title: Lymphatic migration of unconventional T cells promotes site-specific immunity in distinct lymph nodes

city: Würzburg

first: Antoine-Emmanuel

last: Saliba

address: Josef-Schneider-Straße 2 / D15

institute: Helmholtz Institute for RNA-based Infection Research

summary: The adaptive limb of the immune system consists of antibody producing B cells and CD4 T helper and CD8 cytotoxic T cells. Besides these classical lymphocyte subsets, unconventional T cells exist that are characterized by a more limited repertoire of their TCR chains. Gamma delta (gd), mucosal-associated invariant (MAIT) and natural killer T cells (NKT) are the major cell types comprising this invariant T cell compartment. These cells are found throughout the body in the non- and lymphoid tissues and they recognize antigens that are linked to non-polymorphic antigen-presenting molecules like CD1 and MR1. Functionally, these cells typically recognize lipids, small-molecule metabolites and phosphoantigens that may be pathogen-derived or expressed by tissues in the context of activation or stress responses. Investigating these cell types on functional level as a group, we found that tissue-derived unconventional T cells constantly migrate like dendritic cells to draining lymph nodes. scRNAseq revealed transcriptional homogeneity of these subsets and shared functional outputs that as group, rather than separate entities, are critical to control bacterial infections. Importantly, since every tissue harbors a unique set of invariant T cells with specific differentiation states (Th1-like, Th2-like and Th17-like) every draining lymph node is as well populated by a unique composition of such cells. By comparing different lymph nodes using scRNA in an unbiased manner, we could resolve internodal differences and further demonstrate the functional consequences on humoral and cellular immune responses. The discovery that every lymph node mounts a unique immune response has a direct impact on vaccination strategies and immunotherapy approaches that aims at harnessing invariant T cells.

overall_design: Skin draining lymph nodes from a CXCR6-GFP mouse were isolated and pooled. Single cell suspension was created and incubated with antibody mix for sorting. CXCR6-GFP high and CD44 positive cells were sorted and further processed for sequencing.

first: Konrad

last: Knöpper

first: Antoine

middle: E

first: Wolfgang

last: Kastenmüller


title: Mitochondrial DNA atlas reveals physiopathology of type 2 diabetes

status: Public on Aug 30 2022

city: shenzhen

first: Yueying

last: Xiang

state: Guangdong

address: 1098 Xueyuan Avenue, Nanshan District,Shenzhen 518000,China

department: Shenzhen University General Hospital health management center

country: China

institute: shenzhen university

type: Genome variation profiling by high throughput sequencing

summary: Type 2 diabetes (T2D), one of the most common metabolic diseases, is the result of insulin resistance or impaired insulin secretion by mitochondrial dysfunctions. Mitochondrial DNA (mtDNA) polymorphisms play an important role in physiological and pathological characteristics of T2D, however, their mechanism is poorly understood. To directly identify candidate mtDNA variants associated with T2D at the genome-wide level, we constructed forty libraries from ten patients with T2D and thirty control individuals for deep sequencing. We characterized their mtDNA atlas, and analyzed their single nucleotide polymorphisms (MtSNPs), insertions and deletions (InDels), and screened potential mtDNA mutation sites associated with T2D. We found ten mtDNA polymorphisms at nucleotides 489T > C, 3105AC > A, 3107N > C, 8701A > G, 9540T > C, 10398A > G, 10400C > T, 10873T > C, 12705C > T and 14783T > C that showed a significant difference between patients and control subjects. Therefore, our results characterize mtDNA atlas of patients with T2D, and further demonstrate that mtDNA variants are participated in the pathophysiology of T2D and other diseases. In addition, mtDNA variants may be candidate molecular biomarkers of T2D, and they may be valuable for early diagnosis of T2D in the future.

first: Yong

last: Dai


title: Transcriptome analysis of GIGANTEA gene-edited Chinese cabbage with high-temperature treatment.

city: Gwangju,Korea

first: Jeong-Il

last: Kim

address: Chonnam National University, Gwangju 61186, Korea

department: Department of Integrative Food, Bioscience and Biotechnology, Chonnam National University

country: South Korea

institute: Chonnam national university

type: Expression profiling by high throughput sequencing

summary: We report here NGS RNA-seqencing datasets with wild-type (Chiifu) and GI-edited Chinese cabbage (named gi/CRISPR) grown under optimal temperature (22℃) or treated with high-temperature (37°C for 1 h).  Through these RNA-seq analyses, we found differential expression of genes involved in elongated growth between Chiifu and gi/CRISPR plants.

overall_design: Two different temperature treatment (22,37) to B. rapa whole plant: 22; 2weeks old (non-heat treated), 37; 37°C for 1 h,  Chiifu; wild-type Chinese cabbage, gi; GIGANTEA-edited Chinese cabbage


title: Integrated analysis of  transcriptional and chromatin alteration in Jak2V617F and Asxl1-/-Jak2V617F BM c-kit+ cells

status: Public on Aug 29 2022

city: Tianjin

first: Zhijian

last: Xiao

address: 288 Nanjing Road

department: MDS and MPN center

institute: State Key Laboratory of Experimental Hematology, Institute of Hematology & Blood Diseases Hospital, CAMS & PUMC

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We performed bulk RNA-seq, transposase-accessible chromatin using sequencing (ATAC-Seq) and chromatin immunoprecipitation (ChIP)-seq on Asxl1-/-Jak2VF and Jak2VF BM c-kit+ cells to elucidate the transcriptional and associated epigenetic alterations after Asxl1 deletion. GSEA showed that the upregulated genes in Asxl1-/-Jak2VF were significantly associated with bona fide PcG target genes, as identified by the overlap between H3K27me3 and H2AK119ub1 ChIP-seq experiments . Integrated analysis of RNA-seq and ATAC-seq data showed that there existed a significant increase of chromatin accessibility associated with upregulated genes, and these sites with gained accessibility were enriched with increased levels of H3K4me1 and H3K27ac, histone marks of active enhancers in Asxl1-/-Jak2VF BM c-kit+ cells. Moreover, PcG target gene Egr1 was upregulated in BM c-kit+ cells compared with other three genotypes and considered as key gene for disease phenotypes.

overall_design: Examination of chromatin accessibility, histone modifications and mRNA expression in BM c-kit+ cells.

first: Zhongxun

last: Shi

first: Yingying

last: Zhao

first: Zexing

last: Li

first: Jinqin

last: Liu


title: RBM45 is an m6A-binding protein that affects neuronal differentiation and the splicing of a subset of pre-mRNAs

city: D

first: Kate

last: Meyer

state: NC

address: 307 Research Drive Box 3711

department: Biochemistry

country: USA

institute: Duke University School of Medicine

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: RBM45 is an m6A-binding protein that affects neuronal differentiation and the splicing of a subset of pre-mRNAs III

summary: Here, we identify RNA-binding motif containing protein 45 (RBM45) as a novel m⁶A reader, enriched in the brain, and highly expressed during embryonic neurodevelopment. We demonstrate that RBM45 recognizes intronic m⁶A and that its primary role is as a direct regulator of pre-mRNA splicing for its target genes, many of which are associated with neurodevelopment. Furthermore, we show that RBM45 is absolutely required for proper differentiation in cultured SH-SY5Y neuroblastoma cells, and that RBM45 regulates splicing of neurodevelopmental target genes central to the activation of gene expression programs that regulate differentiation in vitro.

overall_design: in vitro DART-seq m⁶A profiling on subcellular fractionated RNAs (chromatin associated (CA), nuclear, and cytoplasmic RNA) from human embryonic kidney cells (HEK293T)

first: Seung

middle: H

last: Choi

first: Huanyu

last: Zhu

first: Mathieu

last: Flamand

middle: D


title: Polo-like kinase 1 inhibitors differentiate triple-negative breast cancer towards a luminal cell fate

city: Basel

first: Mohamed

last: Bentires-Alj

address: Hebelstrasse 20

department: DBM

country: Switzerland

institute: University Hospital Basel

summary: Transcriptomic profiling was applied to investigate the effects of polo-like kinase 1 inhibition in triple-negative breast cancer.

overall_design: SUM149PT cells were treated in vitro with rigosertib for 3 days with or without the supplementation of estradiol.

first: Atul

last: Sethi


title: A second promoter on the human mitochondrial light strand

city: Gothenburg

first: Claes

middle: M

last: Gustafsson

address: Medcinaregatan 9A

department: Department of Medical biochemistry and cell biology

country: Sweden

institute: Gothenburg University

summary: Human mitochondria contain multiple copies of a circular, double-stranded DNA genome which encode for unique protein components required for the synthesis of ATP. On each strand of mitochondrial DNA (mtDNA), polycistronic transcription is initiated from a dedicated promoter. Transcription from the light strand promoter can produce both near-genome length transcripts and short RNA primers for mtDNA replication. In this way, light strand gene expression and mitochondrial DNA replication are seen as coupled events, although there is no consensus as to how this is regulated. Here we report the discovery of a second light strand promoter (LSP2) in human mtDNA, located immediately upstream of the first gene on the light strand. LSP2 possesses nearly identical characteristics to canonical mitochondrial promoters. Our results challenge the prevailing model in mammals wherein regulation of transcription from the light strand promoter is a prerequisite for the synthesis of light strand genes, and suggests that light strand transcription can be decoupled from replication primer formation.

overall_design: Cappable-seq and tagRNA-seq library preparation and sequencing on RNA isolated from HeLa (human) mitochondria were performed by Vertis Biotechnologie (Freising, Germany) with duplicate.

first: Xie

last: Xie

first: Yonghong

first: Benedict

middle: G

last: Tan

first: Maria

last: Falkenberg


title: PI3K Signaling Specifies Proximal-Distal Fate by Driving a Developmental Gene Regulatory Network in SOX9+ Lung Progenitors

status: Public on Aug 25 2022

city: Cincinnati

first: Daniel

middle: Todd

last: Swarr

state: OH- OHIO

address: 3333 Burnet Ave, ML 7009

department: Division of Neonatology & Pulmonary Biology

institute: Cincinnati Children's Hospital Medical Center


title: Expression data from CD43- B220+ splenocytic B cells of WT and Cd37-/- C57BL/6J mice.

status: Public on Aug 26 2022

city: Nijmegen

first: Annemiek

middle: van

last: Spriel

address: Geert Grooteplein Zuid 28

department: Tumor Immunology Lab

country: Netherlands

institute: Radboud Institute for Molecular Life Sciences

summary:
CD37 is a tetraspanin with important functions in the immune system. Here we have isolated RNA from murine splenocytic B cells of WT and CD37 KO mice to check for differences in gene expression.
These data are used to obtain genes that are differentially expressed in response to loss of CD37.

overall_design:
6 Total samples were analyzed. We generated the following pairwise comparisons WT1>CD37KO1; WT1>CD37KO2; WT1>CD37KO3; WT2>CD37KO1; WT2>CD37KO2; WT2>CD37KO3; WT3>CD37KO1; WT3>CD37KO2; WT3>CD37KO3.
NOTE: The submitters declare that CEL files are not available anymore since these analyses have been performed more than 10 years ago (in 2011).

first: Eugène

last: Verwiel

last: van Spriel


title: Transcriptomic signatures responding to PKM2 activator TEPP-46 in the hyperglycemic human renal proximal epithelial tubular cells

city: Kongens Lyngby

first: Xiao

last: Wang

address: Bagsværdvej 103, ST

country: Denmark

institute: Konge Larsen Aps

summary: Pyruvate kinase M2 (PKM2), as the terminal and last rate-limiting enzyme of the glycolytic pathway, is an ideal enzyme for regulating metabolic phenotype. PKM2 tetramer activation has shown a protective role against diabetic kidney disease (DKD). However, the molecular mechanisms involved in diabetic tubular has not been investigated so far. In this study, we performed transcriptome gene expression profiling in human renal proximal tubular epithelial cell line (HK-2 cells) treated with high D-glucose (HG) for 7 days before the addition of 10-μM TEPP-46, an activator of PKM2 tetramerization, for a further 1 day in the presence of HG. Afterwards, we analyzed the differentially expressed (DE) genes and investigated gene relationships based on weighted gene co-expression network analysis (WGCNA). The results showed that 2,902 DE genes were identified (adjusted P-value ≤ 0.05), where 2,509 DE genes (86.46%) were co-expressed in the key module. Four extremely down-regulated DE genes (HSPA8, HSPA2, HSPA1B and ARRB1) and three extremely up-regulated DE genes (GADD45A, IGFBP3 and SIAH1) enriched in the down-regulated endocytosis (hsa04144) and up-regulated p53 signaling pathway (hsa04115), respectively, were validated by the qRT-PCR experiments. The qRT-PCR results showed that the relative expression levels of HSPA8 (adjusted P-value = 4.45×10-34 & log2(FC) = -1.12), HSPA2 (adjusted P-value = 6.09×10-14 & log2(FC) = -1.27), HSPA1B (adjusted P-value = 1.14×10-11 & log2(FC) = -1.02) and ARRB1 (adjusted P-value = 2.60×10-5 & log2(FC) = -1.13) were significantly different (P-value < 0.05) from case group to control group. Furthermore, the interactions and predicted microRNAs of the key genes (HSPA8, HSPA2, HSPA1B and ARRB1) were visualized in networks. This study identified the key candidate transcriptomic biomarkers and biological pathways in the hyperglycemic HK-2 cells responding to the PKM2 activator TEPP-46. These results will be valuable for further research on PKM2 in DKD.

overall_design: The obtained HK-2 cells were exposed to 25-mM HG medium as the hyperglycemic condition for 7 days. Thereafter, three replicates of cells exposed to HG were treated with (case group) or without (control group) 10-μM TEPP-46 for 1 day, separately. We extracted 2 μg RNA per cell replicate sample for RNA sample preparation, generated the sequencing libraries using NEBNext Library Prep Kit (NEB, USA) following the manufacturer’s recommendations and sequenced on the Illumina Hiseq X ten platform to generate the 150 bp paired-end reads step by step.

first: Zhimin

first: Dan

last: Hao

first: Jiating

last: Yu

first: Xin


title: RBM45 is an m6A-binding protein that affects neuronal differentiation and the splicing of a subset of pre-mRNAs VI

first: Kate

last: Meyer

overall_design: Gene expression profiling of RBM45 depletion and restoration (ribodepleted-RNA-seq) on mHiPPoE-2 stable cell lines

first: Seung


title: The immunogenicity of human kidney organoids

city: Rotterdam

first: Hector

last: Tejeda Mora

address: Doctor Molewaterplein 40

department: Developmental Biology

institute: Erasmus MC

summary: Safety issues of human iPSC-derived kidney organoids as a regenerative therapy need to be evaluated.  Therefore, we studied the immunogenicity of human iPSC-derived kidney organoids. We subcutaneously implanted kidney organoids in immune-deficient IL2Ry-/-RAG2-/- mice for 1 month and hereafter performed adoptive transfer of healthy allogeneic human PBMC.  We used single cell RNA sequencing (scRNA-seq) to analyze the diversity of kidney organoid cells and immune cell profiles. We investigated whether innate and adaptive immune cells invade kidney organoids, evoke an immune response, and influence the kidney organoid differentiation and functional capacity. Understanding the immunogenicity of kidney organoids will advance studies in the applicability of kidney organoids for regenerative medicine. Furthermore, it can serve as an in-vivo transplantation model to study solid organ transplantation.

overall_design: Kidney organoids were harvested in-vivo from mice as described in the methods (two months after implantations and four weeks after human PBMC administration). Organoids were digested by TrypLE Select and stored in ice until sequencing. Control samples (Sample 1-3) are from kidney organoids that were subcutaneosly implanted in mice for 2 month. Samples 4-7  (PBMC 1-4) correspond to kidney organoids subcutaneosly implanted and that had an adoptive transfer of healthy allogenic human PBMC (via intraperitoneal injection).

first: Anusha

middle: S

last: Shankar

first: Eric

last: Bindels


title: Transcriptomic analysis of total RNA from melanoma tumor, originating from control and hyperlipic diet

status: Public on Aug 24 2022

city: Paris

first: Franck

last: Letourneur

address: 22 rue mechain

country: France

institute: INSERM

summary: Cancer and cardiovascular disease (CVD) share common risk factors such as dyslipidemia, obesity and inflammation. However, the role of pro-atherogenic environment and its associated low-grade inflammation in tumor progression remains underexplored. Here we show that two-week feeding of wildtype C57BL/6J mice with a high fat high cholesterol atherogenic diet (HFHCD) increases the pool of circulating Ly6C hi monocytes available for initial melanoma development, under the control of IL-1. Descendants of circulating myeloid cells, which accumulate in the tumor microenvironment of mice under HFHCD, heighten pro-angiogenic and immunosuppressive activities locally. Limiting myeloid cell accumulation or targeting VEGF-A production by myeloid cells decrease tumor growth acceleration under HFHCD. Reverting the HFHCD to a chow diet at the time of tumor implantation slows down tumor growth. Together, these data shed light on cross-disease communication between cardiovascular pathologies and cancer.

overall_design: The aim of the analysis is to compare the transcriptomic profile of primary solid tumors isolated from mice fed with a chow diet (CD) or a high fat high choleterol diet (HFHCD). C57BL/6J mice were fed a CD or HFHCD for 2 weeks before subcutaneous injection of B16-F10 melanoma cells. 4 tumors per group were harvested from mice fed with CD or HFHCD at day 9 post injection of B16-F10 melanoma cells to perform trasncriptomic arrays. Tumors were minced and lysed into RLT buffer containing 1% β-mercaptoethanol. RNA was extracted using Qiagen RNeasy kit according to the manufacturer’s instructions.  Analysis of gene was done by affymetrix microarrays at de GENOM’IC core facility (Cochin Institute). Microarray analysis help to understand the molecular pathways involved in aggravation of tumor growth under HFHCD.

first: Thi

last: Tran

first: Jean-Remi

last: Lavillegrand

first: Cédric

last: Lerévérend

first: Bruno

last: Esposito

first: Lucille

last: Cartier

first: Melanie

last: Montabord

first: Jaouen

last: Tran Rajau

first: Marc

last: Diedisheim

first: Nadege

last: Gruel

first: Khadija

last: Ouguerram

first: Lea

last: Paolini

first: Olivia

last: Lenoir

first: Emmanuel

last: Pinteaux

first: Eva

last: Brabencova

first: Corinne

last: Tanchot

first: Pauline

last: Urquia

first: Jacqueline

last: Lehmann-Che

first: Richard

last: Le Naour

first: Yacine

last: Merrouche

first: Christian

last: Stockmann

first: Hafid

last: Ait-Oufella

first: Ziad

last: Mallat

first: Alain

last: Tedgui

last: Tartour

first: Stephane

last: Potteaux


title: Endocytosis of the thrombopoietin receptor Mpl regulates megakaryocyte and erythroid maturation in mice

city: Milwaukee

first: Herve

last: Falet

state: WI

address: 8727 W Watertown Plank Rd

institute: Versiti Blood Research Institute

summary:
Dnm2fl/fl Pf4-Cre (Dnm2Plt-/-) mice lacking the endocytic GTPase dynamin 2 (DNM2) in platelets and megakaryocytes (MKs) develop hallmarks of myelofibrosis. At the cellular level, the tyrosine kinase JAK2 is constitutively active but decreased in expression in Dnm2Plt-/- platelets. Additionally, Dnm2Plt-/- platelets cannot endocytose the thrombopoietin (TPO) receptor Mpl, leading to elevated circulating TPO levels. Here, we assessed whether the hyperproliferative phenotype of Dnm2Plt-/- mice was due to JAK2 constitutive activation or elevated circulating TPO levels. In unstimulated Dnm2Plt-/- platelets, STAT3 and to a lower extent STAT5 were phosphorylated, but their phosphorylation was slowed and diminished upon TPO stimulation. We further crossed Dnm2Plt-/- mice in the Mpl-/- background to generate Mpl-/- Dnm2Plt-/- mice lacking Mpl ubiquitously and DNM2 in platelets and MKs.
Mpl-/- Dnm2Plt-/- platelets had severely reduced JAK2 and STAT3 but normal STAT5 expression. Mpl-/- Dnm2Plt-/- mice had severely reduced bone marrow MK and hematopoietic stem and progenitor cell numbers. Additionally, Mpl-/- Dnm2Plt-/- mice had severe erythroblast maturation defects, decreased expression of hemoglobin and heme homeostasis genes, increased expression of ribosome biogenesis and protein translation genes, and developed anemia with grossly elevated plasma erythropoietin levels, leading to early fatality by postnatal day 25. Mpl-/- Dnm2Plt+/+ mice had impaired EB development at three weeks of age, which normalized with adulthood. Together, the data shows that DNM2-dependent Mpl-mediated endocytosis in platelets and MKs is required for steady-state hematopoiesis and provides novel insights into a developmentally controlled role for Mpl in normal erythropoiesis, regulating hemoglobin and heme production.

overall_design: Isolated erythroblast RNA from a total of 12 mice: 3 wildtype, 3 Dnm2Plt-/- , 3 Mpl-/- , and 3 Mpl-/- Dnm2Plt-/-   were analyzed for differential gene expression using RNA-seq.

first: Nathan

last: Eaton

first: Emily

middle: K

last: Boyd

first: Ratnashree

last: Biswas

first: Melissa

last: Lee-Sundlov

first: Theresa

middle: A

last: Dlugi

first: Haley

last: Ramsey

first: Shikan

last: Zheng

first: Robert

middle: T

last: Burns

first: Martha

middle: C

last: Sola-Visner

first: Karin

last: Hoffmeister


title: Cross-amplified Barcodes on Slides for Spatial Transcriptomics Sequencing

city: Changchun

first: Zhengyang

last: Jin

state: Jilin

address: No. 1163 Xinmin Street

institute: Jilin University

summary: Using microfluidics, well-defined barcodes were generated on the slide surface by cross-amplification, followed by high-throughput sequencing using Novaseq to detect spatial transcriptomic information in the mouse brain.

overall_design: Applying new approach – cross-amplified barcodes on slides for spatial transcriptomics sequencing to mouse brain tissues.

first: Z

middle: Y

first: N

middle: Z

last: Yu

first: J

last: Bai

last: Liu

first: H

middle: R

last: Zhang

first: C

last: Liang


title: Dosage effect of TGF-beta on EMT induction of MCF10A cells

city: Yokohama

first: Kazuhide

last: Watanabe

state: Kanagawa

address: 1-7-22 Suehiro-cho Tsurumi-ku

department: Center for Integrative Medical Sciences

country: Japan

institute: Riken

summary: Epithelial-mesenchymal transition (EMT) is a key cellular process involved in development and disease progression. Single-cell transcriptomes can reveal intermediate EMT states observed in tumors, but previous studies focused on time-dependent responses in epithelial cells after stimulation with EMT signals as an in vitro model for understanding EMT progression, so it was unclear how transcriptomes can change in response to different levels of EMT inducer. Here, we performed single-cell RNA-sequencing with human mammary epithelial cells treated with various concentrations of TGF-β. We found that the dose-dependent EMT harbors multiple intermediate states at the single-cell level after two weeks of treatment, suggesting a stable EMT continuum. After correcting batch effects from experiments, we performed comparative analyses of the dose- and time-dependent EMT. We found that the dose-dependent EMT shows a stronger anti-correlation between epithelial and mesenchymal transcriptional programs and a better resolution of stages of EMT compared to the time-dependent progress. These properties enable more power to detect genes whose expressions are associated with EMT. Nonetheless, our analyses showed cell clusters unique to the time-dependent EMT, which correspond to en route cell populations that do not appear at steady states. Furthermore, combining dose- and time-dependent cell clusters gave rise to more accurate prognosis for cancer patients compared to individual EMT spectrum.

overall_design:
Normal human mammary epithelial cell MCF10A was stimulated with TGFbeta  at concentrations of 0, 12.5, 25, 50, 100, 200, 400, 800 pM for 14 days and all samples were pooled before loading into 10x Genomics. Single-cell 3'RNA-seq library was prepared according to the manufacturer's instructions.
RNA-Seq library was prepared using 10x Chromium Single Cell 3' –end Reagent Kits User Guide (v2 Chemistry).  Libraries were sequenced using paired-end sequencing (26bp Read 1 and 98bp Read 2) with a single sample index (8bp) on the Illumina HiSeq 2500.

first: Masataka

last: Takahashi

first: Hong

last: Tian

first: Panchy

last: Nicolas


title: Mettl3-dependent m6A modulation of mRNAs in the Drosophila brain attenuates the stress response

city: Philadelphia

first: Emily

last: Shields

state: Pennsylvania

address: 3400 Civic Center Blvd

institute: University of Pennsylvania

type: Methylation profiling by high throughput sequencing

summary: We observe the increase of stress resilience in Drosophila in which the m6A methyltransferase Mettl3 is knocked down. Mettl3-dependent m6A is present in the 5' UTR of genes enriched for neurogenesis and signaling pathways, and determine that Mettl3-dependent m6A regulates the decay of heat shock chaperones. Expression levels of genes with Mettl3-dependent m6A are increased with the reduction of Mettl3 and with heat shock stress in the brain.

overall_design: Brain RNA-seq and head m6A-IP used to investigate the role of the m6A RNA modification in heat shock stress and Mettl3 knockdown. Total 40 samples used for m6A-IP analysis and 47 samples for brain RNA-seq analysis

first: Alexandra

last: Perlegos

middle: J

first: Nancy

last: Bonini


title: The mitochondrial single-stranded DNA binding protein directs RNA primer formation for mtDNA replication

first: Claes

last: Gustafsson

summary: Single-stranded DNA (ssDNA) binding proteins protect regions of ssDNA formed during processes such as DNA replication and repair. We here devise a genetic screen and identify the mitochondrial ssDNA-binding protein (mtSSB) as a key regulator of mtDNA levels. In mitochondria, RNA synthesis from the light-strand promoter (LSP) is required for transcription as well as for generating the primers for initiation of mtDNA synthesis. We find that mtSSB is essential for mtDNA replication initiation, as transcription is strongly upregulated from the LSP in a mtSSB knockout mouse model, but cannot support the switch to replication. Using deep sequencing as well as biochemical reconstitution experiments, we find that mtSSB is also necessary to restrict transcription initiation and primer formation to specific promoters and origins of replication both in vitro and in vivo. Pathological mutations in human mtSSB cannot efficiently support primer maturation and origin specific initiation of mtDNA replication in vitro.

overall_design: Cappable-seq library preparation and sequencing on RNA isolated from HeLa (human) or heart (mouse) mitochondria were performed by Vertis Biotechnologie (Freising, Germany) with duplicate or triplicate.

first: Xie

last: Xie

first: Xuefeng

last: Zhu

first: Yonghong

last: Shi

first: Maria

last: Falkenberg


title: RBM45 is an m6A-binding protein that affects neuronal differentiation and the splicing of a subset of pre-mRNAs V

first: Kate

last: Meyer

overall_design: Gene expression profiling of RBM45 depletion and restoration (ribodepleted-RNA-seq) on basal, undifferentiated (DMSO), and differentiated (RA) SH-SY5Y stable cell lines

first: Seung

last: Choi


title: Transcriptome analysis of Tbx18-induced pacemaker cells (TBX18-iPMs) in comparison to ventricular cardiomyocytes

city: Atlanta

first: Gregory

last: Tharp

state: GA

address: 954 Gatewood Dr

department: Developmental and Cognitive Neuroscience

institute: Yerkes National Primate Research Center

summary: To understand the transcriptional landscape of Tbx18-induced cardiac pacemaker cells. The control cells were ventricular myocytes expressing GFP.

overall_design: Neonatal rat ventricular myocytes (NRVMs) were transduced with Adeno-Tbx18 or Adeno GFP. The myocytes were harvested for RNAseq on day 4 after gene transfer.

first: Hee

last: Cho

first: Jun

last: Li

first: Jinmo

last: Gu


title: Intratumor heterogeneity and T cell exhaustion in primary CNS lymphoma

status: Public on Aug 23 2022

city: Muenster

first: Michael

last: Heming

address: Albert-Schweitzer-Campus 1, Bldg A1

department: Department of Neurology with Institute of Translational Neurology

country: Germany

institute: University Hospital Muenster

summary: Single-cell transcriptomic atlas of leukocytes of biopsy fluid (n = 2), blood (n = 2) and CSF (n = 1), and spatial transcriptomics of biopsy tissue (n = 4) from primary CNS B cell lymphoma patients.

overall_design: Single-cell RNA and BCR sequencing of paired biopsy fluid and blood and spatial transcriptomics in primary CNS lymphoma.

first: I-Na

last: Lu

first: Oliver

last: Grauer

first: Gerd

last: Meyer zu Hörste


title: Altered Circular RNA Expression in Patients with Repeated Implantation Failure

status: Public on Aug 28 2022

city: Lanzhou

first: Lin

last: Liu

address: 1547 Duanjiatan Road, Chengguan District

institute: Reproductive Center of the First Hospital of Lanzhou University

summary: To evaluate differentially expressed circRNAs in patients with recurrent implantation failure, we used circRNA microarrays to screen circRNA expression profiles in endometrial biopsies from 6 women with recurrent implantation failure and controls. Our study revealed for the first time a unique set of circRNA expression signatures in the endometrial tissue of patients with repeated implantation failure, which may provide new molecular candidates for the diagnosis and clinical treatment of embryo implantation failure.

overall_design: We performed circRNA microarray analysis on endometrial tissue samples from six implantation windows and applied bioinformatics analysis to investigate these differentially expressed circRNAs. In addition, quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed to confirm these results.


title: PI3K Signaling Specifies Proximal-Distal Fate by Driving a Developmental Gene Regulatory Network in SOX9+ Lung Progenitors [RNA-seq]

first: Daniel

last: Swarr

summary: In order to understand the epigenetic mechanisms directing development and differentiation of SOX9+ lung epithelial progenitor cells (EPCs), paired RNA-Seq and ATAC-Seq was performed on SOX9+ EPCs sorted using flow cytometry from a Sox9-GFP reporter mouse.

overall_design: GFP+ positive lung epithelium was isolated by flow cytometry by selecting GFP+, CD326+, CD31-, CD45-, 7-AAD-negative cells into Trizol. Total RNA was isolated using RNEasy Micro Kits, and poly A RNA was sequenced as described below.

first: D

last: Khattar

first: S

last: Fernandes

first: J

last: Snowball

first: M

last: Guo

last: Gillen

last: Sinner

last: Jain

first: W

last: Zacharias


title: Central FGF21 Production Regulates Memory but Not Peripheral Metabolism

city: Iowa City

first: Kyle

middle: Harrington

last: Flippo

state: IA

address: 169 Newton Road, 3320 PBDB

department: Neuroscience and Pharmacology

institute: University of Iowa

summary: Fibroblast growth factor 21 (FGF21) is a liver-derived endocrine hormone that functions to regulate energy homeostasis and macronutrient intake. Recently, FGF21 was reported to be produced and secreted from hypothalamic tanycytes, to regulate peripheral lipid metabolism. However, rigorous investigation of FGF21 expression in the brain has yet to be accomplished. Utilizing a novel mouse model that drives CRE recombinase in FGF21-expressing cells, we demonstrate that FGF21 is not expressed in the hypothalamus, but instead is produced from the retrosplenial cortex (RSC), an essential brain region for spatial learning and memory. Furthermore, we find that central FGF21 produced in the RSC enhances spatial memory but does not regulate energy homeostasis or sugar intake. Finally, our data demonstrate that administration of FGF21 prolongs the duration of long-term potentiation in the hippocampus and enhances activation of hippocampal neurons. Thus, endogenous and pharmacological FGF21 appear to function in the hippocampus to strengthen the formation of spatial memory.

overall_design: Single cell suspensions derived from the hippocampus of KLB-Cre::Ai14 mice were subjected to FACs sorting and tdTomato positive cells were collected and processed for single cell RNA sequencing. Three conditions were used in this experiment to test the effects of FGF21 administriation on gene expression in the hippocampus; 1) Vehicle, 2) 1 day of FGF21 administration, and 3) 3 days of FGF21 administration.

first: Bolu

last: Zhou

first: Matthew

last: Potthoff


title: PI3K Signaling Specifies Proximal-Distal Fate by Driving a Developmental Gene Regulatory Network in SOX9+ Lung Progenitors [ATAC-seq]

overall_design: GFP+ positive lung epithelium was isolated by flow cytometry by selecting GFP+, CD326+, CD31-, CD45-, 7-AAD-negative cells into cell culture media. Nuclei were isolated, the transposase reaction was performed with Nextera Tn5 transposase (Illumina), and PCR amplification was performed according to previously published protocols (Buenrostro et al. 2015a).


title: LncRNA CHROMR regulates antiviral immunity in humans [ChIRP-Seq]

city: New York City

first: Coen

last: van Solingen

state: New York

address: 435 E30th street, Science Building 723T, Science Building 723T

department: Cardiology/Medicine

institute: NYU School of Medicine

summary: Long non-coding RNAs (lncRNAs) are important regulators of the immune response. Here we identify a novel function for the primate-specific lncRNA, CHROMR. We established that CHROME is most highly transcribed in human macrophages treated with influenza virus and/or polyIC.

overall_design: We used three replicate samples of THP1 macrophages and performed ChIRP-Seq

first: Kathryn

last: Moore


title: RNA sequencing data from murine KP230 undifferentiated pleomorphic sarcoma cells treated with N-Benzylquinazolin-4-amine (NB4A)

city: Cambridge

first: Anne

middle: E

last: Carpenter

state: MA

address: 415 Main Street

institute: The BROAD Institute of MIT and Harvard

summary: The "Cell Painting Assay", an image profile-based drug-discovery platform, identified NB4A as a putative small molecule inhibitor of Yes-associated protein 1 (Yap1). Treatment of KP230 cells with 10 µm NB4A daily for 3 days reduced Yap1 protein levels. Thus, we used RNA sequencing to investigate NB4A-mediated global gene expression changes in these cells.

overall_design: KP230 cells were treated with 10 µM NB4A every 24 hours for 3 days.

first: Ashley

last: Fuller

first: TS

last: Eisinger-Mathason


title: Cryoablation reshapes the immune microenvironment in the distal tumor and enhances the anti-tumor immunity

city: Guangzhou

first: YING

last: WU

state: CHINA

address: 651 Dongfeng East Rd

department: Department of Minimally Invasive Interventional Therapy

institute: Sun Yat-sen University Cancer Center

summary: As one of the local treatments, cryoablation plays an increasingly important role in the comprehensive treatment of malignant tumors with its advantages of less trauma, high reproducibility, and minimally invasive. Activation of anti-tumor immunity, another characteristic of cryoablation, has attracted more and more attention with the extensive application of immunotherapy. Unfortunately, the mechanism by which cryoablation enhances anti-tumor immunity is still unclear. In this study, we used a multi-omics approach to investigate the effects of local cryoablation on the distal tumor microenvironment. In this study, we proved that large amounts of tumor antigens are released post-cryoablation, leading to a sterile inflammatory response in distant tumors. During this period, lysosome-related pathways are activated, resulting in overexpression of SNAP23 and STXBP2, activation of immune effector cells, suppression of the release of immunosuppressive factors, and finally enhancement of anti-tumor immunity. Finally, cryoablation turning “Cold tumors” into “Hot tumors”, which shows a broad prospect in the combined immunotherapy.

overall_design:
C57BL/6J mice  xenografts were generated in 6-8-week-old C57BL/6J mice by subcutaneous implantation of 0.5 × 106 cells (one injection site per mouse/ both side). Animals were randomized in two groups (control VS cryoablation) for collection and analysis.
On the 7th day, the subcutaneous tumor of the mice grew to about 5-6mm in diameter. After anesthesia, one side of the tumor was subjected to cryoablation . The cryoprobes were inserted into the targeted lesions, and two 20-sec freezing cycles, separated by a passive 20-sec and an active 20-sec thawing session, were performed.In order to further study the molecular mechanism of the enhancement of anti-tumor immunity by cryoablation, the mice were sacrificed on the D5 day after freezing and then the contralateral tumors were immediately frozen in liquid nitrogen for 15 min and stored at -80 °C, parallel Bulk RNA sequencing.

first: Ying

last: Wu

first: Fei

last: Cao

first: Danyang

first: Shuanggang

last: chen

first: Han

last: Qi

first: Tao

last: Huang

first: Hongtong

last: Tan

first: Lujun

last: Shen

first: Weijun

last: Fan


title: LncRNA CHROME regulates the interferon response to viral challenge [ChIP-Seq]

summary: Long non-coding RNAs (lncRNAs) are important regulators of the immune response. Here we identify a novel function for the primate-specific lncRNA, CHROME. We established that CHROME is most highly transcribed in human macrophages treated with bacterial lipopolysaccharide and RNA viruses.

overall_design: We used three replicate samples of THP cells that were treated with gapmer targeting CHROME compared to two replicate control samples. In these cells we performed H3k27ac ChIP-Seq

first: Michael

last: Gildea


title: LncRNA CHROME regulates the interferon response to viral challenge [RNA-Seq]

summary: Long non-coding RNAs (lncRNAs) are important regulators of the immune response. Here we identify a novel function for the primate-specific lncRNA, CHROME. We established that CHROME is most highly transcribed in human macrophages treated with influenza virus and/or polyIC.

overall_design: We used three replicate samples of THP cells that were treated with gapmer targeting CHROME and subsequently treated these cells with polyIC.


title: MliR, a novel regulator of the MerR family involved in iron uptake

city: Buenos Aires

first: Sieira

last: Rodrigo

address: Av. Patricias Argentinas 435

country: Argentina

institute: Fundacion Instituto Leloir

summary: The MerR family of transcriptional regulators are dimeric proteins with an N-terminal helix-turn-helix DNA binding domain (DBD), followed by an antiparallel coiled-coil subunit interaction region, and usually by a C-terminal effector binding domain (EBD). This family is distinguished by the high amino acid similarity in the DBD and low similarity in the EBD, consistent with the role of this domain in each protein in sensing effector molecules. Most members of the family are activators, and act at promoters with long spacer regions and respond to the binding of inducers by distorting the promoter DNA to allow open complex formation and transcriptional activation. In MerR proteins the EBD contains a metal binding pocket formed by three cysteines. This Cys center characteristic of MerR regulators is primarily suited for metal binding and mediates activation of metal detoxification systems in bacteria. Over the last decade, it has become clear that the MerR family of regulators is more diverse than originally recognized. The availability of completed bacterial genome sequences has enabled the search for additional types of MerR-like regulators. We here present a novel MerR-like transcription factor lacking an EBD involved in iron uptake regulation (MliR) present in the marine bacterium Bizionia argentinensis (JUB59). An in silico analysis revealed that homologues of the MliR protein are widely distributed among different bacterial species. Deletion of the mliR gene led to decreased cell growth, increased cell adhesion and filamentation. RNA sequencing analysis showed that expression of several iron uptake-related mRNAs were downregulated in mliR-deletion mutant. Through NMR-based metabolomics, ICP-MS, fluorescence microscopy and biochemical analysis we evaluated metabolic and phenotypic changes associated with mliR deletion. This work provides the first evidence of a MerR-family regulator involved in iron uptake and contributes to expanding our current knowledge on iron metabolism.in bacteria.

overall_design: RNA samples corresponding to three biological replicates from the wild-type or the ΔmliR deletion mutant were analyzed

first: Martin

last: Aran

first: Leonardo

last: Pellizza

first: Rodrigo

last: Sieira

first: Magali

last: Bialer


title: Control of immediate early gene expression by CPEB4-mediated mRNA degradation [CLIP-seq]

city: Mannheim

first: Johanna

middle: Daniela

last: Schott

address: Franz-Volhard-Straße 6

department: Mannheim Institute for Innate Immunoscience

institute: Heidelberg University

summary: Immediate early genes (IEGs) represent a unique class of transcription units with rapid induction kinetics and transient expression patterns, which require IEG mRNAs to be short-lived. Here, we establish cytoplasmic polyadenylation element-binding protein 4 (CPEB4) as a major determinant of IEG mRNA instability. We identified human CPEB4 as an RNA-binding protein (RBP) with enhanced association to poly(A) RNA upon inhibition of class I histone deacetylases (HDACs), which is known to cause widespread degradation of poly(A)-containing mRNA. Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) analysis using endogenously tagged CBEP4 in HeLa cells revealed that CPEB4 preferentially binds to the 3' untranslated region (UTR) of IEG mRNAs, at U-rich sequence motifs located in close proximity to the poly(A) site. By transcriptome-wide mRNA decay measurements, we found that the strength of CPEB4 binding correlates with short mRNA half-lives, and that loss of CPEB4 expression leads to the stabilization of IEG mRNAs. Further, we demonstrate that CPEB4 mediates mRNA degradation by recruitment of the evolutionarily conserved CCR4-NOT complex, the major eukaryotic deadenylase. While CPEB4 is primarily known for its ability to stimulate cytoplasmic polyadenylation, our findings establish an additional function for CPEB4 as an RBP that enhances the degradation of short-lived IEG mRNAs.

overall_design: PAR-CLIP of CPEB4 in HeLa cells upon normal condition and treatment with HDAC inhibitor RMD. 3 replicates in each condition, of which one is with radioactive (p32) CLIP protocol, two with infrared (irCLIP) protocol.

first: Svetlana

last: Lebedeva


title: Transcriptome analysis of human aortic valvular endothelial cells with PPARG knockdown and oxLDL treatment.

status: Public on Aug 22 2022

city: Seoul

first: Jae-Hoon

last: Choi

address: 222 Wangsimniro, Seongdong Gu

department: Department of Life Science

institute: Hanyang University

summary: We isolated and cultured human aortic valvular endothelial cells from patients, and performed RNA sequencing of human aortic valvular endothelial cells transfected with siRNA targeting PPARG or negative control siRNA and treated with or without oxLDL.

overall_design: mRNA profiles of human aortic valvular endothelial cells transfected with siRNA targeting PPARG or negative control siRNA and treated with or without oxLDL.


title: Transcriptome analysis of aortic valve tissues from hyperlipidemic and control mice with normal chow diet (chow) or western diet (WD)

summary: We performed RNA sequencing of the aortic valves of C57BL/6J (the wild type with chow diet), Ldlr-/- (1. with chow diet, 2. with WD), and Apoe-/- mice (1. with chow diet, 2. with WD) to compare the overall transcriptomic characteristics within genetic differences (wild type, Ldlr-/-, Apoe-/-) or in dietary differences (chow diet versus WD).

overall_design: mRNA profiles of aortic valve tissues from control mice (C57BL/6J) with normal chow diet (chow) and hyperlipidemic mice (Ldlr-/- and Apoe-/-) with normal chow diet (chow) or western diet (WD).


title: Ribo-Seq performed in parental or CPEB4 knock-out HeLa cells

summary: Immediate early genes (IEGs) represent a unique class of genes with rapid induction kinetics and transient expression patterns, which requires IEG mRNAs to be short-lived. Here, we establish cytoplasmic polyadenylation element-binding protein 4 (CPEB4) as a major determinant of IEG mRNA instability. We identified human CPEB4 as an RNA-binding protein (RBP) with enhanced association to poly(A) RNA upon inhibition of class I histone deacetylases (HDACs), which is known to cause widespread degradation of poly(A)-containing mRNA. Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) analysis using endogenously tagged CBEP4 in HeLa cells revealed that CPEB4 preferentially binds to the 3' untranslated region (UTR) of IEG mRNAs, at U-rich sequence motifs located in close proximity to the poly(A) site. By transcriptome-wide mRNA decay measurements, we found that the strength of CPEB4 binding correlates with short mRNA half-lives, and that loss of CBEP4 expression leads to the stabilization of IEG mRNAs. Further, we demonstrate that CPEB4 mediates mRNA degradation by recruitment of the evolutionarily conserved CCR4-NOT complex, the major eukaryotic deadenylase. While CPEB4 is primarily known for its ability to stimulate cytoplasmic polyadenylation, our findings establish an additional function for CPEB4 as an RBP that enhances the degradation of short-lived IEG mRNAs.

overall_design: Preparation of ribosome-protected fragments (RPFs) followed by RNA-Seq to determine ribosome densities.

first: Fabian

last: Poetz

first: Doris

last: Lindner

first: Georg

last: Stoecklin


title: LncRNA CHROME regulates the interferon response to viral challenge  [COVID]

first: Coen

last: van Solingen

overall_design: Whole blood RNAseq performed on subjects with COVID and controls.

first: Tessa

last: Barrett

first: Jeffrey

last: Berger

first: Kathryn

last: Moore


title: Differential gene expression (DGE) analysis in HeLa cells treated with 20 nM RMD or an equal volume of solvent (DMSO) for 16 hours

overall_design: RNA-Seq to identify differentially expressed genes in HeLa cells following HDAC inhibition with 20 nM RMD for 16 hours.


title: LncRNA CHROME regulates the interferon response to viral and bacterial challenge [RNA-Seq]

overall_design: We used three replicate samples of THP cells that were treated with gapmer targeting CHROME compated to three replicate control samples. We repeated this experiment in which we treated the cells with LPS.


title: MTG8 interacts with LHX6 to specify cortical interneuron subtype identity

status: Public on Aug 21 2022

city: LONDON

first: Nicoletta

last: Kessaris

state: LONDON

address: Gower Street

department: WIBR

country: United Kingdom

institute: University College London

summary: In the study we compared migrating embryonic cortical interneurons from control mouse embryos and ones carryng homozygous deletions of either Mtg8 or Lhx6. The aim was to identify genes that are co-regulated by LHX6 and MTG8.

overall_design: RNA was isolated from fluorescent cells that had been sorted from control and mutant embryonic cortices. The cDNA was used to prepare libraries using the Nextera XT kit (Illumina). These were run on a NextSeq 500 sequencer (lllumina). Raw sequence data were adapter and quality trimmed (fastp v0.20.1), aligned to the genome with STAR (v2.7b) and reads per transcript determined with FeatureCounts (v1.6.4).

first: Agata

last: Stryjewska

first: Zeinab

last: Asgarian

first: Mohammadmersad

last: Ghorbani


title: Paternally inherited H3K27me3 affects chromatin accessibility in mouse embryos produced by round spermatid injection

city: Kofu

first: Mizuki

last: Sakamoto

institute: University of Yamanashi

summary:
Round spermatid injection (ROSI) results in a lower birth rate than
intracytoplasmic sperm injection, which has hampered its clinical
application. Inefficient development of ROSI embryos has been
attributed to epigenetic abnormalities. However, the chromatin-based
mechanism that underpins the low birth rate in ROSI remains to be
determined. Here, we show that a repressive histone mark
H3K27me3 persists from mouse round spermatids into zygotes in
ROSI and that round spermatid-derived H3K27me3 is associated
with less accessible chromatin and impaired gene expression in ROSI
embryos. These loci are initially marked by H3K27me3 but undergo
histone modification remodelling in spermiogenesis, resulting in
reduced H3K27me3 in normal spermatozoa. Therefore, the absence
of the epigenetic remodelling, presumably mediated by histone
turnover during spermiogenesis, leads to dysregulation of chromatin
accessibility and transcription in ROSI embryos. Thus, our results
unveil a molecular logic, in which chromatin states in round
spermatids impinge on chromatin accessibility and transcription in
ROSI embryos, highlighting the importance of epigenetic remodelling
during spermiogenesis in successful reproduction.

overall_design:
ATAC-seq and RNA-seq were performed with mouse ROSI and ICSI embryos at
the one- and two-cell stages.


title: ActD timecourse in HeLa cells, HeLa cells transiently transfected with control (C2) or CPEB4-targeting siRNAs S140 and S181

overall_design: Transcriptional shut-off with ActD followed by RNA-Seq to determine mRNA half-lives on a transcriptome-wide scale in HeLa cells, HeLa cells transiently transfected with control (C2) or CPEB4-targeting siRNAs S140 and S181 in one biological replicate. Cells were treated with 20 nM RMD or equal amount of solvent (DMSO) for 16 hours.


title: c-Myb instructs both stemness and functional exhaustion of CD8+ T cells during chronic infection [bulkRNA-seq]

city: Munich

first: Lorenz

last: Kretschmer

state: Bavaria

address: Trogerstr. 30

department: Institute for Medical Microbiology, Immunology and Hygiene

institute: TU Munich

summary: Chronic viral infections and tumours are associated with CD8+ T cell exhaustion, which is characterized by high expression of inhibitory receptors such as programmed cell death protein 1 (PD-1) and impaired effector function. Understanding the molecular regulation of T cell exhaustion is critical for the development of new immunotherapies. Chronically activated T cells are maintained by precursors of exhausted T (TPEX) cells that express the transcription factor TCF1, self-renew and give rise to TCF-1– exhausted effector T (TEX) cells; this process is currently, however, poorly understood. Here, we reveal that TPEX cells are heterogenous and contain a population of CD62L+ stem-like exhausted T cells that selectively preserve long-term self-renewal capacity and multipotency of antigen-specific T cells during chronic infection. Furthermore, we show that the transcription factor c-Myb is essential for the development of CD62L+ TPEX cells, self-renewal of TPEX cells and the induction of key features of T cell exhaustion. Consequently, Myb-deficient CD8+ T cells show uninhibited cytokine production resulting in fatal immunopathology in response to chronic but not acute LCMV infection and fail to be maintained long-term. Finally, we show that c-Myb-dependent CD62L+ TPEX cells exclusively mediate T cell proliferation during PD-1 checkpoint inhibition and show enhanced antiviral properties, making them attractive targets for new immunotherapeutic strategies. Thus, our findings identify CD62L+ TPEX cells as central to the maintenance of exhausted T cell responses and reveal c-Myb as a key transcriptional orchestrator that combines two central aspects of T cell exhaustion, limitation of T cell function and long-term maintenance during chronic infection.

overall_design: 3 distinct P14 T cell subsets defined by differential expression of CD62L and Ly108 were isolated from LCMV Clone 13 or LCMV Armstrong infected mice on day 28p.i., as well as naive P14 cells from the spleen of an uninfected donor mouse. 10000 cells were sorted from each population and bulk RNA-Seq was performed of all samples.

first: Carlson

last: Tsui

first: Svenja

last: Rapelius

first: Sarah

last: Gabriel

first: David

last: Chisanga

first: Konrad

last: Knöpper

first: Daniel

last: Utzschneider

first: Simone

last: Nüssing

first: Yang

last: Liao

first: Teisha

last: Mason

first: Santiago

last: Torres

first: Stephen

last: Wilcox

first: Krystian

last: Kanev

first: Sebastian

last: Jarosch

middle: L

last: Nutt

first: Dietmar

last: Zehn

first: Ian

last: Parish

first: Wolfgang

last: Kastenmüller

first: Wei

last: Shi

first: Veit

last: Buchholz

first: Axel

last: Kallies


title: LncRNA CHROME regulates the interferon response to viral challenge

first: Coen

last: van Solingen

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

overall_design: Refer to individual Series


title: Comparative expression of embryonic medial- and lateral/caudal-ganglionic eminence cells migrating in the mouse cerebral cortex

first: Nicoletta

last: Kessaris

summary: Cortical interneurons originate in the medial and caudal ganglionic eminence and migrate into the cortex during embryogenesis. We purified cells migrating within the cortex at different embryonic stages and compared their transcriptome to identify transcriptional programmes underlying distinct cortical interneuron fates.

overall_design: We used mice expressing YFP or VENUS in migrating MGE- and CGE-derived cells to purify the respective populations by FACS before extracting RNA and hybridiing on Affymetrix microarrays. We purified cells from two different stages of development: E14.5 and E18.5 for MGE cells and E16.5 and E18.5 for CGE cells.

first: Anna

middle: N

last: Rubin

first: Lorenza

last: Magno


title: Enhancer looping protein LDB1 regulates hepatocyte gene expression by cooperating with liver transcription factors

city: Bethesda

first: GUOYOU

last: LIU

state: Maryland

address: 50 South Dr, Building 50, Room 3150

institute: NIH/NIDDK

summary: To study LDB1 function in liver cell, gene expression changes and LDB1 binding and H3K27ac modification were detected by RNA-seq and ChIP-seq.

overall_design: To study how LDB1 affect gene expression in liver cells, RNA-seq were performed in control and LDB1 knockout human HepG2 cell and primary mouse liver cell. LDB1 ChIP-seq was performed in HepG2 cell to obtain genome-wide LDB1 binding. H3K27ac ChIP-seq was performed in control and LDB1 knockout HepG2 cell to study how LDB1 knockout affect enhancer mark.

first: Guoyou

last: Liu

first: Guoyun

last: Yu

first: Cameron

last: Palmer

first: Ann

last: Dean


title: KCTD9 inhibits the Wnt/β-catenin pathway by decreasing the level of β-catenin in colorectal cancer

status: Public on Aug 19 2022

city: Hefei

first: Qidong

last: Li

state: Anhui

address: 443 Huangshan Road, Shushan district

institute: Lianxin Liu Lab

summary: Colorectal cancer (CRC) is the second leading cause of cancer mortality worldwide. However, the molecular mechanisms underlying CRC progression remain to be further defined to improve patient outcomes. In this study, we found that KCTD9, a member of the potassium channel tetramerization domain-containing (KCTD) gene family, was commonly downregulated in CRC tissues and that KCTD9 expression was negatively correlated with the clinical CRC stage. Survival analysis showed that patients whose tumors expressed low KCTD9 levels had poorer outcomes. Functional analyses revealed that KCTD9 overexpression inhibited CRC cell proliferation and metastasis, whereas KCTD9 knockdown promoted CRC cell proliferation and metastasis in both in vitro and in vivo models. Manipulating KCTD9 levels in CRC cells via overexpression or knockdown showed KCTD9 expression positively influenced the degradation of β-catenin levels leading to inhibition of Wnt signaling and reductions in Wnt pathway target gene expression. Mechanistically, we found KCTD9 associated with ZNT9 (Zinc Transporter 9), a coactivator of β-catenin-mediated gene transcription. The overexpression of KCTD9 or knockdown of ZNT9 in CRC cells increased the polyubiquitination and proteasomal degradation of β-catenin. In turn, the KCTD9-ZNT9 interaction disrupted interactions between β-catenin and ZNT9, thereby leading to decreased β-catenin target gene expression and the inhibition of Wnt signaling. In conclusion, our findings propose that KCTD9 functions as a tumor suppressor that inhibits CRC cell proliferation and metastasis by inactivating the Wnt/β-catenin pathway. Moreover, its frequent downregulation in CRC suggests KCTD9 as a potential prognostic and therapeutic target in CRC.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for adjacent normal colonic mucosa, primary colon adenocarcinoma and distant liver metastasis

first: Hanhui

last: Yao

first: Yichun

last: Wang

first: Lianxin


title: c-Myb instructs both stemness and functional exhaustion of CD8+ T cells during chronic infection [scRNA-seq]

first: Lorenz

last: Kretschmer

overall_design: single cell RNA-sequencing of TCF7-GFP+ P14 T cells obtained from LCMV Clone 13 infected mice on day 28p.i.

first: Carlson

last: Tsui

first: Svenja

last: Rapelius

first: Sarah

last: Gabriel

first: David

last: Chisanga

first: Konrad

last: Knöpper

first: Daniel

last: Utzschneider

first: Simone

last: Nüssing

first: Yang

last: Liao

first: Teisha

last: Mason

first: Santiago

last: Torres

first: Stephen

last: Wilcox

first: Krystian

last: Kanev

first: Sebastian

last: Jarosch

last: Nutt

first: Dietmar

last: Zehn

first: Ian

last: Parish

first: Wolfgang


title: Profiling mouse cochlear cell maturation using 10X Genomics single-cell transcriptomics

city: Omaha

first: Jian

last: Zuo

state: Nebraska

address: 1006 N 20th St, Criss II, 313

department: Biomedical Sciences

institute: Creighton University School of Medicine

summary: Juvenile and mature mouse cochleae contain various low-abundant, vulnerable sensory epithelial cells embedded in the calcified temporal bone, making it challenging to profile the dynamic transcriptome changes of these cells during maturation at the single-cell level. Here we performed the 10X Genomics single-cell RNA sequencing (scRNA-seq) of mouse cochleae at postnatal days 14 (P14) and 28. We attained the transcriptomes of multiple cell types, including hair cells, supporting cells, spiral ganglia, stria fibrocytes, and immune cells. Our hair cell datasets are consistent with published transcripts from bulk RNA seq and scRNA-seq. We also mapped known deafness genes to corresponding cochlear cell types. Importantly, pseudotime trajectory analysis revealed that inner hair cells peak their maturation at P14 while outer hair cells continue to develop until P28. We further identified and confirmed a long noncoding RNA gene Miat expressed during maturation in cochlear hair cells and spiral ganglia neurons. Our transcriptomes of juvenile and mature mouse cochlear cells provided the sequel to those previously published at late embryonic and early postnatal ages and will be valuable resources to investigate cochlear maturation at single-cell resolution.

overall_design: We characterized the transcriptome profiles of various cochlear cell types in the calcified inner ear of P14 and P28 C57BL/6 mice using the 10x single-cell RNA-sequencing platform.

first: Zhenhang

last: Xu

first: Shu

last: Tu

first: Caroline

last: Pass

first: Yan

last: Zhang

first: Huzhan

last: Liu

first: Yusi

last: Fu

last: He


title: Perturbation-informed signatures from crosswise integration of transcriptome and chromatin accessibility analyses predict susceptibility to candidate anticancer drugs (cisplatin validation dataset)

status: Public on Aug 17 2022

city: Bonn

first: Joachim

last: Schultze

state: NRW

address: Carl-Troll-Strasse 31

department: Genomics and Immunoregulation

institute: LIMES (Life and Medical Sciences Center Genomics and Immunoregulation)


title: Perturbation-informed signatures from crosswise integration of transcriptome and chromatin accessibility analyses predict susceptibility to candidate anticancer drugs (cisplatin validation dataset) [RNA-seq]

summary: Transcriptome and chromatin state of cell lines with high (Jurkat) and low (BxPC-3) sensitivity to cisplatin was acquired after 6 h 3 uM treatment with the anticancer drug, in order to validate a drug prediction pipeline based on the construction of a perturbation-informed signature.

overall_design: Transcriptomic profile of BxPC-3 and Jurkat treated with cisplatin 3 uM for 6 h.

first: Caterina

last: Carraro

first: Lorenzo

last: Bonaguro

first: Barbara

last: Gatto


title: c-Myb instructs both stemness and functional exhaustion of CD8+ T cells during chronic infection

first: Lorenz

last: Kretschmer


title: MYB orchestrates T cell exhaustion and response to checkpoint inhibition

city: Clayton

first: Wei

last: Shi

state: Victoria

address: 15 Innovation Walk

department: Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute

country: Australia

institute: Monash University

overall_design: RNA sequencing of CD62Lpos TPEX (Ly108+TIM3-CD62L+), CD62Lneg TPEX (Ly108+TIM3-CD62L-) and TEX (Ly108-TIM3+) P14 T cells obtained from LCMV Docile infected mice on day 7 p.i..

first: Carlson

last: Tsui

first: Lorentz

first: Svenja

last: Rapelius

first: Sarah

last: Gabriel

first: David

last: Chisanga

first: Konrad

last: Knöpper

first: Daniel

last: Utzschneider

first: Simone

last: Nüssing

first: Yang

last: Liao

first: Teisha

last: Mason

first: Santiago

last: Torres

first: Stephen

last: Wilcox

last: Kastenmüller

first: Veit

last: Buchholz

first: Axel

last: Kallies


title: RNA-seq of ITPR3 knockout in SW480 colorectal cancer metastases

city: New York

first: Ryan

last: Moy

state: NY

address: 1230 York Avenue

institute: The Rockefeller University

summary: Here we analyze the transcriptional profiles of liver metastases formed by sgCtrl versus sgITPR3 SW480 colorectal cancer cells. We find downregulated genes in sgITPR3 SW480 metastases to be enriched for pathways related to immune response and non-canonical NFkB signaling.

overall_design: Differential gene expression and pathway analysis of sgITPR3 versus sgCtrl SW480 liver metastases.

middle: H


title: Control of immediate early gene expression by CPEB4-mediated mRNA degradation

first: Johanna

last: Schott


title: RNAseq from pig heart slices cultured in CTCM for 12 days

status: Public on Aug 15 2022

city: Houston

first: Tamer

last: Mohamed

state: TX

address: 6770 Bertner Avenue

institute: Baylor College of Medicine

summary: we conducted temporal bulk RNAseq on heart slices Biomimetic Cardiac Tissue Culture Model (CTCM) for 12 days

middle: M

first: Jessica

last: Miller


title: Liver Transcriptomics Reveals Features of the Host Response in a Mouse Model of Dengue Virus Infection

status: Public on Aug 18 2022

first: zheng

last: wenjiang

state: Guangdong

department: The First Clinical Medical School

institute: Guangzhou University of Chinese Medicine

summary:
Background: Dengue virus (DENV) infection induces various clinical manifestations and even causes organ injuries, leading to severe dengue haemorrhagic fever and dengue shock syndrome. Hepatic dysfunction was identified as a risk predictor of progression to severe disease during the febrile phase of dengue. However, the underlying mechanisms of hepatic injury remain unclear.
Methods: A model of dengue disease was established in IFNAR−/− C57BL/6 mice by challenge with DENV-2. Body weight, symptoms, haematological parameters and liver pathological observations in mice were used to determine the effects of DENV infection. Liver transcriptome sequencing was performed to evaluate the features of the host response FNAR  mice challenged with DENV. Functional enrichment analysis and analysis of significantly differentially expressed genes (DEGs) were used to determine the critical molecular mechanism of hepatic injury.
Results: We observed haemoconcentration, leukopenia and liver pathologies in mice, consistent with findings in clinical dengue patients. Some differences in gene expression and biological processes were identified in this study. Transcriptional patterns in the liver indicated that antiviral responses to DENV and tissue damage via abnormal expression of proinflammatory cytokines were induced. Further analysis showed that the upregulated DEGs were significantly enriched in the leukocyte transendothelial migration, complement and coagulation cascades, and cytokine-cytokine receptor interactions signalling pathways, which are considered to be closely associated with the pathogenic mechanism of dengue. IL6, IL 10, ICAM-1, VCAM-1, MMP9 and NLRP3 were identified as biomarkers of progression to severe disease.
Conclusions: The interactions of these cytokines, which activate inflammatory signalling, may lead to organ injury and haemoconcentration and even to vascular leakage in tissues, including the mouse liver. Our study identifies candidate host targets that could be used for further functional verification.
In this study, we aimed to evaluate the transcriptomics features of liver injury in a mouse model of dengue virus infection based on a lethal animal model, and then perform a series of functional analyses to partially elucidate the underlying pathogenesis of DENV-induced liver injury.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for the liver tissues of dengue virus-infected mice and control mice.

first: Wenjiang

last: Zheng

first: Qian

last: Yan

first: Zonghui

last: Li

first: Xianyang

last: Wang

first: Peng

last: Wu

first: Feng

first: Zizhao

last: Lao

first: Yong

last: Jiang

first: Xiaohong

last: Liu

first: Shaofeng

last: Zhan

first: Geng


title: Modelling aggressive prostate cancers of young men in immune-competent mice, driven by isogenic Trp53 alterations and Pten loss

status: Public on Aug 16 2022

city: Melbourne

first: Sue

last: Haupt

address: 305 Grattan St

institute: Peter MacCallum Cancer Centre

summary: Understanding prostate cancer onset and progression in order to rationally treat this disease has been critically limited by a dire lack of relevant pre-clinical animal models. We have generated a set of genetically engineered mice that mimic human prostate cancer, initiated from the gland epithelia. We chose driver gene mutations that are specifically relevant to cancers of young men, where aggressive disease poses accentuated survival risks. An outstanding value of our models are their intact repertoire of immune cells, which provide invaluable insight into the importance of immune responses in prostate cancer and offer scope for studying treatments, including immunotherapies. Our prostate cancer models strongly support the role of tumour suppressor p53 in functioning to critically restrain the emergence of cancer pathways that drive cell cycle progression; alter metabolism and vasculature to fuel tumour growth; and mediate epithelial to mesenchymal-transition, as vital to invasion. Importantly, we also discovered that the type of p53 alteration dictates the specific immune cell profiles most significantly disrupted, in a temporal manner, with ramifications for disease progression. These new orthotopic mouse models demonstrate that each of the isogenic hotspot p53 amino acid mutations studied (R172H and R245W, the mouse equivalents of human R175H and R248W respectively), drive unique cellular changes affecting pathways of proliferation and immunity. Our findings support the hypothesis that individual p53 mutations confer their own particular oncogenic gain of function in prostate cancer.

overall_design: Temporal (12 weeks old vs end point) comparative gene expression profiling analysis of 3' RNA-seq data for anterior prostate tumour tissue of mice with different genotypes. Pten(-/-)Trp53(+/+): 12-weeks old mice n=3 and endpoint mice n=4; Pten(-/-)Trp53(-/-): 12-weeks old mice n=3 and endpoint mice n=3; Pten(-/-)Trp53(R172H/R172H): 12-weeks old mice n=3 and endpoint mice n=3; Pten(-/-)Trp53(R245W/R245W): 12-weeks old mice n=3 and endpoint mice n=3.

first: Javier

middle: O

last: Mejia-Hernandez

first: Simon

middle: P

last: Keam

first: Reem

last: Saleh

first: Fenella

last: Muntz

first: Stephen

middle: B

last: Fox

first: David

last: Byrne

first: Arielle

last: Kogan

first: Lokman

last: Pang

first: Jennifer

last: Huynh

first: Cassandra

last: Litchfield

first: Franco

last: Caramia

first: Guillermina

last: Lozano

first: Hua

last: He

first: James

last: You

first: Shahneen

last: Sandhu

first: Scott

middle: G

last: Williams

first: Ygal


title: RNA-seq of SW480 colorectal cancer cells transfected with siRNAs targeting different putative metastasis promoting genes

first: Ryan

last: Moy

summary: Here we analyze the transcriptional profiles of SW480 colorectal cancer cells transfected with siRNAs targeting 26 different candidate genes potentially promoting metastasis. UMAP analysis of the transcriptional profiles of these cells revealed clusters potentially indicating some genes to operate in related pathways.

overall_design: Differential gene expression and clustering analysis of SW480 cells transfected with siRNAs targeting 26 putative metastasis promoters.


title: Next Generation Sequencing Facilitates Quantitative Analysis of the effects of PD-L2 on pTreg transcriptome

city: Los Angeles

first: Benjamin

middle: Pierre

last: Hurrell

state: CA

address: 1450 Biggy Street

department: MMI

institute: University of Southern California

summary: PD-1/PD-L2 axis controls peripherally induced regulatory T-cells

overall_design: mRNA profiles of sorted mouse spleen pTregs from either Foxp3GFP WT or Foxp3GFP PD-L2 KO mice were generated by deep sequencing, in duplicates, using a NextSeq 500 (Illumina) system.

last: P Hurrell


title: Study of TUT tailing on miRNA

city: Frederick

first: Xavier

last: Bofill-De Ros

state: MD

address: 1050 Boyles St., blg 560

department: RNA Biology Laboratory

institute: National Cancer Institute

type: Non-coding RNA profiling by high throughput sequencing

summary: Many microRNAs (miRNAs) exist alongside abundant miRNA isoforms (isomiRs), most of which arise from post-maturation sequence modifications, such as 3’ uridylation and adenylation. However, the ways in which these sequence modifications affect miRNA function remain poorly understood. To this aim, we have generated single knock-out cell lines of TUT4, TUT7 and TENT2 (TUT2), as well as double knock-out (DKO) and triple knock-out (TKO) cell lines. Here, using these different cell lines, we have discovered that some of the redundant functions and specific functions of each tailing enzyme. Our study provides a comprehensive characterization of tailing on miRNAs.

overall_design: Briefly, HEK293 cells and corresponding knock-outs were transfected with several wild-type and catalytic-dead constructs. Transfections were performed using PolyJet™ DNA Transfection Reagent (SignaGen) according to the manufacturer's instructions. 48 hours after transfection, either total or AGO-bound (IP) RNA was obtained using a chloroform-isopropanol extraction. RNA was used to prepare small RNA-seq libraries using QIAseq miRNA Library Kit (Qiagen). Constructs were purified in a 6% (w/v) native acrylamide gel based on the expected product size and purified by ethanol precipitation. Libraries quality was assessed by using Qubit dsDNA HS Assay Kit (ThermoFisher) and Agilent High Sensitivity DNA kit (Agilent). Libraries were pulled together, denaturalized and prepared at a final concentration of 12pM and run on MiSeq or NovaSeq 500 according to the manufacturer specifications.  Fastq files were analyzed using a Python-based custom software, QuagmiR (https://cgc.sbgenomics.com/public/apps#nikola_tesic/quagmir/quagmir-1-0/) designed for the optimal mapping and analysis of isomiRs.

first: Acong

last: Yang

first: Shuo

last: Gu


title: Porcine intestinal innate lymphoid cells and lymphocyte spatial context revealed through single-cell RNA sequencing

city: Columbia

first: Sathesh

middle: K

last: Sivasankaran

state: MO

address: Bond Life Sciences Center Suite 106, 1201 Rollins Street

institute: University of Missouri

summary: Intestinal lymphocytes are crucial members of the mucosal immune system with impact over outcomes of intestinal health versus dysbiosis. Resolving intestinal lymphocyte complexity and function is a challenge, as the intestine provides cellular snapshots of a diverse spectrum of immune states. In pigs, intestinal lymphocytes are poorly described relative to humans or traditional model species. Enhanced understanding of porcine intestinal lymphocytes will promote food security and improve utility of pigs as a biomedical model for intestinal research. Single-cell RNA sequencing (scRNA-seq) was performed to provide transcriptomic profiles of lymphocytes in porcine ileum, with 31,983 cells annotated into 26 cell types. Deeper interrogation revealed previously undescribed cells in porcine intestine, including SELLhi gd T cells, group 1 and group 3 innate lymphoid cells (ILCs), and four subsets of B cells. Single-cell transcriptomes in ileum were compared to those in porcine blood, and subsets of activated lymphocytes were detected in ileum but not periphery. Comparison to scRNA-seq human and murine ileum data revealed a general consensus of ileal lymphocytes across species. Lymphocyte spatial context in porcine ileum was conferred through differential tissue dissection prior to scRNA-seq. Antibody-secreting cells, B cells, follicular CD4 ab T cells, and cycling T/ILCs were enriched in ileum with Peyer’s patches, while non-cycling gd T, CD8 ab T, and group 1 ILCs were enriched in ileum without Peyer’s patches. scRNA-seq findings were leveraged to develop advanced toolsets for further identification of ILCs in porcine ileum via flow cytometry and in situ staining. Porcine ileal ILCs identified via scRNA-seq did not transcriptionally mirror peripheral porcine ILCs (corresponding to natural killer cells) but instead had gene signatures indicative of tissue- and activation-specific functions, indicating potentially similar roles to intestinal ILCs identified in humans. Overall, the data serve as a highly-resolved transcriptomic atlas of the porcine intestinal immune landscape and will be useful in further understanding intestinal immune cell function.

overall_design: Ileum was collected from each of two seven-week-old pigs and dissected into three sample types: (1) ileum containing only regions with Peyer’s patches (PP), (2) ileum containing only regions without Peyer’s patches (non-PP), and (3) ileum representative of a transverse cross section containing both regions with and without Peyer’s patches (whole ileum). From each of the six ileum samples, cells were isolated from combined epithelium, lamina propria, and submucosa and enriched for viable lymphocytes. In parallel, PBMC samples were collected for the same two pigs and processed to obtain single-cell suspensions. Sequences used in htis stduy were deposited to the NCBI SRA and can be fount using the identifier PRJNA802582.

first: Jayne

last: Wiarda

first: Julian

last: Trachsel

first: Christopher

last: Tuggle

first: Crystal

last: Loving


title: Activation of TLR-like pathways in cDCs involving TRIF and the type I IFNs leads to a protective adaptive response against influenza infection

status: Public on Aug 12 2022

city: Farmington

first: Djamel

last: Nehar-Belaid

state: CONNECTICUT

address: 10 discovery drive

institute: THE JACKSON LABORATORY

summary: Transcriptomic analyses of rASP-1-activated human blood cDCs revealed that rASP-1 induces upregulation of genes associated with DC maturation, type I IFN and type II IFN signaling involving the TLR4-TRIF transcript components. This transcriptome data in conjunction with the mouse in vitro and and in vivo data suggest the role of TRIF and type I IFNs as critical players in the mechanism of rASP-1 in eliciting a protective response against influenza infection.

overall_design: Human PBMCs, used as source of cDCs, were isolated from leukopaks obtained from three healthy adult volunteers (New York Blood Center) and stimulated with rASP-1 or medium (control)

first: Parakkal

last: Jovvian George

first: Radu

last: Marches

first: Jacques

last: Banchereau

first: Sara

last: Lustigman


title: Perturbation-informed signatures from crosswise integration of transcriptome and chromatin accessibility analyses predict susceptibility to candidate anticancer drugs (cisplatin validation dataset) [ATAC-seq]

first: Joachim

last: Schultze

overall_design: Chromatin accessibility profile of BxPC-3 and Jurkat treated with cisplatin 3 uM for 6 h.

first: Caterina

last: Carraro

first: Lorenzo

last: Bonaguro

first: Barbara

last: Gatto


title: The Human UPAT and the Arabidopsis APOLO lncRNA regulated a comparable set of genes in Arabidopsis

status: Public on Aug 11 2022

city: Gif-sur-Yvette

first: Thomas

last: Blein

address: Avenue des Sciences

institute: CNRS

summary: The overexpression of the Arabidopsis APOLO lncRNA or the human UPAT lncRNA in Arabidopsis seedlings show a significant overlap between APOLO and UPAT deregulated genes, strongly suggesting they exert similar molecular mechanisms.

overall_design: 4-day-old Col, 35S:APOLO or 35S:UPAT seedlings growing at 23°C were analyzed for RNA-Seq in triplicates

first: Camille

last: Fonouni-Farde

first: Aurélie

last: Christ

first: Federico

last: Ariel


title: scRNA-seq or Arabidopsis leaves

city: La Jolla

first: Travis

middle: Andrew

last: Lee

address: 10010 N Torrey Pines Rd

institute: Salk Institute for Biological Studies

summary: scRNA-seq of mature Arabidopsis leaf cells.

overall_design: Protoplasts were generated from 17 d-old first true leaves of Arabidopsis and subjected to scRNA-seq (10X Genomics).

first: Carl

last: Procko

first: Joanne

last: Chory

first: Joseph

last: Ecker


title: Study of TUT tailing on miRNA (Rescue TUT activity)

first: Xavier

last: Bofill-De Ros

summary: Many microRNAs (miRNAs) exist alongside abundant miRNA isoforms (isomiRs), most of which arise from post-maturation sequence modifications, such as 3’ uridylation and adenylation. However, the ways in which these sequence modifications affect miRNA function remain poorly understood. To this aim, we have generated knock-out cell lines of TUT4, TUT7 and TENT2 (TUT2), and rescued with plasmids expressing the cDNA of such enzymes. Here, using these different cell lines, we have discovered that some of the redundant functions and specific functions of each tailing enzyme. Our study provides a comprehensive characterization of tailing on miRNAs.

overall_design: Briefly, HEK293 cells and corresponding knock-outs were transfected with several wild-type and catalytic-dead constructs. Transfections were performed using PolyJet™ DNA Transfection Reagent (SignaGen) according to the manufacturer's instructions. 48 hours after transfection, either total or AGO-bound (IP) RNA was obtained using a chloroform-isopropanol extraction. Total RNA was used to prepare small RNA-seq libraries using QIAseq miRNA Library Kit (Qiagen). Constructs were purified in a 6% (w/v) native acrylamide gel based on the expected product size and purified by ethanol precipitation. Libraries quality was assessed by using Qubit dsDNA HS Assay Kit (ThermoFisher) and Agilent High Sensitivity DNA kit (Agilent). Libraries were pulled together, denaturalized and prepared at a final concentration of 12pM and run on MiSeq or NovaSeq 500 according to the manufacturer specifications.  Fastq files were analyzed using a Python-based custom software, QuagmiR (Bofill-De Ros et al. Bioinformatics 2019) designed for the optimal mapping and analysis of isomiRs.

last: Bofill-Ros

first: Acong

last: Yang

first: Shuo

last: Gu


title: Atrial myocytes-derived exosomal microRNA contributes to atrial fibrosis in atrial fibrillation

city: Harbin

first: Yue

last: Li

state: Heilongjiang

address: Youzheng Street 23#

institute: The First Affiliated Hospital of Harbin Medical University

summary:
Background
Atrial fibrosis plays a critical role in the development of atrial fibrillation (AF). Exosome is a promising cell-free therapeutic approach for the treatment of AF. The purpose of this study was to explore the mechanisms underlying exosomes derived from atrial myocytes regulated atrial remodeling and ask whether their manipulation allows for therapeutic modulation of fibrosis potential abnormalities during AF.
Methods
We isolated exosomes from atrial myocytes and patients serum, microRNA (miRNA) sequencing analyzed the exosomal miRNAs in atrial myocytes-exosomes and patients serum-exosomes. mRNA sequencing and bioinformatics analysis corroborate the key gene as direct targets of miR-210-3p.
Results
The miRNAs sequencing analysis identified that miR-210-3p expression significantly increased in exosomes of tachypacing atrial myocytes and serum of AF patients. In vitro, the analysis showed that miR-210-3p inhibitor reversed tachypacing-induced proliferation and collagen synthesis in atrial fibroblasts. Accordingly, KO miR-210-3p could reduce the incidence of AF and ameliorate atrial fibrosis induced by Ang Ⅱ. The mRNA sequencing analysis and Dual-Luciferase reporter assay proved that glycerol-3-phosphate dehydrogenase 1-like (GPD1L) is the potential target gene of miR-210-3p. The functional analysis suggests that GPD1L regulated atrial fibrosis via PI3K/AKT signaling pathway. Besides, silencing GPD1L in atrial fibroblasts induced cells proliferation and these effects could be reversed by PI3K inhibitor (LY294002).
Conclusion
We demonstrate that atrial myocytes-derived exosomal miR-210-3p promoted the proliferation and collagen synthesis via inhibiting GPD1L in atrial fibroblasts. Preventing pathological crosstalk between atrial myocytes and fibroblasts may be as a novel target to improve atrial fibrosis in AF.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for atrial fibroblasts.

first: Hongting

last: Hao

first: Sen

last: Yan

first: Xinbo

last: Zhao


title: Targeting histone methylation to reprogram the transcriptional state that drives survival of drug-tolerant myeloid leukemia persisters

city: Amsterdam

first: Linda

last: Smit

address: De Boelelaan 1117

department: Hematology

institute: Amsterdam UMC

summary: Although chemotherapy induces complete remission in the majority of acute myeloid leukemia (AML) patients, many face a relapse. This relapse is caused by survival of chemotherapy-resistant leukemia (stem) cells, called measurable residual disease (MRD). Here, we demonstrate that the anthracycline doxorubicin epigenetically reprograms leukemia cells by inducing tri-methylation of histone 3 lysine 27 (H3K27) and H3K4. Moreover, within a doxorubicin-sensitive leukemia cell population, we identified a subpopulation of reversible anthracycline-tolerant cells (ATCs) with leukemic stem cell (LSC) features lacking upregulation of doxorubicin-induced H3K27me3 or H3K4me3. These ATCs have a distinct transcriptional landscape than the leukemia bulk and could be eradicated by inhibition of KDM6. In primary AML, reprogramming the transcriptional state by targeting KDM6 reduced MRD load and survival of LSCs residing within MRD, and enhanced the response to chemotherapy in vivo. Together, our results reveal plasticity of anthracycline resistance in AML cells and highlight the potential of transcriptional reprogramming by epigenetic-based therapeutics to target chemotherapy-resistant AML cells.

overall_design: K562 cells were seeded in 480 wells with 10.000 cells/well and treated with an increasing concentration of doxorubicin (up to 225 ng/ml). After 4 weeks, cells in 21 wells survived chemotherapy treatment. Cells from 4 wells could be maintained in culture in the presence of 225 ng/ul doxorubicin (ATC#1-4). To elucidate the transcriptomic state and regulatory circuits involved in the drug-tolerant condition of the persisters, we performed gene expression profiling analysis of RNA sequencing data for the sensitive K562 parental cells, parental cells shortly treated (48 hours) with 225 ng/ml doxorubicin, and ATC#1-4.

first: Noortje

last: van Gils

first: Han

last: Verhagen


title: Epigenetic regulation of FLT3 by ZNF384-fusion protein

city: Memphis

first: Xujie

state: TN

address: 246 Danny Thomas Place, MS 313

institute: St. Jude Children's Research Hospital

summary: Profiling global gene expression in large ALL cohorts, we identified unique over-expression of FLT3 in ZNF384-rearranged ALL, consistently across cases harboring different fusion partners with ZNF384. Mechanistically, we discovered an intergenic enhancer element at the FLT3 locus that is exclusively activated in ZNF384-rearranged ALL, with the enhancer-promoter looping directly mediated by the fusion protein. There was also a striking global enrichment of active enhancers within ZNF384 binding sites across the genome in ZNF384-rearranged ALL cells.

overall_design: Characterize the local chromatin status of FLT3 gene locus

first: Jun

first: Wenjian


title: Effect  of MCL-1 antagonist in breast cancer associated fibroblasts

status: Public on Aug 14 2022

city: Nantes

first: Frederique

last: Souaze

address: Quai Moncousu

institute: INSERM

summary: Cancer associated fibroblasts (CAF) are a major cellular component of epithelial tumors. In breast cancers in particular these stromal cells have numerous tumorigenic effects in part due to their acquisition of a myofibroblastic phenotype. Breast CAFs (bCAFs) typically express MCL-1. We show here that pharmacological inhibition or knock down of this regulator of mitochondrial integrity in primary bCAFs directly derived from human samples mitigates myofibroblastic features. This decreases expression of genes involved in actomyosin organization and contractility (associated with a cytoplasmic retention of the transcriptional regulator, Yes-Associated Protein - YAP) and decreases bCAFs ability to promote cancer cells invasion in 3D co-culture assays. Our findings underscore the usefulness of targeting MCL-1 in breast cancer ecosystems, not only to favor death of cancer cells but also to counteract the tumorigenic activation of fibroblasts with which they co-evolve.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for breast cancer associated fibroblasts treated or not by MCL1 antagonsit (S63845, 500 nM, 18h).

first: Frédérique

last: Souazé

first: Thomas

last: Bonneaud

first: Agnes

last: Basseville

first: Philippe

last: Juin


title: Gene expression of Tfh cells in the lessions of IgG4-related disease.

status: Public on Aug 10 2022

city: Sapporo

first: Ippei

last: Ikegami

state: Hokkaido

address: S1 W17, Chuo-ku

department: Department of Human Immunology

institute: Research Institute for Frontier Medicine, Sapporo Medical University School of Medicine

summary: To address a possible role of Tfh (CD3+CD4+CXCR5hiPD-1hi) cells in IgG4-related disease(IgG4-RD), we investigated Tfh cells as well as other lymphocyte subsets residing within the affected tissue lesions of IgG4-RD in this study. When compared to gene expression profiles of Tfh cells within tonsils, inflammatory lesions in IgG4-RD preferentially contained Tfh cells with the expression of signature genes of  cytotoxic T lymphocytes. Whereas the expression levels of Tfh-related genes like IL-21, CD200, and Pou2af1 (also named as Bob1) in GC-type Tfh cells of IgG4-RD seemed to be relatively lower than those of GC-type Tfh cells within tonsils. GC-type Tfh cells of IgG4-RD expressed low levels of genes related to Th2 cells, Th17 cells, and Treg cells, albeit the genes related to Th1 cells and interaction of T cells and B cells.

overall_design: T follicular helper (Tfh) cells shape humoral immunity by helping B cell responses at initial and recall phases. Recent studies indicate a possible involvement of Tfh cells in the process of chronic inflammation; however, the functional role of Tfh cells in persistent immune settings still unclear. Here we report a unique capacity of CD4+CD8+ (double positive, DP: CD3+CD4+CD8+PD-1hiCXCR5hi) Tfh cells, which abundantly resided in the fibroinflammatory lesions of IgG4-related disease (IgG4-RD). According to transcriptome analyses, DP-Tfh cells in the lesions of IgG4-RD preferentially expressed the signature genes identifying cytotoxic CD8+ T cells. These findings may illustrate a potential feedback loop for immune tolerance operated by such Tfh cell species in chronic inflammatory conditions.

first: Kosuke

last: Murayama

first: Shingo

last: Ichimiya


title: Fin ray branching is defined by TRAP+ osteolytic tubules in zebrafish

city: Caparica

first: Paulo

middle: Jorge Couto

last: Almeida

address: Monte da Caparica

department: NGS

country: Portugal

institute: STAB VIDA

summary: Study and define how caudal fin rays mineralize and are shaped during regeneration, providing a detailed perspective about its formation and bifurcation and identifying key roles of OLTs.

overall_design: Comparative gene expression analysis of regenerating zebrafish fins between 1 and 3 days post-amputation

first: João

last: Cardeira-da-Silva

first: Anabela

last: Bensimon-Brito

first: Marco

last: Tarasco

first: Ana

middle: S

last: Brandão

first: Joana

last: Rosa

first: Jorge

last: Borbinha

first: P

first: António

last: Jacinto

first: M

last: Cancela

last: Gavaia

first: Didier

middle: Y

last: Stainier

first: Vincent

last: Laizé


title: Study of TUT tailing on miRNA (Endogenous and AGO-IP)

first: Xavier

last: Bofill-De Ros

summary: Many microRNAs (miRNAs) exist alongside abundant miRNA isoforms (isomiRs), most of which arise from post-maturation sequence modifications, such as 3’ uridylation and adenylation. However, the ways in which these sequence modifications affect miRNA function remain poorly understood. To this aim, we have generated knock-out cell lines of TUT4, TUT7 and TENT2 (TUT2), and obtained miRNA profiles from total RNA, AGO1, and AGO2 immunoprecipitations. Here, using these different cell lines, we have discovered that some of the redundant functions and specific functions of each tailing enzyme. Our study provides a comprehensive characterization of tailing on miRNAs.

overall_design: Briefly, HEK293 cells and corresponding knock-outs were transfected with several wild-type and catalytic-dead constructs. Transfections were performed using PolyJet™ DNA Transfection Reagent (SignaGen) according to the manufacturer's instructions. 48 hours after transfection, either total or AGO-bound (IP) RNA was obtained using a chloroform-isopropanol extraction. Total RNA was used to prepare small RNA-seq libraries using QIAseq miRNA Library Kit (Qiagen). Constructs were purified in a 6% (w/v) native acrylamide gel based on the expected product size and purified by ethanol precipitation. Libraries quality was assessed by using Qubit dsDNA HS Assay Kit (ThermoFisher) and Agilent High Sensitivity DNA kit (Agilent). Libraries were pulled together, denaturalized and prepared at a final concentration of 12pM and run on MiSeq or NovaSeq 500 according to the manufacturer specifications.  Fastq files were analyzed using a Python-based custom software, QuagmiR (https://cgc.sbgenomics.com/public/apps#nikola_tesic/quagmir/quagmir-1-0/) designed for the optimal mapping and analysis of isomiRs.

first: Acong

last: Yang

first: Shuo

last: Gu


title: Meso-Seq for in-depth transcriptomics in ultra-low amounts of FACS-purified neuronal nuclei

city: Rehovot

first: Daniella

last: Apelblat

address: Hertzl

department: Neuroscience

country: Israel

institute: Weizmann Institute of Science

summary: Profiling of gene expression in sparse populations of genetically defined neurons is essential for dissecting the molecular mechanisms that control the development and plasticity of neural circuits. However, current transcriptomic approaches are ill suited for detailed mechanistic studies in sparse neuronal populations, as they either are technically complex and relatively expensive (e.g., single-cell RNA sequencing [RNA-seq]) or require large amounts of input material (e.g., traditional bulk RNA-seq).

sample_organism:
Macaca fascicularis
Mus musculus

platform_organism:
Macaca fascicularis
Mus musculus

overall_design: Visual cortex mRNA profiles of adult mice' nuclear RNA, including (1) RNA-Seq of neurons sparsely labeled with AAV-constructs, (2) RNA-Seq from low amounts of antibody-labeled neuronal nuclei in the cortex of mice and of non-human primates, and (3), analyses of experience-regulated gene programs in sparse visual cortex neurons.

first: Ori

last: Roethler

first: Lidor

last: Bitan

first: Hadas

last: Keren-Shaul

first: Ivo

last: Spiegel


title: Spatial transcriptomics of TSC angiomyolipoma

status: Public on Aug 09 2022

city: Boston

first: Yan

last: Tang

address: 20 Shattuck street

institute: Brigham and Women's Hospital

summary: We performed spatial transcriptomics on one IS-dominant tumor and on SLS-dominant tumor. Our goal was to determin different immune infiltration in these two subtypes.

overall_design: NanoString Digital Spatial Profiling (DSP) with Whole Transcriptome Atlas (WTA) was performed on one IS-dominant and one SLS-dominant tumor, with each tumor profiled for 12 ROIs.


title: Gene expression of CD4+CD8+ Tfh cells and CD4+CD8- Tfh in tonsil.

first: Ippei

last: Ikegami

summary: To understand physiological characteristics of CD4+CD8+ Double-positive (DP)-Tfh cells, we continued to conduct comparative transcriptome analysis of tonsillar Tfh cell populations, including DP-Tfh cells and CD4+CD8- single-positive (SP)-Tfh cells  as a control. Results showed that DP-Tfh cells preferentially expressed transcripts related to CTLs, such as CD8 (CD8A and CD8B), Eomes, and granzymes, suggesting a possible cytotoxic attribute of DP-Tfh cells. Th1-cell related signature genes were also presented in DP-Tfh cells and cytokines like interferon (IFN)-gamma and interleukin (IL)-10 were found to be highly presented in DP-Tfh cells rather than SP-Tfh cells. Of note, the expression profile of authentic Tfh-cell (i.e. SP-Tfh cell) related genes like IL-4, IL-21, Bcl6, and Pou2af1 seemed to be shared with DP-Tfh cells.

overall_design: T follicular helper (Tfh) cells shape humoral immunity by helping B cell responses at initial and recall phases.Recent studies indicate a possible involvement of Tfh cells in the process of chronic inflammation; however, the functional role of Tfh cells in persistent immune settings still unclear. Here we report a unique capacity of CD4+CD8+ (double positive, DP: CD3+CD4+CD8+PD-1hiCXCR5hi) Tfh cells, which abundantly resided in the fibroinflammatory lesions of IgG4-related disease (IgG4-RD). According to transcriptome analyses, DP-Tfh cells in the lesions of IgG4-RD preferentially expressed the signature genes identifying cytotoxic CD8+ T cells. These findings may illustrate a potential feedback loop for immune tolerance operated by such Tfh cell species in chronic inflammatory conditions.

first: Kosuke

last: Murayama

first: Shingo

last: Ichimiya


title: Integrated of transcriptome and plant growth substance profiles to reveal the floral sex differentiation in Zanthoxylum armatum DC.

city: Beijing

first: Wenkai

last: Hui

state: Beijing

address: Qinghua East Street

institute: Beingjing Forestry University

summary: Zanthoxylum armatum is a prominent plant for food industries. However, its male flowers often occurred in gynogenesis plants, the potential mechanism remains poorly understood. Herein, a total of 26 floral sex differentiation stages were observed to select four vital periods to reveal key factors using RNA-seq, phytohormones and carbohydrates investigation. Male flower was significantly associated with cytokinin and sucrose signals, whereas female flower was initiated by SA, GA1, and ABA pathways. The floral organ identity genes were attributed to ABCDE model. Eleven ZaAP3/PI and four ZaSTK/SHP were exclusively up-regulated in male and female floral development, respectively. 14 DEGs were obtained to regulate the transition from vegetative to reproductive growth. Finally, 21 crucial factors were identified in co-expression network from a total of 83 important members. This study provided new insights to explore the regulatory mechanism of floral sex differentiation, which would be benefited to cultivate high-yield varieties in Z. armatum.

overall_design:
a total of 26 floral sex differentiation stages were observed to select four vital periods to reveal key factors using RNA-seq, phytohormones and carbohydrates investigation.
Please note that total 26 floral samples were collected to carry out the morphological observation, and then four vital stages of male and female floral differentiation were conduct RNA-seq with three replicates. So there are 24 transcriptomic profiles.


title: Generation of full-length circRNA libraries for Oxford Nanopore long-read sequencing

city: TOULOUSE

first: FABIENNE

last: MEGGETTO

address: 2 AVENUE HUBERT CURIEN

institute: CRCT

summary: Preparation of libraries for circular RNA sequencing with Oxford Nanopore long-read sequencing

overall_design: Long-read sequencing of 4 ALCL cell lines after digestion of ribosomal and linear RNA to enrich for circRNAs.

first: Steffen

last: Fuchs

first: Loelia

last: Babin

first: Christine

last: Gaspin

first: Fabienne

last: Meggetto