title: Brca1 haploinsufficiency promotes early tumor onset and epigenetic alterations in a mouse model of hereditary breast cancer (scRNA-Seq)

city: Boston

first: Joan

middle: S.

last: Brugge

address: 240 Longwood Avenue

department: Department of Cell Biology

institute: Harvard Medical School

summary: Carriers of germline BRCA1 mutations face a high risk of developing breast cancer; however, the mechanisms underlying this tumor predisposition are not completely understood. Here, using a newly developed genetically engineered mouse model for germline Brca1 heterozygosity that allows spatiotemporal control of full Brca1 inactivation, we demonstrate that the early tumor onset in a Brca1-heterozygous background cannot be fully explained by the conventional “two-hit” hypothesis. Specifically, when both Brca1-heterozygous and wild-type conditional mice are induced to Brca1 nullizygosity simultaneously, heterozygous mice develop tumors sooner, suggesting the existence of inherent tumor-promoting alterations in the Brca1-heterozygous state prior to tumor formation. Single-cell RNA-seq and ATAC-seq analyses uncover a unique set of differentially accessible chromatin regions in ostensibly normal Brca1-heterozygous mammary epithelial cells, distinct from wild-type cells and partially mimicking the chromatin and RNA-level changes in tumor cells, thereby pointing to an epigenetic priming mechanism for promoting early tumor onset in the Brca1-heterozygous background. Transcription factor analyses of the altered chromatin regions identify loss of ELF5 and gain of AP-1 binding sites in association with these epigenetically primed chromatin regions, and in vivo experiment reveals that Brca1-heterozygous mice are reliant on the AP-1 pathway for accelerated tumorigenesis. Finally, analysis of downstream target genes identifies Wnt10a as a candidate driver of early tumor onset, and functional experiments demonstrate that it activates the WNT pathway and is sufficient to substantially accelerate tumor formation. Taken together, these findings provide the first in vivo demonstration of a role of BRCA1 haploinsufficiency in accelerating tumor formation and implicate AP-1 and WNT10A as promoters of tumor formation in BRCA1-related breast cancer, advancing our understanding of the disease and informing potential strategies for therapeutic intervention.

overall_design: Single-cell RNA-seq (scRNA-seq) analysis of tumors isolated from WT mice (n=3 tumors) and Brca1-HET mice (n=4 tumors) of the Brca1/p53/Pten (BPP) model, as well as normal mammary glands of uninjected, non-tumor-bearing mice at young age (3-4 months; n=3 WT and 3 Brca1-HET) and old age (13-14 months; n=4 WT and 4 Brca1-HET).

first: Carman

last: Li

first: Laura

last: Selfors

first: Francesca

last: Silvestri


title: Brca1 haploinsufficiency promotes early tumor onset and epigenetic alterations in a mouse model of hereditary breast cancer (ATAC-Seq)

status: Public on Sep 06 2024

state: MA

country: USA

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: ATAC-seq analysis performed on (1) AV cells isolated from normal mammary glands of uninjected, non-tumor-bearing WT mice (n=4 samples, 3-4 months of age) and Brca1-HET mice (n=7 samples, 3-4 months of age), and (2) tumor cells isolated from WT mice (n=3 samples) and Brca1-HET mice (n=3 samples) of the BPP model.

middle: M

middle: S


title: Possible involvement of a MEG3-miR-21-SPRY1-NF-kB feedback loop in spermatogenic cells proliferation, autophagy and apoptosis

status: Public on Sep 14 2024

city: Zhengzhou

first: Xingyu

last: Fang

address: 1st Jianshe West street

country: China

institute: The First Affiliated Hospital of Zhengzhou University

type: Expression profiling by high throughput sequencing

type: Non-coding RNA profiling by high throughput sequencing

summary: Non-obstructive azoospermia (NOA) is the most incurable male infertility with complex etiology. The molecular mechanism that determines the amount of spermatogenesis in NOA patients is still unclear. LncRNAs have been verified to associate with the regulation of spermatogenesis. However, the specific function and molecular mechanism remain to be confirmed. We use RNA-seq analysis for preliminary screening of different lncRNAs between NOA and normal male and find that MEG3 was upregulated in NOA testicular tissues and inhibit cell proliferation and promote cell autophagy and apoptosis in vitro and vivo. Through RNA immunoprecipitation (RIP), biotin pull-down assays and Dual Luciferase Reporter assays, MEG3 is proved to act as competing endogenous RNA of miR-21 and thus influence the SPRY1/ERK/mTOR signal pathway. In addition, SPRY1 positively regulates MEG3 through SPRY1/NF-κB/MEG3 feedback loop. These findings firstly demonstrate that LncRNA MEG3-miR-21-SPRY1-NF-κB acts as a new feedback loop leading to the occurrence of azoospermia through inhibiting the proliferation, promoting autophagy and apoptosis of spermatogenic cells, which provides a new target and theoretical basis for diagnosis and treatment of non-obstructive azoospermia.

overall_design: To explore different expression genes and its function in NOA and normal spermatogenesis patients.

first: Wenyan

last: Song

first: Xiaotong

last: Lu

first: Yujie

last: Ma


title: Dynamics of RNA localization to nuclear speckles are connected to splicing efficiency

status: Public on Sep 05 2024

city: Chicago

first: Jingyi

last: Fei

state: IL

address: GCIS W142, 929 E 57th Street

institute: The University of Chicago

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

overall_design: Refer to individual Series


title: Mesenchymal Stem Cells Generated from Human Urine-derived iPSCs Exhibit Low Immunogenicity and Im-munomodulatory Profile

status: Public on Sep 07 2024

city: Changsha, Hunan, China

first: Peiyun

last: Wang

address: 110 Xiangya Road

institute: Central South University

summary:
We conducted transcriptome analysis to determine the differences between umbilical cord-derived mesenchymal stem cells (named UCMSC) and iMSCs generated from human urine-derived iPSC (named B6MSC) at the transcriptomic level.
The expression of genes related to aging, rejuvenation,  proliferation or immune regulatory factors in iMSCs was different significantly from that of UCMSCs.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for UCMSCs and iMSCs generated from human urine-drived iPSC.

first: Ying

last: Zhang


title: Effect of embryo lysates treatment on gene expression.

city: Beijing

last: Liu

address: 5 Summer Palace Road, Haidian District, Beijing

department: College of Future Technology

institute: Peking University

summary: To systematically evaluate the effect of embryo lysates feeding, we conducted RNA sequencing (RNA-seq) on day 7 N2 worms treated with embryo lysates. Our findings revealed that embryo lysates affected the expression of genes related to fatty acid metabolism, degradation of branched-chain amino acids, and lysosome functions. Notably, worms treated with embryo lysates showed increased fatty acid degradation gene expression and decreased fatty acid elongation gene expression.

overall_design: Day 7 N2 worms were treated with embryo lysates for 24h. Then worms were collected for mRNA seq and gene expression profiliing analysis.

first: Liankui

last: Zhou

first: Wanqiu

last: Ding


title: Developmental emergence of first- and higher-order thalamic neuron molecular identities

status: Public on Sep 12 2024

city: Genève 4

first: Quentin

last: Lo Giudice

state: switzerland

address: 1 rue Michel Servet

department: Department of Basic Neurosciences

country: Switzerland

institute: University of Geneva

summary: The thalamus is organized into nuclei that have distinct input and output connectivities with the cortex. While first-order (FO) nuclei – also called core nuclei – relay input from sensory organs on the body surface and project to primary sensory areas, higher-order (HO) nuclei – matrix nuclei – instead receive their driver input from the cortex and project to secondary and associative areas within cortico-thalamo-cortical loops. Input-dependent processes have been shown to play a critical role in the emergence of FO thalamic neuron identity from a ground state HO neuron identity, yet how this identity emerges during development remains unknown. Here, using single-nucleus RNA sequencing of the developing embryonic thalamus we show that FO thalamic identity emerges after HO identity, and that peripheral input is critical for the maturation of excitatory, but not inhibitory FO-type neurons. Our findings reveal that subsets of HO neurons are developmentally co-opted into FO-type neurons, providing a mechanistic framework for the diversification of thalamic neuron types during development and evolution.

overall_design: Diencephalon of the embryos were collected in ice-cold Hank's Balanced Salt Solution (HBSS) and segments were microdissected under a stereomicroscope at different time points. Thalamic chunks were frozen and stored at -80°C until further processing for the nucseq sqmples, or immediatly processed for the sing-cell ones.

first: Robin

last: Wagener

first: Philipp

last: Abe

last: Frangeul

first: Denis

last: Jabaudon


title: Dynamics of RNA localization to nuclear speckles are connected to splicing efficiency [RNA-Seq]

summary: Nuclear speckles, a type of membraneless nuclear organelle in higher eukaryotic cells, play a vital role in gene expression regulation. Using the reverse transcription-based RBP binding sites sequencing (ARTR-seq) method, we study human transcripts associated with nuclear speckles. We identify three gene groups whose transcripts demonstrate different speckle localization properties and dynamics – stably enriched in nuclear speckle post-transcriptionally, transiently enriched in speckles at the pre-mRNA stage co-transcriptionally, and not enriched in speckles. We show that nuclear speckles specifically facilitate splicing of speckle-enriched transcripts. We further reveal RNA sequence features contributing to transcript speckle localization, underscoring a tight interplay between genome organization, RNA cis-elements, and transcript speckle enrichment, and connecting transcript speckle localization with splicing logic. Finally, we show that speckles can act as hubs for the regulated retention of introns during cellular stress. Collectively, our data highlight a role of nuclear speckles in both co- and post-transcriptional splicing regulation.

overall_design: To accurately assess RNA abundance, we conducted RNA-Seq for poly A RNA and nuclear RNA in HeLa or HepG2 cells subjected to heat shock or Pladienolide B treatment. Additionally, we utilized RNA-Seq of nuclear RNA specifically for the computation of Method 2.

first: Jinjun

last: Wu

first: Yu

last: Xiao

first: Yunzheng

first: Li

last: Wen

first: Chuanyang

last: Jin

first: Shun

first: Sneha

last: Paul

first: Chuan

last: He

first: Oded

last: Regev


title: Dynamics of RNA localization to nuclear speckles are connected to splicing efficiency [ARTR-seq]

first: Jingyi

last: Fei

overall_design: We employed ARTR-seq, utilizing pAG-RTase, a fusion of Protein A/G and reverse transcriptase, to map the nuclear speckle-enriched transcriptome. This method combines Protein A/G's IgG binding domains to target IgG subclasses, facilitating binding to antibodies against nuclear speckle scaffold proteins (SON or SRRM2). Reverse transcription was initiated with exogenous biotin dNTPs, followed by enrichment of biotinylated cDNAs using streptavidin beads and library preparation for sequencing.


title: Brca1 haploinsufficiency promotes early tumor onset and epigenetic alterations in a mouse model of hereditary breast cancer

first: Joan

last: Brugge

first: Carman

last: Li

first: Laura

last: Selfors

first: Francesca

last: Silvestri


title: Elevated choline drives KLF5-dominated transcriptional reprogramming to facilitate liver cancer progression

status: Public on Sep 04 2024

city: Shanghai

first: Xianghuo

address: No 270, Dong An Road

institute: Fudan University

summary: We provide evidence highlighting  a novel mechanism by which choline remodels CRC and upregulate the SE-driven key transcription factor KLF5 via the methyl donor pathway, implicating potential therapeutic targets in HCC.

overall_design: Whole-transcriptome sequencing (RNA-Seq) was perfemed to identify the downstream target genes regulated by choline in Huh7 cells. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) was performed using indicated antibodies to study the H3K27ac and H3K4me1 status and binding sites of transcription factor in Huh 7 cells under 0μM and 15μM choline treatment. Cleavage Under Targets and Tagmentation (CUT&Tag) was performed to explore whether  the super-enhancers or promoters within downstream target genes are directly regulated by KLF5.

first: Xinrong

first: Shenglin

last: Huang

first: Zhiao

last: Chen


title: Identification of differentially expressed genes in tumorigenesis of cervical cancer via next generation sequencing

city: Tianjin

first: Jia

last: Xu

address: No.94 Weijin Road

department: School of Medicine

institute: Nankai University

summary: The incidence and mortality of cervical cancer remain high in female malignant tumors worldwide. There is still a lack of diagnostic and prognostic markers for cervical carcinoma. This study aimed to screen differentially expressed genes (DEGs) between normal and cervical cancer tissues to identify candidate genes for further research.

overall_design: To screen differentiated expressed genes (DEGs) between cervical cancer tissues and its paired adjacent normal cervical tissues through RNA sequencing.

first: Xiangshu

first: Wen

last: Yang

first: Yuanguang

last: Meng


title: Decoding Transcriptional Identity in Developing Human Sensory Neurons and Organoid Modeling

first: xiaoqun

last: wang

state: Beijing

address: 15 Datun Road, Chaoyang District

department: Institute of Biophysics

institute: Chinese Academy of Sciences

summary: Dorsal root ganglia (DRG) play a crucial role in processing sensory information, making it essential to understand their development. Here, we construct a single-cell spatiotemporal transcriptomic atlas of human embryonic DRG. This atlas reveals the diversity of cell types and highlights the extrinsic signaling cascades and intrinsic regulatory hierarchies that guide cell fate decisions, including neuronal/glial lineage restriction, sensory neuron differentiation and specification, and the formation of neuron-satellite glial cell (SGC) unit. Additionally, we identify a human-enriched NTRK3+/DCC+ nociceptor subtype, which is involved in multimodal nociceptive processing. Mimicking the programmed activation of signaling pathways in vivo, we successfully establish functional human DRG organoids and underscore the critical roles of transcriptional regulators in the fate commitment of unspecialized sensory neurons (uSNs). Overall, our research elucidates the multilevel signaling pathways and transcription factor (TF) regulatory hierarchies that underpin the diversity of somatosensory neurons, emphasizing the phenotypic distinctions in human nociceptor subtypes.

overall_design: we sampled and performed snRNA-seq on DRG samples from gestational weeks (GW) 7-21 with biological replicates

first: Xiaoqun

last: Wang

first: Lu

last: Tian

first: Wei

last: Zhou

first: Mengdi


title: Septo-dentate gyrus cholinergic circuits modulate function and morphogenesis of adult neural stem cells through granule cells in mice

first: Jeremy

last: Simon

address: 450 Brookline Ave

department: Department of Data Science

institute: Dana-Farber Cancer Institute

summary: Dentate gyrus (DG) is a critical structure for spatial memory and adult neurogenesis, and both processes are dynamically regulated by neural circuit activity. It remains unknown how neural circuits orchestrate the molecular landscape of the highly heterogeneous DG to regulate these distinct functions. Here we first report that activation of a cholinergic circuit from Diagonal Band of Broca to DG promotes both neural stem cell (NSC) activation and spatial memory. Furthermore, using single-nucleus RNA-sequencing of DG, we report broad transcriptomic changes across multiple types of mature and adult-born cells in response to cholinergic circuit activation. Notably, various neuronal populations exhibit abundant molecular changes that support synaptic functions crucial for spatial memory; while NSCs exhibit molecular changes that support their structural development and neurogenic proliferation. Interestingly, our electrophysiology and NicheNet analyses reveal granule cells, endothelial cells, and astrocytes as potential intermediaries for cholinergic activity-dependent NSC regulation. Our findings provide cell-type specific molecular mechanisms underlying cholinergic circuit activity-dependent regulation of distinct DG functions.

overall_design: Single-nucleus RNA-seq (SPLiT-seq) of mouse dentate gyrus from stimulated (Chr2) and sham (yfp) stimulated animals. FASTQs represent two sublibraries comprised of cells from all 10 biological samples

first: Luis

last: Quintanilla

first: Juan

last: Song


title: Epigenomic and Functional Convergence Between Glucocorticoid- and IL4-Driven Macrophage Programming

status: Public on Sep 03 2024

city: New york

first: Yurii

last: Chinenov

state: New York

address: 535 E 71 str, Hospital for Special Surgery

department: Research Division

institute: Hospital for special surgery

summary: Macrophages adopt distinct phenotypes in response to environmental cues, with type-2 cytokine interleukin-4 (IL4) promoting a tissue-repair homeostatic state (M2IL4). Glucocorticoids, widely used anti-inflammatory therapeutics, reportedly impart a similar phenotype (M2GC), but how or to what extent these populations functionally converge is unknown. We show that M2IL4 and M2GC transcriptomes share a striking overlap mirrored by a shift in chromatin landscape in both common and signal-specific gene subsets. This core homeostatic program is enacted by transcriptional effectors KLF4 and the GC receptor, whose genome-wide occupancy and actions are integrated in a stimulus-specific manner by the nuclear receptor cofactor GRIP1. Indeed, many of the M2IL4:M2GC-shared transcriptomic changes were GRIP1-dependent. Consistently, GRIP1 loss attenuated phagocytic activity of both populations in vitro and macrophage tissue-repair properties in the murine colitis model in vivo. These findings provide a mechanistic framework for homeostatic macrophage programming by distinct signals, to better inform anti-inflammatory drug design.

overall_design: Mouse primary bone marrow-derived macrophages from C57BL/6  mice  were exposed to IL4 or Dexamethasone  for 24 h

first: Dinesh

middle: K

last: Deochand

first: Marija

last: Dacic

first: Michael

middle: J

last: Bale

first: Andrew

middle: W

last: Daman

first: Steven

middle: Z

last: Josefowicz

first: Inez

last: Rogatsky


summary: Macrophage polarization is implicated in acute and chronic inflammatory settings, and central to diseases such as obesity, atherosclerosis and cancer. However, the transcription programs driving macrophage polarization are only partially known. Macrophage phenotypes exist on a spectrum, with anti-inflammatory (M2-like) and pro-inflammatory (M1-like) states viewed as polar opposite and relatively stable phenotype. The main driver of the M2-like program in response to IL4 is the Krüppel-like factor 4(KLF4), which uses transcriptional coregulator GRIP1 as a co-activator. Coincidentally, glucocorticoid hormone (GC)-activated GC receptor (GR) also induces the M2-like state, and GRIP1 is a well-known co-factor of GR. We are investigating if GRIP1 facilitates transcriptional and phenotypic convergence of the M2-like state via the GR and KLF4-regulated pathways. Using RNA-seq on bone marrow-derived macrophages (BMDMs) from WT and GRIP1-cKO mice, we have shown that GR and KLF4-regulated pathways share common and distinct transcriptomes. A lot of the key M2 genes are GRIP1-dependent, and some key pro-inflammatory targets need GRIP1 for suppression in the M2–like state. Similarly, H3K27ac ChIP and ATAC-seq, which profile active and open chromatin regions respectively, revealed both specific and shared enhancers for these two pathways in the M2-like state. GR recruitment to the genomic binding sites was shown to be strictly GC-specific, whereas GRIP1 is recruited in both signal-specific and combinatorial manner. It is plausible that GRIP1 facilitates the binding of GR and KLF4 to the shared enhancers, thus leading to overlapping changes in gene expression. At the cellular level, GC- and particularly IL4-polarized GRIP1-cKO BMDMs exhibit reduced phagocytosis, an M2-characteristic activity, compared to the WT. Furthermore, in vivo, in the acute inflammatory setting of DSS-induced colitis, GRIP1-cKO mice exhibit greater colonic inflammation and tissue destruction, and fail to up-regulate M2 genes to the same degree as the WT. GRIP1 is therefore a crucial regulator of macrophage polarization to the anti-inflammatory state. The specific contribution of GRIP1 to the shared transcriptional phenotype is currently being assessed.

overall_design: Mouse primary bone marrow-derived macrophages from BL6(wild type) and LysM-Cre+/+; GRIP1fl/fl  mice  were exposed to IL4 or glucocorticoids corticosterone  and dexamethasone  for 24 h


title: Epigenomic and Functional Convergence Between Glucocorticoid- and IL4-Driven Macrophage Programming (ChIP-Seq)

overall_design: Mouse primary bone marrow-derived macrophages from C57BL/6  mice  were exposed to IL4 or glucocorticoids corticosterone  and dexamethasone  for 24 h


title: Effect of TGF-B1 on primary human lung fibroblasts growing in media with physiological glucose and glutamine

status: Public on Sep 02 2024

city: London

first: Greg

last: Contento

state: Greater London

address: Rayne Institute, 5 University Street

department: UCL Respiratory

country: United Kingdom

institute: University College London

summary: The interplay between metabolism and gene expression is primarily known through epigenetic modifications which use metabolites such as acetyl-CoA. Glucose and glutamine are critical for the synthesis of these metabolites but the concentrations of these two major nutrients are not standardized in conventional tissue culture media. The aim of this study was to investigate the transcriptomic effect of TGF-B1 after 24 hours in primary human lung fibroblasts (pHLFs) growing in DMEM with glucose and glutamine set at physiological concentrations (5 and 0.7 mM, respectively). We were then able to compare these gene  expression patterns to a previously published dataset of pHLFs growing in DMEM with higher concentrations of glucose and glutamine (25 and 2 mM, respectively) (GSE102674). This revealed a dramatic difference of roughly 3-fold differentially expressed genes dependent on DMEM formulation as well as just half of the top 20 enriched pathways induced by TGF-B1 shared, as analysed with GSEA. The conclusion was that the concentrations of glucose and glutamine have profound effects on gene expression changes.

overall_design: Primary human lung fibroblasts were treated with TGFβ1 for 24h in DMEM with 5mM glucose and 0.7mM glutamine. Each treatment was done in triplicate, for a total of 6 samples. Untreated cells were used as control. Paired-end RNA sequencing was performed with the NextSeq sequencing platform (Illumina, San Diego, CA, USA). Each sample was run on 4 different lanes.

first: Delphine

last: Guillotin

first: Rachel

last: Chambers

first: Jo-Anne

last: Wilson

first: Manuela

last: Plate


title: Immune Response of Transplanted Kidney Tissues Assembled from Organoid Building Blocks

first: Leonardo

middle: V.

last: Riella

address: 149 13th Street

department: Medicine

institute: Massachusetts General Hospital

summary: The transcriptional profile of kidney organoid-based tissues resembled the gene signature observed in human kidney transplant rejection when transplanted into mice reconstituted with an allogeneic immune system.

overall_design: Gene expression profiles of kidney tissues assembled from organoid building blocks harvested from NSG mice with no immune system, or reconstituted with allogeneic human immune cells treated or not with Rapamycin

first: Riella

last: Leonardo


title: Small RNA Profiling of Lesional Psoriatic and Adjacent Normal Skin Tissues

city: Kolkata

first: Raghunath

last: Chatterjee

state: WB

address: 203 BT Road, HGU, Indian Statistical Institute

department: Human Genetics Unit

country: India

institute: Indian Statistical institute

summary:
Purpose: To identify the differentially expressed miRNA in lesional skin of psoriasis patients compared to adjacent normal tissue by Next  Generation Sequencing.
Methods: Small RNA isolated from 24 paired lesional and adjacent normal tissue sample and Next Generation Sequencing was performed. The sequences reads were trimmed of adapter sequences and quality trimmed using CutAdapt, followed by alignment to the human genome via 'Novoalign'. The aligned files were sorted based on chromosomal location and HTSeq-count was used to obtain the raw read counts for all known miRNAs. Differnetial expression analysis was performed with the raw reads using R-package 'edgeR'.  MiRNA expression was quantified in tissue and serum by qRT-PCR method using SYBR-Green assay. Correlation analysis was done between miRNA expression in individual samples with their respective PASI scores to identify the miRNA associated with disease severity.
Results:  We identified 75 differentially expressed miRNA with at least 2-fold deregulation that were statistically significant. qRT-PCR validation showed approx 90% concordance with NGS data. Fold change of 4 miRNAs - miR-30b-5p, miR-7-1-3p, miR-33a-3p and miR-4742-3p were found to be asssociated with disease severity in psoriasis skin. Another miRNA let-7d-5p expression was found to be associated with disease severity in serum of HLA-Cw6 positive patients.
Conclusions: Our study identified several unqiue deregulated miRNAs in psoriatric skin, which had not been previously reported. We also identified miRNA whose deregulated expression in psoriasis is associated with disease severity and the biological pathways through which they regulate disease pathogenesis.

overall_design: Small RNA profiling in 24 paired lesional psoriatic skin and adjacent normal skin

first: Torsa

last: Ganguly

first: Sayantan

last: Laha

first: Aditi

last: Chandra

first: Swapan

last: Senapati

first: Gobinda


title: Effect of beta cell ablation on pancreatic ductal cells in adult zebrafish

status: Public on Sep 01 2024

city: Liège

first: Isabelle

last: Manfroid

address: Avenue de l'Hopital 1

department: GIGA-Stem Cell

country: Belgium

institute: Université de Liège

summary: To determine the change at the transcriptomic level of ductal cells during beta cell regeneration, we performed RNA-seq on pancreatic ductal cells 3 days after beta cell destruction and compared with non ablated control

overall_design: Comparative gene expression profiling analysis of RNA-seq data with or without beta cell ablation

first: David

last: Bergemann

first: Arnaud

last: Lavergne

first: Laura

last: Massoz


title: Bulk RNAseq of FNE1 cells with ectopic expression of YAP1 and HPV E6/E7 oncogenes

city: boston

first: Jiyuan

last: Liu

state: Massachusetts

address: 55 fruit st.

institute: Mass Gen Hospital

summary: High grade serous ovarian carcinoma (HGSOC) is the most common and aggressive ovarian malignancy. Accumulating evidence indicates that HGSOC may originate from human fallopian tube epithelial cells (FTECs), although the exact pathogen(s) and/or molecular mechanism underlying the malignant transformation of FTECs is unclear. Here we show that human papillomavirus (HPV), which could reach FTECs via retrograde menstruation or sperm-carrying, interacts with the yes-associated protein 1 (YAP1) to drive the initiation and progression of high grade serous cancer (HGSC). HPV prevents FTECs from natural replicative and YAP1-induced senescence, thereby promoting YAP1-induced malignant Transformation of FTECs. HPV also stimulates proliferation and drives metastasis of YAP1-transformed FTECs. YAP1, in turn, stimulates the expression of the putative HPV receptors and suppresses the innate immune system to facilitate HPV acquisition. These findings provide critical clues for developing new strategies to prevent and treat HGSOC.

overall_design: FNE1 cells were cultured to 40% confluent and then transfected with retrovirus-based empty control vector (MXIV), or vectors expressing wild type of YAP1 (YAP), or constitutively active YAP1 (YAPS127A, a replacement of Serine at residue 127 with Alanine resulting in the constitutive activation of YAP1 protein [66]). All transfected cells were selected with G418 (200-400μg/ml). YAP1 expression in these cells was confirmed by RT-PCR and Western blot.

first: Chunbo

last: He

first: Xingmin

last: Lv

first: Cheng

last: Wang


title: Relaxin Modulates the Genomic Actions and Biological Effects of Estrogen in the Myometrium [hTERT RNA-seq]

status: Public on Aug 31 2024

city: Dallas

first: W. Lee

last: Kraus

state: TX

address: 5323 Harry Hines Blvd.

institute: UT Southwestern Medical Center

summary: Estradiol (E2) and relaxin (Rln) are steroid and polypeptide hormones, respectively, with important roles in the female reproductive tract, including myometrium.  Some actions of Rln, which are mediated by its membrane receptor RXFP1, require or are augmented by E2 signaling through its cognate nuclear steroid receptor, estrogen receptor alpha (ER⍺).  In contrast, other actions of Rln act in opposition to the effects of E2.  Here we explore the molecular and genomic mechanisms that underlie the functional interplay between E2 and Rln in the myometrium.  We used both ovariectomized female mice and immortalized human myometrial cells expressing wild type or mutant ER⍺ (hTERT-HM-ER⍺ cells).  Our results indicate that Rln attenuates the genomic actions and biological effects of estrogen in the myometrium and myometrial cells by reducing phosphorylation ER⍺ on serine 118 (S118).  Interestingly, we observed a potent inhibitory effect of Rln on the E2-dependent binding of ER⍺ across the genome.  The reduction in ER⍺ binding was associated with changes in the hormone-regulated transcriptome, including a decrease in the E2-dependent expression of neighboring genes.  The inhibitory effects of Rln cotreatment on the E2-dependent phosphorylation of ER⍺ required the nuclear dual-specificity phosphatases DUSP1 and DUSP5.  Moreover, the inhibitory effects of Rln were reflected in a concomitant inhibition of the E2-dependent contraction of myometrial cells.  Collectively, our results identify a pathway that integrates Rln/RXFP1 and E2/ER⍺ signaling, resulting in a convergence of membrane and nuclear signaling pathways to control genomic and biological outcomes.

overall_design: RNA-seq of hTERT cells with Dox-inducible expression of ER⍺(WT,S118E) were generated.RNA-seq experiment was performed in two biological replicates of wild-type and S118E mutant hTERT-HM cells treated with Veh, E2, Rln and E+R at 1h. 0h treatment is taken as the control samples.

first: Sudeshna

last: Tripathy

first: Anusha

last: Nagari

first: Shu-Ping

last: Chiu

first: Tulip

last: Nandu

first: Cristel

last: Camacho

first: Mala

last: Mahendroo

first: W

middle: L


title: Single-cell transcriptomic atlas of enteroendocrine cells along the murine gastrointestinal tract.

city: Cambridge

first: Fiona

last: Gribble

state: Cambridgeshire

address: Box 289 Addenbrooke's Hospital, Hills Road

department: Medical Research Laboratories

institute: University of Cambridge

summary: Enteroendocrine cells (EECs) produce over 20 gut hormones which contribute to intestinal physiology, nutrient metabolism and the regulation of food intake. The objective of this study was to generate a comprehensive transcriptomic map of mouse EECs from the stomach to the rectum.

overall_design:
EECs were purified by flow-cytometry from the stomach, upper small intestine, lower small intestine, caecum and large intestine of NeuroD1-Cre mice, and analysed by single cell RNA sequencing.
Neurod1-Cre, large intestine, scRNA-seq; re-analysis of GSE137572

first: Christopher

middle: A

last: Smith


title: Fucosylated glycoproteins and fucosylated glycolipids play opposing roles in cholera intoxication

first: Jennifer

last: Kohler

address: 5323 Harry Hines Blvd

institute: UT Southwestern

summary: Cholera toxin (CT) is the etiological agent of cholera. Here we report that multiple classes of fucosylated glycoconjugates function in CT binding and intoxication of intestinal epithelial cells. In Colo205 cells, knockout of B3GNT5, the enzyme required for synthesis of lacto- and neolacto-series glycosphingolipids (GSLs), reduces CT binding but sensitizes cells to intoxication. Overexpressing B3GNT5 to generate more fucosylated GSLs confers protection against intoxication, indicating that fucosylated GSLs act as decoy receptors for CT. Knockout (KO) of B3GALT5 causes increased production of fucosylated O-linked and N-linked glycoproteins, and leads to increased CT binding and intoxication. Knockout of B3GNT5 in B3GALT5 KO cells eliminates production of fucosylated GSLs but increases intoxication, identifying fucosylated glycoproteins as functional receptors for CT. These findings provide insight into molecular determinants regulating CT sensitivity of host cells.

overall_design: Colo205 cells infected with lentivirus encoding the Brunello genome-wide CRISPR KO pooled library (Addgene catalog no. 73178) were incubated with biotinylated cholera toxin B subunit and streptavidin-DTAF.  Cells with the highest 1% fluorescence and lowest 1% fluorescence were collected by fluorescence-activated cell sorting. sgRNA enrichment was determined relative to the input cell population.

first: Atossa

middle: C

last: Ghorashi

first: Andrew

last: Boucher

first: Stephanie

last: Archer-Hartmann

first: Nathan

middle: B

last: Murray

first: Rohit

middle: R

last: Konada

first: Xunzhi

last: Zhang

first: Chao

last: Xing

first: Parastoo

last: Azadi

first: Ulf

last: Yrlif


title: DNA methylation profiling of salivary gland tumors

city: München

first: Philipp

last: Jurmeister

address: Thalkirchner Straße 36

department: Institute of Pathology

country: Germany

institute: Ludwig Maximilians University Hospital Munich

type: Methylation profiling by genome tiling array

summary: Tumors of the major and minor salivary gland encompass a diverse spectrum of diagnostically challenging neoplasms. Recent studies have identified several gene fusions and somatic mutations that are specific or highly enriched in certain salivary gland tumor entities and can assist histopathological diagnosis. Still, there is an unmet need to identify additional diagnostic biomarkers for entities lacking specific alterations. In this study, we collected a comprehensive cohort of 363 cases encompassing 20 different salivary gland tumor entities and explored the potential of DNA methylation to classify these tumors. We were able to show that most entities show specific epigenetic signatures and present a machine learning algorithm that can be used to classify diagnostically challenging cases.

overall_design: 363 specimens were analyzed using the MethylationEPIC BeadChip


title: GLABRA2 Transcription Factor integrates Arsenic tolerance with cell differentiation in Epidermal Cells

city: Madrid

first: Jose M.

last: Franco-Zorrilla

state: Madrid

address: Darwin, 3

department: Plant Molecular Genetics

country: Spain

institute: Centro Nacinal de Biotecnologia-CSIC

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

overall_design: Refer to individual Series


title: FGF7 and FGF10 Promote Fate Transition of Human Epidermal Cell-derived Organoids to an Eccrine Gland Phenotype

city: Hangzhou

first: Liang

last: Gong

address: WenYi West Rd 1369

institute: Liangzhu Laboratory

summary: Epidermal appendages play a key role in thermoregulation, which is crucial in maintaining the equilibrium of body temperature. The normality of eccrine sweat glands (SwGs) and hair follicles (HFs) assures homeostatic thermoregulation. However, the fate commitments of SwGs and HFs development and their key regulators are still poorly understood. To elucidate the regulatory networks and the major regulators in SwGs and HFs development, we generated a transcriptomic atlas within three tissues (dorsal back skin, front and hinder ventral foot skin) across four early development stages (embryonic (E) days E15.5 and E17.5, postnatal (P) days P1 and P3) of Mus musculus. We then applied a second-order polynomial model to fit time-course expression of genes and used likelihood ratio test (LRT) to identify temporal dynamic expressed genes. We further constructed temporal regulatory networks to investigate the master regulators during different developmental stages. We found the distinct temporally developmental patterns between dorsal back and ventral foot skin. Eight temporally dynamic expression modes were inferred, which are comprised of over 90% of the dynamic genes. Shh, Eda, Bmp2 and other previously identified factors were noted in our dominant dynamic patterns as well and temporally dynamic expressed in dorsal back and ventral foot. Combined with differentially expressed genes analysis, we highlighted 501 genes that were common in front and hinder foot, albeit uniquely enrich or depleted in dorsal back. Moreover, the integration of temporal expression models and genes regulatory networks accentuated master transcription factors functioning at specific stages in different tissues. Our findings demonstrated the temporally dynamic determinants during SwGs and HFs fate commitments and can contribute to restoring impaired SwGs and HFs clinically after hypohidrosis or burn injury.

overall_design: To understand the early fate determination of hair follicles (HFs) and eccrine sweat glands (ESGs), we performed RNA-sequencing on mice dorsal back tissues and  ventral front/hinder foot tissue at two embryonic (E) days 15.5 and 17.5 and two postnatal (P) days P1 and P3.

first: Zhen

last: You

first: Haihong

last: Li

first: Yonghong


title: TRβ-agonist Sobetirome(GC-1) Attenuates Pulmonary Fibrosis via Promoting MAS-AT2s Differentiation into AT1s.

status: Public on Aug 30 2024

city: Xinxiang

first: Guoying

last: Yu

state: Henan

address: East of Construction Road

institute: Henan Normal University

summary: Idiopathic pulmonary fibrosis (IPF) is a progressive and lethal lung disease, characterized by hyperplasia and blocked differentiation of alveolar type-2 cells (AT2s). Here, we found that sobetirome (GC-1), a thyroid hormone receptor β (TRβ) specific agonist, exhibits striking efficiency and safety to treat PF in two mouse models (bleomycin and silica) and one rat model. We proposed that most IPF is a type of non-thyroidal illness syndrome (NTIS) based on hormone level tests. Using lineage tracing mice and co-culture, we confirmed that the anti-fibrosis effect of GC-1 is primarily through modulating the maladaptive activation state AT2s (MAS-AT2s) cell fate, promoting differentiation and inhibiting proliferation. Luciferase and ChiP assays reveal that TRβ with GC-1 directly regulate KLF2 and CEBPA, and further drive the AT1 genes expression. AT2s-KLF2-knockout or CEBPA AAV-knockdown mice nullified the GC-1 effect of anti-fibrosis and showed an abnormal regeneration in pneumonectomy, indicating that KLF2 and CEBPA are the anti-fibrotic targets of GC-1.

overall_design: 3 conditions were evaluated in mouse: Mouse + Bleomycin, sacrificed 21 days after exposure, mouse + Bleomycin + GC1, sacrificed 21 days after exposure, GC1 was given from day 11 to day 21 for mice, and control mouse + PBS, sacrificed 21 days after exposure. Each treatment had three replicates for a total of 9 RNA samples. Each replicate was composed of three mice pooled together.

first: Lan

last: Wang

first: Xin

last: Pan

first: Zhongzheng


title: Deletion of Mgat2 in Spermatogonia Blocks Spermatogenesis

city: Bronx

first: Deyou

last: Zheng

state: NY

address: 1301 Morris Park Avenue

department: Department of Genetics

institute: Albert Einstein College of Medicine

summary: Identifying factors required for spermatogenesis is important for understanding mechanisms of male fertility. Inactivation of the Mgat1 or Man2a2 gene leads to a block in spermatogenesis causing infertility in male mice. In both cases multi-nucleated germ cells (MNC) are formed and there are no sperm in the epididymis. MGAT1 GlcNAc-transferase initiates complex N-glycan synthesis and MAN2A2 mannosidase generates the substrate for MGAT2 GlcNAc-transferase which transfers GlcNAc to form a biantennary complex N-glycan. Deletion of Mgat2 can leave its substrate, a hybrid N-glycan terminating in a single GlcNAc, to accumulate or the single GlcNAc may be extended to with Gal and GlcNAc to form polylactosamine. In this paper we show that conditional deletion of Mgat2 in spermatogonia via Stra8-iCre caused a block in spermatogenesis prior to the formation of round spermatids. This phenotype differs from deletion of Mgat1 by Stra8-iCre which generates MNC of round and elongated spermatids. In addition, RNA-seq analysis of germ cells prepared at 15 days after birth revealed a unique transcriptomic landscape in Mgat2[-/-] germ cells. Bioinformatic analyses predicted potential roles for ERK and AKT activities which were validated by western analyses. By contrast, Mgat1[-/-] germ cells have reduced ERK activity and unchanged AKT activity. Therefore, the defective spermatogenesis observed following removal of MGAT1 or MGAT2 must arise from the different immature N-glycans that accumulate rather than from the loss of complex N-glycans which is common to both.

overall_design: To determine gene expression changes in male germ cells from 15-day mice in which Mgat2 was conditionally deleted using Str8-iCre compared to control germ cells. Four control Mgat2[F/F] and 4 mutant Mgat2[F/F]:Stra8-iCre male pups were sacrificed, germ cells were prepared and frozen at -80ºC. Total RNA was isolated using a pellet pestle and RNAzol (Invitrogen) according to the manufacturer's instructions. Total RNA was frozen at -80ºC and sent on dry ice to Novogene for the preparation of poly A+ RNA and cDNA prior to performing RNA-seq. Non-stranded mRNA-Seq libraries (poly A enriched) were prepared by Novogene for 150 base pair paired-end (PE) sequencing on an Illumina HiSeq platform (NovaSeq Illumina PE150 with 6 G raw data per sample).

first: Mohd Shamoon

last: Asmat

first: Xiang Yu

first: Mohd

last: Nauman

first: Pamela

last: Stanley


title: Untargeted mutagenesis of brassinosteroid receptor SbBRI1 confers drought tolerance by altering phenylpropanoid metabolism in Sorghum bicolor

city: Cerdanyola del Vallès

first: Veredas

last: Coleto-Alcudia

state: Barcelona

address: Edifici CRAG - Campus UAB

department: Molecular Genetics

institute: Center for Research in Agricultural Genomics (CRAG)

summary: Drought is a critical issue in modern agriculture, therefore there is a need to create crops with drought resilience. The complexity of plant responses to abiotic stresses, particularly in the field of brassinosteroid (BR) signaling, has been the subject of extensive research. In this study, we unveil compelling insights indicating that the BRASSINOSTEROID INSENSITIVE 1 (BRI1) receptor in Arabidopsis and Sorghum plays a critical role as a negative regulator of drought responses. Introducing untargeted mutation in the sorghum BRI1 receptor (SbBRI1) effectively enhances the plant ability to withstand osmotic and drought stress. Through DNA Affinity Purification sequencing (DAP-Seq) we show that the sorghum BRI1-EMS-SUPPRESSOR 1 (SbBES1) transcription factor, a downstream player of the BR signaling, binds to a conserved G-box binding motif, and it is responsible for regulating BR homeostasis, as its Arabidopsis ortholog AtBES1. We further characterized the drought tolerance of sorghum bri1 mutants and decipher SbBES1-mediated regulation of phenylpropanoid pathway. Our findings suggest that SbBRI1 signaling serves as a dual purpose: under normal conditions, it regulates lignin biosynthesis by SbBES1, but during drought conditions, BES1 becomes less active, allowing the activation of the flavonoid pathway. This adaptive shift improves the photosynthetic rate and photoprotection, reinforcing crop adaptation to drought.

overall_design: To identify the early transcriptional targets of SbBES1 in drought sensing, we performed RNA sequencing (RNA-Seq) experiments of sorghum plants (root tissue) in control and drought stress conditions (30% field capacity). BC2F3 EMS-mutagenized plants carrying or lacking the SNP in the extracellular region of SbBRI1,  assuming  they are near isogenic lines(NIL-bri1-72 and NIL-WT-72), were used. RNA was extracted with Maxwell® 16 LEV Plant RNA Kit (Promega) and quality checked using Bioanalyzer.  Three biological replicates were obtained per genotype and condition.

first: Juan

last: Fontanet-Manzaneque

first: Ana

middle: I

last: Caño-Delgado


title: Targeting axonal guidance dependencies in glioblastoma with ROBO1 CAR T cells

city: Toronto

first: Jason

last: Moffat

state: Ontario

address: 686 Bay Street

department: Genetics and Genome Biology

country: Canada

institute: Hospital for Sick Children

summary: Resistance to genotoxic therapies and tumor recurrence are hallmarks of glioblastoma (GBM), an aggressive brain tumor. Here, we explore functional drivers of post-treatment recurrent GBM. By conducting genome-wide CRISPR-Cas9 knockout screens in patient-derived GBM models, we uncover distinct genetic dependencies in recurrent tumor cells absent in their patient-matched primary predecessors, accompanied by increased mutational burden and differential transcript and protein expression. These analyses map a multilayered genetic response to drive tumor recurrence, identifying protein tyrosine phosphatase 4A2 (PTP4A2) as a novel modulator of self-renewal, proliferation and tumorigenicity at GBM recurrence. Genetic perturbation or small molecule inhibition of PTP4A2 activity represses axon guidance activity through a dephosphorylation axis with roundabout guidance receptor 1 (ROBO1), exploiting a functional dependency on ROBO signaling. Importantly, engineered anti-ROBO1 single-domain antibodies mimic effects of PTP4A2 inhibition. Since a pan-PTP4A inhibitor was limited by poor penetrance across the blood brain barrier (BBB) in vivo, a second-generation chimeric antigen receptor (CAR)-T cell therapy was engineered against ROBO1 that elicits specific and potent anti-tumor responses in vivo. A single dose of anti-ROBO1 CAR-T cells doubles median survival in patient-derived xenograft (PDX) models of recurrent glioblastoma, and also eradicates tumors in ~50% of mice in PDX models of adult lung-to-brain metastases and pediatric relapsed medulloblastoma. We conclude that functional reprogramming drives tumorigenicity and dependence on a multi-targetable PTP4A-ROBO1 signaling axis at GBM recurrence, with potential in other malignant brain tumors.

overall_design: Gene expression profiling was performed in triplicate by RNA sequencing in cell lines derived from primary and recurrent glioblastoma.  Along with the baseline expression, differential expression was determined after transduction with CRISPR sgRNAs targeting the AAVS1 locus or two individual guides targeting the PTP4A2 gene.

first: Chirayu

last: Chokshi

first: Benjamin

last: Brakel

first: Kevin

last: Brown

first: Sheila

last: Singh


title: Distinctive CD39+CD9+ lung interstitial macrophages suppress IL-23/Th17-mediated neutrophilic asthma by inhibiting NETosis [bulk RNA-Seq]

city: Seoul

first: Do Young

last: Hyeon

state: Select State

address: 1 Gwanak-ro, Gwanak-gu

department: School of Biological Sciences

country: South Korea

institute: Seoul National University

summary: IL-23-Th17 signaling axis is responsible for neutrophilic inflammation in various barrier tissues. However, mechanistic links between IL-23 and Th17 activation remain unclear, despite of their critical contribution to chronic inflammatory diseases. Here, we discovered a novel signaling pathway linking IL-23 to Th17 activation in our neutrophil-dominant asthma (NDA) model. Through single-cell multi-omics analysis, we identified distinctive CD39+CD9+ interstitial macrophages (IMs) suppressed by IL-23. CD39+CD9+ IMs increased by IL-23p19 inhibitor suppressed NETosis, which inhibited Th17 activation. To inhibit NETosis, CD39+CD9+ IMs first attach to neutrophils in a CD9-dependent manner, and then remove ATP molecules near neutrophils in a CD39-dependent manner. Thus, our discovery of CD39+CD9+ IMs provides previously unrecognized signaling component linking IL-23 and NETosis for controlling Th17 activation/neutrophilic inflammation. We finally demonstrated the decreased number of CD39+CD9+ immune cells in patients with severe chronic rhinosinusitis and inflammatory bowel disease, suggesting CD39+CD9+ IMs as potential therapeutic targets for IL-23-Th17-mediated inflammatory diseases.

overall_design: RNA sequencing of CD39+CD9+ interstitial macrophages (IMs) and CD39-CD9- monocytes isolated from the lungs in neutrophil-dominant asthma mice under OVA+LPS/OVA+aIL-23p19 condition.

first: Ji-Hwan

last: Ryu

first: Daehee

last: Hwang


title: Distinctive CD39+CD9+ lung interstitial macrophages suppress IL-23/Th17-mediated neutrophilic asthma by inhibiting NETosis [scRNA-Seq]

overall_design: Single-cell RNA sequencing of CD45+ cells isolated from the lungs in neutrophil-dominant asthma mice under OVA+LPS/PBS (PBS), OVA+LPS/OVA (OVA), OVA+LPA/OVA+Dex (Dex), and OVA+LPS/OVA+aIL-23p19 (p19) conditions.


title: MAVS signaling shapes microglia responses to neurotropic virus infection

status: Public on Aug 29 2024

city: Hannover

first: Ulrich

last: Kalinke

state: Lower Saxony

address: Feodor-Lynen-Str. 7

department: Institute for Experimental Infection Research

institute: Twincore, Centre for Experimental and Clinical Infection Research

summary: Viral encephalitis is characterized by a series of immunological reactions that can control virus infection in the brain, but dysregulated responses may cause excessive inflammation and brain damage. Microglia are brain-resident myeloid cells that are specialized on surveillance of the local environment and in case of viral brain infection they contribute to control of the infection and restriction of viral dissemination. Here, we report that after exposure to neurotropic vesicular stomatitis virus (VSV), murine in vitro microglia cultures showed rapid upregulation of a broad range of pro-inflammatory and antiviral genes, which were stably expressed over the entire 8 hours infection period. Additionally, a set of immunomodulatory genes was upregulated between 6 and 8 hours post infection. In in vitro microglia, the induction of several immune response pathways including cytokine responses was dependent on mitochondrial antiviral-signaling protein (MAVS). Consequently, in Mavs-deficient microglia control of virus infection failed as indicated by augmented accumulation of viral transcripts. Thus, in the analyzed in vitro system, MAVS signaling is critically required to achieve full microglia activation and to mediate profound antiviral effects. In Mavs-deficient mice, intranasal VSV instillation caused higher disease severity than in WT mice and the virus disseminated beyond the olfactory bulb and entered more distal brain regions. Virus spread to inner regions of the olfactory bulb, i.e., the granular cell layer, correlated with infiltration of highly inflammatory non-microglial myeloid cells into the olfactory bulb. Furthermore, increased cytokine levels were detected in the nasal cavity, the olfactory bulb and in other brain regions. Thus, microglial MAVS signaling is critically needed for virus sensing, full microglia activation, and for orchestration of protective immunity in the virus-infected CNS.

overall_design: Brains from C57BL/6 and B6.STOCK-Mavs (tm1Tsc) (Mavs-/-) newborns were dissected at day 3 post birth, meninges and blood vessels were removed using a stereomicroscope. Next, the brains were minced and digested with 0.1% trypsin and 0.25% DNAse, single cell suspensions were seeded in pre-coated poly-L-lysine flasks in DMEM with 1% penicillin/streptomycin supplemented with 10% FCS. Medium was changed on day 1 and 5. After 10 days of mixed glial culture, microglia were harvested by shaking the flasks (125 rpm) for 40 min at 37°C. Microglial cells were re-plated at a density of 2.0 x 105 cells per well in 24 well plates. The cultured microglia were inoculated at a MOI of 0.5 of VSV diluted in DMEM medium for 1 hr at 37°C in 5% CO2. At 2, 4, 6 and 8 h post infection, the supernatants were removed and the adhering cells were then harvested and lysed by TRIzol. RNA were extracted from cell lysates and sequenced on Illumina NovaSeq-6000 with 50 bp paired-end reads configuration.

first: Felix

last: Mulenge

first: Olivia

last: Gern

first: Andreas

last: Pavlou


title: CBFA2T3-GLIS2 Mediates Transcriptional Regulation of Developmental Pathways through a Gene Regulatory Network

city: Palo Alto

first: Tanja

last: Gruber

state: CA

address: 1000 Welch Road Suite 300

department: Pediatrics

institute: Stanford University

summary: CBFA2T3-GLIS2 is a fusion oncogene found in pediatric acute megakaryoblastic leukemia (AMKL) that is associated with a poor prognosis. We established a model of CBFA2T3-GLIS2 driven AMKL that allows the distinction of fusion specific changes from those that reflect the megakaryoblast lineage of this leukemia. Using this model, we mapped genome wide binding that in turn imparts the characteristic transcriptional signature. A network of transcription factor genes bound and upregulated by the fusion were found to have downstream effects that result in dysregulated signaling of developmental pathways including NOTCH, Hedgehog, TGF, and WNT. Transcriptional regulation is mediated by homo-dimerization and binding of the ETO transcription factor through the nervy homology region 2 (NHR2). Loss of NHR2 abrogated the development of leukemia and led to the downregulation of JAK/STAT and NOTCH transcriptional signatures. These data contribute to the understanding of CBFA2T3-GLIS2 mediated leukemogenesis and identify potential therapeutic vulnerabilities for future studies.

overall_design: Human CD34+ stem cells were isolated from commerically available cord blood units, transduced with CBFA2T3-GLIS2 lentiviral construct and differentiated to megaryoblasts with TPO and IL1β. After 7-9 days of differentiation, cells were sorted for GFP purity, and transplanted into immunodeficient recipient mice. Bone marrow was collected from moribund mice, mouse CD45 cells were depleted, and the recovered leukemic cells were used for RNA sequencing and cleavage under targets and release using nuclease with sequencing (CUT&RUN-seq) for the marks TY, ETO, CtBP1, p300, and H3K27ac. Cells transduced with the CBFA2T3-GLIS2 construct with deletion of nervy homology region 2 (NHR2) were only cultured in vitro due to their lack of expansion in vivo. Wild type megakaryoblasts were cultured in vitro for normal control samples.

first: Pratima

last: Nallagatla

first: Elias

last: Salfati


title: Targeting IRE1α reprograms the tumor microenvironment and enhances anti-tumor immunity in prostate cancer [scRNA-seq2]

city: Oslo

first: Fahri

last: Saatcioglu

address: Blindernveien 31

department: Department of Biosciences

country: Norway

institute: University of Oslo

summary: Unfolded protein response (UPR) is a central intracellular stress response pathway that is hijacked by tumor cells for their survival. However, how activation of UPR in cancer cells may shape the tumor microenvironment (TME) remains largely unexplored. Here, we investigated the potential role of IRE1α signaling on modulation of TME dynamics in prostate cancer (PCa). We found that IRE1α is increased in PCa patient tumors and genetic inhibition of IRE1α in syngeneic mouse PCa models, as well as in an orthotopic model, dramatically reduced tumor growth. Multiomics analysis suggested that IRE1α ablation in cancer cells significantly potentiated interferon (IFN) response and activation of immune system related pathways in the TME. Single-cell RNA-sequencing (scRNA-seq) revealed that the abundance of immunosuppressive cells, such as tumor-associated macrophages (TAMs), were markedly reduced in the IRE1α deficient tumors. Geneset Enrichment Analysis demonstrated that IFN response was significantly enriched in TAMs, cancer cells, and dendritic cells. Notably, the small molecule IRE1α inhibitor MKC8866 (ORIN1001), currently in clinical trials, reprogrammed the TME and enhanced the response to anti-PD-1 blockade therapy in syngeneic PCa mouse models. Furthermore, a novel scRNA-seq-derived TAM gene signature is strongly associated with poor PCa survival, which is reduced by the MKC8866 + anti-PD-1 combination therapy. Our findings indicate that activation of IRE1α signaling not only promotes cancer cell growth and survival, but it also interferes with anti-tumor immunity in the TME. Thus, IRE1α targeting could present a novel approach for improving anti-PD-1 immunotherapy in PCa.

overall_design: scRNA-sequencing (scRNA-seq) was performed on PTEN KO Myc-CaP tumors from animals treated with indicated treatments. Freshly dissected tumor samples were washed with PBS and transferred into GEXSCOPE Tissue Preservation Solution (Singleron), and stored on ice. Six tumor samples from each group were pooled for scRNA-seq. The tissue samples were then washed with Hanks balanced salt solution three times and minced into small pieces which were digested with Tissue Dissociation Solution (Singleron) at 37 °C for 15 min with agitation. After digestion, cells were passed through a 40 μm strainer to remove cell debris and centrifuged at 300 × g for 5 min. The supernatant was removed and cell pellet was resuspended in 1 ml PBS, followed by red blood cell lysis using RBC lysis buffer. Next, cells were centrifuged at 500 × g for 5 min and resuspended in PBS. The single cell suspension was adjusted to a concentration of 1 × 105 / ml for library preparation. This was loaded onto microfluidics chips (GEXSCOPE Single Cell RNA-seq Kit, Singleron Biotechnologies) and GEXSCOPE 3´SD scRNAseq libraries were prepared according to manufacturer instructions (Singleron Biotechnologies). Individual libraries were diluted and pooled for next generation sequencing on Illumina NovaSeq with 150 bp paired-end reads.

first: Bilal

last: Unal

first: Omer

middle: F

last: Kuzu

first: Yang

last: Jin


title: Epigenomic and Functional Convergence Between Glucocorticoid- and IL4-Driven Macrophage Programming [ATAC-seq]

city: New york

first: Yurii

last: Chinenov

address: 535 E 71 str, Hospital for Special Surgery

institute: Hospital for special surgery

summary: Macrophages adopt distinct phenotypes in response to environmental cues, with type-2 cytokine interleukin-4 (IL4) promoting a tissue-repair homeostatic state (M2IL4). Glucocorticoids - widely used anti-inflammatory therapeutics - reportedly impart a similar phenotype (M2GC), however how or to what extent these populations functionally converge is unknown. We show that M2IL4 and M2GC transcriptomes share a striking overlap mirrored by a shift in chromatin landscape in both common and signal-specific gene subsets. This core homeostatic program is enacted by transcriptional effectors KLF4 and the GC receptor (GR), whose genome-wide occupancy and actions are integrated in a stimulus-specific manner by the nuclear receptor cofactor GRIP1. Indeed, 35% of the shared IL4- vs. GC-elicited transcriptomic changes were GRIP1-dependent. Consistently, GRIP1 loss attenuated M2IL4 and M2GC phagocytic activity in vitro and macrophage tissue-repair properties in the murine colitis model in vivo. These findings provide a mechanistic framework for homeostatic macrophage programming by distinct signals, to better inform GR-targeting drug design.

overall_design: Mouse primary bone marrow-derived macrophages from C57BL/6  mice  were exposed to IL4 or Dexamethasone  for 24 h

first: Dinesh

last: Deochand

first: Marija

last: Dacic

first: Inez

last: Rogatsky


title: Seq-Scope profiling of mouse colon sections with biopsy-induced injuries

city: Ann Arbor

first: Jun Hee

last: Lee

state: MI

address: 109 Zina Pitcher Pl

institute: University of Michigan

summary: We used Sequence-Scope (Seq-Scope), which can generate ultra-high definition images of sequence-based molecular signatures resolved at a submicrometer scale, for profiling spatial transcriptome associated with biopsy-associated colon injury. Raw 1st-Seq FASTQ with intact tile and coordinate information could be retrieved from https://doi.org/10.5281/zenodo.13118097

sample_organism:
Mus musculus
synthetic construct

platform_organism:
Mus musculus
synthetic construct

overall_design: The colon samples were obtained from C57BL/6 mice after 24 hrs of biopsy-induced physical wounding. The Seq-Scope procedure was performed on the injured colon sample as previously described with minor modifications involving the use of the Illumina HiSeq 2500 flow cells and chemical strategies to increase RNA sequence capture.

first: Yichen

last: Si

first: Chun-Seok

last: Cho

first: Miguel

last: Quiros

first: Asma

last: Nusrat

first: Jun

middle: H

first: Hyun

last: Kang


title: Three-dimensional chromatin architecture in DRG neurons following sciatic nerve injury in WT and Rad21 KO mice.

city: Columbus

first: Ilaria

last: Palmisano

state: Ohio

address: 460 West 12th Avenue

institute: Ohio State University

summary: Axonal regeneration after injury requires active gene transcription. Whether chromatin 3D genome architecture is required for the neuronal regenerative transcriptional programme is unexplored. Here we addressed this question in a model of sciatic nerve injury by combinatorial Hi-C, promoter-capture Hi-C, CUT&Tag for H3K27ac and RNA-seq in wild-type and cohesin-deficient sensory dorsal root ganglia neurons.

overall_design: Neuronal nuclei were isolated from mouse sciatic dorsal root ganglia 3 days after sciatic nerve crush (SNC) or sham  or in naïve conditions from WT (INTACT-AdvillinCre) and Rad21 KO (INTACT-Scc1flox/flox-AdvillinCre) mice, 14 days after tamoxifen injection. Neuronal nuclei were then used for RNA-seq (N=3 replicates), Hi-C (N=3 or 2 replicates), promoter Hi-C (N=3 replicates), and H3K27ac Cut&Tag (N=3 replicates).

first: Tong

last: Liu

first: Simone

last: Di Giovanni

first: Zheng

last: Wang


title: Inflammatory mediators suppress FGFR2 expression in human keratinocytes to promote skin inflammation

city: ZÜRICH

first: Luca

last: Ferrarese

state: Zürich

address: Otto-Stern-Weg 7, HPL F23.2

department: Institute of Molecular Health Sciences (IMHS)

institute: ETH Zürich

summary: FGFs are key orchestrators of development, tissue homeostasis and repair. FGFR deficiency in mouse keratinocytes causes an inflammatory skin phenotype resembling atopic dermatitis (AD). To determine the human relevance of this finding, we generated human keratinocytes with a CRISPR/Cas9-induced knockout of FGFR2. Loss of this receptor promoted the expression of interferon-stimulated genes and other genes involved in inflammation under homeostatic conditions and in particular in response to inflammatory mediators without affecting cell viability. Bioinformatics analyses and functional studies showed strong down-regulation of FGFR2 expression in cultured keratinocytes exposed to various pro-inflammatory stimuli and in keratinocytes of lesional AD skin, which is likely to further aggravate the inflammatory phenotype. These results reveal a key function of FGFR2 in human keratinocytes in the suppression of skin inflammation and suggests a role of FGFR2 down-regulation in AD pathogenesis.

overall_design: To identify differentially expressed genes (DEG) in the presence and absence of FGFR2 signaling, we generated FGFR2 KO HaCaT cell lines and performed RNA sequencing (RNA-seq) of 3 KO and 3 WT cell lines.

first: Sabine

last: Werner

first: Michael

last: Koch


title: i-motif studies

city: Darlinghurst

first: Cristian

middle: David

last: Pena Martinez

state: NSW

address: 384 Victoria St, Garvan Institute of Medical Research Lvl 10 immunology

department: St Vincent’s Clinical School, Faculty of Medicine, Department of Immunology

country: Australia

institute: Garvan Institute of Medical research


title: Natural killer cells are required for the recruitment of CD8+ T cells and the efficacy of immune checkpoint blockade in melanoma brain metastases

city: Leeds

first: Mihaela

last: Lorger

address: Becket street

department: School of Medicine

institute: University of Leeds

summary: This study investigated how the depletion of natural killer (NK) cells in mice treated with a combined PD-1/CTLA-4 blockade affects the molecular profiles of intracranial tumors in a two-site B16-OVA melanoma brain metastases model. This model contains concomitant intracranial and extracranial tumors, to mimic the presence of extracranial metastases in melanoma patients with brain metastases, and intracranial responses to the combined PD-1/CTLA-4 blockade that are observed in the clinic can be reproduced in this model.

overall_design: C57Bl6 mice were implanted with B16-OVA murine melanoma cells on the flank under the skin, and 3 days later B16-OVA/Fluc melanoma cells were implanted into the brain (striatum). Mice were treated with IgG control antibodies (IgG_B group) or a combined anti-PD-1 / anti-CTLA-4 therapy on days 5 and 7 post-intracranial cancer cell implantation, and intracranial tumors were isolated on day 8 post-intracranial cancer cell implantation. Mice in the therapy group were further split into those with high (PCcd8H_B group) and low (PCcd8L_B group) intra-tumoral CD8+ T cell abundance, corresponding to therapeutic responders and non-responders, respectively. In the fourth group, NK cells were depleted throughout the experiment using anti-Asialo-GM2 antibody concomitant to therapy with a combined anti-PD-1 / anti-CTLA-4 therapy (PCdNK_B group). mRNA was isolated from bulk tumors obtained from 3 separate mice per group (replicates 1-3) and subjected to mRNAseq and data analysis.

first: Christopher

last: Fife

first: Williams

last: Jennifer


title: Targeting IRE1α reprograms the tumor microenvironment and enhances anti-tumor immunity in prostate cancer [RNA-Seq]

first: Fahri

last: Saatcioglu

overall_design: RNA-sequencing (RNA-seq) was performed on Myc-CaP IRE1α WT and KO cells, as well as corresponding xenografted tumor samples from FVB mice. Briefly, RNA was isolated from the cells grown in vitro or the tumor samples using Trizol and purified using RNeasy columns (Qiagen). After RNA isolation, TruSeq stranded RNA-seq libraries were prepared according to manufacturer instructions (Illumina) and 50 bp paired end sequencing was performed using Illumina NovaSeq (Illumina, Inc) at the NorSeq Sequencing Core (Ullevål).

first: Bilal

last: Unal

first: Omer

last: Kuzu

first: Yang

last: Jin


title: Targeting IRE1α reprograms the tumor microenvironment and enhances anti-tumor immunity in prostate cancer

overall_design: Refer to individual Series


title: RNA-seq expression profiling of Xenopus tropicalis blastula embryos upon Tet2 and Tet3 knockdown

city: Mainz

first: Emil

last: Karaulanov

address: Ackermannweg 4

institute: Institute of Molecular Biology

summary: To analyze the role of Tet proteins on global gene expression, we profiled the transcriptome of Xenopus tropicalis blastula embryos upon knockdown of Tet2 and Tet3 using Morpholino antisense oligonucleotides.

overall_design: Gene expression profiling by RNA-seq of blastula stage 9 Xenopus tropicalis embryos injected with Tet2 and Tet3 Morpholinos in biological triplicates.


title: Pol III chromatin immunoprecipitation-sequencing (ChIP-seq) of Xenopus tropicalis blastula embryos

summary: To correlate 5fC clusters with RNA polymerase III occupancy genome-wide, we carried out Pol III ChIP-seq profiling in Xenopus tropicalis blastula embryos.

overall_design: Chromatin from blastula stage 9 Xenopus tropicalis embryos was analyzed by ChIP-seq using RNA polymerase III antibody.


title: MCF7 wild-type RNA profiling for i-motif DNA studies

summary: mRNA sequencing of MCF7 wild-type cells for the study of i-Motif DNA in the human genome

overall_design: MCF7 cells were grown, harvested and pelleted following conventional protocols. Total RNA wa collected using RNeasy Plus Mini Kit followed by sequencing using illumina NovaSeq 6000. Reads trimmed and mapped to hg19 reference genome and read counts obtained using FeatureCounts and the NCBI RefSeq gtf for hg19.

middle: D

last: Peña Martinez


title: An Efficient Direct Conversion Strategy to Generate Functional Astrocytes from Human Adult Fibroblasts

city: San Antonio

first: Uchit

last: Bhaskar

state: Texas

address: One, UTSA Circle

department: Neuroscience, Developmental and Regenerative Biology

institute: University of Texas at San Antonio

summary: Direct reprogramming approaches offer an attractive alternative to stem-cell-derived models, allowing the retention of epigenetic information and age-associated cellular phenotypes. To explore such age-related phenotypes, several groups have generated multiple neuronal subtypes, neural progenitor cells, oligodendrocytes, and other cell types directly from fibroblasts. Other groups have had success at the efficient conversion of embryonic fibroblasts to astrocytes but have not yet achieved similar conversion efficiency for adult human fibroblasts. In order to generate astrocytes for the study of age-related diseases, we developed an improved direct conversion strategy employing a combination of small molecules to activate specific pathways that induce trans-differentiation of human adult fibroblasts to astrocytes. We demonstrate that this method produces mature GFAP+/S100β+ cells at high efficiency (40-45%), comparable to previous studies utilizing embryonic fibroblasts. Further, Fibroblast-derived induced Astrocytes (FdiAs) are enriched for markers of astrocyte functionality, including ion-channel buffering, gap-junction communication, and glutamate uptake; and exhibit astrocyte-like calcium signaling and neuroinflammatory phenotypes. RNA-Seq analysis indicates an adult rather than fetal astrocytic gene expression signature, with a greater correlation to temporal lobe astrocytes. Fibroblast-derived induced astrocytes provide a useful tool in understanding age-associated disease processes and complement existing in vitro models of induced neurons (iNs), providing an additional platform to study late-stage brain disorders.

overall_design: We generated astrocytes directly from human adult fibroblasts, and verified astrocyte specificity using standard immuno-staining assays, astrocyte-functionality assays, and gene expression profiling. Additionally, we tested neuroinflammatory properties of these FdiAs by stimulating the cells with cytokines, and performing gene expression analysis.

first: Jason

last: Ayo

first: Melanie

last: Carless


title: EGFR and EGFRvIII coopt host defense pathways, promoting progression in glioblastoma

city: San Francisco

first: Zhenyi

last: An

department: Neurology

institute: UCSF

summary: To investigate the function of TLR2 in EGFR and EGFRvIII co-expressing glioblastoma cells, we knocked out TLR2 by CRISPR.

overall_design: We then performed gene expression profiling analysis using data obtained from RNA-seq of the cells.

first: William

last: Weiss


title: Human genomic DNA is widely interspersed with i-motif structures

summary: DNA i-motif structures are formed in the nuclei of human cells and are believed to provide critical genomic regulation. While the existence of i-motif structures in human cells has been demonstrated by immunofluorescent staining and by the characterisation of select model genes, the abundance and distribution of such structures in the human genome have remained unclear. Here we utilise high affinity i-motif immunoprecipitation followed by sequencing to map i-motifs in the genomic DNA of human MCF7, U2OS and HEK293T cells. Validated by biolayer interferometry and circular dichroism spectroscopy, our approach identified i-motif structures that are widely distributed throughout the human genome and are common in genes upregulated in G0/G1 cell cycle phases.

overall_design: MCF7, HEK293T and U2OS cells were grown, harvested and pelleted following conventional protocols. Protein and RNA depleted genomic DNA was extracted, purified and fragmented by sonication. Antibody incubation was perfomed at 4 °C using an i-motif specific antibody (Biotinylated iMab scFv) to pull-down putative i-Motif forming sequences followed by sequencing using illumina NovaSeq 6000 and NovaSeq 6000 SP reagent kit in a paired-end 300 cycles mode following the manufacturer protocols . Data was finally collected from two biological replicates for each cell line and sequenced input DNA from each replicate was used for peak calling analysis. Additinally, DNA derived from MCF7 cells by a modified protocol incubating purified DNA and the iMab antibody at 16 °C was also sequenced to inviestigate temperature bias.

first: Mahdi

last: Zeraati


title: Differences in cell subsets between sun-exposed and unexposed skin: preliminary single-cell sequencing and biological analysis from a single case

status: Public on Aug 28 2024

city: Guangzhou

first: zhou

last: feng

state: None Selected

address: 600# Tianhe Road, Guangzhou, Guangdong P.R, China

institute: The Third Affiliated Hospital of Sun Yat-sen University

summary: The microcosmic and specific changes of human photoaging skin without chronologic aging is still unclear and rare-ly analyzed. We aimed to assess  biological processes of the cell-subgroups alteration  and mechanisms of skin pho-toaging using single cell sequencing and biological analysis  were used between sun-exposed forearm skin  and un-exposed buttock skin in same individual via skin punch biopsy.

overall_design: Forearm skin and buttock skin were obtained from the same healthy individual via skin punch biopsy. After enzymatically and mechanically disrupting the tissue and stringent cell filtration, dead cells were thoroughly removed. Single cell RNA sequencing (scRNA-seq) was used to test the cell populations and subsets. Seurat and other packages in R version was used for data analysis.

first: Feng

last: zhou

first: Yu

last: Sun

first: Xinling

last: Chen

first: Wenyi

last: Hou

first: Li

last: Li

first: Yao

last: Qing

first: Qingfang

last: Xu

first: Xinyao

last: Zheng

first: Jing

last: Shen

first: Wei

last: Lai

first: Kai

last: Han

first: Yue


title: Single nucleus RNA-seq of rat nucleus accumbens

city: Philadelphia

first: Richard

last: Crist

state: PA

address: 125 South 31st St, Room 2207

institute: University of Pennsylvania

summary: Single nucleus RNA-seq of nucleus accumbens from male Brown Norway rats after a single acute or repeated saline injections

overall_design: We performed the 10x Genomics 3' gene expression assay (v3.1) on nuclei isolated from frozen Brown Norway (BN) rat nucleus accumbens brain punches. A cohort of male BN rats received an experimenter delivered dose of saline (n=5) and were sacrificed two hours after injection. A second cohort of male BN rats received repeated saline injections as yoked controls in a study of morphine for three hours daily for 10 days (n=6), and animals were sacrificed two hours after the final session. Brains were removed and snap-frozen in dry ice-cold isopentane. Brains were coronally sectioned and the nucleus accumbens core and shell were removed by micro-punch bilaterally, and the punches were stored at-80°Cuntil further processing. Nuclei were isolated by differential centrifugation and microfluidically captured using the 10x Genomics Chromium controller. Sequencing libraries were constructed and sequenced on an Illumina NovaSeq 6000.

first: Benjamin

last: Reiner

first: Wade

last: Berrettini

first: Heath

last: Schmidt


title: Developmental molecular controls over arealization of descending cortical motor pathways

status: Public on Aug 27 2024

city: Geneva

first: Denis

last: Jabaudon

address: Rue Michel-Servet 1

institute: University of Geneva

summary: Layer 5 extratelencephalic (ET) neurons are a main class of neocortical projection neurons that predominate in the motor cortex and send their axon to multiple proximal and distal targets including the thalamus, pons, medulla and spinal cord1-6. Precise connectivity of ET neurons is critical for fine motor control; they are central to loss of function upon spinal cord injury and specifically degenerate in amyotrophic lateral sclerosis7-10. ET neurons consist of several subtypes of cells with distinct laminar and areal locations, molecular identities, connectivities, and functions11,12. Two cardinal subtypes of ET neurons have been identified: neurons that express Nprs1 or Hpgd and project proximally to the pons and thalamus (ETprox), and neurons that express Slco2a1 and project more distally to the pons, medulla and spinal cord (ETdist)11. Despite their critical function, how these neuronal subtypes emerge during development and acquire their area-specific distributions remains unaddressed. Here, using combinations of anatomical labeling, MAPseq mapping13, and single- nucleus transcriptomics across developing cortical areas, we reveal that these two subtypes of ET neurons are present at birth along opposite antero-posterior cortical gradients. We first characterize area-specific developmental axonal dynamics of ETprox and ETdist neurons and find that the former can emerge by pruning of subsets of ETdist neurons. We next identify area- and ET neuron subtype-specific developmental transcriptional programs to identify key target genes in vivo and predict their gene regulatory networks. Finally, we reprogram ET neuron subtype- specific connectivity from motor to visual-like, by generating more proximal connections through postnatal in vivo knockdown of three subtype-specific transcription factors. Together, these findings delineate the functional transcriptional programs controlling ET neuron diversity across cortical areas and provide a molecular blueprint to investigate and direct the developmental emergence of corticospinal motor function.

overall_design: In order to study the molecular development of extratelencephalic neurons, we performed MapSeq sindbis virus injections in motor, visual and somatosensory cortical areas and collected cortical and subcortical targets. These experiments were conducted on wild type control animals and enucleated animals at P0. They consist of anterogradely transported unique barcodes from the injection site to the different collected areas. The data provided here contains fastq files with original read as well as a R object file with annotated barcode counts.


title: Developmental molecular controls over arealization of descending cortical motor pathways [AbeP_GEO]

overall_design: In order to study the molecular development of extratelencephalic neurons, we performed single nuclei RNA sequencing of retrogradely traced cortical neurons at different postnatal developmental stages and micro-dissecting primary motor and somatosensory areas.


title: Comparison of ILC1s and NK cells transcriptomes in subcutaneous tumor model

city: Hefei

first: XiqQiong

last: Wang

state: Anhui

address: School of Life Sciences, No. 443 Huangshan Road

institute: University Of Science And Technology Of China

summary: ILC1s are tissue-resident lymphocytes, their recruitment, migration and infiltration into subcutaneous tumors is a big concern. In addition, as NK cells and ILC1s are phenotypically similar, it is difficult to assess the relative contribution of these two cell types to anti-tumor function. Here, we compared the transcriptomes of ILC1s and NK cells from MC38 tumor-bearing mice to understand the biological/phenotypic changes of ILC1s.

overall_design: Tumor tissue was obtained 14 days after subcutaneous transplantation of MC38 cells in WT mice and tumor infiltrating cells were isolated. ILC1s and NK cells  were directly sorted into 1.5 ml RNase-free tubes containing 700 μl Trizol (Takara). After sort, samples were flash frozen. Three ILC1s and three NK cell samples were processed and sequenced.

first: Rongbin

last: Zhou

first: Jiaxian

last: Yan

first: Zonghui

last: Huang


title: Single-cell transcriptomics reveal distinctive patterns of fibroblast activation in heart failure with preserved ejection fraction

city: Heidelberg

first: Florian

last: Leuschner

address: INF 669

institute: Uni Heidelberg

summary: Inflammation, fibrosis and metabolic stress critically promote heart failure with preserved ejection fraction (HFpEF). Exposure to high-fat diet and nitric oxide synthase inhibitor N[w]-nitro-l-arginine methyl ester (L-NAME) recapitulate features of HFpEF in mice. To identify disease specific traits during adverse remodeling, we profiled interstitial cells in early murine HFpEF using single-cell RNAseq (scRNAseq). Diastolic dysfunction and perivascular fibrosis were accompanied by an activation of cardiac fibroblast and macrophage subsets. Integration of fibroblasts from HFpEF with two murine models for heart failure with reduced ejection fraction (HFrEF) identified a catalog of conserved fibroblast phenotypes across mouse models. Moreover, HFpEF specific characteristics included induced metabolic, hypoxic and inflammatory transcription factors and pathways, including enhanced expression of Angiopoietin-like 4 (Angptl4) next to basement membrane compounds, such as collagen IV (Col4a1). Fibroblast activation was further dissected into transcriptional and compositional shifts and thereby highly responsive cell states for each HF model were identified. In contrast to HFrEF, where myofibroblast and matrifibrocyte activation were crucial features, we found that these cell states played a subsidiary role in early HFpEF. These disease-specific fibroblast signatures were corroborated in human myocardial bulk transcriptomes. Furthermore, we identified a potential cross-talk between macrophages and fibroblasts via SPP1 and TNFɑ with estimated fibroblast target genes including Col4a1 and Angptl4. Treatment with recombinant ANGPTL4 ameliorated the murine HFpEF phenotype and diastolic dysfunction by reducing collagen IV deposition from fibroblasts in vivo and in vitro. In line, ANGPTL4, was elevated in plasma samples of HFpEF patients and particularly high levels associated with a preserved global longitudinal strain. Taken together, our study provides a comprehensive characterization of molecular fibroblast activation patterns in murine HFpEF, as well as the identification of Angiopoietin-like 4 as central mechanistic regulator with protective effects.

overall_design: Comparing effects of HFpEF treamtent (L-NAME +HFD) mouse model with control. We sequenced ventricular tissue of 2x control and 2x HFpEF mice.

first: Laura

middle: M

last: Wienecke


title: CCL2-mediated Endothelial Injury Drives Cardiac Dysfunction in Long COVID

first: Yu

last: Liu

address: 1215 Welch Road

department: CVI

summary: Chronic endotheliitis and various cardiovascular co-morbidities are more likely to develop in patients who are recovering from a post-acute SARS-CoV-2 infection. Despite a growing body of clinical data suggesting that the endothelium could be the cause of both cardiac injury and the multi-organ damage found in COVID-19 patients, there is no clear link between endothelial (EC) dysfunction and increased cardiac risk during long COVID. Here, we studied long COVID-19-associated endotheliitis and its implications on cardiac dysfunction. Thrombotic vascular tissues from long COVID patients were harvested and profiled to identify the different mechanisms of viral-induced EC pathogenesis. Human induced pluripotent stem cell (iPSC)–derived ECs were leveraged to model endotheliitis in-a-dish after exposure to SARS-CoV-2, which showed similar EC dysfunction and upregulation of specific cytokines such as CCL2 and IL6, as seen in the primary ECs of long COVID patients. 3D fabricated cardiac organoids generated from iPSC-ECs and iPSC-derived cardiomyocytes (iPSC-CMs) were utilized to understand the pathological influence of endotheliitis on cardiac dysfunction. Notably, cardiac dysfunction was observed only in cardiac organoids that were fabricated with both iPSC-CMs and iPSC-ECs after exposure to SARS-CoV-2. Simultaneous profiling of chromatin accessibility and gene expression dynamics via integration of ATAC-seq and RNA-seq at a single cell resolution revealed CCL2 as the prime cytokine responsible for the non-endothelial “phenotype switching” and the impending cardiac dysfunction in cardiac organoids. This was further validated by high-throughput proteomics that showed CCL2 to be released only by cardiac organoids that were fabricated with iPSC-CMs and iPSC-ECs after SARS-CoV-2 infection. Lastly, disease modeling of the cardiac organoids as well as exposure of human ACE2 transgenic mice to SARS-CoV-2 spike proteins uncovered a putative mechanism for the cardiac dysfunction involving posttranslational modification of cardiac proteins driven by oxidative stress and inflammation. These results suggest that EC-released cytokines can contribute to the pathogenesis of long COVID-associated cardiac dysfunction, and thus a thorough clinical profiling of vascular health could help identify early signs of heart disease in COVID-19 patients.

overall_design: Bulk was conduced on iPSC-EC generated from Long COVID patients. scRNA-seq was conducted on human arteries that were collected from Long COVID patients that had undergone below-knee amputation. Similarly, scRNA-seq was conducted on arteries collected from NON-COVID patients that suffered from peripherial artery disease (PAD) and undergone below-knee amputation due to complete occlusion of their arteries.

first: Nazish

last: Sayed


title: Transcription factors Sox8 and Sox10 contribute differentially to maintenance of the differentiated state in oligodendrocytes

city: Erlangen

first: Michael

last: Wegner

state: Bavaria

address: Fahrstr. 17

department: Institut of Biochemistry

summary: To identify target genes of the transcription factors Sox8 and Sox10 in oligodendrocytes in a genome-wide and unbiased approach, we induced oligodendocyte-specific deletion of Sox8, Sox10 or a combination thereof (Sox8 knockout, Sox10 knockout, double knockout) in 2 months-old mice, dissected the corpus callosum of control and mutant mice after 21 days and isolated total RNA for ensuing RNA-Seq studies. Among others we detected substantial changes in the expression of genes associated with differentiation of oligodendrocytes and myelination in Sox10 knockout mice as well as double knockout mice but not in Sox8 knockout mice.

overall_design: We generated mRNA profiles of control, Sox8 knockout, Sox10 knockout and double knockout mice using the Illumina NovaSeq 6000, PE 2x150 platform.

first: Lisa

last: Joerg

first: Elisabeth

last: Sock


title: Effect of ageing on gene expression in the African turquiose killifish retina

city: Leuven

first: Lieve

last: Moons

address: Naamsestraat 61

department: Biology

institute: KU Leuven

summary: The mammalian central nervous system (CNS) is susceptible to age-related pathologies, resulting in progressive, irreversible disease. Neurodegenerative eye conditions, like glaucoma and age-related macular degeneneration (AMD), are on the rise due to increased life expectancy. Despite this, there are currently no long-term therapies to prevent degeneration and vision loss. The short-lived African turquoise kililfsh (Nothobranchius furzeri GRZ-AD) is an ideal genetic model for ageing studies, exhibiting rapid ageing phenotypes within its four to six-mont lifespan. Investigating the molecular consequences of ageing in the retina, we conducted bulk RNA-sequencing, revealing dysregulation of genetic pathways associated with ageing CNS and retinal diseas in the aged killifish retina.

overall_design: To investigate the impact of ageing on the retina of the killifish, we performed bulk RNA-sequencing on the retina of three different age groups: 6- (young), 12- (middle-aged), and 18-week-old fish (old). Per condition (age group) we sequenced 10 biological replicates. Each biological replicated consisted of the right and left retina of one female killifish.

first: Steven

last: Bergmans

first: Nicole

last: Noel

first: Luca

last: Masin

first: Ellen

last: Harding

first: Aleksandra

last: Krzywańska

first: Julie

last: De Schutter

first: Ayana

last: Rajagopal

first: Chi-Kuo

last: Hu

first: Lut

last: Arckens

first: Philip

last: Ruzycki

first: Ryan

last: MacDonald

first: Brian

middle: S

last: Clark


title: Generation of human region-specific brain organoids with medullary spinal trigeminal nuclei

city: Shanghai

first: Jinkui

last: Zhu

state: Shanghai

address: 393 Middle Huaxia Road, Pudong, Shanghai, 201210, China.

department: School of Life Science and Technology

institute: ShanghaiTech University

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


title: RNA-seq Analysis of Round and Sirloin Muscle Tissues from Hanwoo Neonatal Calves Born to Mothers with Overnutrition during Mid to Late Pregnancy

city: Pyeongchang-gun

first: Borhan

last: Shokrollahi

state: Gangwon-do

address: 국립축산과학원 한우연구소 강원도 평창군 대관령면 경강로 4937, 25340

institute: National Institute of animal science

summary: This study investigates gene expression in round and sirloin muscle tissues from neonatal calves using RNA-seq.

overall_design: RNA was extracted from round and sirloin muscle tissues of control and treated groups, sequenced using Illumina NovaSeq.

first: Myung

last: Park

first: Youl

last: Baek

first: Shil

last: Jin

first: Ki

last: Jang

first: Sung

last: Moon

first: Kyung-Hwan

last: Um

first: Sun

first: Hyun-Jeong

last: Lee


title: Bulk RNA-sequencing from SVBP or TTL gene knock-out and WT engineered heart tissues

city: Münster

first: Marisol

last: Herrera-Rivero

state: NRW

address: Albert-Schweitzer-Campus 1

institute: University of Münster

summary: For this study, we used commercially available hiPSCs (Allen Institute, AICS-0031) and created knock-out lines for small vasohibin binding protein (SVBP) and tubulin tyrosine ligase (TTL), respectively. These hiPSCs were then differentiated into cardiomyocytes using a monolayer protocol and then further used for generation of engineered heart tissues (EHTs) to more closely model human physiology. EHTs were either cultured on control silicone posts or stiffer silicone posts to achieve enhanced afterload.

overall_design: We casted EHTs composed of 1 Mio cells from freshly differentiated hiPSC-cardiomyocytes (cTnT+ cells >85%) in a fibrin-based matrix and cultured them in low-glucose DMEM. WT EHTs were exposed to control silicone posts (0.28 mN/mm) or stiffer silicone posts for afterload enhancement (0.8 mN/mm). SVBP-KO and TTL-KO EHTs were only cultured on stiff silicone posts. After 60 d, EHTs were harvested, snap-frozen and RNA extracted for bulk RNAseq. n=3 EHTs per group were included, however, for SVBP-KO no successful library preparation was achieved, reducing the n number in this group.

first: Niels

last: Pietsch

first: Lucie

last: Carrier


title: Bulk RNA-sequencing from whole heart lysates of a mouse model of hypertrophic cardiomyopathy after TTL overexpression.

summary: Reducing microtubule detyrosination has been shown to inprove contractility in heart failure cardiomyocytes. To evaluate the feasibility of targeting the detyrosinated microtubule network for treatment of cardiomyopathy in an in-vivo context we overexpressed tubulin tyrosine ligase (TTL) by AAV9 delivery in a mouse model of HCM (Mybpc3-/-) and an empty vector as control in HCM and WT mice.

overall_design: We overexpressed AAV9-TTL or AAV9-empty in p21 mice (2.25E11 vg/g) and monitored heart function for 12 weeks by echocardiography. Animals were then subjected to hemodynamic measurements and sacrificed, before heart tissue was harvested, snap frozen and powdered. RNA was extracted with the Promega SV total RNA kit. n=3 animals were included for RNAseq.

first: Birgit

last: Geertz

first: Lisa

last: Krämer

first: Giulia

last: Mearini


title: Succinate dehydrogenase deficiency-driven succinate accumulation induces drug resistance in acute myeloid leukemia via ubiquitin-cullin regulation

city: Hangzhou

first: Meidan

last: Ying

address: No.866 Yuhangtang Road

department: College of Pharmaceutical Sciences

institute: Zhejiang University

summary: Drug resistance is vital for the poor prognosis of acute myeloid leukemia (AML) patients, but the underlying mechanism remains poorly understood. Given the unique microenvironment of bone marrow, we reasoned that drug resistance of AML might rely on distinct microenvironment-associated metabolic processes. Here, we identified SDH deficiency and over-cumulative succinate as typical features in AML, with a marked function in causing the resistance of AML cells to a wide range of anti-cancer therapies. Mechanistically, succinate promoted the accumulation of oncogenic proteins in a manner that precedes transcriptional activation. This function was mediated by succinate-triggered upregulation of UBC12 phosphorylation, which impaired its E2 function in cullins neddylation. Notably, decreasing succinate levels by fludarabine could effectively restore the drug sensitivity of SDH-deficient AML PDX. Together, we uncover a novel function of succinate in driving drug resistance by regulating p-UBC12/cullin activity, and indicate reshaping succinate metabolism as a promising treatment for SDH-deficient AML.

overall_design: HL-60 cells were treated with 6 mM succinate for 3 h or 24 h and then harvested. SDHA is knocked down in HL-60 cells using lentiviral infection, followed by two rounds of puromycin selection to establish stable SDHA knockdown cell lines.  RNA sequencing was then actualized and analyzed.

first: Yifan

last: Chen

first: Miao

last: Xian

first: Wenwen


title: TAS-102, Irinotecan, and Bevacizumab in Pre-treated Metastatic Colorectal Cancer (TABAsCO), A Phase II Clinical Trial. ClinicalTrials.gov NCT04109924

status: Public on Aug 26 2024

city: Buffalo

first: Jianxin

last: Wang

state: NY

address: 665 Elm Street

department: Molecular and Cellular Biology

institute: Roswell Park Comprehensive Cancer Center

summary: Background: The efficacy of FOLFIRI plus an antiangiogenesis biologic agent as 2nd line therapy for metastatic colorectal adenocarcinoma is limited. TAS-102 is a novel oral antimetabolite with a distinct mechanism of action from fluoropyrimidines. We evaluated the antitumor efficacy of TAS-102, irinotecan and bevacizumab in patients with pre-treated, advanced colorectal adenocarcinoma in a multicenter, phase II, single-arm study. Methods: Patients with advanced colorectal adenocarcinoma who had progressed after oxaliplatin and fluoropyrimidine and were eligible for treatment with bevacizumab were treated with irinotecan, bevacizumab, and TAS-102 in 28-day cycles. The primary endpoint was progression-free survival (PFS). Results: We enrolled 35 evaluable patients. The study was positive. The median PFS was 7.9 (90% CI 6.2-11.8) months (vs. 6 months in historical control, p=0.018). The median overall survival was 16.5 (90% CI 9.8-17.5) months. Sixty-seven percent of patients experienced grade 3 or higher treatment-related adverse events. The most common toxicities were hematological (neutropenia) and gastrointestinal (diarrhea, nausea, and vomiting). Conclusions: Irinotecan, TAS-102 and bevacizumab is an active 2nd line therapy for patients with metastatic colorectal adenocarcinoma. Neutropenia is common and can affect dose density/intensity mandating use of G-CSF. A randomized study versus standard of care therapy is warranted.

overall_design:
We evaluated the significance of expression of genes involved in different types of DNA repair, trifluorothymidine metabolism, cell cycle control, and replication in treatment outcomes. We identified 12 patients treated at Roswell Park with available archival tumor tissue. Under a separate de-identified protocol (BDR 155422), we extracted RNA from these tumors for whole transcriptome sequencing (see Supplementary Methods). Additionally, we identified 21 patients treated with TAS-102 monotherapy with available archival tumor samples for similar analyses to be used as controls (BDR 151721). Genes of interest were grouped in the following categories: HR, BER, MMR, nucleotide excision repair (NER), proofreading polymerases, non-homologous end joining (NHEJ), DNA damage response, cell cycle checkpoints, replication, and trifluorothymidine/nucleotide metabolism.
Clinical data for samples is available from authers by request.

first: Christos

last: Fountzilas

first: Patrick

last: Boland

first: Andrei

last: Bakin