9 very high). WS exposure resulted in significantly increased ROS in bro-ALI (11.2%) and bro-ALI-CB (25.7%) along with correspondingly increased NFkB levels (bro-ALI: 35.6%; bro-ALI-CB: 18.1%). A total of 1262 (817-up and 445-down), 329 (141-up and 188-down), and 102 (33-up and 69-down) genes were differentially regulated in the WS-exposed bro-ALI, bro-ALI-CB, and alv-ALI models respectively. The enriched pathways included the terms acute phase response, mitochondrial dysfunction, inflammation, oxidative stress, NFkB, ROS, xenobiotic metabolism of AHR, and chronic respiratory disorder. The enrichment of the ‘cilium’ related genes was predominant in the WS-exposed bro-ALI (180-up and 7-down). The pathways primary ciliary dyskinesia, ciliopathy, and ciliary movement were enriched in both WS-exposed bro-ALI and bro-ALI-CB. Interleukin-6 and tumor necrosis factor-α were reduced (p<0.05) in WS-exposed bro-ALI and bro-ALI-CB.

overall_design: We analysed bronchial, chronic bronchitis-like, and alveolar ALI cell cultures in the presence or absence of wood smoke

first: Swapna

last: Upadhyay

first: Mizanur

last: Rahman

first: Selina

last: Rinaldi

first: Jeremy

last: Koelmel

first: Elizabeth

middle: Z

last: Lin

first: Padukudru

last: Anand Mahesh

first: Gunnar

last: Johanson

first: Krystal

last: Godri Pollitt

first: Lena

last: Palmberg

first: Martin

last: Irmler

first: Koustav

last: Ganguly


title: Elemental sulfur enhances the anti-fungal effect of Lacticaseibacillus rhamnosus Lcr35

city: Clermont-ferrand

first: sylvie

last: miquel

address: 28 place henri Dunant

institute: UCA

summary: Lacticaseibacillus rhamnosus Lcr35 is a well-known bacterial strain whose efficiency in preventing recurrent vulvovaginal candidiasis has been largely demonstrated in clinical trials.  The presence of sodium thiosulfate (STS) has been shown to enhance its ability to inhibit the growth of C. albicans strains. In this study, we confirmed that Lcr35 has a fungicidal effect not only on the planktonic form of C. albicans but also on other life forms such as hypha and biofilm. Transcriptomic analysis showed that the presence of C. albicans induced a metabolic adaptation of Lcr35 potentially associated with a competitive advantage over yeast cells. However, STS alone had no impact on the global gene expression of Lcr35, which is not in favor of the involvement of an enzymatic transformation of STS. Comparative gas chromatography- mass spectrometry analysis of the organic phase from cell-free supernatant (CFS) fractions obtained from Lcr35 cultures performed in the presence and absence of STS identified elemental sulfur (S0) in the samples initially containing STS. In addition, the anti-candida activity of CFS from STS-containing cultures was shown to be pH-dependent and occurred at acidic pH lower than 5. We next investigated the antifungal activity of lactic acid and acetic acid, the two main organic acids produced by Lactobacillus spp. The two molecules affected the viability of C. albicans but only at pH 3.5 and in a dose-dependent manner, an antifungal effect that was enhanced in samples containing STS in which the thiosulfate was decomposed into S0. In conclusion, the use of STS as an excipient in the manufacturing process of Lcr35 exerted a dual action since the production of organic acids by Lcr35 facilitates the decomposition of thiosulfate into S0, thereby enhancing the bacteria’s own anti-fungal effect.

overall_design:
To investigate the transcriptomic impact of Sodium thiosulphate (STS) and C. albicans ATCC MYA-2876 on Lcr35.
We then performed RNA-sequencing comparing Lcr35 vs Lcr35 + STS; and Lcr35 vs Lcr35 + C. albicans ATCC MYA-2876.

first: Manjyot

last: Kaur

first: Sylvie

last: Miquel

first: Laurence

last: Olivier-Nakusi

first: Claudia

last: Thoral

first: Marjolaine

last: Vareille-Delarbre

first: Clara

last: Bekirian

first: Christophe

last: d’Enfert

first: Thierry

last: Fontaine

first: Karine

last: Roget

first: Christiane

last: Forestier


title: Human fetal brain self-organizes into long-term expanding organoids (bulk RNA-Seq)

first: Delilah

last: Hendriks

overall_design:
Bulk RNA-sequencing of human fetal brain organoids
-------------------------------------
Authors state "The raw data are not publicly available due to donor privacy concerns."

first: Benedetta

last: Artegiani


title: SRSF1 is crucial for male meiosis through alternative splicing during homologous pairing and synapsis in mice [CLIP-seq]

first: Jiali

last: Liu

summary: Homologous pairing and synapsis are essential in meiotic prophase I during spermatogenesis. However, the underlying mechanisms of how alternative splicing (AS) functions in homologous pairing and synapsis remain largely unclear. We reveal that SRSF1 is essential for gene expression and AS in homologous pairing and synapsis. Conditional knockout (cKO) of Srsf1 in mouse germ cells impaired homologous pairing and synapsis, leading to non-obstructive azoospermia (NOA). SRSF1 was required for initial homology recognition, telomere-led chromosome movement, and synaptonemal complex (SC) assembly. Moreover, SRSF1 interacted with TRA2B and U2AF2, directly binding and regulating the expression of Dmc1, Sycp1, Sun1, and Majin via AS to implement homologous pairing and synapsis during the meiotic prophase I program. Altogether, our data reveal the critical role of an SRSF1-mediated post-transcriptional regulatory mechanism in homologous pairing and synapsis during meiotic prophase I, providing a framework for elucidating the molecular mechanisms underlying the post-transcriptional network of male meiosis.

overall_design: Total cells were isolated from adult WT C57BL/6N mouse testes, and then the cells were cross-linked by ultraviolet light (254 nm) to maintain the covalent binding of RBPs to their cognate RNA. Subsequently, SRSF1 and cross-linked RNAs were immunoprecipitated with an anti-SRSF1 antibody and digested with micrococcal nuclease (EN0181, Thermo Fisher Scientific). An IR800-biotin adapter was ligated to the 3′ ends of the RNA fragments. Then, the SRSF1/RNA complexes were separated by SDS‒PAGE and transferred to a nitrocellulose membrane (HATF00010, Millipore). These RNA and protein complexes from approximately 47 to 62 kDa were extracted from the nitrocellulose membrane, after which proteinase K (9034, Takara) digestion was performed. RNA was isolated with saturated phenol (AM9712, Ambion), ligated with adaptors, and converted to cDNA with a SuperScript III First-Strand Kit (18080-051, Invitrogen). The cDNA was amplified by PCR to prepare the corresponding libraries and then sequenced on illumina NovaSeq 6000.

first: Longjie

last: Sun

first: Juan

last: Chen

first: Rong

last: Ye

first: Zheng

last: Lv


title: RNA-seq data of human scalp hair follicles following SEP1 phage treatment

city: Muenster

first: Markus

last: Fehrholz

address: Mendelstr. 17

institute: Monasterium Laboratory GmbH

summary: Human scalp hair follicles were snapfrozen directly after surgery or organ-cultured in the presence/absence of SEP1 phages. After treatment total RNA was isolated and prepared for RNA-seq

overall_design: Human anagen VI hair follicles were snap frozen after surgery or organ cultured for 30h in the presence/absence of SEP1 phages and total RNA was isolated for human trancriptomic analysis by RNA sequencing

first: Marta

last: Lousada

first: Janin

last: Edelkamp

first: Tim

last: Lachnit

first: Irena

last: Pastar

first: Francisco

last: Jimenez

first: Hanieh

last: Erdmann

first: Thomas

middle: C

last: Bosch

first: Ralf

last: Paus


title: ONTOGENY SHAPES THE ABILITY OF ETV6-RUNX1 TO ENHANCE HEMATOPOIETIC STEM CELL SELF-RENEWAL AND DISRUPT EARLY LYMPHOPOIESIS [in vitro]

first: Mohamed

last: Eldeeb

overall_design: FL (E14.5) and BM (8 weeks old) HSCs were sorted from WT or iE/R and cultured for three days in ex vivo HSC expansion conditions (Wilkinson et al., 2019) with 1 μg/mL DOX (Sigma-Aldrich)

first: Anna

last: Konturek-Ciesla

first: Qinyu

last: Zhang

first: Shabnam

last: Kharazi

first: Johanna

last: Tingvall-Gustafsson

first: Jonas

last: Ungerbäck

first: Mikael

last: Sigvardsson

first: David

last: Bryder


title: Identification of genes enriched in Nkx2-1-positive neurons in the Arc, VMH, and DMH of adult mice

city: St Louis

first: Kyohei

last: Tokizane

state: MO

address: 660 S Euclid Ave

department: Developmental Biology

institute: Washington University School of Medicine

summary: We aimed to identify genes enriched in Nkx2-1-positive neurons in the dorsomedial hypothalamus (DMH). Using Nkx2-1-CreERT2 mice crossed with Rosa-ZsGreen mice after tamoxifen induction, we collected ZsGreen-marked Nkx2-1-positive neurons from the DMH, ventromedial hypothalamic nucleus (VMH), and arcuate nucleus (Arc) by laser microdissection and compared their gene expression profiles.

overall_design: Nkx2-1-positive neurons (marked by ZsGreen) in the Arc, VMH and DMH were collected from 2-month-old Nkx2-1-CreERT2 mice crossed with ROSA-loxSTOPlox-ZsGreen1 mice after tamoxifen injection by laser micro-dissection using the Leica LMD 6000 system (n=2-3 each group).

first: Cynthia

middle: S

last: Brace

first: Shin-ichiro

last: Imai


title: Effect of depletion of Ppp1r17 on gene expression in the dorsomedial hypothalamus (DMH)

summary: To investigate the function of Ppp1r17 in the DMH, we established laser microdissection to obtain the DMH specifically which Ppp1r17 has been knocked down by lentivirus carrying shRNA using stereotactic injection system.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for DMH and its KD derivatives (shPpp1r17).

first: Tokizane

last: Kyohei

first: Imai

last: Shin-ichiro


title: Autonomous transposons tune their sequences to ensure somatic suppression [polyA_HeLa]

first: IBRAHIM

last: ILIK

overall_design: Gene expression profiling of HeLa cells subject to knockdown of three SAFB proteins (SAFB1, SAFB2 and SLTM) at the same time, and a control treatment. poly(A)+ RNAs were sequenced in three biological replicates per treatment (6 libraries in total).

first: Ibrahim

last: Ilik

first: Petar

last: Glažar

first: Tugce

last: Aktas


title: Splicing factor SRSF1 is essential for homing of precursor spermatogonial stem cells in mice

first: Jiali

last: Liu

summary: Spermatogonial stem cells (SSCs) provide a continuous spermatogenesis and male fertility. However, the underlying mechanisms of alternative splicing (AS) in mouse SSCs are still largely unclear. We demonstrated that SRSF1 is essential for gene expression and splicing in mouse SSCs. Specific deletion of Srsf1 in mouse germ cells impairs self-renewal and homing of SSCs leading to male infertility. Whole-mount staining data showed the absence of germ cells in the testes of adult cKO mice, which indicates severe non-obstructive azoospermia (NOA) in cKO mice. Expression of SSC-related genes (Gfra1, Pou5f1, Plzf, Dnd1, Stra8, and Taf4b) was significantly reduced in the testes of conditional knockout (cKO) mice. CLIP-seq data found that SSC-related genes (Plzf, Id4, Setdb1, Stra8, Tial1, Bcas2, Ddx5, Srsf10, Uhrf1, and Bud31) were bound by SRSF1 in the mouse testes. Moreover, multi-omics analysis suggests that SRSF1 directly binds and regulates the expression of Tial1 via AS to implement SSCs self-renewal and differentiation. Collectively, our data reveal the critical role of an SRSF1-mediated AS in SSCs self-renewal and differentiation, which may provide a framework to elucidate the molecular mechanisms of the post-transcriptional network underlying SSCs.

overall_design: Testes were isolated and prepared from 5 dpp mouse. Total RNA was extracted from 5 dpp mouse testes

first: Longjie

last: Sun

first: Zheng

last: Lv


title: Autonomous transposons tune their sequences to ensure somatic suppression [polyA_3T3]


title: iPSC-Derived PSEN2 (N141I) astrocytes and microglia exhibit a primed inflammatory phenotype

status: Public on Jan 07 2024

city: DARLINGTON

first: Andre

last: Mckenzie

state: New South Wales

address: J03 Level 1 Mylab HubElectrical Engineering buildi

country: Australia

institute: University of Sydney

summary: Background: Widescale evidence points to the involvement of glia and immune pathways in the progression of Alzheimer’s disease (AD). AD-associated iPSC-derived glial cells show a diverse range of AD-related phenotypic states encompassing cytokine/chemokine release, phagocytosis and morphological profiles, but to date studies are limited to cells derived from PSEN1, APOE and APP mutations or sporadic patients. The aim of the current study was to successfully differentiate iPSC-derived microglia and astrocytes from patients harbouring an AD-causative PSEN2 (N141I) mutation and characterise the inflammatory and morphological profile of these cells. Methods: iPSCs from three healthy control individuals and three familial AD patients harbouring a heterozygous PSEN2 (N141I) mutation were used to derive astrocytes and microglia-like cells and cell identity and morphology were characterised through immunofluorescent microscopy. Cellular characterisation involved the stimulation of these cells by LPS and Aβ42 and analysis of cytokine/chemokine release was conducted through ELISAs and multi-cytokine arrays. The phagocytic capacity of these cells was then indexed by the uptake of fluorescently labelled fibrillar Aβ42. Results: AD-derived astrocytes and microglia-like cells exhibited an atrophied and less complex morphological appearance than healthy controls. AD-derived astrocytes showed increased basal expression of GFAP, S100β and increased secretion and phagocytosis of Aβ42 while AD-derived microglia-like cells showed decreased IL-8 secretion compared to healthy controls. Upon immunological challenge AD-derived astrocytes and microglia-like cells show exaggerated secretion of the pro-inflammatory IL-6, CXCL1, ICAM-1 and IL-8 from astrocytes and IL-18 and MIF from microglia.Conclusion: Our study showed, for the first time, the differentiation and characterisation of iPSC-derived astrocytes and microglia-like cells harbouring a PSEN2 (N141I) mutation. PSEN2 (N141I)-mutant astrocytes and microglia-like cells presented with a ‘primed’ phenotype characterised by reduced morphological complexity, exaggerated pro-inflammatory cytokine secretion and altered Aβ42 production and phagocytosis.

overall_design: To investigate the formation of induced pluripotent stem cell derived astrocytes. RNA-seq was completed for iPSC-derived astrocytes neuroal progenitor cells and primary astrocytes. RNA-seq data was then analsyed for differential gene expression comparing cell types

first: Michael

last: Sullivan

first: Samuel

middle: D

last: Lane

first: André

last: McKenzie

first: Sarah

middle: R

last: Ball

first: Margaret

last: Sunde

first: Gregory

last: Neely

first: Cesar

middle: L

last: Moreno

first: Alexandra

last: Maximova

first: Eryn

last: Werry

last: Kassiou


title: Autonomous transposons tune their sequences to ensure somatic suppression [FLASH]

first: IBRAHIM

last: ILIK

sample_organism:
Drosophila melanogaster
Homo sapiens
Mus musculus

overall_design: FLASH libraries of 33 RBPs and 4 negative controls in HEK293 cells, 1 RBP and matching negastive control in mouse 3T3 cells, and 1 RBP and matching negastive control in Drosophila S2 cells (82 in total)

last: Ilik


title: Cardiovascular risk in patients with type 2 diabetes is associated with alteration of blood monocyte phenotype and plasma metabolome

first: Nicolas

last: Venteclef

address: 156 rue de Vaugirard

institute: Institut Necker Enfants Malades, INSERM U1151

summary: To gain insight into the relationship between circulating monocytes and cardiovascular risk (CV) progression in patients with type 2 diabetes (T2D), we collected blood monocytes (CD14 positive selection) from 92 people with type 2 diabetes and coronary artery calcium score (CAC-score). Gene expression profiles of circulating monocytes were assessed by RNA sequencing.

overall_design:
We included RNA-Seq samples from circulating monocytes of 92 patients with type 2 diabetes, stratified into four groups according to their CV risk assessed by CAC-score: very low with CAC-score = 0 (CV0 group), low with CAC-score between 1 and 100 (CV1 group), intermediate with CAC-score between 101 and 400 (CV2 group), and high with CAC-score greater than 400 (CV3 group).

**Submitter declares that the raw data files were not submitted because the RNA-seq data are the property of the Assistance Publique - Hôpitaux de Paris, who did not provide permission for the deposit of the raw sequence data**

first: Jean

last: Julla

first: Diane

last: Girard

first: Marc

last: Diedisheim

first: Charline

last: Potier

last: Riveline

first: Sophie

last: Lemoine

first: Henri

last: Garchon

first: Louis

first: Ronan

last: Roussel

first: Claire

last: Vandiedonck

first: Fawaz

last: Alzaid

middle: F

last: Gautier


title: Autonomous transposons tune their sequences to ensure somatic suppression [polyA_HEK293]

overall_design: Gene expression profiling of HEK293 cells subject to knockdown of individual SAFB proteins - SAFB1, SAFB2 and SLTM, all three SAFB proteins at the same time, and a control treatment. poly(A)+ RNAs were sequenced in three biological replicates per treatment (15 libraries in total).

first: Ibrahim

first: Petar

last: Glažar

first: Tugce

last: Aktas


title: Autonomous transposons tune their sequences to ensure somatic suppression.

platform_organism:
Drosophila melanogaster
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Single cell RNA-seq of myeloid cells in human age-related macular degeneration (AMD)

city: Durham

first: Daniel

last: Saban

state: NC

address: 2351 Erwin Rd, Duke Eye Center

institute: Duke University

summary: This study aims to characterize myeloid cells collected from neuroretinas and RPE/choroid tissues of human donors at the age of over 70.

overall_design: Profiling of myeloid cells in human using single cell RNA-seq

first: Chen

last: Yu

first: Sejiro

last: Littleton


title: Autonomous transposons tune their sequences to ensure somatic suppression [polyA_S2]

first: IBRAHIM

last: ILIK

overall_design: Gene expression profiling of Saf-B-depleted D. melanogaster S2 cells, and a control treatment. poly(A)+ RNAs were sequenced in three biological replicates per treatment (6 libraries in total).

last: Ilik