title: Runx transcription factors regulate early T-developmental progression by inducing T-lineage identity and innate lymphoid cell program genes via gene network regulation

city: Pasadena

first: Boyoung

last: Shin

address: 1200 E. California Blvd

institute: California Institute of Technology

summary: Runx transcription factors are essential for generating functional T cells starting from the earliest stages in thymic T-development.  To evaluate how Runx factors instruct T cell development, we perturbed Runx activities using 1) CRISPR-Cas9, simultaneously deleting two predominant, redundant Runx factors, Runx1 and Runx3, and 2) Runx1-overepxression at a stage before pro-T cells commit to a T cell fate. Then single-cell transcriptome analysis was performed.    Runx1/Runx3 knockout (KO) vs. Runx1 overexpression (OE) resulted in contrasting deviations from the normal developmental trajectory. The separation of both perturbation conditions from the controls implied that Runx factors regulate gene networks in individual cells to control separate pathways instead of changing distributions of cells among different stages along the control pathway.  Notably, a common core set of genes responded to both KO and OE, in opposite directions.  Runx activation target genes were selectively enriched for T-identity program and innate lymphoid cell program genes, whereas inhibition targets were significantly associated with stem/progenitor program and myeloid lineage pathways.  Pseudotime inference analysis suggested that Runx perturbations also substantially regulate developmental speed, as Runx KO significantly slowed down developmental progression, whereas Runx1 OE resulted in pronounced acceleration.  This interpretation was largely supported by Runx activities regulating key transcription factors involved in establishing distinct gene network modules during T cell development. Indeed, further analysis showed that Runx target genes get combinatorial inputs from these Runx-regulated transcription factors as well as from Runx factors themselves.   Together, our data suggest that Runx transcription factors drive early T developmental progression by launching gene expression modules associated with T-identity and innate lymphoid cells through mediating other transcription factor cooperativities.

overall_design: To recapitulate early T cell development in a perturbation-accessible way, the OP9-Dll1 in vitro culture system was exploited.   For Runx1 overexpression conditions, freshly obtained bone-marrow-derived progenitor cells from 8-10 week old B6.Bcl2 transgenic mice were cultured on OP9-Dll1 cells with 10ng/ml of IL-7 and Flt3-ligand.  On day 2, MSCV-Runx1-mCherry or MSCV-empty-mCherry vectors were delivered using spinfection.  Two days after delivering retrovirus (total 4 days of culture) or 4 days post-infection (total 6 days of culture), each of the experimental replicates was stained with a distinct hashtag oligo and live Lineage marker-negative, mCherry+, CD45+ cells were FACS sorted for transcriptome analysis.     For Runx1/Runx3 double knockout, slightly more time was allowed for the disruption of the targeted genes to reduce protein levels before gene expression was measured. Bone-marrow-derived progenitor cells from B6.Cas9;Bcl2 transgenic mice were co-cultured with OP9-Dll1 cells in medium supplemented with 10ng/ml each of IL-7 and Flt3-ligand. On day 2, E42-gRunx1-humanNGFR (hNGFR) and E42-gRunx3-mTurquoise vectors were retrovirally introduced.  Three gRNA pairs were utilized for each Runx1 and Runx3 targeting.   For control conditions, E42-gControl-humanNGFR (hNGFR) and E42-gControl-mTurquoise vectors were utilized, where control gRNA targets luciferase sequences. At 3 days post-infection (total 5 days of culture) or 6 days post-infection (total 8 days of culture), each of the experimental replicates was stained with a different hashtag oligo and live Lineage-, mTurquoise+, hNGFR+, CD45+  cells were FACS sorted.      Equal numbers of FACS sorted cells from each of the experimental replicates from Runx1 OE and Runx1/Runx3 double KOs and controls were pooled and subjected to single-cell RNA-seq using 10X Chromium v3.

first: Fan

first: Wen

first: Ellen

last: Rothenberg


title: Unreprogrammed H3K9me3 prevents minor zygotic genome activation and lineage commitment in SCNT embryos

status: Public on Jul 10 2023

city: Shanghai

first: Qianshu

last: Zhu

state: Shanghai

address: Siping Road

department: School of Life Science and Technology

country: China

institute: Tongji University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Somatic cell nuclear transfer (SCNT) can be used to reprogram differentiated somatic cells to a totipotent state but has poor efficiency in supporting full-term development. H3K9me3 is considered to be an epigenetic barrier to zygotic genomic activation in 2-cell SCNT embryos. However, the mechanism underlying the failure of H3K9me3 reprogramming during SCNT embryo development remains elusive. Here, we perform genome-wide profiling of H3K9me3 in cumulus cell-derived SCNT embryos. We find redundant H3K9me3 marks are closely related to defective minor zygotic genome activation. Moreover, SCNT blastocysts show severely indistinct lineage-specific H3K9me3 deposition. We identify MAX and MCRS1 as potential H3K9me3-related transcription factors and are essential for early embryogenesis. Overexpression of Max and Mcrs1 significantly benefits SCNT embryo development. Notably, MCRS1 partially rescues lineage-specific H3K9me3 allocation, and further improves the efficiency of full-term development. Importantly, our data confirm the conservation of deficient H3K9me3 differentiation in Sertoli cell-derived SCNT embryos, which may be regulated by alternative mechanisms.

overall_design: H3K9me3 in somatic cell nuclear transfer (NT) embryos

first: Ruiming

last: Xv


title: Single-cell multi-omics sequencing of human spermatogenesis reveals a DNA demethylation event associated with male meiotic recombination

status: Public on Jul 09 2023

city: Guangzhou

first: Xinyan

last: Yang

state: Guangdong

address: No.1023,South Shatai Road, Baiyun District

department: Developmental Biology

institute: Southern Medical University

type: Methylation profiling by high throughput sequencing

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Perturbed epigenetic transcriptional regulation in AML with IDH mutations causes increased susceptibility to NK cells [methylation]

city: Huddinge

first: Anna

last: Palau

address: Hälsovägen 9

country: Sweden

institute: Karolinska Institutet

summary:
IDH mutations are found in 20% of acute myeloid leukemia (AML) patients. IDH inhibitors (IDHi) have emerged as a novel treatment option. However, only 30-40% of patients respond to single drug treatment and there is an unmet need for improvement as well as identifying alternative treatment options. The overall aim of this study was therefore to gain deeper insights into the molecular signatures of IDH mutations, the effects of IDHi as well as to identify a molecular vulnerability to tailor novel therapies for AML patients with IDH mutations. Here, we have characterized the transcriptional and epigenetic landscape before and after treatment with IDH2i AG-221, using an IDH2 mutated AML cell line model and AML patient cohorts.    We discovered decreased DNA hydroxymethylation in IDH2 mutated AML cells, particular in enhancers and a perturbed transcriptional regulatory network involving myeloid transcription factors. In addition, hypermethylation of the HLA I cluster caused a dramatic down-regulation of HLA genes, triggering an enhanced natural killer (NK) cell activation, and displayed an increased susceptibility to NK cell mediated killing. These responses were reverted when the AML cells were pre-exposed to IFN-gamma, resulting in up-regulation of cell surface HLA class I. Finally, analyses of DNA methylation data from IDHi-treated patients showed that non-responders continued to harbor hypermethylation in HLA class I genes, suggesting maintained susceptibility to NK cells. In conclusion, this study provides new insights into the perturbed epigenetic and transcriptional regulation in IDH mutated AML and shed light on a potential strategy for personalized medicine in AML.
Methylation/hydroxymethylation profiling by array

overall_design: Bisulfite and oxidative bisulfite converted DNA from the 32 samples were hybridised to the Illumina Infinium MethylationEPIC microarray for 850,000 CpG

first: Filip

last: Segerberg

first: Michael

last: Lidschreiber

first: Katja

first: Aonghus J

last: Naughton

first: Maria

last: Needhamsen

first: Lisa

last: Jung

first: Maja

last: Jagodic

first: Patrick

last: Cramer

first: Sören

last: Lehmann

first: Mattias

last: Carlsten

first: Andreas

last: Lennartsson


title: Single-cell multi-omics sequencing reveals the epigenomic landscapes in human spermatogenesis [STRT-seq; scCOOL-seq]

summary: Human spermatogenesis is a well-ordered dynamic process, which ensures the long-term production of functional spermatozoa; however, the roles of DNA methylation and chromatin accessibility in this process remain largely unknown. Here, by simultaneously investigating the chromatin accessibility, DNA methylome and transcriptome landscapes of human spermatogenesis using the modified single-cell chromatin overall omic-scale landscape sequencing approach (Modified scCOOL-seq), we revealed that the transcriptional changes throughout human spermatogenesis were correlated with the dynamic alternations of chromatin accessibility; particularly, several potential transcription factors and a set of regulatory elements involved in such process were identified. Remarkably, a round of DNA demethylation was uncovered upon meiosis initiation in human spermatogenesis, which was associated with male meiotic recombination and conserved between human and mouse. And aberrant DNA hypermethylation at recombination hotspots could be detected in leptotene spermatocytes of certain nonobstructive azoospermia patients. Functionally, the intervention of DNA demethylation affected male meiotic recombination. Collectively, this study provides multi-omics landscapes of human spermatogenesis at single-cell resolution, and offers insights into the association between DNA demethylation and male meiotic recombination.

overall_design:
In total, we analyzed 1097 scCOOL-seq data of normal human adult testicular cells and 306 scCOOL-seq data of testicular cells from one nonobstructive azoospermia (NOA) donor. Samples 1-32 are RNA-seq data (RNA part), samples 33-1435 are scCOOL-seq data (DNA part).
*** All the raw fastq data have been uploaded to GSA for Human as access controlled data under accession HRA000148 (https://bigd.big.ac.cn/gsa-human/browse/HRA000148)***

first: Xiao-Yang

last: Zhao

first: Shuai

last: Gao

first: Gang

last: Chang

first: Yaping

last: Huang

first: Lin

last: Li


title: Concentration- and co-factor-dependent Runx DNA binding site choices during early T-development

first: Boyoung

last: Shin

state: CA

country: USA

summary: Runx1 and Runx3 function redundantly in early T-development, and together drive T-lineage developmental progression by regulating distinct sets of genes in different stages.  Context-specific Runx target genes are particularly enriched near Runx binding sites that dynamically shift from pre-T-commitment stages (Phase 1) to post-T-commitment stages (Phase 2).  As total Runx activities (Runx1+Runx3) are maintained stably throughout early T-development stages, yet Runx factors physically interact with multiple collaborators, we hypothesized that different Runx-collaborating transcription factors compete to recruit a limited pool of Runx transcription factors.  To test whether increasing Runx availability can alter Runx DNA-binding site choices, Runx1 overexpression vectors were introduced to two different systems. First, we increased Runx1 expression in a DN3-like cell line (representing a post-commitment, Phase 2 stage) in the presence or absence of exogenous Phase 1 co-factor, PU.1. Second, we increased Runx1 expression in Phase 1 primary cells which naturally express PU.1 and other Phase 1 collaborators.  Then, Runx1 binding behaviors were measured using ChIP-seq or CUT&RUN (C&R).      A modest increase of Runx1 levels (about 2-3 fold increase) substantially increased the number of Runx binding sites seen and the intensity of occupancy in both systems. When Runx expression was at physiological levels, PU.1 dominated Runx1 site choice before T commitment by recruiting Runx1 to PU.1 sites. The Phase 2 cell line system showed that it did this while depleting Runx1 from alternative high quality Runx motif sites.  However, when Runx1 was overexpressed, Runx1 was still recruited to PU.1 sites, but this recruitment did not evacuate Runx1 occupancy from default preferred sites. Notably, Runx1 overexpression in primary Phase 1 cells caused precocious occupancy of post-commitment, Phase 2-specific sites. We found that these are often co-occupied with TCF1, E2A, and HEB, but have minimal co-binding with PU.1. In addition, Runx1 overexpression resulted in new binding sites that were not normally observed in pro-T cells, which are mostly sequestered by closed chromatin normally although they harbor more numerous Runx motifs.  Thus, these data suggest that Runx DNA binding site choices are sensitive to Runx concentration and co-factors during early T-development.

overall_design:
CUT&RUN (C&R), ChIP-seq, ATAC-seq
To measure Runx binding sites in unperturbed DN1 cells, freshly obtained bone-marrow-derived progenitor cells from 8-10 week old C57BL/6J wild-type (WT) animals were cultured on OP9-Dll1 cells with cytokines (IL-7 and Flt3-ligand, 10 ng/ml each).  On day 5, live lineage (TCRb, TCRgd, CD19, NK1.1, CD11b, CD11c, Ly6G/C) negative, CD45+, cKithi, and CD25- cells were FACS sorted. Approximately 250K -500K cells were utilized to conduct CUT&RUN (C&R).  For unperturbed thymic DN3 C&R, thymus was obtained from 5-6 weeks old C57BL/6J WT mice and live Lineage marker-negative DN3 cells (cKitlow, CD44low, and CD25-) were sorted.  500K cells were subjected to C&R analysis.

To detect Runx1 occupancies with Runx1 overexpression in Phase 1, freshly obtained bone-marrow progenitor cells from B6.Bcl2 transgenic mice were co-cultured with OP9-Dll1 cells in medium supplemented with 10ng/ml of IL-7 and Flt3-ligand.  On day 2, MSCV-Runx1-mCherry (Runx1 OE) or MSCV-empty control-mCherry vectors were delivered using spinfection.  Two days after delivering retrovirus (total 4 days of culture), live Lineage-, mCherry+, CD45+, cKithi Phase 1 cells were FACS sorted. 180-250K cells were utilized to measure Runx1 binding using C&R.

ChIP-seq
DN3-like Scid.adh.2c2 (P2C2) cells were retrovirally infected with pMXs-PU.1-human NGFR (hNGFR) and MSCV-Runx1-HA-mCherry vectors.  For control conditions, empty vectors for pMXs-hNGFR and MSCV-mCherry vectors were introduced. On day 2 post-infection, P2C2 cells were stained with anti-NGFR-PE antibody and hNGFR-expressing P2C2 cells were selected using anti-PE microbeads (Miltenyi Biotec).  Approximately 10 million cells were fixed at RT in 1 mg/ml disuccinimidyl glutarate (DSG) and 1% formaldehyde and subjected to chromatin immunoprecipitation (ChIP) using anti-HA antibody.

ATAC-seq
To assess chromatin accessibility changes upon Runx1 overexpression, control- or Runx1 OE introduced-pro-T cells (the same as the C&R samples) were FACS sorted and washed with 1X HBSS with 10mM HEPES twice.  Then, chromatin accessibility profiles were measured using ATAC-seq.

first: Jue

last: Wang

first: Ellen

last: Rothenberg


title: Endocrine treatment potentiates antigen presentation and in combination with SMAC mimetics enhances an anti-tumor immune response in hormone receptor positive breast cancer (RNA-rela-KOs)

status: Public on Jul 08 2023

city: Boston

first: Yingtian

last: Xie

state: MA

address: 450 BROOKLINE AVE

institute: DANA FARBER CANCER INSTITUTE INC

type: Expression profiling by high throughput sequencing

summary: To investigate the effects of endocrine treatment in combination with SMAC mimetics in ER+ breast cancer on tumor cell response to IFNg

overall_design: Comparative gene expression profiling analysis of RNA-seq data from MCF7 Control, MCF7 RELA KO1, MCF7 KO2 treated for 72h with: Hormone deprived conditions, estradiol 10nM, vehicle (DMSO) or Fulvestrant 10nM with and without stimulation of IFNg 10ng/mL for 6h.

first: Francisco

last: Hermida Prado

first: Rinath

last: Jeselsohn


title: Endocrine treatment potentiates antigen presentation and in combination with SMAC mimetics enhances an anti-tumor immune response in hormone receptor positive breast cancer (ATAC-seq WH/E2)

overall_design: Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) in MCF7 cells grown in HD and E2, with and without stimulation of IFNg 10ng/mL for 1h.


title: N6-methyladenosine modification is essential for prostate cancer cells

city: Chicago

first: Xiaolong

last: Cui

state: IL

address: 680 N Lake Shore Drive

department: Department of Preventive Medicine

institute: Northwestern University

summary: N6-methyladenosine (m6A) modification of messenger RNAs (mRNAs) is a pivotal mechanism controlling mRNA fate in cells. RNA m6A modification is regulated by the functional balance between methyltransferases and demethylases. Here we demonstrated that FTO-IT1 enhancer RNA (eRNA), a long non-coding RNA (lncRNA) transcribed from the last intron of FTO gene is significantly upregulated in CRPC and aggressive tumors compared to primary tumors. FTO-IT1 knockout by CRISPR/Cas9 almost completely blocks growth and G1-S cell cycle transition of both androgen-sensitive and castration-resistant prostate cancer cells. Meanwhile, the mRNA m6A was dramatically increased in FTO-IT knockout PCa cells and we identified FTO-IT1 as a binding partner of FTO. From m6A-seq, we unexpectedly found hypermethylated m6A associated with upregulated levels of the mRNAs for p53 signaling pathway genes in 22Rv1 prostate cancer cells. Mechanistic study showed that FTO-IT1 recruits FTO to the P53 target mRNA to promote their m6A demethylation, which leads to their degradation.

overall_design: MeRIP-seq was performed on both wild-type and FTO KO prostate cancer cells

first: Jiangbo

last: Wei

first: Jianong

last: Zhang

first: Chuan

last: He

first: Haojie


title: Endocrine treatment potentiates antigen presentation and in combination with SMAC mimetics enhances an anti-tumor immune response in hormone receptor positive breast cancer


title: Endocrine treatment potentiates antigen presentation and in combination with SMAC mimetics enhances an anti-tumor immune response in hormone receptor positive breast cancer (orig)

overall_design: Comparative gene expression profiling analysis of RNA-seq data from MCF7 cells treated with vehicle, Fulvestrant (10nM), Birinapant (100nM) and the combination for 72h with and without the presence of IFNg for the last 24h..


title: A lncRNA from the FTO locus acts as an inhibitor of mRNA m6A methylation and p53 tumor suppression

state: Illinois

address: 929 E 57th St, Room E319B

institute: University of Chicago

summary: The FTO gene locus has been linked to cancer and obesity through encoded N6-methyladenosine (m6A) demethylase FTO or inherited genomic variants (e.g. intronic single-nucleotide polymorphisms). Here we demonstrate that FTO-IT1, a long noncoding RNA (lncRNA) transcribed from a FTO gene intron, is upregulated during prostate cancer (PCa) progression and positively correlated with poor survival of patients with tumors only expressing wild-type p53. We show that RBM15, a mRNA/substrate binding subunit of the m6A methyltransferase complex binds and increases mRNA m6A methylation and stability of p53 transcriptional target genes; however, FTO-IT1 overexpression abolishes these effects by blocking RBM15 binding of p53 target gene mRNAs. Therapeutic targeting of FTO-IT1 restores mRNA m6A level and p53 signaling and inhibits PCa tumor growth in mice. Our study identifies FTO-IT1 lncRNA as a bono fide inhibitor of m6A methylation and p53 tumor suppression and nominates FTO-IT1 as a potential biomarker and therapeutic target of cancer.

overall_design: [Dataset 1] 4 samples. Duplicates for CLIP-Seq samples with an antibody against RBM15 in 22Rv1 (CRL-2505) human prostate carcinoma cell line

first: Zhongyu

last: Zou


title: Endocrine treatment potentiates antigen presentation and in combination with SMAC mimetics enhances an anti-tumor immune response in hormone receptor positive breast cancer (RNA-seq)

overall_design: Comparative gene expression profiling analysis of RNA-seq data from MCF7 cells grown in Hormone deprived conditions or in the presence of estradiol and stimulated with IFNg for 6, 12, 24 and 48h.


title: Effect of Pf-Rel/poI-κB RNA interference on gene expression in mantle tissue in Pinctada fucata

status: Public on Jul 07 2023

city: Beijing

first: Bowen

last: Shuai

state: Beijing

address: Tsinghua University, Haidian District, Beijing

department: School of Life Science

institute: Tsinghua university

summary: Pinctada fucata is an important pearl production shellfish in aquaculture. The formation of shells and pearls is a hot research topic in biomineralization, and matrix proteins secreted by the mantle tissues play the key role in this process. However, upstream regulatory mechanisms of transcription factors on the matrix protein genes remain unclear. Previous studies have shown that NF-κB signaling pathway regulated biomineralization process through expression regulation of specific matrix proteins.In this study, we systematically investigated the effect of the NF-κB signaling pathway key factor Pf-Rel and inhibitory protein poI-κB on the biomineralization and shell regeneration process. RNA-seq was conducted based on RNA interference animal model to identify potential regulatory genes by transcription factor Pf-Rel. We extracted total RNA from the whole tissue samples of the pericardium 7 days after dsRNA injection, and named them as the Rel-RNAi group, IκB-RNAi group and control group, respectively. We first calculated the Pearson correlation coefficient between each pair of samples, which reflects the similarity in overall gene expression patterns among the samples. The calculation results showed a high correlation coefficient of 0.954 between the Rel-RNAi group and IκB-RNAi group. We performed Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses on differentially expressed genes (DEGs) from two groups. The top three enriched GO terms for DEGs were: extracellular region, peptidase inhibitor activity and peptidase regulator activity. In addition, immune receptor activity and integrin-mediated signaling pathways terms were significantly enriched. The KEGG Pathway classification showed that a large number of DEGs were classified as signal transduction and immune system terms. Biomineralization is also an important natural immune pathway in shellfish, which suggests that the NF-κB signaling pathway in P. fucata is closely related to natural immunity. KEGG pathway enrichment analysis indicated that substantial DEGs were attributed to PI3K-Akt signaling pathway term, which plays a vital role in cell survival

overall_design: To investigate the regulatory effcet of NF-κB signaling pathway on biomineralization, we established RNA interference animial model in pearl oysters, P. fucata. Healthy adult oysters with similar size and weight were injected with 30 μg different dsRNAs into the adductor muscle, and each treatment group included five oysters. Control group was injected with 30 μg gfp dsRNA. After 7 days, the mantle tissues from five individuals were collected for total RNA extraction and RNA-seq.

first: Rongqing


title: Conserved transcriptional programming across sex and species after peripheral nerve injury predicts treatments for neuropathic pain

city: Toronto

first: Shahrzad

last: Ghazisaeidi

state: Ontario

address: 250 College Street

department: Krembil Centre for Neuroinformatics

country: Canada

institute: Centre for Addiction and Mental Health

summary:
Background and Purpose
Chronic pain is a devastating problem affecting one in five individuals around the globe, with neuropathic pain the most debilitating and poorly treated type of chronic pain. Advances in transcriptomics have contributed to cataloguing diverse cellular pathways and transcriptomic alterations in response to peripheral nerve injury but have focused on phenomenology and classifying transcriptomic responses.
Key Results
We identified 93 transcripts in the dorsal horn that were increased by peripheral nerve injury in male and female mice and rats. Following gene ontology and transcription factor analyses, we constructed a pain interactome for the proteins encoded by the differentially expressed genes, discovering new, conserved signalling nodes. We investigated the interactome with the Drug‐Gene database to predict FDA‐approved medications that may modulate key nodes within the network. The top hit from the analysis was fostamatinib, the molecular target of which is the non‐receptor spleen associated tyrosine kinase (Syk), which our analysis had identified as a key node in the interactome. We found that intrathecally administrating the active metabolite of fostamatinib, R406 and another Syk inhibitor P505‐15, significantly reversed pain hypersensitivity in both sexes.
Conclusions and Implications
Thus, we have identified and shown the efficacy of an agent that could not have been previously predicted to have analgesic properties.

sample_organism:
Mus musculus
Rattus norvegicus

platform_organism:
Mus musculus
Rattus norvegicus

overall_design: To identifying new types of pain‐relieving agents, we compared transcriptional reprogramming changes in the dorsal spinal cord after spared nerve injury in male and female mice and rats, and imputed commonalities, as well as differences in cellular pathways and gene regulation.

first: Milind

middle: M

last: Muley

first: YuShan

last: Tu

first: David

middle: P

last: Finn

first: Mahshad

last: Kolahdouzan

first: Graham

last: Pitcher

first: Doyeon

last: Kim

first: Ameet

middle: S

last: Sengar

first: Arun

middle: K

last: Ramani

first: Michael

last: Brudno

middle: W

last: Salter


title: Boosting NAD preferentially blunts TH17 inflammation via arginine biosynthesis and redox regulatory control in control and psoriasis subjects [healthy or psoriasis]

status: Public on Jul 18 2023

city: Bethesda

first: Komudi

last: Singh

address: Building 10

institute: NIH

summary: To evaluate whether NAD+-boosting modulates adaptive immunity, primary CD4+ T cells from healthy control and psoriasis subjects were exposed to vehicle or nicotinamide riboside (NR) supplementation. NR blunts IFNg and IL-17 secretion with greater effects on TH17 polarization. RNA-seq analysis implicates NR blunting of sequestosome 1 (SQSTM1/p62)-coupled oxidative stress. NR administration increases SQSTM1 and reduces reactive oxygen species (ROS) levels. Furthermore NR activates NRF2, and genetic knockdown of NRF2 and of the NRF2-dependent gene, SQSTM1 diminish NR amelioratory effects. Metabolomic analysis identify that NAD+-boosting increases arginine and fumarate biosynthesis and genetic knockdown of argininosuccinate lyase ameliorates NR-effects on IL-17 production. Hence, NR via amino acid metabolites orchestrate NRF2 activation, augments CD4+ T cell antioxidant defenses and attenuates TH17 responsiveness. Oral NR supplementation in healthy volunteers similarly increase serum arginine, SQSTM1 and antioxidant enzyme gene expression and blunts TH17 immune responsiveness, supporting evaluation of NAD+-boosting in CD4+ T cell linked inflammation.

overall_design: CD4 T cells isolated were from human blood. RNA was extracted from CD4 T cells by miRNeasy Micro Kit (Qiagen). RNA was quantified using a Qubit 3.0 fluorometer (Thermo Fisher Scientific) and its integrity confirmed using an Agilent Technologies 2100 Bioanalyzer (Agilent).

first: Kim

last: Han

middle: N

last: Sack


title: Single cell sequencing reveals the landscape of the brain metastatic microenvironment

city: Winston-Salem

first: Qianqian

last: Song

state: NC

address: 1 Medical Center Boulevard

department: Cancer Genomics and Precision Oncology

institute: Wake Forest Baptist Comprehensive Cancer Center

summary: Our analysis identifies intratumoral components that are predominantly tumor cells, fibroblasts, myeloid cells, stromal cells that express neural stem cell (NSC) markers, with minor populations of oligodendrocytes (oligo) and T cells. Different samples present diverse cell compositions, indicating the underlying cellular interactions that affect different cellular infiltration within TME. Importantly, we identify the tumor-associated fibroblasts in both in-house and external scRNA-seq datasets that 1) highly expresses type I collagen genes; 2) dominates the cell-cell interactions in TME through the type I collagen signaling axis; and 3) remodels the TME to a collagen-I-rich extracellular matrix (ECM) similar to the original TME at the primary sites. We also observe the M1 activation of the native microglial cells and the infiltrated macrophages, which may create a proinflammatory TME and be responsible for the high expression of collagen type I in fibroblasts. Moreover, the tumor cell-specific receptors are significantly associated with patients’ survival in both brain metastasis and the native glioblastoma cases. Overall, our analyses identify the type I collagen secreting tumor-associated fibroblasts as a key mediator in metastatic brain tumors and reveal the involved tumor receptors associated with patients’ survival. Our discoveries provide potential biomarkers for effective therapeutic targets and intervention strategies.

overall_design: Using single-cell RNA sequencing, we have profiled five brain tumor tissue samples that are metastasis from breast and lung cancer.

first: Jing

last: Su

first: Jimmy

last: Ruiz

middle: D

last: Chan


title: Intraperitoneal paclitaxel is a safe and effective therapeutic strategy for treating mucinous appendiceal adenocarcinoma

city: HOUSTON

first: John Paul

last: Shen

state: TX

address: 1515 Holcombe Blvd

department: Gastrointestinal Medical Oncology

institute: The University of Texas MD Anderson Cancer Center

summary: Appendiceal adenocarcinomas (AAs) are a rare and heterogeneous mix of tumors for which few preclinical models exist. The rarity of AA has made performing prospective clinical trials difficult, which has partly contributed to AA remaining an orphan disease with no chemotherapeutic agents approved by the FDA for its treatment. AA has a unique biology in which it frequently forms diffuse peritoneal metastases but almost never spreads via a hematogenous route and rarely spreads to lymphatics. Given the localization of AA to the peritoneal space, intraperitoneal (IP) delivery of chemotherapy could be an effective treatment strategy. Here, we tested the efficacy of paclitaxel given by IP administration using three orthotopic patient-derived xenograft (PDX) models of AA established in immunodeficient NSG mice. Weekly IP paclitaxel treatment dramatically reduced AA tumor growth in all three PDX models. Comparing the safety and efficacy of intravenous (IV) to IP administration, IP delivery of paclitaxel was more effective with reduced systemic side effects in mice. Given the established safety record of IP paclitaxel in gastric and ovarian cancers, and lack of effective chemotherapeutics for AA, these data showing the activity of IP paclitaxel in orthotopic PDX models of mucinous AA support the evaluation of IP paclitaxel in a prospective clinical trial.

overall_design: Comprehensive analyses of RNA-seq profiles comparing patient-derived xenograft (PDX) models of appendiceal cancer treated with intraperitoneal injections of paclitaxel versus those treated with saline control

first: John

first: Ichiaki

last: Ito

first: Saikat

last: Chowdhury


title: Effects of Adjudin on gene expression of db/db mouse pancreatic islets

city: Solna

first: Olov

last: Andersson

state: Stockholm

address: Biomedicum Quarter 5D, Tomtebodavagen 16

department: Department of Cell and Molecular Biology

summary: To investigate effects of Adjudin on gene expression of islets from db/db mouse, islets from 12 to 13 weeks old male db/db mice were isolated, cultured in incubator for overnight recovery, and treated with either DMSO or 10 µM Adjudin for 1 day before RNA sequencing.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for islets treated with DMSO and Adjudin

first: Lipeng

last: Ren


title: Effects of Adjudin on gene expression of postnatal day 0 mouse pancreatic islets

summary: To investigate effects of Adjudin on gene expression of postnatal day 0 mouse islets (P0 islets), islets from postnatal day 0 mouse (regardless of sex) were isolated, cultured in incubator for overnight recovery, and treated with either DMSO or 10 µM Adjudin for 1 day before RNA sequencing.


title: Cryobiopsy: A Breakthrough Strategy for Clinical Utilization of Lung Cancer Organoids

city: Daejeon

first: Jong-Eun

last: Park

address: 291, Daehak-ro, Yuseong-gu

country: South Korea

institute: Korea Advanced Institute of Science and Technology

summary: The biggest problem with lung cancer organoid (LCO) is that most lung cancer organoids are derived from surgical specimens of early-stage lung cancer patients. In fact, patients who initially need chemotherapy are patients with advanced lung cancer who cannot undergo a surgery. So, making lung organoids from biopsy specimens successfully is an urgent task. The success rate of culturing LCO from biopsy tissues is very low because conventional lung biopsies such as transthoracic needle biopsy and forcep biopsy only get a small amount of lung tissue. The possibility of critical complications such as bleeding and pneumothorax makes it difficult to obtain enough specimens. Overgrowth of normal lung cells in late passages is also a critical problem in LCO and optimized culture conditions for LCO remain to be identified. To overcome the hurdles of lung cancer organoid, we made LCOs from cryobiopsy specimens of all stages of lung cancer patients. Transbronchial cryobiopsy can obtain over 10 times greater volume of tissue compared to bronchoscopic forceps biopsy and it can be applied to even peripheral lesions with radial endobronchial ultrasonography. The success rate of LCO culture was also markedly improved and recapitulated the characteristics of primary tumors. LCOs derived from cryobiopsy specimens can overcome the critical limitations of present lung cancer organoids. We expect that cryobiopsy will be a breakthrough strategy of clinical application of LCO in all stages of lung cancer.

overall_design: We performed scRNA-seq on normal (n=1) and cancer (n=1) lung organoids that we established from cryo-biopsy specimens.

first: Dahye

last: Lee

first: Yongjun

last: Koh

first: Chaeuk

last: Chung


title: High-throughput profiling of antigen-specific antibody secreting cells using an antibody secretion trap (TRAPnSeq)

city: Tarrytown

first: Joseph

middle: Cooper

last: Devlin

state: NY

address: 777 Old Saw Mill River Rd

department: Molecular Profiling and Data Science

institute: Regeneron Pharmaceuticals

summary: Following activation by cognate antigen, B cells undergo fine-tuning of their antigen receptors and may ultimately differentiate into antibody-secreting cells (ASCs). While antigen-specific antibodies from B cell receptor (BCR) expressing B cells can be readily cloned and sequenced following flow sorting, antigen-specific plasma cells that lack surface BCR cannot be easily profiled in a high-throughput way. Here, we report an approach, TRAPnSeq (antigen specificity mapping through Ig secretionTRAPandSequencing), that allows capture of secreted antibodies on the surface of ASCs, which in turn enables high-throughput screening of single ASCs against large antigen panels and recovery of paired VH:VL antibody sequences. This approach incorporates flow cytometry, standard microfluidic platforms and DNA barcoding technologies to isolate and characterize antigen-specific ASCs through single cell V(D)J, RNA and antigen barcode sequencing. We show the utility of TRAPnSeq by profiling antigen-specific IgG and IgE plasma cells from mouse and humans and validate antigen binding by ELISA. TRAPnSeq can easily be combined with existing B cell platforms to accelerate antibody discovery from ASCs and can further be expanded to any protein secreting-cells.

overall_design:
The TRAPnSeq protocol was performed on bone marrow from hIL-4Rα immunized mice to identify and sort IgG+/hIL-4Rα binding plasma cells.  The spleen and draining lymph nodes were also processed from these mice, stained using conventional B cell surface markers, and IgG+/hIL-4Rα-binding B cells were sorted. All sorted cells were processed using the 10x genomics platform.
>>> Submitter statement concerning raw data: The VELOCIMMUNE V3 mouse strain is a patented and proprietary genome. We have been advised by our legal counsel not to release raw sequencing (fastq) files. However, we are depositing the processed single cell count tables to enable reproducibility of our analysis. <<<<

middle: C

first: Seblewongel

last: Asrat

first: Andrea

last: Vecchione

first: Brian

last: Klotz


title: Adjudin improves beta cell maturation, hepatic glucose uptake and glucose homeostasis

overall_design: Refer to individual Series


title: Study of the changes on MAIT cell transcriptome after exposure to Klebsiella pneumoniae, interferon alpha and influenza virus

city: London

first: Patricia

last: Barral

address: London

country: United Kingdom

institute: King's College London

summary: To investigate the molecular mechanisms mediating activation of human circulating MAIT cells during infection with Klebsiella pneumoniae and compare the transcriptomics with cells activated by type-I interferons or influenza virus

overall_design: Comparative gene expression profiling analysis of RNA-seq data for human circulating MAIT cells after exposure to 3 conditions: Klebsiella pneumoniae, IFNα2A and influenza virus

first: J

last: Lopez-Rodriguez

first: C

last: Barrington

first: P


title: Targeted high throughput mutagenesis of the human spliceosome reveals its in vivo operating principles [RNA-Seq]

city: San Francisco

first: Irene

last: Beusch

state: California

address: 600 16th Street

department: Biochemistry and Biophysics

institute: UCSF

summary: A major impediment to studying the human spliceosome in vivo has been the inability to program point mutations in endogenous genes on a large scale. CRISPR-Cas9 technology now provides such opportunities. Due to its scalability and ability to introduce point mutations, we chose CRISPR-Cas9 base editing for a forward genetic screen of the spliceosome in an eHAP haploid human cell line background. We maintained eHAP FNLS cells as haploid so that we could subsequently assign genotype-phenotype relationships. We designed a single guide RNA (sgRNA) library targeting a hand-curated list of 153 human spliceosomal proteins which engage at various steps of the splicing cycle. After mutagenesis, we interrogated the spliceosome using the potent inhibitor pladienolide B (PB), which targets U2 snRNP by binding to a pocket between the SF3B1 and PHF5A subunits of the SF3b complex, preventing stabilization of the U2-branchpoint RNA duplex. Validation and genomic sequencing revealed resistance mutations in SF3B1 and PHF5A in residues adjacent to the compound binding pocket. We mapped hypersensitive mutants to U2 snRNP components, but also to factors that act as late as the second chemical step, after SF3b has dissociated. Strikingly, we obtained resistance mutants in SUGP1, a spliceosomal G-patch protein of unknown function that lacks orthologs in yeast and is also a newly proposed tumor suppressor whose loss underpins the splicing changes induced by cancer-associated SF3B1 mutations.

overall_design: RNA-seq to investigate the effect of identified mutations in the spliceosomal proteins PHF5A, SF3B1 and SUGP1 (linked to resistance to the small molecule spliceosome inhibitor pladienolide B) on pre-mRNA splicing.

first: Hiten

last: Madhani


title: GDF15 gene suppression by BET inhibition and gene signature based personalized chemotherapy to counteract cachexia [Quant-seq]

city: Vienna

first: Daniel

last: Gerlach

address: Dr.-Boehringer-Gasse 5-11

department: Global Computational Biology and Digital Sciences

country: Austria

institute: Boehringer Ingelheim RCV GmbH & Co KG

summary: Elevation of growth differentiation factor 15 (GDF15) has recently emerged as a key driver of cancer cachexia, a syndrome of extreme weight loss that affects patients with various types of cancer. Chemotherapy, as well as the tumour itself have been reported to contribute to increased GDF15 secretion in both cancer and host tissues. Bromodomain and extraterminal (BET) domain proteins have been shown to regulate GDF15 expression in pancreatic cancer cells. With currently neither preventive strategies nor specific therapeutic options for cachexia available, antibodies targeting GDF15 or its receptor are being developed. However, the mechanism of chemotherapy-induced GDF15 upregulation in tumour and host cells is still unknown, leaving uncertainty about safety and efficacy of therapies targeting retrospective reduction of GDF15 in cancer cachexia. We tested various cardiotoxic drugs for their effect on GDF15 regulation in cardiomyocyte cell culture and analysed gene expression, mutational status, and pathway activities of the NCI-60 cancer cell line panels to get further insights into the regulation of GDF15 expression in both cancer and host cells upon treatment with anticancer drugs. To investigate the regulatory effect of BET inhibition on GDF expression more extensively, we screened BET inhibitors in doxorubicin treated human cardiomyocyte cell culture as well as in 21 cancer cell lines and performed differential analysis of gene expression, metabolites, and pathway activities. Our results reveal that DNA interacting compounds induce GDF15 expression in human cardiomyocytes up to 20-fold, whereas cardiotoxic drugs with other modes of action led, if at all, to less than 3-fold induction. We discovered that BET inhibitors suppress doxorubicin-induced GDF15 overexpression in human cardiomyocytes up to 7.4-fold (p=0.0051). In cancer cell lines, we identified compound-specific gene signatures that correlate with the extent of GDF15 induction upon treatment in the espective cell line. BET inhibition also reduces GDF15 expression in a defined subset of cancer cell lines that differentiate from the other tested cell lines in a less active PI3K/Akt axis and significantly higher extracellular pantothenate concentration (p=0.00095). We identified compound- and cell-line-specific gene signatures that predict GDF15 upregulation upon treatment, providing indication that the risk for drug-induced GDF15 overexpression and concomitant cachexia can be reduced by a biomarker-driven selection of patient-specific anticancer agents. Our results suggest that BET inhibitors could counteract cachexia at the transcriptional level in tumour and host tissues by reducing GDF15 expression. We identified characteristic gene and metabolite expressions of responsive cancer cell lines that can serve as biomarkers for patient selection.

overall_design: Colon cancer cell lines were grown in media and treated with either DMSO (control) or compound. BI 894999 (WO2014076237) is the clinical compound synthesized by Boehringer Ingelheim. The experiment was performed in biological duplicates per condition/cell lines. Information about sampling time points, compounds, compound concentrations, and replicates are detailed out in the respective sample descriptions. All samples were subjected to RNA isolation, followed by library preparation (QuantSeq 3' mRNA-Seq Library Prep Kit FWD for Illumina, Lexogen), and messenger RNA sequencing on an Illumina NextSeq 500 system.

first: Ulrike

last: Tontsch-Grunt

first: Schweifer

last: Norbert


title: Targeted high throughput mutagenesis of the human spliceosome reveals its in vivo operating principles [editing]

overall_design: We transduced eHAP FNLS with lentivirus carrying individual sgRNAs, grew cells for six days (= t0), isolated genomic DNA, amplified the edited locus and subjected the amplicons to deep sequencing. This approach revealed the consequences of base editing at the amino acid level. For sgRNAs conferring resistance to PB we also collected cells at t8 and t15 in the presence and absence of 2 nM PB treatment.


title: Intra-tumoural Microbiota Remodels the Tumour Immune

city: chengdu

first: hong

last: wu

address: 55 renmin south road

institute: sichuan cancer hospital

summary: Esophageal squamous cell carcinoma (ESCC) accounts for ~90% of all cases of esophageal cancer and the sixth most common cause of cancer related death worldwide. 1, 2 It remains a globally challenging disease and mostly diagnosed cases requires a multidisciplinary approach with extensive treatments including surgery, chemoradiotherapy and/or chemotherapy.  ESCC has a relatively high tumor mutational burden,3 suggesting that it could benefit from PD-1 blockade.  Recently, three phase III clinical trials reported that PD-1 blockade significantly improved progression-free survival (PFS) and overall survival (OS) when combined with chemotherapy as first-line therapy in advanced/metastatic ESCC.4, 5 Moreover, in 2021 ASCO annual meeting,  two phase II clinical trials reported that neoadjuvant chemo-immunotherapy (NACI) induced an obvious improvement of the pathologic complete response (pCR) for resectable ESCC (35.  3% and 42.5%, respectively).  Several phase III trials on this topic are ongoing (NCT04807673, NCT04848753, NCT04280822) (https://clinicaltrial s.gov/ct2/show/NCT04807673), indicating that neoadjuvant NACI would become promising treatment for locally advanced ESCC.  Nevertheless, not all ESCC patients could respond to NACI. Patients who responded well or poorly to neoadjuvant immunotherapy were treated with surgery. We collected samples from these patients and conducted single-cell sequencing to analyze the tumor immune microenvironment of patients with different therapeutic effects, compare the differences in immune cell composition in the immune microenvironment, and explore more effective neoadjuvant therapy methods.

overall_design:
Patients who received 2 cycles of neoadjuvant immunotherapy were assessed by imaging, followed by surgical removal of the tumor. Tumor samples from patients with good and bad efficacy were collected, digested into single cells, and analyzed by single-cell sequencing.
3 human Esophageal Squamous Cell Carcinoma tissue samples; ESCC2-NACI NR; ESCC1-NACI R; ESCC3-NACI R.

[UPDATE] 10-26-2023: ESCC2 samples were corrected from responder to non-responder; ESCC3 and ESCC1 samples were corrected from non-responder to responder.

first: Hong

last: Wu

first: Chuan

last: Xu


title: Targeted high throughput mutagenesis of the human spliceosome reveals its in vivo operating principles [screen]

overall_design: We generated a monoclonal stable cell line expressing FNLS, a cytosine base editor, in an eHAP haploid cell background. A CRISPR-Cas9 base editing screen was performed using a library of 43618 sgRNAs (including 100o non-targeting sgRNAs) targeting 153 proteins. After six days to allow for editing and selection of transduced cells, we then split the selected pool into treatment arms (control/DMSO vs. 2 nM pladienolid B (PB), which approximates its EC50).We cultured cells for two weeks while maintaining a representation of 500 cells per sgRNA. On days 0, 8 and 14 we isolated genomic DNA and amplified and sequenced the sgRNA inserts; we also did so for the input plasmid library. Libraries were sequenced with paired-end to identify both sgRNA sequence as well as a unique barcode linked to each sgRNA.


title: Profiling transcriptomic responses of human stem cell-derived medium spiny neuron-like cells to exogenous phasic and tonic neurotransmitters

status: Public on Jul 06 2023

city: Raleigh

first: Ryan

last: Tam

state: North Carolina

address: 840 Main Campus Drive, Building Partners II, Room 3700

department: Chemical and Biomolecular Engineering

institute: North Carolina State University

summary: Transcriptomic responses to neurotransmitters contribute to the complex processes driving memory and addiction. Advances in both measurement methods and experimental models continue to improve our understanding of this regulatory layer. Here we focus on the experimental potential of stem cell derived neurons, currently the only ethical model that can be used in reductionist and experimentally perturbable studies of human cells. Prior work has focused on generating distinct cell types from human stem cells, and has also shown their utility in modeling development and cellular phenotypes related to neurodegeneration. Here we seek an understanding of how stem cell derived neural cultures respond to perturbations experienced during development and disease progression. This work profiles transcriptomic responses of human medium spiny neuron-like cells with three specific goals. We first characterize transcriptomic responses to dopamine and dopamine receptor agonists and antagonists presented in dosing patterns mimicking acute, chronic, and withdrawal regimens. We also assess transcriptomic responses to low and persistent tonic levels of dopamine, acetylcholine, and glutamate to better mimic the in vivo environment. Finally, we identify similar and distinct responses between hMSN-like cells derived from H9 and H1 stem cell lines, providing some context for the extent of variability these types of systems will likely pose for experimentalists. The results here suggest future optimizations of human stem cell derived neurons to increase their in vivo relevance and the biological insights that can be garnered from these models.

overall_design: We investigate responses to tonic and phasic neurotransmitters (dopamine, its antagonists and agonists and acetylcholine, and glutamate) in hESC-derived medium spiny neurons. We then perform gene expression profiling analysis using data obtained from RNA-seq of H1 and H9 derived hMSN-like cells exposed to a variety of exposures. Comparative gene expression analyses were compared against PBS dosed hMSN-like cells.

first: Albert

middle: J

last: Keung


title: Intriniscally disordered domain of transcription factor TCF-1 is requried for T cell developmental fidelity [CUT&RUN]

city: Philadelphia

first: Naomi

last: Goldman

state: PA

address: 324 BRB II/III, 421 Curie Blvd

institute: University of Pennsylvania

summary: The transcription factor TCF-1 has long been demonstrated to be essential for normal T cell development, with a critical role at the earliest stages of lineage specification downstream of Notch signaling in the thymus. Recent work has shown that TCF-1 targets and is essential for the opening of chromatin in T cells to orchestrate the gene regulatory cascade that occurs during development. However, besides the high mobility group DNA binding domain, no portion of the TCF-1 protein has been characterized to determine its role in hematopoiesis. In this study, we fully characterize all regions of TCF-1 and assess their importance in TCF-1-driven epigenetic and transcriptional regulation as well as in endowing developmental competency to the T cell lineage. Surprisingly, we identified an intrinsically disordered N-terminal region of TCF-1 termed “L1” that is necessary for efficient transition from early thymic progenitors to double-negative (DN)2 T cells. The few T cells that develop from progenitor cells expressing mutant TCF-1 lacking the L1 region exhibit lineage infidelity that was distinct and unique from the lineage diversion of TCF-1-deficient cells. Transcriptional and epigenetic profiling indicated that the L1 region was required for efficient target gene transactivation which correlated with the de-repression of a set of Gata2-driven genes normally restricted to the mast cell and dendritic cell lineages. DL1 mutant DN2 cells showed aberrant opening of previously silent chromatin at mast cell genes and Gata2-bound enhancers. These data support an essential role for TCF-1’s intrinsically disordered N-terminus in maintaining fidelity to the T cell lineage through carefully balanced activation of partner transcription factors that are required to repress alternative lineages.

overall_design: CUT&RUN in DN3 TCF-1KO cell line and in primary developing T cells from Tcf7eGFP fl/fl Cre+ progenitors cultures on OP9-DLL1 cells.

first: Aditi

last: Chandra

first: Abhijeet

middle: R

last: Patil

first: Ashley

last: Vanderbeck

first: Hai-Hui

last: Xue

first: Ivan

last: Maillard

first: Golnaz

last: Vahedi


title: Endocrine treatment potentiates antigen presentation and in combination with SMAC mimetics enhances an anti-tumor immune response in hormone receptor positive breast cancer (RNA-seq WT_Talen)

first: Yingtian

last: Xie

address: 450 BROOKLINE AVE

summary: To investigate the effects of endocrine treatment in combination with SMAC mimetics in ER+ breast cancer on tumor cell response to IFNg

overall_design: Comparative gene expression profiling analysis of RNA-seq data from MCF7 ER wildtype or Y537S knock/in (Talen) were grown in Hormone deprived conditions or in the presence of estradiol and stimulated with IFNg for 24h.

first: Francisco

last: Hermida Prado

first: Rinath

last: Jeselsohn


title: Comparative Transcriptomic Profiling of Differentially Expressed Genes and Related Metabolic Pathways in a Saudi Wheat Cultivar (Najran) under Salinity Stress

city: NEWCASTLE UPON TYNE

first: Norah

middle: Abdullah

last: Alyahya

address: Drummond WEB REQ Tyne

institute: Newcastle University

summary: A comparative RNA-Seq analysis was done in root and shoot of Najran wheat cultivar between plants grown under two conditions: control (0 mM NaCl) and salt treatment (200 mM NaCl). The current study revealed differentially expressed genes and various associated biological pathways involved in plant responses to salt stress between the two conditions in the root and shoot plant tissues, providing important insights into the molecular mechanisms underlying salt tolerance in wheat.

overall_design: Global profiling of wheat gene expression (Najran cultivar) was analysed under control and salt treatment in three biological replicates of root and shoot tissues.

first: Tahar

last: Taybi


title: Gene Editing of Hematopoietic Stem Cells Restores T Cell Response in a Familial Hemophagocytic Lymphohistiocytosis Model

city: Freiburg

first: Gianni

last: Monaco

address: Breisacher Straße

country: Germany

institute: Freiburg Medical Center

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


title: Keratin 19 binds and regulates cytoplasmic HNRNPK mRNA targets in triple-negative breast cancer

first: Markus

last: Hafner

state: MD

address: 50 South Drive

department: NIAMS

summary: This study identifies how a cytoskeletal protein can directly regulate gene expression by controlling the subcellular localization of RNA-binding proteins to support pathways involved in cancer progression.

overall_design: We used parental and K19 KO cell lines to study the impact of cytoskeleton in gene expression regulation.

first: Arwa

last: Fallatah

first: Dimitrios

middle: G

last: Anastasakis

first: Amirhossein

last: Manzourolajdad

first: Pooja

last: Sharma

first: Xiantao

last: Wang

first: Alexis

last: Jacob

first: Sarah

last: Alsharif

first: Ahmed

last: Elgerbi

first: Pierre

middle: A

last: Coulombe

first: Byung

last: Chung


title: Effects of cannabidiol on miRNA expression in naïve mice

city: Dublin

first: Mona

last: Heiland

address: 123 St. Stephen's green

country: Ireland

institute: RCSI

type: Non-coding RNA profiling by high throughput sequencing

summary: The non-psychotomimetic phytocannabinoid cannabidiol (CBD) has shown anticonvulsant effects in several seizure models in rats and mice. However, the mechanisms by which CBD exerts its antiepileptic effects are not fully elucidated. MicroRNAs (miRNA) are short non-coding RNAs which regulate protein expression. Dysregulation of miRNAs can contribute to the pathology of various diseases, including epilepsy. This could indicate that the modulation of miRNAs is a potential mechanism of action for the therapeutic effects of CBD. Therefore, we sequenced hippocampal samples from CBD or vehicle treated naive mice to see the effects of CBD on miRNA expression.

overall_design: Ventral hippocampi from CBD or vehicle treated naïve mice (n=10/group) were processed for small RNA sequencing.

first: Ngoc

middle: T

last: Nguyen

first: David

last: Henshall


title: Intriniscally disordered domain of transcription factor TCF-1 is requried for T cell developmental fidelity  [ATAC-Seq]

first: Naomi

last: Goldman

overall_design: ATAC-seq and RNA-seq in a DN3 TCF-1KO cell line and in primary developing T cells from Tcf7eGFP fl/fl Cre+ progenitors cultures on OP9-DLL1 cells.

first: Aditi

last: Chandra

first: Abhijeet

last: Patil

first: Ashley

last: Vanderbeck

first: Hai-Hui

last: Xue

first: Ivan

last: Maillard

first: Golnaz

last: Vahedi


title: Intriniscally disordered domain of transcription factor TCF-1 is requried for T cell developmental fidelity


title: Intriniscally disordered domain of transcription factor TCF-1 is requried for T cell developmental fidelity [RNA-Seq]


title: Dysfunction of 5-methylcytosine oxidases and the Polycomb protein RYBP leads to widespread DNA hypermethylation and cell transformation [WGBS]

city: Grand Rapids

first: Gerd

last: Pfeifer

state: MI

address: 333 Bostwick Ave. NE

institute: Van Andel Research Institute

summary: We inactivated RYBP, YAF2, TET1, TET2, and TET3 in bronchial cells, individually and in combination, and observed widespread DNA methylation by performing whole genome bisulfite sequencing.

overall_design: Comparative DNA methylation analysis of WGBS data in HBEC3 defective cell (Control cells, RKO, YKO, TET2 KO, TET3KO, TET2/TET3 DKO, TET123 TKO and TET123/RYBP QKO )

first: Wei

last: Cui

first: Zhijun

last: Huang


title: Dysfunction of 5-methylcytosine oxidases and the Polycomb protein RYBP leads to widespread DNA hypermethylation and cell transformation [ChIP-seq]

summary: We assessed the levels of the PRC1 Polycomb mark, RYBP, H2AK119Ub1, and the PRC2 Polycomb mark, H3K27me3, considering the extensive crosstalk between PRC1 and PRC2 activities.

overall_design: Chromatin immunoprecipitation sequencing (ChIP-seq) for RYBP, H2AK119ub1 and H3K27me3 in HBEC3 defective cell (Control cells, RYBP KO, YAF2 KO, RYBP/YAF2 DKO).


title: Dysfunction of 5-methylcytosine oxidases and the Polycomb protein RYBP leads to widespread DNA hypermethylation and cell transformation


title: IFNg-producing Tfh cells control the differentiation of lung resident memory B cells

city: Birmingham

first: Alexander

last: Rosenberg

state: AL

address: 1720 2nd Avenue South, Suite 276/11

institute: University of Alabama at Birmingham

summary: T follicular helper (Tfh) cells are a specific subset of CD4 T cells that localize in the B cell follicle and are essential for the Germinal Center (GC) response. Under polarizing environments, Tfh cells produce IFN-g. Beyond promoting class switching, the role played by IFN-g-producing Tfh cells remains largely unexplored. Using an influenza infection model, we show here that interactions with IFN-γ producing Tfh cells during the peak of the response skew the GC B cell response towards the memory differentiation pathway. Consequently, lung-memory B cells fail to differentiate without IFN-γ producing Tfh cells. Collectively, our results support a model in which temporary changes in cytokine production by Tfh cells dynamically control the outcome of the GC response.

overall_design: RNAseq was performed for the following (in triplicate for each sample type): 1. C57BL/6 mice were irradiated and reconstituted with a 50:50 mix of bone marrow from CD45.1+ B6 (WT) and CD45.2+ IfngR1-/- donors. Eight weeks later, reconstituted mice were infected with PR8 influenza virus, and WT and IfngR1-/- GC B cells were sorted from the lung-draining mediastinal lymph node on day 12 after infection; 2. C57BL/6 mice were infected with PR8, and GC B cells from the lung-draining mediastinal lymph node and CXCR3+ memory B cells from the lungs were sorted on day 30; and 3. C57BL/6 mice were infected with PR8 and CXCR3+, and CXCR3- GC B cells were sorted from the lung-draining mediastinal lymph node on day 12.

first: Nicole

last: Arroyo

first: Holly

last: Bachus

first: Amber

last: Papillion

first: Beatriz

last: León

first: André

last: Ballesteros-Tato


title: Dysfunction of 5-methylcytosine oxidases and the Polycomb protein RYBP leads to widespread DNA hypermethylation and cell transformation [RNA-seq]

summary:
To investigate the function of RYBP, YAF2, TET1, TET2, TET3 or the cooperative function in the human bronchial cells, we established defective clone cell lines in which each target gene has been knocked out by guide RNA
We then performed gene expression profiling analysis from mRNA-seq.

overall_design: Comparative gene expression profiling analysis of mRNA-seq data in HBEC3 defective cell (Control cells, RYBP KO, YAF2 KO, RYBP/YAF2 DKO , TET1/2/3 TKO and TET1/2/3/RYBP QKO ).


title: The effect of GLP-1 receptor agonist Lixisenatide on experimental diabetic retinopathy

status: Public on Jul 05 2023

city: Frankfurt

first: Paulus

last: Wohlfart

state: Hessen

address: Industriepark Höchst, K703

department: MSAT M&I

institute: Sanofi Aventis Deutschland GmbH

summary: Glucagon-like peptide-1 receptor agonists (GLP-1 RAs) are effective treatments for type 2 diabetes, effectively lowering glucose without weight gain and with low risk for hypoglycemia. However, their influence on the retinal neurovascular unit remains unclear. In this study we analyzed the effects of the GLP-1 RA lixisenatide in a rat model of type-1 like diabetic retinopathy.  Vasculo- and neuroprotective effects were assessed in experimental diabetic retinopathy and high glucose-cultivated C. elegans, respectively. In STZ-diabetic Wistar rats acellular capillaries and pericytes (quantitative retinal morphometry), neuroretinal function (mfERG), macroglia (GFAP western blot) and microglia (immunohistochemistry) quantification, methylglyoxal (LC-MS/MS) and retinal gene expressions (RNA-sequencing) were determined. The antioxidant properties of lixisenatide were tested in C. elegans. Lixisenatide had no effect on glucose metabolism. Lixisenatide preserved the retinal vasculature and neuroretinal function. The macro- and microglial activation was mitigated. Lixisenatide normalized some gene expression changes in diabetic animals to control levels. Ets2 was identified as a regulator of inflammatory genes. In C. elegans lixisenatide showed the antioxidative property. In summary, these data suggest that lixisenatide has a protective effect on the diabetic retina, most likely due to a combination of neuroprotective, anti-inflammatory and antioxidative effects of lixisenatide on the neurovascular unit.

overall_design: First to investigate gene expression changes in whole retinae from rat with hyperglycemia but not hyperinsulinemia. And second, to identify differences and underlying signalling pathways induced by the treatment with the GLP1 receptor agonist Lixisenatide

first: Kuebra

last: Oezer

first: Matthias

last: Kolibabka

first: Johann

last: Gassenhuber

first: Hans-Peter

last: Hammes


title: Chromatin regulation of transcriptional enhancers and cell fate by the Sotos syndrome gene NSD1 [TT-Seq]

city: New York

first: Zhen

last: Sun

address: 408 E 69th street, Z619

department: Human Oncology and Pathogenesis Program

institute: Memorial Sloan Kettering Cancer Center

summary: Nuclear-receptor-binding SET-domain protein 1 (NSD1), a methyltransferase that catalyzes H3K36me2, is essential for mammalian development and frequently dysregulated in diseases, including Sotos syndrome. Despite impacts of H3K36me2 on H3K27me3 and DNA methylation, a direct role of NSD1 in transcriptional regulation remains largely unknown. Here, we show that NSD1 and H3K36me2 are enriched at cis-regulatory elements, particularly enhancers. NSD1 enhancer association is conferred by a tandem quadruple PHD-PWWP module, which recognizes p300-catalyzed H3K18ac. By combining acute NSD1 depletion with time-resolved epigenomic and nascent transcriptomic analyses, we demonstrate that NSD1 promotes enhancer-dependent gene transcription by facilitating RNA polymerase II pause release. Notably, NSD1 can act as a transcriptional coactivator independent of its catalytic activity. Moreover, NSD1 enables activation of developmental transcriptional programs associated with Sotos syndrome pathophysiology and controls ESC multilineage differentiation. Collectively, we have identified NSD1 as an enhancer-acting transcriptional coactivator that contributes to cell fate transition and Sotos syndrome development.

overall_design: TT-seq was performed with NSD1-dTAG mESCs after 0, 1, and 6h of dTAG-13 treatment.

first: Kristian

last: Helin


title: Endocrine treatment potentiates antigen presentation and in combination with SMAC mimetics enhances an anti-tumor immune response in hormone receptor positive breast cancer (RELA ChIP)

city: Boston

first: Yingtian

last: Xie

institute: DANA FARBER CANCER INSTITUTE INC

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for RELA in MCF7 cells grown in HD and E2, with and without stimulation of IFNg 10ng/mL for 1h.

first: Francisco

last: Hermida Prado

first: Rinath

last: Jeselsohn


title: First-line durvalumab and tremelimumab with chemotherapy in RAS-mutated metastatic colorectal cancer: a phase 1b/2 trial

city: DIJON

first: François

last: Ghiringhelli

address: 1 rue du Professeur Marion

country: France

institute: UMR INSERM 1231

overall_design: Refer to individual Series


title: Microglial MHC-I induction with aging and Alzheimer’s is conserved in mouse models and humans

city: Oklahoma City

first: Willard

last: Freeman

address: 825 NE 13th St

institute: Oklahoma Medical Research Foundation

summary: Major Histocompatibility Complex I (MHC-I) CNS cellular localization and function is still being determined after previously being thought to be absent from the brain. MHC-I expression has been reported to increase with brain aging in mouse, rat, and human whole tissue analyses. Neuronal MHC-I has been proposed to regulate developmental synapse elimination and tau pathology in Alzheimer’s disease (AD). Here we report that across newly generated and publicly available ribosomal profiling, cell sorting, and single-cell data, microglia were the primary source of classical and non-classical MHC-I in mice and humans. Translating Ribosome Affinity Purification (TRAP)-qPCR analysis of 3-6 and 18-22 month old (m.o.) mice revealed significant age-related microglial induction of MHC-I pathway genes B2m, H2-D1, H2-K1, H2-M3, H2-Q6, and Tap1 but not in astrocytes and neurons. Across a timecourse (12-23 m.o.), microglial MHC-I gradually increased until 21 m.o. and then accelerated. MHC-I protein was also enriched in microglia and increased with aging. Microglial expression, and absence in astrocytes and neurons, of MHC-I binding Leukocyte Immunoglobulin-like (Lilrs) and Paired immunoglobin-like type 2 (Pilrs) receptor families could enable cell-autonomous MHC-I signaling and increased with aging in mice and humans. Increased microglial MHC-I, Lilrs, and Pilrs were observed in AD mouse models and human AD data is evident across numerous mouse models, methods, and studies. MHC-I expression correlated with p16INK4A, suggesting an association with cellular senescence. Conserved induction of MHC-I, Lilrs, and Pilrs with aging and AD opens the possibility of cell-autonomous MHC-I signaling to regulate microglial reactivation with aging and neurodegeneration.

overall_design: RNA-Seq and qPCR of old and young mice to compare MHC-I expression with human data

first: Collyn

last: Kellogg

first: Kevin

last: Pham

first: Adeline

middle: H

last: Machalinski

first: Hunter

middle: L

last: Porter

first: Harris

middle: E

last: Blankenship

first: Kyla

last: Tooley

first: Michael

middle: B

last: Stout

first: Heather

last: Rice

first: Amanda

last: Sharpe

last: Beckstead

first: Ana

last: Chucair-Elliott

first: Sarah

last: Ocañas


title: AoMedA has complex regulatory relationship with AoBrlA, AoAbaA and AoWetA in conidiation, trap formation, and secondary metabolism in nematode-trapping fungus Arthrobotrys oligospora

city: Kunming

last: Bai

address: No. 2, Cuihu North Road

institute: Yunnan University

summary: Transcriptomic analysis revealed that differentially expressed genes in the absence of AomedA and AoWetA were involved in mycelial growth, development process,conidiation, cell nucleus, trap formation, peroxisome, endocytosis and lipid metabolism.

overall_design: WT (represented by AO and AOB) and ΔAomedA (represented by MedA) and  ΔAowetA (represented by WetA) strains grown on cellophane-overlaid CMY plates at 28°C for 5  days, three replicates were used for each sample. After collecting the samples, RNA was extracted with the AxyPrep Multisource RNA Miniprep Kit (Axygen, Jiangsu, China), and then sent to Majorbio Bio-Pharm Technology Co., Ltd. (Shanghai, China) to construct the transcriptome of WT and ΔAomedA and ΔAowetA strains. The final cDNA library was sequenced on an Illumina Novaseq 4000 platform.

first: Meihua

last: Xie

first: Qianqian

last: Liu

first: Yingmei

last: Zhu

first: Xuewei

last: Yang

first: Keqin

last: Zhang

first: Jinkui


title: SARS-CoV-2 Niches in Human Placenta Revealed by Spatial Transcriptomics

city: Houston

last: Jochum

address: 1 Baylor Plaza

department: Obstetrics and Gynecology

institute: Baylor College of Medicine

summary: Diverse placental functions are compartmentalized to separate maternal-fetal antigens and restrict vertical transmission of pathogens. We hypothesized a high-resolution map, with single-cell and spatial resolution, would identify previously undetectable microbe immune microenvironments. To test this hypothesis, we utilized Visium Spatial Transcriptomics paired with H&E staining to generate 17,927 spatial transcriptomes and integrated these data with 273,944 published placenta single-cell and single-nuclei transcriptomes to generate a term placenta atlas of the maternal decidua, fetal chorionic villi, and chorioamniotic membranes. Comparisons of healthy control placentae (n=4), SARS-CoV-2 asymptomatic (n=4; no COVID-19 symptoms), and symptomatic (n=5; pneumonia, respiratory failure) placentae identified distinct thresholds of SARS-CoV-2 detection including (i) no detectable virus, (ii) sparse virus (limit of detection determined to be 1/7,000 cells), and (iii) highly positive SARS-CoV-2 in syncytiotrophoblasts, rigorously cross-validated by RT-qPCR, RNAscope, and immunohistochemistry. High levels of SARS-CoV-2 in placentae were associated with histiocytic intervillositis and perivillous fibrin deposition. Spatial transcriptomics revealed that this led to a significant shift from anti-inflammatory M2 macrophage populations toward intermediate pro-inflammatory M1 macrophages. In conclusion, we identified immune microenvironments representing host-pathogen battlegrounds where SARS-CoV-2 detection in the placenta was not dependent on maternal symptoms. Additional studies utilizing this approach are warranted, especially in the placenta.

sample_organism: Severe acute respiratory syndrome coronavirus 2

overall_design: Spatial transcriptomics. Human placentae from distinct regions including the chorionic villi, decidua, and chorioamniotic membranes, or cross-sections from the parenchyma, were fresh-frozen in optimal cutting temperature solution (FF-OCT). Blocks were cryosectioned and H&E stained directly on 10X Genomics Visium Gene Expression slides (v2) and imaged using a Nikon Eclipse SE Ni microscope using a Nikon DS-Ri1 camera, Nikon Plan Apo objective at 10x magnification (0.45 aperture, 0.91 μm/pixel resolution). Tissues were permeabilized and RNA was subject to spatial transcriptomics library preparation including poly(dT) reverse transcription. Libraries were sequenced on the Illumina NovaSeq S4 platform with 2% PhiX.    Single-cell and spatial transcriptomics analyses. Reads were demultiplexed and aligned to a custom human (GRCh38) and SARS-CoV-2 reference genome (NC_045512.2) using SpaceRanger (v1.3.0) and custom bash scripts. Downstream analyses were done using the package Seurat (v4.0.3) in rStudio (v4.1.1). Counts matrices were filtered iteratively to exclude low-quality transcriptomes and clusters defined by quality control metrics (e.g. mitochondrial or hemoglobin gene expression). Spatial transcriptomes were normalized and scaled using a negative binomial model (SCTransform) and the top 3,000 most variable transcripts were used for to principal component analysis dimension reduction. The first 30 PCA dimensions were used for K-nearest neighbors’ analysis, clustered using a Louvain algorithm with the default resolution parameter (0.6), and visualized by unique manifold approximation and projection (UMAP) in two dimensions. Significantly upregulated transcripts were manually examined and compared to the term placenta atlas in addition to using EnrichR(58), PlacentaCellEnrich(110), and the Human Protein Atlas(111) to annotate clusters. Pseudotime trajectory analysis was done using Monocle3 (v1.0.0) utilizing gene sets and starting points denoted in the text. Spatial and scRNA-seq datasets were integrated using reciprocal PCA of 3,000 reference transcripts prior to clustering.

first: Kjersti

last: Aagaard

first: Enrico

last: Barrozo


title: Epigenome-wide association study on peripheral blood DNA of bronchopulmonary dysplasia in preterm infants

city: Research Triangle Park

first: Xuting

last: Wang

address: 111 TW Alexander Dr

institute: NIEHS/NIH

summary: Bronchopulmonary dysplasia (BPD) is a lung disease in premature infants characterized by impaired pulmonary development which persists into later life. While advances in neonatal care have improved survival rates of premature infants, cases of BPD haves been increased. Therapeutic options are limited for prevention and treatment. This study was designed to explore the relationship between gestational age (GA), birth weight and estímate blood cell-type composition in premature infants and to elucidate early epigenetic biomarkers associated with BPD. Peripheral blood DNA (at days 14 and 28) from preterm neonates that went on to develop BPD (n = 14) or not (nonBPD, n = 93)  was applied to Illumina EPIC methylation arrays. Using DNA methylation analysis of cord blood DNA, we investigated association of GA and birth weight with the estimated distribution of cord blood cell types, particularly the nucleated red blood cell (NRBC) in a pilot-size cohort of preterm infants with or without BPD. We describe changes in methylation-based estimates of blood cell-type composition in relation to GA and birth weight. After adjusting for covariates (GA, birth weight, cell type proportions, etc.) we identify differentially methylated CpGs and genes associated with BPD at different time points.

overall_design: Peripheral blood DNA at days 14 and 28 from preterm neonates that went on to develop BPD (n = 12) or not (nonBPD, n = 84)  was applied to Illumina EPIC methylation arrays. We identify differentially methylated CpGs associated with BPD.

first: Douglas

last: Bell

first: Michelle

last: Campbell

first: Hye-Youn

last: Cho


title: A Th1-like CD4+ T-cell Cluster that Predicts Disease-free Survival in Early-Stage Lung Cancer

city: Hidaka-City, Saitama

first: Satoshi

last: Yamasaki

department: Department of Respiratory Medicine

country: Japan

institute: Saitama Medical University

summary: We performed scRNAseq of T cells in pre- and postoperative PBMCs and TILs of lung cancer patients, and analyzed gene expression pattern, TCR clonotypes, and numerical correlations among them.

overall_design:
Investigating how differences in gene expression and TCR repatoires of T cells in PBMCs and TILs affect the postoperative disease-free survival
Please note that the processed data generated from both scRNAseq and Feature Barcoding(ADT) samples are linked to the corresponding scRNA-seq sample records.

first: Akitoshi

last: Yanagihara

first: Kosuke

last: Hashimoto

first: Ryo

last: Taguchi

first: Tetsuya

last: Umesaki

first: Hisao

last: Imai

first: Kyoichi

last: Kaira

first: Hiroyuki

last: Nitanda

first: Hirozo

last: Sakaguchi

first: Hironori

last: Ishida

first: Kunihiko

last: Kobayashi

first: Katsuhisa

last: Horimoto

first: Hiroshi

last: Kagamu


title: De-differentiation and Proliferation of Artery Endothelial Cells Drive Coronary Collateral Development

city: Bangalore

first: Soumyashree

last: Das

address: Rajiv gandhi nagar

country: India

institute: National Centre for Biological Sciences

summary: We have earlier observed that upon myocardial infarction, pre-existing coronary arteries give rise to new collateral arteries via Artery Reassembly (AR) in neonatal mice, but not in adults. To understand this age dependent phenomenon, Gja5 (Cx40), an established artery endothelial cell marker, was used to lineage trace artery cells with TdTomato expression. For the neonatal mice, Tamoxifen was induced at postnatal (P)0, Myocardial Infarction (MI)/sham was performed at P2 and hearts were harvested at either P4 or P5. Whereas, for the adults, mice aged ~2months were used. Tamoxifen was induced at Day (D)0, MI/sham was performed at D2 and hearts were harvested at either D9 or D11. Ventricles from hearts were digested and FACS sorted for DAPI-, Ter199- and TdTomato+ cells.

overall_design: Single Cell RNA sequencing data was obtained from neonatal and adult mice heart ventricles using 10X genomics, and further processed on Cell Ranger.

first: Karen

middle: M

last: Gonzalez

first: Hanjay

last: Wang

first: Pam

last: Rios Coronado

first: Joseph

middle: Y

last: Woo

first: Kristy

last: Red-Horse


title: Chromatin regulation of transcriptional enhancers and cell fate by the Sotos syndrome gene NSD1 [SLAM-seq]

first: Zhen

last: Sun

summary: Nuclear-receptor-binding SET-domain protein 1 (NSD1), a methyltransferase that catalyzes H3K36me2, is essential for mammalian development and frequently dysregulated in diseases, including a spectrum of cancers and the genetic disorder Sotos syndrome. Despite its function in modulating the chromatin landscape, a direct role of NSD1 in transcriptional regulation remains largely unknown. Here, we show that NSD1 and H3K36me2 are enriched at cis-regulatory elements, particularly enhancers, in a cell type-specific manner. NSD1 enhancer association is conferred by its tandem quadruple PHD-PWWP domain, which is a hotspot for Sotos syndrome missense mutations. By combining acute NSD1 depletion with temporally resolved epigenomic and nascent transcriptomic analysis, we demonstrate that NSD1/H3K36me2 plays a critical role in facilitating enhancer-dependent gene transcription by promoting promoter pause release of RNA polymerase II. Moreover, NSD1 regulates ESC multilineage differentiation through facilitating transcriptional activation of critical developmental programs implicated in Sotos syndrome pathogenesis. Collectively, we have identified NSD1 as a novel enhancer-acting transcriptional coactivator and provided mechanistic insights into its contribution to cell fate transition and association with a human developmental disorder.

overall_design: SLAM-seq was performed with NSD1-dTAG mESCs after 0, 6, and 24h of dTAG-13 treatment (batch 1) and after 0 and 1h of dTAG-13 treatment (batch 2). SLAM-seq was performed with NSD1-dTAG mESCs expressing WT or N1751Q catalytically-dead FL NSD1 after 0 and 6h of dTAG-13 treatment (batch 3) and after 0 and 1h of dTAG-13 treatment (batch 4). SLAM-seq was also performed with SETD2-dTAG mESCs after 0, 6 and 24h of dTAG-13 treatment.

first: Kristian

last: Helin


title: First-line durvalumab and tremelimumab with chemotherapy in RAS-mutated metastatic colorectal cancer: a phase 1b/2 trial [scRNA-Seq]

first: François

last: Ghiringhelli

summary: While patients with microsatellite instable, metastatic colorectal cancer (CRC) benefit from immune checkpoint blockade, chemotherapy with targeted therapies remains the only therapeutic option for microsatellite stable (MSS) tumors. The single arm, phase IB/II MEDITREME trial evaluates safety and efficacy of durvalumab plus tremelimumab in combination with mFOLFOX6 chemotherapy in first line, in 57 patients with RAS-mutant unresectable mCRC. Safety was primary objective of phase IB and no safety issue was observed. The primary objective of phase II was efficacy in terms of 3-month PFS in MSS patients, which was met with a 3 month PFS of 90.7% [95% CI: 79.2-96%]. For secondary objective, response rate was 64.5%, median PFS was 8.2 months [95% CI: 5.9–8.6] and overall survival was not reached in MSS patients.  Higher tumor mutational burden and a lower degree of genomic instability in responder patients. Integrated transcriptomic analysis underlined that high immune signature, and low epithelio-mesenchymal transition were associated with better outcome. Immunomonitoring showed induction of neoantigen and NY-ESO1 and TERT blood tumor specific T cell response associated with better PFS. The combination of durvalumab-tremelimumab with mFOLFOX6 was tolerable with promising clinical activity in MSS mCRC.

overall_design: We performed gene expression profiling analysis using RNA-seq data from 34 patients splited into responders and non-responders

last: Ghiringelli

first: Marion

last: Thibaudin

first: Caroline

last: Truntzer

first: Elise

last: Ballot


title: Chromatin regulation of transcriptional enhancers and cell fate by the Sotos syndrome gene NSD1 [RRBS]

overall_design: RRBS was performed with NSD1-dTAG mESCs after 0, 6, and 24h of 500 nM dTAG-13 treatment and with wild type and Nsd1 knockout mESCs.


title: Chromatin regulation of transcriptional enhancers and cell fate by the Sotos syndrome gene NSD1 [CUT&RUN]

overall_design: CUT&RUN for indicated proteins and histone PTMs, including NSD1 and H3K36me2, was performed with mESCs, EpiSCs and embryoid bodies (EBs) in the steady state and/or with NSD1 manipulations (degradation in NSD1-dTAG cells, knockout or rescue). CUT&RUN for SETD2 and H3K36me3 was performed in SETD2-dTAG mESCs in the presence or absence of dTAG-13.


title: Chromatin regulation of transcriptional enhancers and cell fate by the Sotos syndrome gene NSD1 [ATAC-seq]

overall_design: ATAC-seq was performed with NSD1-dTAG mESCs after 0-24h of 500 nM dTAG-13 treatment and in embryoid bodies (EBs) differentiated for 6 days from NSD1-dTAG mESCs in the presence or absence of 500 nM dTAG-13.


title: Chromatin regulation of transcriptional enhancers and cell fate by the Sotos syndrome gene NSD1 [ChIP-seq]

overall_design: ChIP-seq for Pol II NTD (total Pol II), Pol II S5P, Pol II S2P, NELF-C, SPT5, PAF1 and SPT6 was performed with NSD1-dTAG mESCs with or without 6h of 500 nM dTAG-13 treatment. ChIP-seq for H3K18ac was performed with NSD1-dTAG mESCs and EpiSCs. ChIP-seq for H3K27ac was performed with NSD1-dTAG mESCs and embryoid bodies (EBs) differentiated for 6 days from NSD1-dTAG mESCs in the presence or absence of 500 nM dTAG-13.


title: Transcriptional profiling of mutants of FdC2 (At1g32550) in Arabidopsis thaliana

status: Public on Jul 04 2023

city: London

first: Guy

last: Hanke

address: Mile End

department: School of Biological and Behavioural Sciences

institute: Queen Mary University of London

summary:
Two RNAi knock-down lines for At1g32550 were generated and protein content confirmed by western blotting. Individuals of wild type, one severe (line 8) and one mild (line 1) knock-down line were grown to four weeks old and then total RNA extracted from 2 h into the light period.
Total RNA was extracted and then sent to OakLabs for hybridization and analaysis

overall_design: The Agilent Arabidopsis chip was enriched for chloroplast transcripts in a custom array

first: Tatjana

last: Goss


title: IRX2 regulates angiotensin II-induced cardiac fibrosis by transcriptionally activating Egr1 in mice [ChIP-Seq]

city: Wuhan

first: Zhen-Guo

last: Ma

state: Hubei

address: No.238 Jiefang Road

institute: Renmin Hospital fo Wuhan University

summary: Cardiac fibrosis is a common feature of chronic heart failure. Iroquois homeobox (IRX) family of transcription factors plays important roles in heart development; however, the role of IRX2 in cardiac fibrosis has not been clarified. Here we report that IRX2 expression is significantly upregulated in the fibrotic hearts. Increased IRX2 expression is mainly derived from cardiac fibroblast (CF) during the angiotensin II (Ang II)-induced fibrotic response. Using two CF-specific Irx2-knockout mouse models, we show that deletion of Irx2 in CFs protect against pathological fibrotic remodelling and improve cardiac function in mice. In contrast, Irx2 gain of function in CFs exaggerate fibrotic remodelling. Mechanistically, we find that IRX2 directly binds to the promoter of the early growth response factor 1 (EGR1) and subsequently initiates the transcription of several fibrosis-related genes. Our study provides the evidence that IRX2 regulates the EGR1 pathway upon Ang II stimulation and drives cardiac fibrosis.

overall_design: Human CFs were infected with an adenovirus carrying IRX2 for 4 h and then subjected to Ang II stimulation for 24 h. ChIP-Seq was performed with an anti-IRX2 antibody (#PCRP-IRX2-1B7, Developmental Studies Hybridoma Bank, Houston, USA; or #PCRP-IRX2-1C1, Developmental Studies Hybridoma Bank).


title: Chromatin regulation of transcriptional enhancers and cell fate by the Sotos syndrome gene NSD1 [RNA-seq]

overall_design: RNA-seq was performed with wild-type mESCs nontreated or treated with 500 nM dTAG-13 for 72h. RNA-seq was also performed with nontreated NSD1-dTAG mESCs and embryoid bodies (EBs) differentiated for 6 days from NSD1-dTAG mESCs in the presence or absence of 500 nM dTAG-13.


title: IRX2 regulates angiotensin II-induced cardiac fibrosis by transcriptionally activating EGR1 in mice [RNA-seq]

overall_design: CFs were infected with an adenovirus carrying Irx2 or Gfp and then subjected to Ang II stimulation for 24 h. After that, RNA-seq was performed.


title: IRX2 regulates angiotensin II-induced cardiac fibrosis by transcriptionally activating Egr1 in mice

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

overall_design: Refer to individual Series


title: Oxidized phospholipid oxPAPC alters regulatory T cell differentiation and decreases their protective function in atherosclerosis

city: Nashville

first: Amy

last: Major

address: 1161 21st Ave S

institute: Vanderbilt University Medical Center

summary: Background: Regulatory T cells (Tregs) are protective in atherosclerosis but reduced during disease progression due to cell death and loss of stability. However, the mechanisms of Treg dysfunction remain unknown. Oxidized phospholipids (oxPLs) are abundant in atherosclerosis and can activate innate immune cells, but little is known regarding their impact on T cells. Given Treg loss during atherosclerosis progression and oxPL levels in the plaque microenvironment, we investigated whether oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (oxPAPC), an oxPL associated with atherosclerotic plaques, alters Treg differentiation and function. Methods: CD4+ T cells were polarized to Treg, Th1, and Th17 cells with or without oxPAPC and assessed by flow cytometry. Gene expression in oxPAPC-treated Tregs was analyzed by bulk RNA sequencing. Functional studies of oxPAPC-induced Tregs were performed by co-culturing Tregs with CTV-labeled cells in vitro, and by adoptively transferring Tregs to hyperlipidemic Ldlr-/- mice to measure atherosclerosis progression. Results: Compared to controls, oxPAPC-treated Tregs were less viable, but surviving cells expressed higher levels of the Th1-associated markers T-bet, CXCR3, and IFN-. Th1 and Th17 skewing cultures were unaltered by oxPAPC. IFN- is linked to Treg instability, thus Treg polarization experiments were repeated using Ifngr1-/- CD4+ T cells. IFNR1 deficiency did not improve cell viability in oxPAPC-treated Tregs, however, T-bet and IFN- expression was not increased in surviving cells suggesting a role for IFN-signaling. OxPAPC-treated Tregs were less suppressive in vitro, and adoptive transfer studies in hyperlipidemic Ldlr-/- mice showed that oxPAPC-induced Tregs possessed altered tissue homing and were insufficient to inhibit atherosclerosis progression.  Conclusions: OxPAPC elicits Treg-specific changes altering Treg differentiation and inducing a Th1-like phenotype in surviving cells partially through IFN- signaling. This is biologically relevant as oxPAPC-treated Tregs do not reduce atherosclerosis progression in Ldlr-/- mice. This study supports the role for oxPLs in negatively impacting Treg differentiation and atheroprotective function.

overall_design: To investigate oxPAPC-induced alterations in Tregs, we polarized Tregs from mouse naïve CD4+ T cells for three days in vitro in the presence or absence of oxPAPC. We then analyzed differential gene expression using data obtained through RNA-seq. We compared control Tregs (4 replicates) to oxPAPC-treated Tregs (4 replicates).

first: Brenna

last: Appleton

first: Sydney

last: Palmer

first: Harrison

middle: P

last: Smith

first: Lilly

last: Stephens

middle: S


title: Combinatorial immunotherapy with agonistic CD40 activates dendritic cells to express IL-12 and overcome PD-1 resistance II

city: New Haven

first: Miller-Jensen

last: Lab

address: 55 Prospect St

department: Biomedical Engineering

institute: Yale University

summary: Checkpoint inhibitors have revolutionized cancer treatment, but resistance remains a significant clinical challenge. Myeloid cells within the tumor microenvironment can modulate checkpoint resistance by either supporting or suppressing adaptive immune responses. Using an anti-PD-1 resistant mouse melanoma model, we show that targeting the myeloid compartment via CD40 activation and CSF1R blockade in combination with anti-PD-1 results in complete tumor regression in a majority of mice. This triple therapy combination is primarily CD40 agonist-driven in the first 24 hours post-therapy and shows a similar systemic cytokine profile in human patients as in mice. Functional single-cell cytokine secretion profiling of dendritic cells (DCs) using a novel microwell assay identified a CCL22+CCL5+ IL-12-secreting DC subset as important early-stage effectors of triple therapy. CD4+ and CD8+ T cells are both critical effectors of treatment, and systems analysis of single-cell RNA-sequencing data supports a role for DC-secreted IL-12 in priming T cell activation and recruitment. Finally, we show that treatment with a novel IL-12 mRNA therapeutic alone is sufficient to overcome PD-1 resistance and cause tumor regression. Overall, we conclude that combining myeloid-based innate immune activation and enhancement of adaptive immunity is a viable strategy to overcome anti-PD-1 resistance.

overall_design: C57BL6/J mice were injected with 0.5x10^6 YUMMER1.7 cells on day 0, and then were either left untreated or treated with the combination of anti-PD-1, CSF1R blockade, and agonistic CD40 (i.e., TTx) by i.p. injection on day 7. To assess cellular and molecular heterogeneity in the tumor in response to treatment, we performed scTCR-seq on cells isolated from untreated and TTx-treated tumors 24 hours after treatment on day 8. One tumor was used per treatment condition.

first: Irina

last: Krkbaeva

first: Kate

last: Bridges

first: William

last: Damsky

first: Gabriela

last: Pizzurro

first: Amanda

middle: F

last: Alexander

first: Meaghan

middle: K

last: McGeary

first: Viswanathan

last: Muthusamy

first: James

last: Eyles

first: Nadia

last: Luheshi

first: Noel

last: Turner

first: Sarah

last: Weiss

first: Kelly

last: Olino

first: Susan

last: Kaech

first: Harriet

last: Kluger

first: Kathryn

last: Miller-Jensen

first: Marcus

last: Bosenberg


title: The histone acetyl transferases CBP and p300 regulate stress response pathways in synovial fibroblasts

city: Bern

first: Kerstin

last: Klein

state: Bern

address: Murtenstrasse 28

department: Department of Rheumatology and Immunology

country: Switzerland

institute: University Hospital Bern

summary: The activation of stress response pathways in synovial fibroblasts (SF) is a hallmark of rheumatoid arthritis (RA). CBP and p300 are two highly homologous histone acetyl transferases and writers of activating histone 3 lysine 27 acetylation (H3K27ac) marks. We investigated individual functions of CBP and p300 using a silencing strategy, followed by RNA-sequencing, and pathway enrichment analysis. We have selected stress-related pathways for a further in-depth investigation of individual functions of CBP and p300 in SF. Pathway enrichment analysis pointed to a profound role of CBP and/ or p300 in regulating stress response-related gene expression, with an enrichment of pathways associated with oxidative stress, hypoxia, autophagy and proteasome function. We silenced CBP or p300, and performed confirmatory experiments on transcriptome, protein and functional levels. We have identified some overlap of CBP and p300 target genes in the oxidative stress response pathway, however, with several genes being regulated in opposite directions. The majority of stress response genes was regulated by p300, with a specific function of p300 in regulating hypoxia response genes and genes encoding proteasome subunits. Silencing of p300 suppressed proteasome enzymatic activities. CBP and p300 regulated autophagy on transcriptome and functional levels. Whereas CBP was indispensable for autophagy synthesis, silencing of p300 affected late-stage autophagy. In line with impaired autophagy and proteasome function, poly-ubiquitinated proteins accumulated after silencing of p300.

overall_design: To study individual functions of CBP and p300 in RA synovial fibroblasts, we silenced their expression in synovial fibroblasts using transfection of gapmeRs. 24h after silenecing, cells were stimulated with TNF or left untreated. Cells were harvested 24h after stimulation.


title: Interference with the HNF4-dependent gene regulatory network diminishes ER stress in hepatocytes

city: Iowa City

first: David

middle: Thomas

last: Rutkowski

state: IA

address: 51 Newton Rd.

department: Anatomy and Cell Biology

institute: University of Iowa Carver College of Medicine

summary: In all eukaryotic cell types, the unfolded protein response (UPR) upregulates factors that promote protein folding and misfolded protein clearance to help alleviate endoplasmic reticulum (ER) stress. Yet ER stress in the liver is uniquely accompanied by the suppression of metabolic genes, the coordination and purpose of which is largely unknown. Here, we combined in silico machine learning, in vivo liver-specific deletion of the master regulator of hepatocyte differentiation HNF4α, and in vitro manipulation of hepatocyte differentiation state to determine how the UPR regulates hepatocyte identity, and toward what end. Machine learning identified a cluster of correlated genes that were profoundly suppressed by persistent ER stress in the liver. These genes, which encode diverse functions including metabolism, coagulation, drug detoxification, and bile synthesis, are likely targets of the master regulator of hepatocyte differentiation HNF4α. The response of these genes to ER stress was phenocopied by liver-specific deletion of HNF4α. Strikingly, while deletion of HNF4α exacerbated liver injury in response to an ER stress challenge, it also diminished UPR activation and partially preserved ER ultrastructure, suggesting attenuated ER stress. Conversely, pharmacological maintenance of hepatocyte identity in vitro enhanced sensitivity to stress. Together, our findings suggest that the UPR regulates hepatocyte identity through HNF4α to protect ER homeostasis even at the expense of liver function.

overall_design: Hnf4a floxed (fl/fl) male mice of variable age (8 - 12 weeks old) were injected with AAV-TBG-iCre or AAV-TBG-eGFP (2.5 x 10^11 GC/mice). A week later, Hnf4a -/- or fl/fl mice were injected intraperitoneally with 0.5 mg/kg tunicamycin (TM) or vehicle control (DMSO). Each experimental group has 3-4 mice. 14h post tunicamycin injection, mice were sacrificed, and total RNA was prepared from resected livers.

first: Anit

last: Shah

first: Thomas


title: Extracellular Vesicles (EVs) derived from Hemangioma stem cells (HemSC)

city: Shanghai

first: Enhao

last: Lu

address: 826 Zhangheng Road

institute: Fudan University

summary: In order to investigate the function and mechanism of HemSC-EVs, we performed RNA-seq of HemSC-EVs which were extracted from cell culture supernatant of HemSC.

overall_design: HemSC-EVs were extracted form cell culture supernatant of HemSC by ultra-centrifuge method.  Then RNA-seq was conducted for HemSC-EVs

first: Xi

last: Yang

first: Xianyi

last: Sha

first: Xiaoxi

last: Lin


title: An adipokine derived from subcutaneous adipose tissue

city: Daejeon

first: Ju Hee

last: Lee

address: Munhwa-ro 266

institute: Chungnam National University

summary: Adipokines are proteins secreted by the adipose tissue and are associated with obesity-related metabolic disorders; however, most studies have focused on adipokines expressed by visceral adipose tissue. This study aimed to identify the adipokine potentially derived from subcutaneous adipose tissue and its clinical significance.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for visceral adipose tissue and subcutaneoua adipose tissue in human.

first: Ju

first: Ji

last: Kim

first: Seok

first: Sun

middle: J

last: Han