title: Apparent RNA bridging between PRC2 and chromatin is an artifact of non-specific chromatin precipitation upon RNA degradation

city: London

last: Hall Hickman

state: Please select

address: Flat A, 75 Dunsmure Road, Dunsmure Road

department: Cancer Institute

institute: University College London

summary: Polycomb repressive complex 2 (PRC2) modifies chromatin to maintain repression of genes specific for other cell lineages and is essential for development. In vitro, RNA inhibits PRC2 catalytic activity and interaction with nucleosomes but the effect of RNA on PRC2 in cells has been unclear, with studies concluding that RNA either antagonises or promotes the association of PRC2 with chromatin. Suggesting the existence of an RNA bridge that connects PRC2 to chromatin, treatment of formaldehyde-crosslinked, sonicated nuclear extracts with RNase A reduces co-precipitation of polycomb target sites with PRC2. Here, we show that this apparent loss of PRC2 chromatin association after RNase A treatment is due to non-specific chromatin precipitation. RNA degradation causes chromatin to precipitate out of solution, thereby reducing the enrichment of specific DNA sequences in chromatin immunoprecipitation reactions. Maintaining chromatin solubility by the addition of poly-L-glutamic acid (PGA) rescues detection of PRC2 chromatin association upon RNA degradation.  These findings undermine support for the model that RNA bridges PRC2 and chromatin in cells.

overall_design: Chromatin Immunoprecipitation and DNA-sequencing (ChIP-seq) for histone mark H3K27me3 and chromatin binding proteins RPB1 and SUZ12, in mESCs and following addition of RNaseA and or poly-glutamic acid.

first: Alexander

first: Richard

last: Jenner


title: Targeted demethylation and activation of NLRC5 augments cancer immunogenicity through MHC class I

status: Public on Feb 22 2024

city: Sapporo

first: Koichi

middle: S.

last: Kobayashi

state: Hokkaido

address: Kita 15, Nishi 7, Kita-ku

department: Graduate school of medicine

country: Japan

institute: Hokkaido University

type: Expression profiling by high throughput sequencing

summary: Impaired expression of MHC class I constitutes a major mechanism of immune evasion of cancers, leading to poor prognosis and resistance to checkpoint blockade therapies. Existing drugs for MHC class I have limited applicability due to severe side effects. Here we show a novel approach of robust and specific induction of MHC class I by targeting an MHC class I transactivator (CITA), NLRC5, using a CRISPR/Cas9 based gene-specific targeted demethylaion (TDM) system and targeted demethylation and activation (TDMa) system. The TDMa system specifically recruits a demethylating enzyme and transcriptional activators, providing efficient demethylation and transactivation of the NLRC5 promoter. TDMa in mouse and human cancer cells induced MHC class I antigen presentation and accelerated CD8+ T cell activation with tumor suppression effects both in vitro and in vivo. Moreover, enhanced immunogenicity by NLRC5 TDMa boosted efficacy of anti-PD1 therapy. Therefore, NLRC5 targeting by the TDMa system confers an attractive therapeutic approach against cancer.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for B16F10-TDM and B16F10-TDMa cells transduced with scramble sgRNA or Nlrc5 targeting sgRNA.

first: Xin

last: Sun

first: Toshiyuki

last: Watanabe

first: Oda

last: Yoshitaka

first: Weidong

last: Shen

first: Alaa

last: Ahmad

first: Ryota

last: Ouda

first: Paul

last: de Figueiredo

first: Hidemitsu

last: Kitamura

first: Shinya

last: Tanaka

middle: S


title: Single-Cell RNA-seq reveals transcriptomic modulation of Alzheimer’s Disease by Activated Protein C (APC)

city: Tampa

first: Ji

last: Li

state: FL

address: 12901 Bruce B. Downs Blvd

department: Surgery

country: USA

institute: University of South Florida

summary: Activated Protein C is an anti-inflammatory with proven treatment of cardiac and neurological disorders. We studied the transcriptional effects of APC on Alzheimers Disease transgenic mice.

overall_design: Daily i.p. injection of exogenous APC in WT and 5xFAD mice for 5 months. Daily saline injection were given as controls to WT and 5xFAD mice.

first: Mohammad

middle: K

last: Fatmi


title: Effect of ezetimibe treatment on gene expression of TGFβ1-activated primary human lung fibroblasts

city: Seodaemun-gu

first: Chanho

last: Lee

state: Seoul

address: 50-1 Yonsei-ro

department: Department of Internal Medicine

country: South Korea

institute: Yonsei University College of Medicine

summary: To verify whether ezetimibe inhibits myofibroblast differentiation at the transcriptome level and to reveal the mechanism of action of ezetimibe, we sequenced primary human lung fibroblasts that we isolated and cultured.

overall_design: Comparative gene expression analysis of mRNA-Seq data from primary human lung fibroblasts treated with TGFβ1 and/or ezetimibe (Negative control/ TGFβ1-treated positive control / TGFβ1&ezetimibe-treated group).

first: Se Hyun

last: Kwak

first: Jisu

last: Han

first: Ju Hye

last: Shin

first: Yu Seol

first: Jeong Su

last: Park

first: Beom Jin

last: Lim

first: Jin Gu

first: Young Sam

last: Kim

first: Song Yee

first: Soo Han

last: Bae


title: Maturation of Long Lived Plasma cells

city: Bronx

first: David

middle: Robert

last: Fooksman

state: NY

address: 1300 Morris Park

department: Pathology

institute: Albert Einstein College of Medicine

summary: To analyze global gene transcriptional changes and BCR clonal differences associated with long lived plasma cell specification, we sorted LLPCs and bulk PCs using Blimp1-ERT2-cre rosa26-LSLYFP expressing mice and analyzed them using bulk RNAsequencing

overall_design: on day 90 or day 5 after tamoxifen, YFP+ and YFP- CD138high B220- PCs were FACS-purified from BM, SPLEEN from young mice (age 6-8  for treatment) and middle-aged mice.   (age 24 weeks at tamoxifen treatment), with 3-4 mice per group.  Day 5 mice were aged 24 weeks before treatment

first: Zhixin

last: Jing

first: Deyou

last: Zheng

first: Phillip

last: Galbo

first: Thomas

last: MacCarthy


title: A cellular model of the oncogenic FOXL2 somatic pathogenic variant C134W provides new insights in pathogenicity

city: PARIS

first: Anne Laure

last: Todeschini

address: 15 rue hélène brion

country: France

institute: CNRS/Université Paris Cité

summary: To investigate the  dysfunction of FOXL2-C134W in human granulosa cells, we established homozygous cell lines for FOXL2 WT by CRISPR/Cas9 technologies.

overall_design: We then performed gene expression profiling analysis using data obtained from RNA-seq of 2 different clones (FOXL2 WT/WT vs FOXL2 WT/Mut).

first: Herman

last: Laetitia

first: Todeschini

last: Anne Laure

first: Veitia

last: Reiner


title: Integrated analysis of miRNA-mRNA regulatory networks in pediatric hepatocellular carcinoma.

status: Public on Feb 21 2024

city: Cincinnati

first: Rebekah

last: Karns

state: OH

address: 3333 Burnet Ave

department: Gastroenterology, Hepatology, & Nutrition

institute: Cincinnati Children's Hospital Medical Center

type: Non-coding RNA profiling by high throughput sequencing

summary: Pediatric hepatocellular carcinoma (HCC) is a group of liver cancers whose mechanisms behind their pathogenesis and progression are poorly understood. We aimed to identify alterations in the miRNAs and putative target mRNAs in not only tumor tissues of patients with pediatric HCC but also in corresponding background livers by using liver tissues without underlying liver disease as a control.

overall_design: We performed miRNA and mRNA profiling of pediatric HCC (consisting of fibrolamellar HCC (FLC) and non-FLC HCC) and paired liver tissues to identify miRNAs whose expression levels differed significantly from control livers. Our analysis generated prioritized lists of miRNAs and mRNAs from the following three sets of analyses: (a) pediatric HCC vs. control; (b) FLC vs. control; and (c) background liver tissue vs. control.

first: Kyung-Won

last: Min

first: Jung-Hyun

last: Cho

first: Je-Hyun

last: Yoon

first: Soona


title: Smart-seq analysis comparing melanocytes from Spry1flox/flox mice tail epidermis to the melanocytes derived from Spry1 epidermis specific knockout mice tail and back epidermis

city: Hangzhou

first: Yingzhe

last: Cui

address: Jiefang road

country: China

institute: Second Affiliated Hospital, Zhejiang University School of Medicine

summary: The function and survival of melanocytes is regulated by an elaborate network of paracrine factors synthesized mainly by epidermal keratinocytes. Keratinocytes and melanocytes respond to UV exposure by eliciting a tanning response. However, how keratinocytes and melanocytes interact in the absence of UV exposure is unknown. Here, we demonstrate that after Spry1 knockout in epidermal keratinocytes, melanocyte stem cells (McSCs) in the hair follicle exit the stem cell niche without depleting the pool of these cells. We also found that McSCs migrate to the epidermis in a Scf/C-kit-dependent manner induced by a tanning-like response resulting from Spry1 loss in epidermal keratinocytes. Once there, these cells differentiate into functional melanocytes. These findings show the potential for developing therapies for skin pigmentation disorders by manipulating McSCs.

overall_design: Cell type: FACS purified melanocytes from mouse epidermis; 3 conditions:Spry1flox/flox mice tail epidermis vs Spry1flox/flox k14-Cre-ERT2 mice tail epidermis vs Spry1flox/flox k14-Cre-ERT2 mice back epidermis ; 3 replicate for each condition.

first: Ying-Zhe

first: Fan

last: Xu


title: Gene expression profiling of BGC-823 cells with or without PCDH9-Cter overexpression

city: Shanghai

first: Yajuan

last: Zhang

address: 320 Yueyang Road

institute: Shanghai Institute of Biochemistry and Cell Biology

summary: Loss of Protocadherin 9 (PCDH9) is associated with the metastasis and the prognosis of gastric cancer patients, while the molecular mechanism of PCDH9-impaired gastric cancer metastasis remains unclear. Here we show that PCDH9 is cleaved in gastric cancer cells and intracellular domain of PCDH9 (PCDH9-Cter) translocates into nucleus. To decipher the mechanism underlying PCDH9-Cter-inhibited tumor cell migration, we performed RNA-Sequencing analysis for BGC-823 cells with or without PCDH9-Cter overexpression.

overall_design:
BGC-823 cells (Flag vector or PCDH9-Cter-Flag overexpression) were harvested and mRNA were extracted for high-throughput sequencing
Flag v.s. PCDH9-Cter-Flag overexpression, in triplicate


title: Redox-sensitive high-mobility group box-1 isoforms contribute to liver fibrosis progression and resolution in mice

city: Chicago

first: Xiaodong

last: Ge

state: IL

address: 909 South Wolcott Ave, COMRB 6140

institute: The University of Illinois at Chicago

summary: Since oxidative stress plays a major role in liver fibrosis and induces post-translational modifications of proteins, we hypothesized that redox-sensitive HMGB1 isoforms contribute to liver fibrosis progression and resolution. This study is significant because a rise in [H] HMGB1 could flag 'patient at risk', the presence of [O] HMGB1 could suggest 'disease in progress or active scarring', while the appearance of [SO3] HMGB1 could point at 'resolution under way'. The latter could be used as a readout for response to pharmacological intervention with anti-fibrotic agents.

overall_design: RNA sequencing was performed in total liver at peak ALD.  RAW264.7 cells were treated with BSA or complex ([O] HMGB1 + IL1β) for 6 hours.

first: Natalia

last: Nieto


title: Gene expression profile at single cell level of T cells from the brain of running wheel tranied or untrained mice after phototrombotic stroke

status: Public on Feb 23 2024

city: Duesseldorf

first: Christopher

last: Nelke

state: NRW

address: UKD, Moorenstraße 5

country: Germany

institute: University Hospital Düsseldorf

summary: We used single cell sequencing to analyze the impact of running wheel training on CD3+ T cells in the brain of mice suffering from stroke.

overall_design: Stroke was induced in C57BL/6 mice by photothrombosis. At day two mice started training on running wheels. Controls did not undergo training. At day 14 mice were sacrificed and CD3+ T cells were isolated from brains by single cell sorting and analyzed by using scRNA-Seq

first: Jens

last: Minnerup

first: Wiebke

last: Hansen


title: Heparan sulfate modifications of betaglycan promote TIMP3-dependent ectodomain shedding to fine-tune TGF-β signaling.

city: COLUMBIA

first: HAO

last: JI

state: SC

address: 715 SUMTER ST CLS RM 109

department: College of Pharmacy

institute: University of South Carolina

summary: Betaglycan/type III TGF-β receptor (TGFBR3) is an established co-receptor for the TGF-β superfamily with direct binding demonstrated for TGF-β 1-3 and inhibin A. Betaglycan can be membrane-bound or have its ectodomain cleaved/ shed to produce soluble-betaglycan that has been demonstrated to sequester ligands.  Extracellular domain of betaglycan is modified with glycosaminoglycan chains at Ser residues S534 and S545, to which heparan sulfate and chondroitin sulfate chains are covalently attached. To delineate the contribution of the heparan and chondroitin sulfate modifications on betaglycan on its shedding and thereby on TGF-β signaling in ovarian cancer biology, we used mutants of betaglycan with alterations to the different glycosaminoglycan modifications. We made the unexpected discovery that the heparan sulfate modifications are essential for maximum ectodomain shedding of betaglycan, which is further essential for the ability of betaglycan to suppress TGF-β signaling and the tumor cells responses to exogenous TGF-β ligand. Using unbiased transcriptomics, we identified TIMP3 as a key regulator of betaglycan shedding and thereby TGF-β signaling.  Taken together, these studies are the first to demonstrate a critical link between the well-known modifications on betaglycan and TGF-β signaling responses.

overall_design: To investigate the role of specific glycosaminoglycan modifications on betaglycan shedding and its impact on TGF-β signaling, we generated HEYA8 cells expressing serine 534 to alanine and Ser 545 to alanine mutants of betaglycan either as single mutations or double mutants ( delat GAG).  We then performed RNA-SEQ collected from the mutant BG expressing cells in three biological replicates and performed gene expression profiling analysis.

first: Alex

last: Choi

first: Mythreye

last: Karthikeyan

first: Hao

last: ji

first: Diego

last: Altomare

first: Michael

last: Shtutman


title: RNA-seq analysis comparing Spry1flox/flox mice back epidermis to the back epidermis derived from Spry1 epidermis specific knockout mice

summary: We found that Spry1 epidermis specific knockout give rise to back epidermis hyperpigmentaiton highly mimicks tanning response in mouse

overall_design: Spry1 epidermis specific knockout were induced by tamoxifen treatment. The epidermis were harvested and compared with the Spry1flox/flox mouse back epidermis for RNA-seq analysis


title: Advanced Complexity and Short-Term Plasticity of Neural Activity in Reciprocally Connected Human Cerebral Organoids

city: Meguro

first: Yoshiho

last: Ikeuchi

state: Tokyo

address: 4-6-1 Komaba

department: Institute of Industrial Science

institute: The University of Tokyo

summary: An inter-regional cortical tract is one of the most fundamental architectural motifs that integrates neural circuits to orchestrate and generate complex functions of the human brain. To understand the mechanistic significance of inter-regional projections on development of neural circuits, we investigated an in vitro neural tissue model for inter-regional connections, in which two cerebral organoids are connected with a bundle of reciprocally extended axons. The connected organoids produced more complex and intense oscillatory activity than conventional or directly fused cerebral organoids, suggesting the inter-organoid axonal connections enhance and support the complex network activity. In addition, optogenetic stimulation of the inter-organoid axon bundles could entrain the activity of the organoids and induce robust plasticity of the macroscopic circuit. These results demonstrated that the projection axons could serve as a structural hub that boosts functionality of the organoid-circuits. This model could contribute to further investigation on development and functions of macroscopic neuronal circuits in vitro.

overall_design:
To access the functional role of the inter-organoid connection through an axon bundle, we compared the ‘connected’ organoids with that of conventional cerebral organoids (‘single’ organoids). The connected organoids contained approximately twice the number of cells of a single organoid, which could affect the neuronal characteristics. We thus also assessed the tissues generated by directly fusing two cerebral organoids (‘fused’ organoids) as an additional control. To fuse organoids, two organoids were cultured together in a low-adhesion surface culture vessel to induce their spontaneous fusion. We assessed how the inter-organoid connections altered the gene expression profiles in the organoids.
Single, fused, and connected organoid at 7 weeks of culture on PDMS-MEA chip were first extracted from PDMS device in warmed maintenance medium and centrifuged at 100 xg for 30 seconds. To get single-cell suspensions, the organoids were dissociated by AccuMax for 10-30 min at 37°C and then centrifuged 200 xg for 5 min. Then, the pelleted cells were re-suspended in DMEM containing 10% FBS to be subjected to 10x Genomics Chromium single-cell RNA-seq library preparation, according to the manufacturer’s protocol. Finally, the library was sequencing with 150-bp paired-end reads on DNBSEQ. The sequencing data were processed using Cell Ranger analysis pipeline v3 with default parameters. Reads were aligned to human reference genome (GRCh38). Cell Ranger output “filtered gene-barcoded” count matrix was loaded into Scanpy (Wolf et al., 2018) with other python packages (scanpy==1.8.1 anndata==0.7.6 umap==0.5.1 numpy==1.19.5 scipy==1.4.1 pandas==1.1.5 scikit-learn==0.22.2.post1 statsmodels==0.10.2 python-igraph==0.9.6 pynndescent==0.5.4) for downstream analysis. Poor-quality cells were excluded based on the following criterion: min_genes > 200 and min_cells < 3, mitochondrial gene percentage < 10%, and nFeatures <4000. Cells with percentage of hemoglobin reads > 5% were discarded. In total, 17,636 cells remained for subsequent analysis. Batch effect correction was performed.

first: Tatsuya

last: Osaki

first: Tomoya

last: Duenki

first: Siu Yu

last: Chow

first: Yasuhiro

last: Ikegami

first: Romain

last: Beaubois

first: Timothee

last: Levi

first: Nao

last: Nakagawa-Tamagawa

first: Yoji

last: Hirano


title: A common allele increases endometrial Wnt4 expression, with antagonistic implications for pregnancy, reproductive cancers, and endometriosis

city: Vienna

first: Mihaela

last: Pavlicev

address: Djerassiplatz 1

department: Evolutionary Biology

country: Austria

institute: University of Vienna

summary: The common human single nucleotide polymorphism rs3820282 is associated with multiple phenotypes ranging from gestational length to likelihood of endometriosis and ovarian cancer and can thus serve as a paradigm for a highly pleiotropic genetic variant. Pleiotropy makes it challenging to assign specific causal roles to particular genetic variants. Deleterious mutations in multifunctional genes may cause either the co-occurrence of multiple disorders in the same individuals (i.e., syndromes), or be repeatedly associated with a variety of disorders in a population. Moreover, the adverse effects can be in combination with advantages in other traits, maintaining high frequencies of deleterious alleles in the population. To reveal the causal role of this specific SNP, we investigated the molecular mechanisms affected by rs3820282 in mice. We have shown previously that rs3820282 introduces a high affinity estrogen receptor 1 binding site at the Wnt4 locus. Here we introduced this nucleotide substitution into the homologous site of the mouse genome by CRISPR/Cas 9 and show that this change causes a specific upregulation of Wnt4 transcription in the endometrial stromal cells (but not that of CDC42, also located close to rs3820282) during the preovulatory estrogen peak in late proestrus. Transcriptomic analysis of the whole uterus reveals broad systemic effects on uterine gene expression, including downregulation of proliferation and induction of many progesterone-regulated pro-implantation genes. The effect on proliferation is limited to the luminal epithelium, whereas other effects involve the uterine stromal compartment. We suggest that in the uterus, these changes could contribute to increased permissiveness to embryo invasion. Yet in other estrogen-responsive tissues, the same changes potentially lead to decreased resistance to invasion by cancer cells and endometriotic foci. A single molecular effect of rs3820282 on Wnt4 expression may thus underlie the various associated phenotypic effects.

overall_design: To investigate the influence the SNP rs3820282 on uterine expression patterns we generated CRISPR/Cas9 knock-in lines with this substitution into BL/6 mice. We then collected uteri from proestrus and 17.5dpc wildtype and transgenic females and conducted RNAseq.

first: Michaela

first: Caitlin

last: McDonough-Goldstein


title: Aging-associated Decline in Vascular Smooth Muscle Cell Mechanosensation is Mediated by Piezo1 Channel

city: Brooklyn

first: Ngoc

last: Luu

address: 6 Metrotech Center

institute: New York University

summary: Aging of the vasculature is associated with detrimental changes in vascular smooth muscle cell (VSMC) mechanosensitivity to extrinsic forces in their surrounding microenvironment. However, how chronological aging alters VSMCs’ ability to sense and adapt to mechanical perturbations remains unexplored. Here, we show defective VSMC mechanosensation in aging measured with ultrasound tweezers-based micromechanical system, force instantaneous frequency spectrum and transcriptome analyses.  The mechanobiological study reveals thataged VSMCs adapt a relatively inert solid-like state with altered actin cytoskeletal integrity, resulting in an impairment in their mechanosensitivity and dynamic mechanoresponse to mechanical perturbations. The aging-associated decline in mechanosensation behaviors is mediated by hyperactivity of Piezo1-dependent calcium signaling. Inhibition of Piezo1 alleviates vascular aging and partially restores the loss in dynamic contractile properties in aged cells. Altogether, our study reveals the novel signaling pathway underlying aging-associated aberrant mechanosensation in VSMC and identifies Piezo1 as a potential therapeutic mechanobiological target to alleviate vascular aging.

overall_design: Single-cell RNA-sequencing libraries of four groups of vascular smooth muscle cells extracted from C57BL/6 mice  young (12 weeks) and old (58-78 weeks) cultured in four different treatments. Young + Angiotensin II and Old + siPiezo1 are the experimental groups, while Young and Old are the control groups.

first: Weiqiang

last: Chen


title: Characterization of two distinct immortalized endothelial cell lines HMEC-1 and EA.hy926 for in vitro studies: exploring the impact of calcium electroporation, Ca2+ signaling and transcriptome profile

city: Ljubljana

first: Tim

last: Bozic

address: Zaloška cesta 2

department: Department of Experimental Oncology

country: Slovenia

institute: Institute of Oncology Ljubljana

summary: The vascular endothelium consists of endothelial cells (ECs) with important biological functions and their impairment is associated with various pathologies. ECs vary based on tissue origin and gene expression, while their functionality depends on calcium (Ca2+) signaling. In tumors, disruption of Ca2+ homeostasis after calcium electroporation (CaEP) has been shown to elicit an enhanced antitumor effect with only minimal effect on normal tissue. The difference in response to CaEP was not only observed between cancer and normal cells, but also between different endothelial cell lines. Although several vascular EC models have been developed, there is a lack of understanding regarding the molecular basis that could help explain different responses of normal tissue to CaEP. Therefore, our study aims to determine the effect of CaEP on established immortalized human EC lines EA.hy926 and HMEC-1 in terms of cytoskeleton, Ca2+ kinetics and differences in gene expression involved in regulation of Ca2+ signaling and homeostasis.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for human immortalized microvascular endothelial cell lines EA.hy926 and HMEC-1.

first: Iva

last: Santek

first: Barbara

last: Lisec

first: Bostjan

last: Markelc

first: Milka

last: Vrecl

first: Robert

last: Frangez

first: Maja

last: Cemazar


title: Differential expression of genes affected by CRY1 or CRY2 in the absence of PER and NR1D proteins

city: Taichung

first: Yi-Ying

last: Chiou

address: No.145, Xingda Rd., South Dist.

department: Graduate Institute of Biochemistry

country: Taiwan

institute: National Chung Hsing University

summary: The transcription-translation feedback loop, the core clock mechanism, is required for circadian rhythm. CRY protein, including CRY1 and CRY2, plays an important repressor role in the regulation of clock genes. However, other proteins, like PER1, PER2, NR1D1 and NR1D2, in the loop mask the transcriptional effects of CRY. This study provides data to find candidate genes specifically affected by CRY1 or CRY2 in mouse embryonic fibroblast (MEF) cells.

overall_design: This study contains three types of samples: 1. MEF cells lacking CRY1, CRY2, PER1, PER2, NR1D1 and NR1D2 proteins (CPN-/-), 2. Exogenous CRY1 expression in the CPN-/- cells , 3.Exogenous CRY2 expression in the CPN-/- cells


title: An Upstream G‐Quadruplex DNA Structure Can Stimulate Gene Transcription

city: Manchester

first: Angela

last: Simeone

address: 200 Hathersage Rd

country: United Kingdom

institute: Genomic Services QIAGEN

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Four-stranded G-quadruplexes (G4s) are prevalent secondary structural features of the human genome. They are formed through stacking of Hoogsteen hydrogen-bonded G-tetrads connected by loops of variable length and sequence. G4 structures are found widely in genome control elements including gene promoters and are tightly linked to active gene expression and the recruitment of transcription factors and chromatin remodellers. To systematically investigate how G4 structures modulate transcription, we have developed a strategy in which different G4 sequences are engineered into the promoter of a synthetic reporter gene and ectopically into the same endogenous position in the genome of human cells. Here, we show that G4 sequences introduced into the host genome can fold into an G4 structure. Crucially, we demonstrate that G4 formation within a gene promoter stimulates transcription compared to G4-negative promoters and that this is not due to inherent G-richness. Finally, we provide robust evidence that transcriptional levels are directly related to the degree of thermodynamic stability of the G4 structure. Taken together, our results provide compelling evidence that G4 structures have a direct and positive regulatory role in gene transcription.

overall_design: DNA G-quadruplexes (G4-ChIP-seq) has been profiled in HEK293 cells (human embryonic kidney cells). 3 biological replicates have been performed, and each of those have been assessed by 3 technical replicates and 1 input library.

first: Yuqi

first: Sergio

middle: M

last: Cuesta

first: Larry

last: Melidis

first: David

last: Tannahill

first: Shankar

last: Balasubramanian


title: Abnormal metabolism of palmitic acid promotes the formation of renal calcium oxalate stones

city: Tianjin

first: Rui

last: Wang

address: Pingjiang Road 23

department: Urology

institute: The Second Hospital of Tianjin Medical University

summary: Individual metabolic factors are involved in the occurrence and development of CaOx stones, and dyslipidemia is an independent risk factor for the formation of urolithiasis. This study have screened out palmitic acid as one of the significantly differential metabolites in the urine of patients with renal CaOx stones compared with healthy controls by metabolomics. This study aims to further clarify the correlation between abnormal palmitic acid metabolism and the formation of renal CaOx stones, confirm the promoting effect of abnormal palmitic acid metabolism on the adhesion of CaOx crystals and the formation of renal CaOx stones

overall_design: Transcriptomics was applied to analyze the changes of genes related to the metabolism of palmitic acid in renal tubular epithelial cells


title: Divergent role of Mitochondrial Amidoxime Reducing Component 1 (MARC1) in human and mouse

city: Tarrytown

first: Gabor

last: Halasz

state: New York

address: 777 old saw mill river road

institute: Regeneron Pharmaceuticals

summary: Recent human genome-wide association studies have identified common missense variants in MARC1, p.Ala165Thr and p.Met187Lys, associated with lower hepatic fat, reduction in liver enzymes and protection from most causes of cirrhosis. Using an exome-wide association study we recapitulated earlier MARC1 p.Ala165Thr and p.Met187Lys findings in 540,000 individuals from five ancestry groups. We also discovered novel rare putative loss of function variants in MARC1 with a phenotype similar to MARC1 p.Ala165Thr/p.Met187Lys variants. In vitro studies of recombinant human MARC1 protein revealed Ala165Thr substitution causes protein instability and aberrant localization in hepatic cells, suggesting MARC1 inhibition or deletion may lead to hepatoprotection. Following this hypothesis, we generated Marc1 knockout mice and evaluated the effect of Marc1 deletion on liver phenotype. Unexpectedly, our study found that whole-body Marc1 deficiency in mouse is not protective against hepatic triglyceride accumulation, liver inflammation or fibrosis. In attempts to explain the lack of the observed phenotype, we discovered that Marc1 plays only a minor role in mouse liver while its paralogue Marc2 is the main Marc family enzyme in mice. Our findings highlight the major difference in MARC1 physiological function between human and mouse.

overall_design:
Marc1-/- mice were generated using Regeneron’s (Tarrytown, NY) VelociGene® technology (ID #20561), and  Wild-type (Marc1+/+) littermates were used as controls. All experiments were performed on 100% C57BL/6NTac background.
Marc2 knockout mice (ID #20646) were generated similarly as Marc1 KO mouse using Regeneron VelociGene® technology, where second exon of Marc2 gene was replaced with LacZ reporter. Marc2 mice were bred on 100% C57BL/6NTac background.
RNA-Seq was performed on RNA purified  from wild-type and homozygous Marc1 or Marc2 knockout mice tissues (heart, kidney, liver, lung, and white adipose).

first: Eriks

last: Smagris

first: Viktoria

last: Gusarova


title: Inter-patient heterogeneity in the hepatic ischemia-reperfusion injury transcriptome: implications for research and diagnostics

city: Graz

first: Silvia

last: Groiss

address: Neue Stiftingtalstraße 6

department: Institute of Pathology

institute: Medical University of Graz

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

overall_design: Refer to individual Series


title: Inter-patient heterogeneity in the hepatic ischemia-reperfusion injury transcriptome: implications for research and diagnostics [EMC data]

summary: Background & Aims: Cellular responses induced by surgical procedure or ischemia-reperfusion injury (IRI) may severely alter transcriptome profiles. We have investigated this effect to obtain insight into molecular ischemia responses during surgical procedures and characterize pre-analytical effects impacting on molecular analyses. Methods: 143 non-malignant liver samples were obtained from 30 patients at different time points of ischemia during surgery from two individual cohorts, treated either with the Pringle maneuver or total vascular exclusion. Transcriptomics profiles were analyzed by Affymetrix microarrays and expression of selected mRNAs was validated by RT-qPCR. Results: Transcriptional profiles of both cohorts displayed 179 genes that were mutually deregulated confirming elevated cytokine signaling, and NFkB as the dominant pathways in ischemia response. Contrary to ischemia, reperfusion induced pro-apoptotic and pro-inflammatory cascades involving TNF, NFkB and MAPK pathways. FOS and JUN were down-regulated in steatosis compared to their up-regulation in normal livers. Surprisingly, molecular signatures of underlying primary and secondary cancers were clearly present in the non-tumor tissue. Conclusions: We identified transcripts mutually deregulated during ischemia and reperfusion injury in both cohorts that can be used to monitor ischemia during liver surgery and highlight the importance of pre-analytical quality control. The marked inter-patient variability might reflect differences in individual stress responses and impact of underlying disease conditions. Furthermore, we provide a comprehensive and pre-analytically highly standardized in vivo transcriptome profile of histologically normal liver and identified 230 genes with substantial pre-analytical robustness (<2 % covariation across both cohorts) that might serve as reference genes and could be particularly suited for future diagnostic applications. Conclusions: We identified transcripts mutually deregulated during ischemia and reperfusion injury in both cohorts that can be used to monitor ischemia during liver surgery and highlight the importance of pre-analytical quality control. The marked inter-patient variability might reflect differences in individual stress responses and impact of underlying disease conditions. Furthermore, we provide a comprehensive and pre-analytically highly standardized in vivo transcriptome profile of histologically normal liver and identified 230 genes with substantial pre-analytical robustness (<2 % covariation across both cohorts) that might serve as reference genes and could be particularly suited for future diagnostic applications.

overall_design: Ischemia related changes of RNA profiles obtained from frozen liver tissue samples collected before, during and after routine surgery and compared between the different time points. Erasmus Medical Center Rotterdam, 69 samples of a total of 9 patients at up to 7 different time points per patient.

first: Christian

last: Viertler

first: Marcel

last: Kap

first: Gerwin

last: Bernhardt

first: Hans

last: Mischinger

first: Anieta

last: Sieuwerts

first: Cees

last: Verhoef

first: Peter

last: Riegman

first: Mogens

last: Kruhoffer

last: Svec

last: Sjöback

first: Karl-Friedrich

last: Becker

first: Kurt

last: Zatloukal


title: RNA-Seq of primary CKD osteoblasts

city: Phoenix

first: Katherine

last: Wesseling-Perry

state: AZ

address: 1919 E. Thomas Rd

department: Nephrology

institute: Phoenix Children's Hospital

summary: Primary osteoblasts from patients with end-stage kidney disease have defective maturation and mineralization characteristics in vivo and in vitro.  This study was designed to evaluate signaling pathways associated with this abnormal maturation phenotype.

overall_design: Cross-sectional RNA-Seq analysis of primary osteoblasts from 9 pediatric end-stage kidney disease patients and 3 healthy controls grown to confluence


title: Sequencing in Over 50,000 Cases Identifies Coding and Structural Variation Underlying Atrial Fibrillation Risk

city: Cambridge

first: Matthew

middle: C

last: Hill

state: MA

address: 75 Aimes Street

institute: Broad Institute of MIT and Harvard

summary: Atrial fibrillation (AF) is a prevalent and morbid abnormality of the heart rhythm with a strong genetic component Here, we meta-analyzed genome and exome sequencing data from 36 studies that included 52,416 AF cases and 277,762 controls In burden tests of rare coding variation, we identified novel associations between AF and the genes MYBPC3, LMNA, PKP2, FAM189A2 and KDM5B We further identified associations between AF and rare structural variants due to deletions in CTNNA3 and duplications of GATA4 We broadly replicated our findings in independent samples from MyCode, deCODE and UK Biobank Finally, we found that CRISPR-knockout of KDM5B in stem cell derived atrial cardiomyocytes led to a shortening of the action potential duration and widespread transcriptomic dysregulation of genes relevant to atrial homeostasis and conduction Our results highlight the contribution of rare coding and structural variants to AF, the genetic link between AF and cardiomyopathies, and expand our understanding of the rare variant architecture for this common arrhythmia

overall_design: To characterize the molecular role of KDM5B in cardiomyocytes we knocked the gene out in iPS-derived atrial cardiomyocytes with a lentiviral CRISPR-Cas9-based approach. For this we independently delivered 2 KDM5B targeting sgRNAs that we found to be highly mutagenic After lentiviral delivery and puromycin selection, we then performed transcriptomic analysis using bulk RNA sequencing

first: Patrick

middle: T

last: Ellinor

first: Sean

middle: J

last: Jurgens

first: Seung

middle: H

last: Choi


title: Single-cell profiling identifies a specific monocyte subset for neuroregeneration

city: Guangzhou

first: xialin

last: liu

address: Jinsui Road

institute: Zhongshan Opythalmic Center, Sun yat-sen University

summary: To elucidate the classification and function of monocyte subsets in the neuroprotective retina, we used single-cell RNA sequencing on isolated retinal immune cells. We identified a novel subpopulation of monocyte named Ly6ClowMo/MΦ, which display neural restorative features.

overall_design: Single cell profiling of retinal immune cells from PBS and MSC-sEV-treated optic nerve injuried mice, using 10X genomic platform.

first: Xialin

last: Liu

first: Wei

last: Yi


title: Retinal Ly6Clow Mo/MΦ protect optic nerve against injury

summary:
We investigate the therapeutic effect of mesenchymal stem cell-derived small extracellular vesicles (MSC-sEVs), a potent immune-modulating therapy for tissue regeneration, in repairing optic nerve damage. Intraocular administration of MSC-sEVs promotes both RGC survival and axon regeneration against optic nerve crush injury. Mechanistically, MSC-sEVs recruits a neural restorative population of hematogenous Ly6Clow monocyte/monocyte-derived macrophage (Mo/MΦ). Gain-of-function by intravitreal administration of the donor Ly6Clow Mo/MΦ markedly improves neurological outcomes in vivo.
We then performed gene expression profiling analysis using data obtained from RNA-seq of 4 different cells at two time points.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for C3H10T1/2 cells and its KD derivatives (shKDM4D, shNFIB, and shMLL1).

first: Ying

last: Xue


title: Inter-patient heterogeneity in the hepatic ischemia-reperfusion injury transcriptome: implications for research and diagnostics [MUG data]

first: Silvia

last: Groiss

overall_design: Ischemia related changes of RNA profiles obtained from frozen liver tissue samples collected before, during and after routine surgery and compared between the different time points. Medical University of Graz, 83 samples of a total of 21 patients at 4 different time points per patient.

first: Christian

last: Viertler

first: Marcel

last: Kap

first: Gerwin

last: Bernhardt

last: Mischinger

last: Sieuwerts

first: David

first: Robert


title: Metagenomic sequencing of adult zebrafish feces after SMZ exposure (Exposure of Danio rerio to environmental sulfamethoxazole may contribute to neurobehavioral abnormalities via gut microbiome disturbance)

first: Qiu

last: Yushu

state: Yangpu

address: Kongjiang Road, No. 1665

institute: Xihua Hospital

summary: The neurotoxic effects and mechanisms of low-dose and long-term sulfamethoxazole (SMZ) exposure remain unknown. This study exposed zebrafish to environmental SMZ concentrations and observed behavioral outcomes. SMZ exposure increased hyperactivity and altered the transcript levels of 17 genes associated with neurological function. It impaired intestinal function by reducing the number of intestinal goblet cells and lipid content. Metabolomic results indicated that the contents of several lipids and amino acids in the gut were altered, which might affect the expression levels of neurological function-related genes. Metagenomic results demonstrated that SMZ exposure substantially altered the composition of the gut microbiome. Zebrafish receiving a transplanted fecal microbiome from the SMZ group were also found to exhibit abnormal behavior, suggesting that the gut microbiome is an important target for SMZ exposure-induced neurobehavioral abnormalities. Multi-omics correlation analysis revealed that gut micrometabolic function was related to differential gut metabolite levels, which may affect neurological function through the gut-brain-axis. Reduced abundance of Lefsonia and Microbacterium was strongly correlated with intestinal metabolic function and may be the key bacterial genera in neurobehavioral changes. This study confirms for the first time that SMZ-induced neurotoxicity in zebrafish is closely mediated by alterations in the gut microbiome.

overall_design: We exposed zebrafish eggs to 5000ng/L SMZ solution. We collected feces from the control group and SMZ group at 120 days. Analysis of bacterial flora changes in feces by metagenomic sequencing. Simultaneously detecting the transcriptome of zebrafish brain tissue and the metabolome of the intestine

first: Kan

last: Yu


title: Sphingosine 1-phosphate maintains naïve T cell by regulating apoptotic signaling

city: New York

first: Dhaval

last: Dixit

address: 550 First Avenue

institute: NYU School of Medicine

summary: Effective immunity requires a large, diverse naïve T cell repertoire circulating among lymphoid organs in search of antigen.  Sphingosine 1-phosphate (S1P) and its receptor S1PR1 contribute by both directing T cell migration and supporting T cell survival. Here, we address how S1P enables T cell survival, and the implications for patients treated with S1PR1 antagonists. Contrary to expectations, we found that S1PR1 limits apoptosis by maintaining the appropriate balance of BCL2 family members via restraint of JNK activity. Interestingly, the same residues of S1PR1 that enable receptor internalization are required to prevent JNK overactivation and limit apoptosis. Findings in mice were recapitulated in ulcerative colitis patients treated with the drug ozanimod, and the loss of naïve T cells limited B cell responses. Our findings highlight an unexpected effect of S1PR1 antagonists on the ability to mount immune responses within lymph nodes, beyond their effect on lymph node egress.

overall_design: We analyzed gene expression difference between control naive CD4 T cells and naive CD4 T cells lacking S1P signaling (either from SPNS2-deficient mice or from S1PR1-deficient mice). For SPNS2: CD45.2 Spns2f/f Lyve1-Cre mice or CD45.2 littermate control mice were lethally irradiated and reconstituted with WT CD45 .1/.2 bone marrow. After reconstitution, naïve CD4 T cells were sorted by FACS and RNA-Seq was performed. For S1PR1: a 50:50 mixture of CD45.2 UBC-GFP:CD45.2 S1pr1f/f UBC-CreERT2 or CD45.2 UBC-GFP:CD45.2 S1pr1f/f littermate control bone marrow was transferred into lethally irradiated CD45.1/.2 WT mice. After reconstitution, mice were treated with tamoxifen. Four weeks after tamoxifen treatment, CD45.2 S1PR1-deficient or CD45.2 control naïve T cells were sorted and RNA-Seq was performed.

first: Dixit

last: Dhaval


title: MSC-sEV regulates Ly6Clow Mo/MΦ in neural restoration


title: Focal adhesion kinase-YAP signaling axis drives drug-tolerant persister cells and residual disease in NSCLC

status: Public on Feb 20 2024

city: San Francisco

last: Wu

state: CA

address: 600 16th street

department: Department of Medicine

institute: University of California, San Franciso

summary: Targeted therapy is effective in many tumor types including lung cancer, the leading cause of cancer mortality. Paradigm defining examples are targeted therapies directed against non-small cell lung cancer (NSCLC) subtypes with oncogenic alterations in EGFR, ALK and KRAS. The success of targeted therapy is limited by drug-tolerant persister cells (DTPs) which withstand and adapt to treatment and comprise the residual disease state that is typical during treatment with clinical targeted therapies. Here, we integrate studies in patient-derived and immunocompetent lung cancer models and clinical specimens obtained from patients on targeted therapy to uncover a focal adhesion kinase (FAK)-YAP signaling axis that promotes residual disease during oncogenic EGFR-, ALK-, and KRAS-targeted therapies. FAK-YAP signaling inhibition combined with the primary targeted therapy suppressed residual drug-tolerant cells and enhanced tumor responses. This study unveils a FAK-YAP signaling module that promotes residual disease in lung cancer and mechanism-based therapeutic strategies to improve tumor response.

overall_design: RNA sequencing in (1) EGFR-mutant PC9 cells, ALK fusion-positive H3122 cells, and KRAS-mutant H358 cells upon drug treatment. The following conditions are available per each cell line: treatment with 0.1 % DMSO for 48 hours in parental cells, treatment with targeted inhibitor for 48 hours in parental cells, treatment with targeted inhibitor for ≥ 9 days in persister cells, treatment with targeted inhibitor in acquired resistant cells. Relates to Fig. 2o, Supplemental Fig. 2d.  (2) Isogenic EGFR-mutant PC9-C2 and H1975-B10 prior to treatment (t0) and in osimertinib persisters (d9).  (3) EGFR-mutant PC9 cells comparing empty vector control cells (EV), YAP-WT overexpressing cells, hyperactive YAP-5SA overexpressing cells, and inactive YAP-S94A cells.  (4) EGFR-mutant PC9 osimertinib persister cells and ALK fusion-positive H3122 alectinib persister cells upon combinatorial treatment with TEAD inhibitor VT104. Relates to Suppl. Figure 4: Gene set enrichment analysis (GSEA) was performed using the YAP-5SA_UP gene set (Supplementary Table 3) on RNAseq expression data from PC9 osimertinib DTPs and H3122 alectinib DTPs, both treated with 10 µM VT104 for 24 h.  (5) EGFR-mutant organoid models TH107 and TH330 comparing cells treated with 0.1% DMSO for 3 days and with 1 µM osimertinib for 11 days; relates to Suppl. Figure 7e: RNAseq expression data of the EGFR-mutant TH107 PDO model, comparing untreated DMSO control (DMSO) versus osimertinib persisters (D9). (6) EGFR-mutant PDX TH021 and ALK fusion-positive LG0812 comparing vehicle and TKI treated tumors at treatment endpoint; relates to Supplemental Fig. 8(d&e): Gene set enrichment analysis for the YAP-5SA_UP gene set (Supplementary Table 3) using RNAseq expression data of the EGFR-mutant TH021 PDX model (d) and ALK fusion-positive LG0812 PDX model (e), comparing vehicle control (VEH) versus treatment group (TKI).

first: Franziska

last: Haderk

first: Yu-Ting

last: Chou

first: Lauren

last: Cech


title: Post-transcriptional regulation of necroptosis gene EIF2AK2 in peripheral blood of bronchopulmonary dysplasia

city: Nanning

first: Wen

middle: Jing

last: Liu

state: Guangxi

address: No 6 Shuangyong Road

institute: The first affiliated hospital of Guangxi Medical University

summary: Bronchopulmonary dysplasia (BPD) remains the most frequent chronic lung disease among infants, which involves multifactorial pathogenesis. Necroptosis represents a caspase-independent mode of programmed cell death, and its deregulation associates with lung diseases, but the mechanisms of necroptosis during BPD are indistinct. This work was conducted for unveiling the post-transcriptional regulatory mechanisms of necroptosis in BPD.

overall_design: We sequenced messenger RNA (mRNA) (BPD, n=4; controls, n=4) and microRNA (miRNA) (BPD, n=5; controls, n=5) profiles in peripheral blood specimens. The GSE125873 dataset composed of mRNA profiling from 11 BPD individuals and 21 controls was gathered. Based upon the circBank and starBase databases, circRNA-miRNA-mRNA networks and a necroptosis-based subnetwork were built. The diagnostic efficacy of eukaryotic translation initiation factor 2 alpha kinase 2 (EIF2AK2) was assessed via receiver operator characteristic curve (ROC). EIF2AK2-relevant molecules were analyzed with weighted correlation network analysis (WGCNA), and then molecular classification was conducted through consensus clustering method.

first: Wen-Jing

first: Yuan-Han

last: Qin


title: Transcriptome-wide 1-Methyladenosine Functional Profiling of Messenger RNA and Long Non-Coding RNA in Bladder Cancer

city: zhengzhou

first: Jianjian

last: Yin

state: Henan

address: No. 100 Science Road

institute: Zhengzhou University

summary: This study aimed to gain a deeper understanding of m1A methylation in BLCA mRNAs and lncRNAs.

overall_design: methylated RNA immunoprecipitation sequencing and RNA sequencing were performed to profile mRNA and lncRNA m1A methylation and expression in BLCA cells, with or without stable knockdown of the m1A methyltransferase tRNA methyltransferase 61A (TRMT61A).

first: Lirong

last: Zhang

first: Dongkui

last: Song


title: Identification of unique genomic signatures in patients with fibromyalgia and chronic pain

city: Chicago

first: Gayatry

last: Mohapatra

address: 840 S Wood Street

institute: University of Illinois at Chicago

summary: Fibromyalgia is a chronic pain syndrome characterized by widespread pain. The pathophysiology of fibromyalgia is not clearly understood and there are no specific biomarkers available for accurate diagnosis. Here we define genomic signatures using high throughput RNA sequencing on 96 fibromyalgia and 93 matched controls. Our findings revealed two major fibromyalgia-associated expression signatures. The first group included 44 patients with a signature enriched for gene expression associated with extracellular matrix and downregulation of RhoGDI signaling pathway. The second group included 31 patients and showed a profound reduction in the expression of inflammatory mediators with an increased expression of genes involved in the CLEAR signaling pathway. These results suggest defective tissue homeostasis associated with the extra-cellular matrix and cellular program that regulates lysosomal biogenesis and participates in macromolecule clearance in fibromyalgia. Further elucidation of these pathways will lead to development of accurate diagnostic markers, and effective therapeutic options for fibromyalgia.

overall_design: High throughput RNA sequencing

first: Fabien

last: Dachet

first: Louis

last: Coleman

first: Bruce

last: Gillias

first: Frederick

middle: G

last: Behm


title: HCAR1 nuclear transportation is critical for cell function

city: minneappolis

first: LiBang

last: yang

state: MN

address: 420 delaware st

department: medicine

institute: University of minnesota

summary: Membrane receptor nuclear translocation play some novel role in cancer pathology. In the current studies, we demonstrate HCAR1 distributes in cancer cell nucleus in Lung cancer; Lactate promote HCAR1 nuclear translocation. Proteomics analysis found there are a set of nuclear proteins binds with HCAR1; interaction of HCAR1 with SFPQ etc other proteins promote cell self-renewal and cell invasion in lung cancer. ChIP sequencing analysis discovered there are a set of genes were targeted by HCAR1.

overall_design: Lung cancer cell with/without lactate treatment were used to immunoprecipitate chromatin fragments with anti-HCAR1 antibodies. The pellet then were used for ChIPseq.

first: Libang

last: Yang

first: Thomas

last: Kono


title: Transcriptome and chromatin accessibility in oral squamous carcinoma with human plasma-like medium culture (RNA-Seq)

city: Chuo-ku

first: Jun

last: Nakayama

state: Osaka-shi / Osaka

address: Otemae 3-1-69

institute: Osaka International Cancer Institute

summary: In recent years, human plasma like medium (HPLM) have been developed that mimic the conditions of the human physiology. In comparison with conventional cell culture medium, cell culture in HPLM had extensive effects on cellular metabolism, including metabolome, redox status, and glucose utilization. However, molecular mechanisms underlying HPLM effects on human cells are only partly understood. Transcriptome and chromatin accessibilities are important regulators of cell states and implicate in a variety of biological processes. We habituated three oral cancer cell lines such as SAS, HSC-3, and HSC-4 in HPLM and evaluated cell growth and drug sensitivity. We perform transcriptome analysis between HPLM and conventional culture condition in HSC-3 cells by mRNA-seq.

overall_design: Assay for mRNA -sequencing to identifying transcriptme of HSC-3 cells cultured in HPLM or DMEM conditions.

first: Karen

last: Yamaguchi

first: Yusuke

last: Yamamoto


title: Transcriptome and chromatin accessibility in oral squamous carcinoma with human plasma-like medium culture (ATAC-Seq)

summary: In recent years, human plasma like medium (HPLM) have been developed that mimic the conditions of the human physiology. In comparison with conventional cell culture medium, cell culture in HPLM had extensive effects on cellular metabolism, including metabolome, redox status, and glucose utilization. However, molecular mechanisms underlying HPLM effects on human cells are only partly understood. Transcriptome and chromatin accessibilities are important regulators of cell states and implicate in a variety of biological processes. We habituated three oral cancer cell lines such as SAS, HSC-3, and HSC-4 in HPLM and evaluated cell growth and drug sensitivity. We perform epigenetic analysis between HPLM and conventional culture condition in HSC-3 cells by ATAC-seq.

overall_design: Assay for Transposase-Accessible Chromatin (ATAC) -sequence to identifying open chromatin regions of HSC-3 cells cultured in HPLM or DMEM conditions.

first: Kenichi

last: Miyata

first: Reo

last: Maruyama


title: Cellular and molecular alterations to muscles and neuromuscular synapses in Megf10-related myopathy

city: Providence

first: Gregorio

last: Valdez

state: RI

address: 70 Ship Street

institute: Brown University

summary: Loss-of-function mutations in MEGF10 lead to a rare and understudied neuromuscular disorder known as MEGF10-related myopathy. There are no treatments for the progressive respiratory distress, motor impairment, and structural abnormalities in muscles caused by the loss of MEGF10 function. In this study, we asked whether mice lacking MEGF10 (MEGF10 KO) could be used as a model to generate preclinical insights to treat MEGF10-related myopathy. We deployed cellular and molecular assays to examine juvenile, young adult, and middle-aged Megf10 constitutive knockout (KO) mice. We found fewer muscle fibers in developing and adult Megf10 KO mice, supporting published studies that MEGF10 regulates myogenesis by affecting satellite cell differentiation. Interestingly, muscle fibers do not exhibit morphological hallmarks of atrophy in either young adult or middle-aged Megf10 KO mice. We next examined the neuromuscular junction (NMJ), in which MEGF10 has been shown to concentrate postnatally, using light and electron microscopy. We found early and progressive degenerative features at the NMJs of Megf10 KO mice. These include increased postsynaptic fragmentation, axon terminals failing to completely appose the postsynaptic region and perisynaptic Schwann cells intruding into the NMJ synaptic cleft. These findings strongly suggest that the NMJ is a site of postnatal pathology in MEGF10-related myopathy. We next sought to identify molecular mechanisms altered in muscles lacking Megf10 using RNA-seq. We discovered genes and pathways associated with myogenesis, skeletal muscle health, and NMJ stability dysregulated in Megf10 KO mice compared to wild-type mice. Altogether, these data support using MEGF10 KO mice to discover and test potential treatments for MEGF10-related myopathy.

overall_design: To investigate the cellular and molecular changes to muscles and NMJs caused by loss of Megf10 in Megf10 global knockout mice.

first: Devin

last: Juron

first: Mary

middle: F

last: Avila

first: Robert

middle: L

last: Hastings

first: Ariane

last: Pendragon

first: Liam

last: Wilson

first: Jeremy

last: Kay


title: Mod5 mediates a molecular trade-off between transcriptional and translational regulation and antifungal resistance.

city: Jena

first: Matthew

middle: George

last: Blango

address: Adolf-Reichwein-Str. 23

department: Junior Resaerch Group RNA Biology of Fungal Infections

institute: Leibniz Institute for Natural Product Research and Infection Biology

summary: Increasing antifungal drug resistance is a major concern associated with human fungal infections. Genetic mutation and epimutation mechanisms are clearly linked to resistance in some fungi, but few studies have investigated RNA modifications (the epitranscriptome) as a mediator of resistance. Here, deletion of the Aspergillus fumigatus tRNA-modifying isopentenyl transferase ortholog, Mod5, led to altered growth and stress response, but also unexpected resistance against the antifungal drug 5-fluorocytosine (5-FC). Simultaneous profiling of the transcriptome and proteome of 5-FC-stressed wild-type and ∆mod5 strains revealed a similar general adaptation to stress; however, the knockout displayed elevated expression of cross-pathway control (CPC) genes. Quantification of CPC transcription factor cpcA and client gene argB showed premature CPC activation in ∆mod5, which was further increased upon treatment with 5-FC. By associating codon usage patterns with proteomics abundances, we observed the number of tRNATyrGΨA-decoded codons to negatively correlate with protein abundance in the knockout, indicative of modification-tunable transcripts. Overexpression of tRNATyrGΨA in the ∆mod5 strain rescued select phenotypes but, surprisingly, failed to reverse 5-FC resistance. Investigation of the purported tRNA gene-mediated silencing function of Mod5 uncovered a negative correlation between tRNA proximity and gene induction under normal growth, suggesting that tRNA gene-mediated silencing is active in A. fumigatus. In conclusion, 5-FC resistance in the absence of bifunctional Mod5 likely originates from multifaceted transcriptional and translational changes that skew the fungus towards starvation responses over optimal growth, potentially offering a mechanism reliant on RNA modification and tRNA gene-mediated silencing capable of facilitating transient antifungal resistance.

overall_design: Comparative gene expression profiling analysis of poly(A) RNA-seq data for 𝚫mod5 knockouts of Aspergillus fumigatus mycelium left unstressed for 24 hours in Aspergillus minimal media or grown for 16 hours unstressed followed by 8 hours stressed with 5-fluorocytosine.

first: Alexander

last: Bruch

first: Valentina

last: Lazarova


title: Sin3B loss enhances the T-cell inflamed tumor microenvironment through the redistribution of H3K27Ac signals in pancreatic cancer [RNA-seq]

city: Shanghai

first: Zhengyan

last: Zhang

address: State Key Laboratory of Systems Medicine for Cancer, Stem Cell Research Center, Ren Ji Hospital, School of Medicine, Shanghai Cancer Institute, Shanghai Jiao Tong University,

summary: Pancreatic ductal adenocarcinoma (PDAC) displays a poor response to immunotherapy. This poor response is predominantly attributed to the highly immunosuppressive tumor microenvironment (TME), characterized by a scarcity and dysfunction of infiltrating CD8+ T cells. Epigenetic regulators have recently been implicated in immune escape and immunotherapy sensitivity, presenting an appealing approach for directly targeting epigenetic factors or combining epigenetic therapies with immunotherapy in PDAC. Our study is the first to delineate the regulatory function of the epigenetic factor Sin3B in the tumor immune microenvironment (TIME) of PDAC. Using murine PDAC models, we discovered that intrinsic Sin3B loss in tumor cells reshapes the TIME, manifested by increased CD8+ T cell infiltration into tumors through enhanced secretion of  CXCL9 and CXCL10. This was accompanied by an increase in the cytotoxicity of CD8+ T cells, as evidenced by elevated Granzyme B (GzmB) and IFNγ production. Furthermore, Sin3B-deficient tumor cells exhibited heightened secretion of CXCL9/10 in response to IFNγ, creating a positive feedback loop via the CXCL9/10-CXCR3 axis and thus intensifying immune response against PDAC. Additionally, loss of Sin3B increased PDAC susceptibility to anti-PD1 therapy in mice. Corroborating our findings in the mouse model, analysis of human PDAC samples demonstrated a significant inverse correlation between Sin3B levels and both CD8+ T cell presence and CXCL9/10 expression. Notably, PDAC patients with lower Sin3B expression exhibited a more favorable response to anti-PD1 therapy. Concerning the mechanism, we systematically uncovered the extensive epigenetic regulation employed by Sin3B loss via the modulation of H3K27Ac redistribution in PDAC cells. Sin3B deficiency resulted in an uptick of H3K27Ac peaks concentrated in the promoter and enhancer regions. Under the treatment of IFNγ, Sin3B loss leads to the enrichment of H3K27Ac peaks in promoter regions of ISG-related genes such as CXCL9 and CXCL10, thereby boosting their expression. Collectively, our research outcomes propose a potential target for enhancing the accessibility of cytotoxic T cells to the tumor site and improving immunotherapy in the challenging landscape of PDAC.

overall_design: To investigate the function of Sin3B regulating TIME of PDAC, wo conducted Sin3B KO and WT KPC1199 cells. Then, we performed RNA sequencing to analyse gene expression profile of Sin3B KO and WT cells.

first: Yingying

last: Tang

first: Junyi

last: Xu

first: Jing

last: Xue


title: Spatial microniches of IL-2 synergize with IL-10 to drive lung migratory Th2 cells in response to inhaled allergen [Curio]

city: Pittsburgh

first: Amanda

middle: Catherine

last: Poholek

state: Pennsylvania

address: 4401 Penn Ave., Rangos Research Center 8129

department: Pediatrics

institute: University of Pittsburgh

summary: The mechanisms that guide Th2 cell differentiation in barrier tissues are unclear but important for the development of allergic asthma. Using temporal, spatial and single cell transcriptomic tracking of house dust mite (HDM) specific T cells, we describe the molecular pathways driving allergen-specific Th2 cells. Early Blimp-1 expression occurs in a subset of CD4 T cells in the lymph node prior to lung migration driven by IL-10 from allergen-specific T cells, but was not required for GATA3 upregulation. Instead, IL-2 via STAT5 was required to directly repress Bcl6 and Bach2 to support GATA3 and Blimp-1 upregulation. Loss of Blimp-1 during priming in the lymph node ablated the formation of Th2 cells that migrate to the lung, indicating early Blimp-1 promotes the population of Th2 cells with migratory capability. Spatial microniches of IL-2 in the lymph node discriminate and support these earliest Blimp-1+ migratory Th2 cells, demonstrating that lymph node localization is a primary driver of differentiation. Our findings illuminate the molecular pathways for inhaled allergens to promote Th2 cells and identify an early requirement for IL-2 mediated spatial microniches and allergen-driven IL-10 from responding T cells that drive allergic asthma.

overall_design: 1DER CD45.1 Tg T cells were adoptively transferred to CD45.2 host, followed by intrasnasal immunization with 25ug of house dust mite daily for three days. Mediastinal lymph nodes (mLN) were isolated on day 4 and day 6, snap frozen and embedded in OCT for Curio Biosciences Slide Seeker Tile (3mm x 3mm).

first: Kun

last: He

first: William

middle: A

last: MacDonald


title: Principles of the Earliest Lineage Specification and X Chromosome Dosage Compensation in Bovine Early Embryos

city: Hangzhou

first: Bingjie

last: Hu

address: Yuhangtang Rd, Hangzhou, Zhejiang , China

institute: Zhejiang University

summary: Lineage specification and X chromosome dosage compensation are two crucial biological processes that occur during preimplantation embryonic development. While these processes have been studied extensively in mice and humans, they are less understood in other species. This study aims to provide fundamental insights into bovine preimplantation development using single-cell RNA-sequencing. The study analyzes the transcriptomes of 286 individual cells and reveals that bovine trophectoderm/inner cell mass transcriptomes diverge at the early blastocyst stage, after cavitation but before blastocyst expansion. The study also identifies transcriptomic markers and provides the timing of lineage specification events in the bovine embryo. Moreover, the study confirms the occurrence of X chromosome dosage compensation from morula to middle blastocyst and reveals that this compensation results from downregulation of X-linked genes in female embryonic cells. The transcriptional atlas generated by this study is expected to be widely useful in improving our understanding of mammalian early embryo development.

overall_design: poly RNA profiles of  bovine embryos

first: Hao

last: Jin


title: Neonatal brain injury unravels transcriptional and signaling changes underlying the reactivation of cortical progenitors

city: Lyon

first: Guillaume

last: Marcy

address: 8 avenue Rockefeller

institute: Universite Lyon 1

summary:
We used chronic neonatal hypoxia as a rodent model of early brain injury, to investigate the reactivation of cortical progenitors at postnatal times.
We performed gene expression profiling analysis using data obtained from RNA-seq of normoxic and hypoxic brains at embryonic, P2 and P12 time points

overall_design: Comparative gene expression profiling analysis of RNA-seq data for progenitors (tdTomato+) cells in normoxic or hypoxic conditions at various time points (E13, E15 and P12)

first: Louis

last: Foucault

first: Olivier

last: Raineteau


title: M.tb adaptation to human alveolar lining fluid (ALF)

city: San Antonio

first: Anna

last: Allue Guardia

state: Texas

address: 8715 Military Dr W

institute: Texas Biomedical Research Institute

summary: Superbugs such as drug-resistant Mycobacterium tuberculosis (DR-M.tb) poses a serious global health threat. Upon infection, M.tb reaches the alveolar space and comes in close contact with soluble components of the human alveolar lining fluid (ALF) for an uncertain period of time prior to and following its encounter with alveolar compartment cells. Homeostatic ALF hydrolases, which main function is maintaining lung health, modify the M.tb cell envelope, driving M.tb-host cell interactions. Still, the contribution of human ALF to M.tb adaptation to the host during infection is poorly understood. Here, we exposed 4 M.tb strains (H37Rv, HN878, CDC1551, W-7642) with different levels of virulence, transmissibility, and drug resistance to human ALF for shorter (15 min) and longer (12 h) periods of time. RNA sequencing was performed to determine the strains’ transcriptional profiles in response to ALF exposure. Gene expression analysis showed a clear temporal and strain-specific adaptation to human ALF. When comparing ALF-exposed vs. unexposed bacteria for each M.tb strain, differential expression (DE) analysis revealed a total of 397 DE genes associated with lipid metabolism, cell envelope and processes, intermediary metabolism and respiration, and regulatory proteins, among others. Most DE was detected at 12 h post-ALF exposure, with DR-M.tb strain W-7642 having the highest number of DE genes. Interestingly, genes from the KstR2 regulon, which controls the degradation of cholesterol C and D rings, were significantly upregulated early in all strains post-ALF exposure. These results indicate that M.tb-ALF contact (15 min to 12 h prior to M.tb recognition by host cells in the alveolar space) drives global metabolic and physiologic changes in M.tb during the initial stages of the infection, with potential implications in infection outcome.

overall_design: RNAseq data of 4 different M.tb strains exposed to human Alveolar Lining Fluid (ALF) for 15min and 12h, 2 replicates each, control sample is unexposed bacteria for each of the strains. Total: 32 samples

last: Allué-Guardia

first: Andreu

last: Garcia-Vilanova

first: Alyssa

last: Schami

first: Angélica

last: Olmo-Fontánez

first: Amberlee

last: Hicks

first: Jay

last: Peters

first: Diego

last: Maselli

first: Mark

middle: D

last: Wewers

first: Yufeng

last: Wang

first: Jordi

middle: B

last: Torrelles


title: CD4+ T cells select unique P65, IRF1 and FOS dependen responses in APC for integration into antiviral CD8+ T cell immunity

city: Parkville

first: Jan

last: Schroeder

state: Victoria

address: Peter Doherty Institute Melbourne

department: DMI

country: Australia

institute: University of Melbourne

summary: Antiviral CD8+ T cell immunity depends on the integration of varying contextual cues, but how antigen presenting cells (APC ) consolidate these signals for decoding by T cells remains unclear. We describe gradual IFN-α/β-induced transcriptional adaptations that endowed APC with the capacity to rapidly activate the transcriptional regulators P65, IRF1 and FOS upon CD4+ T cell-mediated CD40 stimulation. While these responses operated through broadly utilized signaling components, they induced a unique set of costimulatory molecules and soluble mediators that could not be elicited by IFN-α/β or CD40 alone. This response was critical for the acquisition of antiviral CD8+ T cell effector function, and its activity in APC from patients infected with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) correlated with milder disease. These observations uncover a sequential integration process whereby APC rely on CD4+ T cells to select the innate circuits that guide antiviral CD8+ T cell responses.

overall_design: Cleavage under targets and tagmentation (CUT&Tag) for the transcription factor IRF1 in stimulated BM-derived cDC1.

first: Elise

last: Gressier

first: Jonas

last: Schulte-Schrepping

first: Lev

last: Petrov

first: Sophia

last: Brumhard

first: Paula

last: Stubbemann

first: Jasper

last: Spitzer

first: Tomislav

last: Kostevc

first: Paul

last: Whitney

first: Annabell

last: Bachem

first: Alexandru

last: Odainic

first: Carolien

last: van de Sandt

first: Oanh

last: Nguyen

first: Thomas

last: Ashhurst

first: Kayla

last: Wilson

first: Clare

last: Oates

first: Linden

last: Gearing

first: Tina

last: Meischel

first: Katharina

last: Hochheiser

first: Marie

last: Greyer

first: Michele

last: Clarke

first: Maike

last: Kreutzenbeck

first: Sarah

middle: S

last: Gabriel

first: Wolfgang

last: Kastenmüller

first: Christian

last: Kurts

last: Londrigan

first: Axel

last: Kallies

first: Katherine

last: Kedzierska

last: Hertzog

first: Eicke

last: Latz

first: Yu-Chen

middle: E

last: Chen

first: Kristen

last: Radford

first: Michael

last: Chopin

first: Leif

last: Sander

last: Gebhardt

first: Birgit

last: Sawitzki

first: Joachim

last: Schultze

first: Susanne

middle: V

last: Schmidt

first: Sammy

last: Bedoui


title: GENE EXPRESSION PROFILING IN PEDIATRIC APPENDICITIS

city: Vancouver

first: Robert

middle: E. W.

last: Hancock

state: British Columbia

address: 2259 - Lower Mall Research Station

department: Department of Microbiology & Immunology

country: Canada

institute: University of British Columbia

summary: Gene expression profiling of pediatric patients with simple appendicitis (SA) compared to perforated appendicitis (PA) and non-appendicitis abdominal pain (NAAP).

overall_design: RNA-Seq of peripheral whole blood collected from patients with: simple appendicitis (SA; n=35), perforated appendicitis (PA; n=14), and non-appendicitis abdominal pain (NAAP; n=22).

first: Bhavjinder

middle: K

last: Dhillon

first: Simone

last: Kortbeek

first: Arjun

last: Baghela

first: Mary

last: Brindle

first: Dori-Ann

last: Martin

first: Craig

middle: N

last: Jenne

first: Hans

last: Vogel

first: Amy

last: Lee

first: Graham

last: Thompson


title: Spatial microniches of IL-2 synergize with IL-10 to drive lung migratory Th2 cells in response to inhaled allergen [Spatial transcriptomics]

first: Amanda

last: Poholek

overall_design: 1DER CD45.1 Tg T cells were adoptively transferred to CD45.2 host, followed by intrasnasal immunization with 25ug of house dust mite daily for three days. Mediastinal lymph nodes (mLN) were isolated on day 4 and day 6, snap frozen and embedded in OCT for 10X Genomics Visium Spatial Gene Expression.

first: Kun

last: He

middle: C

first: William

last: MacDonald


title: Nfe2l3 promotes neuroprotection and long-distance axon regeneration after injury in vivo

city: Farmington

first: Ephraim

last: Trakhtenberg

state: CT

address: 263 Farmington Ave. RM L4005

department: Neuroscience

institute: University of Connecticut School of Medicine

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

overall_design: Refer to individual Series


title: PHF12 regulates HDAC1 to promote tumorigenesis via EGFR/AKT signaling pathway in non-small cell lung cancer [RNA-seq]

first: Yiru

last: Kong

state: Shanghai

address: 130 Dong 'an Road, Xuhui District, Shanghai

institute: Fudan University

summary: Our study identified for PHF12 an oncogenic role in lung cancer proliferation for the first time. PHF12 transcriptionally regulate HDAC1 and activate EGFR/AKT signaling pathway in NSCLC progression. PHF12 may serve as an important target in lung cancer therapy.

overall_design: To investigate the mechanism of PHF12 regulating signaling pathway, we studied H1299 and H1299 with PHF12 KD.

first: Rongrong

last: Jiang

first: Jing

last: Li

first: Xinli

last: Zhou


title: PHF12 regulates HDAC1 to promote tumorigenesis via EGFR/AKT signaling pathway in non-small cell lung cancer [ChIP-seq]

overall_design: To investigate the mechanism of PHF12 regulating transcription, we studied untreated H1299 cell line through ChIP assay using PHF12 antibody.


title: Spatial transcriptomic profiling of the aging mouse brain [spatial scRNA-seq]

city: Guangzhou

first: Cheng

last: Wu

address: No. 74, Zhongshan 2nd Road, Yuexiu District, Guangzhou

institute: Sun Yat-sen University

summary: The mammalian brain can be divided into distinct structural and functional regions to perform a variety of diverse functions, but during normal aging, exactly how each region is affected, and the information interaction changes between different regions, remains largely unknown. To gain a better insight into these processes, here we generate a single-cell spatial transcriptomic (ST) atlas of young and old mice brains involving cerebrum, brain stem and fiber tracts regions. Based on the unbiased classification of spatial molecular atlas, 27 distinguished brain spatial domains were obtained, which are similar to known anatomical regions, but slightly different. Through differential expression analysis and gene set enrichment analysis (GSEA), we identified aging-related genes and pathways that vary in a coordinated or opposite manner across regions. Combined with single-cell transcriptomic data, we characterized the spatial distribution of cell types, identified an up-regulated gene Ifi27 across regions and cell types in VIS region. Through ligand-receptor interaction analysis, we identified all possible information interaction changes between regions with aging. In summary, we establish a brain spatial molecular atlas (accessible online at https:) to provide a rich resource of spatially differentially expressed genes and information interaction, which may help to understand aging and provide novel insights into the molecular mechanism of brain aging.

overall_design: 8 slices from young (2 months old) and old mice (28 months old) (strain C57BL/6J) were prepared according to Visium spatial protocols of tissue preparation guide (10x genomics). The brains were rapidly extracted from the cranial cavity, and immediately submerged in ice-cold artificial cerebrospinal fluid. The brains were then blotted to remove excess liquid. The brains were subsequently embedded in Optimal Cutting Temperature compound (OCT) and frozen in isopentane (2-methylbutane, Sigma) precooled with dry ice and liquid nitrogen. The brains were cut into 10-um sections.

first: Xie

last: Zhi

first: Wu

last: Cheng


title: RBP Footprint Grand Challenge: An evaluation of novel computational approaches to RNA-binding protein target prediction from structural data

city: LA JOLLA

first: Hsuan-Lin

last: Her

address: 9500 Gilman Dr,

institute: UCSD

summary: The Froehlking team uses ADAR eCLIP data to train an generative model to predict ADAR binding site.

overall_design:
Protein-bound nucleotides have been shown to display high eCLIP, and in some systems and for select nucleotides, low icSHAPE reactivities at the same time. However, obtaining both eCLIP and icSHAPE data is expensive. We thus built a machine learning model that can be used to generate sequences characteristic of protein-bound regions from experimental data, even if some information is missing, and to predict protein binding sites (details can be found in Materials and Methods of associated publication).
ADAR eCLIP in K562 cells.

first: Steven

last: Blue

first: Jasmine

last: Mueller

first: Sam

last: Park

first: Gene

last: Yeo

first: Brian

last: Yee


title: PHF12 regulates HDAC1 to promote tumorigenesis via EGFR/AKT signaling pathway in non-small cell lung cancer


title: The single cell RNAseq analysis of regenerated mouse lung in mouse-rat interspecies

status: Public on Feb 19 2024

city: PHOENIX

first: Bingqiang

last: Wen

address: 475 N 5th st

department: Child Health

institute: The University of Arizona

summary: we report the application of single cell RNAseq analysis of mouse lung from E20.5 Nkx2-1-/- rat (mutant), and mouse-rat chimeric lung from E20.5 wt rat (control).

overall_design: The mouse lung from Nkx2-1-/- rat (mutant) and mouse-rat chimeric lung from wt rat (control) were prepared from E20.5 mouse-rat interspecies. The samples were dissociated into small pieces and digested into single cell suspension which used to prepare single cell RNA 3' v3 library for sequencing with 10X Genomics platform.

first: Vladimir

last: Kalinichenko

first: Enhong


title: Effect of ovariectomy and LCAT overexpression on gene expression of mouse liver cancer

city: Wuhan

first: Wenzhi

last: He

state: None Selected

address: No. 299 Bayi Road

institute: Wuhan University

summary: To unravel the molecular mechanisms behind estrogen-mediated alleviation of liver cancer, we initially established a liver cancer model in ovariectomized mice. Through cross-species comparison of differentially expressed genes under estrogen dominance in human and murine liver cancers, we identified LCAT as a crucial downstream molecule mediating estrogen-induced gender disparities in liver cancer. Subsequent transcriptome sequencing of mouse liver tissues overexpressing the LCAT gene revealed that LCAT plays a pivotal role in inhibiting the cholesterol synthesis pathway, potentially serving as a key molecular mechanism through which it suppresses the occurrence and progression of liver cancer.

overall_design:
To elucidate the molecular mechanisms underlying estrogen-mediated mitigation of liver cancer, we initially induced primary liver cancer in ovariectomized female mice and those subjected to sham surgery. To uncover the molecular mechanisms by which LCAT alleviates liver cancer, we induced primary liver cancer in mice through overexpression of LCAT in the liver, comparing it with a control group.
We then obtained mouse liver tissues at the terminal sampling time point for transcriptome sequencing analysis
Comparative gene expression profiling analysis of RNA-seq data between the group Sham and OVX, or Vec and LCAT.

first: Zan

last: Huang

first: Yongping


title: Gene expression profile at single cell level of human pneumocytes and fibroblasts cultured as organoids

city: Homburg

first: Felix

last: Ritzmann

address: Kirrberger Street

institute: Saarland University

summary: Alveolar type 2 (AT2) are part of the stem cell niche of the lung and their differentiation is required for pulmonary homeostasis and tissue regeneration. A disturbed crosstalk between fibroblasts and epithelial cells contributes to the loss of lung structure in chronic lung diseases. We used single cell RNA sequencing to analyze the interaction of human fibroblasts and the alveolar epithelium modelled in organoid cultures.

overall_design: Human AT2 cells were seeded in Matrigel in co-culture with or without primary fibroblasts. Single cell suspensions were obtained after 21 days and single cell transcriptome profiling was performed by BD Rhapsody™ Single-Cell Analysis System.

first: Christoph

last: Beisswenger

first: Yiwen

last: Yao

first: Robert

last: Bals


title: Topical Ruxolitinib in the Treatment of Necrobiosis Lipoidica: A Prospective, Open-Label Study

city: Rochester

first: Tao

last: Ma

address: 200 1st ST SW

institute: Mayo Clinic

summary:
Necrobiosis Lipoidica (NL) is a rare granulomatous disease. There are few effective treatments for NL. We sought to investigate the efficacy and safety of the JAK 1/2 inhibitor, ruxolitnib, in the treatment of NL and identify biomarkers associated with disease and treatment response. We conducted an open-label, phase 2 study of ruxolitinib in 12 patients with NL.
We performed transcriptomic analysis of tissue samples pre- and post-treatment. At week 12, the mean NL lesion score decreased by 58.2% (SD 28.7%, P=0.003). Transcriptomic analysis demonstrated enrichment of type I and type II interferon pathways in baseline disease. Weighted Gene Co-expression Network Analysis (WGCNA) demonstrated post-treatment changes in interferon pathways with key hub genes IFNG and STAT1. Limitations include small sample size and a study group limited to patients with <10% BSA. In conclusion, ruxolitinib is an effective treatment for NL and targets the key pathogenic mediators of disease.

overall_design: Whole transcriptomic sequencing was performed on lesional and control (normal skin) tissue before therapy (n = 12 and 12, respectively) and lesional responsive and refractory after therapy (n = 8 and 4, respectively).

first: Angelina

last: Hwang

first: Jacob

last: Kechter

first: Xing

last: Li

first: Alysia

last: Hughes

first: Kevin

middle: J

last: Severson

first: Blake

last: Boudreaux

first: Puneet

last: Bhulla

first: Shams

last: Nassir

first: Miranda

last: Yousif

last: Zhang

first: Richard

last: Butterfield

last: Nelson

first: Xianying

last: Xing

first: Lam

last: Tsoi

first: Samantha

last: Zunich

first: Aleksandar

last: Sekulic

first: Mark

last: Pittelkow

first: Johann

last: Gudjonsson

first: Aaron

last: Mangold


title: Multiplex single cell analysis of serotonergic neuron function in planarians reveals widespread effects in diverse cell types

city: Exeter

first: Jordi

last: Solana

address: Northcote House

department: Health and Life Sciences

institute: University of Exeter

summary: We used scRNA-seq to examine serotonergic function in asexual planarians. We profiled 47,292 single cell transcriptomes of planarian of 3 different RNAi categories: gfp(RNAi), lhx1/5-1(RNAi) and pitx(RNAi). We detected changes in cell type frequency and gene expression patterns in planarian of different knock-down conditions, capturing regulatory programs of distinct cell types in response to loss of serotonergic function.

overall_design: We obtained cell dissociations from adult populations of the asexual strain of Schmidtea mediterranea of serotonergic trascription factors (pitx and lhx1/5-1) RNAi conditions.

first: Elena

last: Emili

first: Dianalí

last: Rodríguez-Fernández

first: Alberto

last: Pérez-Posada

first: Helena

last: García-Castro


title: Cell type-specific expression of tRNAs in the brain regulates cellular homeostasis.

first: Carlos

last: Guzman

address: 9262 REGENTS RD

summary: Defects in tRNA biogenesis are associated with diverse neurological disorders, yet our understanding of these diseases has been hampered an inability to determine tRNA expression in individual cell types within a complex tissue. Here, we developed a mouse model in which RNA Polymerase III is conditionally epitope-tagged in a Cre-dependent manner, allowing us to accurately profile tRNA expression in any cell type in vivo. We investigated tRNA expression in eleven diverse nervous system cell types, revealing dramatic heterogeneity in the expression of tRNA genes between populations. We found that while maintenance of levels of tRNA anticodon pools is critical for cellular homeostasis, neurons are differentially vulnerable to insults to distinct tRNA anticodon families. Cell type-specific translatome analysis suggests that the balance between tRNA availability and codon demand may underlie such differential resilience. Our work provides a platform for investigating the complexities of mRNA translation and tRNA biology in the brain.

overall_design: The Polr3a-cKI mouse line was crossed to appropriate Cre driver lines to obtain Polr3a Fl/+, CRE + mice, expressing FLAG-tagged POLR3A in the appropriate cell populations. A total of 11 CNS cell types were analyzed. In addition, ChIP was performed on whole liver (n=2) and cerebellum (n=4) from global knock-in mice (Polr3A FLAG/FLAG) mice, expressing FLAG-tagged POLR3A in every cell-type. FLAG-tagged POLR3A and associated DNAs were immunoprecipitated using FLAG antibody (Sigma, F1804) and protein G dynabeads from the appropriate brain regions of 6-week-old mice. DNA libraries were made using the NEBNext Ultra II Library preparation kit (NEB E7645S) and KAPA Single-Indexed Adapters for Illumina (Roche, KR1317) according to manufacturer’s protocol.The design includes two biological replicates for the immunoprecipitation and one input library for each cell type.

first: Michael

last: Molumby

first: Mridu

last: Kapur

first: Sven

last: Heinz

first: Susan

last: Ackerman


title: A conserved trypanosomatid differentiation regulator controls substrate attachment and morphological development in Trypanosoma congolense

city: Edinburgh

first: Alasdair

last: Ivens

address: Kings Buildings

institute: Centre for Immunity, Infection and Evolution

summary: Trypanosomatid parasites undergo developmental regulation to adapt to the different environments encountered during their life cycle. In Trypanosoma brucei, a genome wide selectional screen previously identified a regulator of the protein family ESAG9, which is highly expressed in stumpy forms, a morphologically distinct bloodstream stage adapted for tsetse transmission. This regulator, TbREG9.1, has an orthologue in Trypanosoma congolense, despite the absence of a stumpy morphotype in that parasite species, which is an important cause of livestock trypanosomosis. RNAi mediated gene silencing of TcREG9.1 in Trypanosoma congolense caused a loss of attachment of the parasites to a surface substrate in vitro, a key feature of the biology of these parasites that is distinct from T. brucei. This detachment was phenocopied by treatment of the parasites with a phosphodiesterase inhibitor, which also promotes detachment in the insect trypanosomatid Crithidia fasciculata. RNAseq analysis revealed that TcREG9.1 silencing caused the upregulation of mRNAs for several classes of surface molecules, including transferrin receptor-like molecules, immunodominant proteins, and molecules related to those associated with stumpy development in T. brucei. Depletion of TcREG9.1 in vivo also generated an enhanced level of parasites in the blood circulation consistent with reduced parasite attachment to the microvasculature. The morphological progression to insect forms of the parasite was also perturbed. We propose a model whereby TcREG9.1 acts as a regulator of attachment and development, with detached parasites being adapted for transmission.

overall_design: Trypanosoma congolense cells were trnasfected to allow doxycycline indiucible gene silencing of TcREG9.1 (TcIL3000.11.14540.1 ) and two clones analysed for their gene expression by RNAseq under conditions of doxycycline for 3 days, or in the absnece of doxycycliune

first: Eleanor

last: Silvester

first: Keith

last: Matthews


title: 14-3-3 binding motif phosphorylation disrupts Hdac4 organized condensates to stimulate cardiac reprogramming

city: ANN ARBOR

first: Liu

last: Liu

state: MICHIGAN

address: 2800 Plymouth Rd,SPC 2800

department: Cardiac Surgery

institute: University of Michigan

summary: Cell fate conversion is associated with extensive post translational modifications (PTMs) and architectural changes of sub-organelles, yet how these events are interconnected remains unknown. We report here the identification of a phosphorylation code in 14-3-3 binding motifs (PC14-3-3) that greatly stimulates induced cardiomyocyte (iCM) formation from fibroblasts. PC14-3-3 is identified in pivotal functional proteins for iCM reprogramming, including transcription factors and epigenetic factors. Akt1 kinase and PP2A phosphatase are key writer and eraser of the PC14-3-3 code, respectively. PC14-3-3 activation induces iCM formation with the presence of only Tbx5. In contrast, PC14-3-3 inhibition by mutagenesis or inhibitor-mediated code removal abolishes reprogramming. We discover that key PC14-3-3 embedded factors, such as Hdac4, Mef2c and Foxo1, form Hdac4 organized inhibitory nuclear condensates. PC14-3-3 activation disrupts Hdac4 condensates to promote cardiac gene expression. Our study suggests that sub-organelle dynamics regulated by a PTM code could be a general mechanism for stimulating cell reprogramming.

overall_design: To investigate the effect of the HDAC4 inhibitor TMP269 on cardiac reprogramming, we utilized TMP269 to treat neonatal cardiac fibroblasts transfected with MGT. Additionally, cells infected with the MGT virus or DsRed were employed as positive and negative controls, respectively. We subsequently conducted gene expression profiling analysis using data obtained from RNA sequencing of three different cell types 14 days after virus infection.

first: Zhong

last: Wang


title: Coordinated excitatory and inhibitory circuits governing unpredictable threat induced anxiety

city: jinan

first: Shuaiwen

last: Teng

address: wenhuaxi road 44

institute: Shandoong University

summary: Anxiety is elicited by excessive apprehension about unpredictable threats. However, the neural circuit governing unpredictable threat induced anxiety remains unclear. Here, we found ventral bed nucleus of the stria terminalis (vBNST) GABAergic neurons displayed selective activation to unpredictable threats by means ofthrough coordinated excitatory input from insular cortex (IC) glutamergic neurons and inhibitory input from lateral nucleus of the amygdala (CeL) somatostatin (SOM) neurons. Using activity-dependent neuronal tagging technology, we found that unpredictable threat responsive cells in vBNST drive freezing and anxiety via projections to ventral lateral periaqueductal grey (vlPAG) and median nucleus of the amygdala (CeM) respectively. Finally, we identified KCNQ3 plays an essential role in hyperactivity of vBNST GABAergic neurons and induced anxiety. These data identified a forward inhibitory circuit that determine the selective activation of vBNST in unpredictable threat and anxiety, and suggest that Kcnq3 KCNQ3 channel acts as a promising target in treatment of anxiety disorder following unpredictable stress.

overall_design: The molecular mechanism underlying the altered activity of vBNST CS- responsive ensemble to unpredictable threats is still unknown. We thus employed phosphorylated translating ribosome affinity purification (pTRAP) combining with RNA sequencing to screen the key molecules involved in vBNST’s response to unpredictable threats (Knight et al., 2012; Zhang et al., 2022). vBNST were prepared from mice 90 min after CS+ or CS- test and the precipitated ribosomal RNA was sequenced following immunoprecipitated (IP) with a phosphoS6 (Ser244, Ser247)-specific antibody

first: Zheyu

last: Chen

first: Xinyue

last: Wei


title: Spatial transcriptomic validation of a biomimetic model of fibrosis enables re-evaluation of a therapeutic antibody targeting LOXL2

city: SOUTHAMPTON

first: Joseph

last: Bell

state: Hampshire

address: Tremona Road

department: Medicine

institute: Clinical and Experimental Sciences, Southampton General Hospital

summary: Matrix stiffening by lysyl oxidase-like 2 (LOXL2) mediated collagen cross-linking is proposed as a core feed-forward mechanism that promotes fibrogenesis. Failure in clinical trials of simtuzumab (the humanized version of AB0023, a monoclonal antibody against human LOXL2) suggested targeting LOXL2 may not have disease relevance, however target engagement was not directly evaluated. We compared the spatial transcriptome of active human lung fibrogenesis sites with different human cell culture models to determine the model with greatest disease relevance. Within the selected model system, we then evaluated AB0023, identifying that it did not inhibit collagen cross-linking or reduce tissue stiffness, nor did it inhibit LOXL2 catalytic activity. In contrast it did potently inhibit angiogenesis consistent with an alternative, non-enzymatic mechanism of action. Thus, AB0023 is anti-angiogenic but does not inhibit LOXL2 catalytic activity, collagen cross-linking or tissue stiffening. These findings have implications for the interpretation of lack of efficacy of simtuzumab in clinical trials of fibrotic diseases.

overall_design: To compare different model systems for fiboblastic foci, we cultured primary fibroblasts in different culture conditions including standard 2D culture, 2D culture with ficoll (scar-in-a-jar), and 3D spheroids, with and without the profibrotic cytokine TGF-β

first: Elizabeth

middle: R

last: Davies

first: Christopher

last: Brereton

first: Milica

last: Vukmirovic

first: James

last: Roberts

first: Kerry

last: Lunn

first: Leanne

last: Wickens

first: Franco

last: Conforti

first: Robert

last: Ridley

first: Aiman

last: Alzetani

first: Sanjay

last: Jogai

first: Ben

last: Marshall

first: Aurelie

last: Fabre

first: Luca

last: Richeldi

first: Phillip

last: Monk

first: Paul

last: Skipp

first: Naftali

last: Kaminski

first: Emily

last: Offer

first: Yihua

first: Donna

first: Mark

last: Jones


title: Cell-type-specific expression of tRNAs in the brain regulates cellular homeostasis

city: La Jolla

first: Mridu

last: Kapur

address: 9500 Gilman Drive

department: Cellular and Molecular Medicine

institute: University of California San Diego

summary: Defects in tRNA biogenesis are associated with multiple neurological disorders, yet our understanding of these diseases has been hampered an inability to determine tRNA expression in individual cell types within a complex tissue. Here, we developed a mouse model in which RNA Polymerase III is conditionally epitope-tagged in a Cre-dependent manner, allowing us to accurately profile tRNA expression in any cell type in vivo. We investigated tRNA expression in eleven diverse nervous system cell types, revealing dramatic heterogeneity in the expression of tRNA genes between populations. We found that while maintenance of levels of tRNA anticodon pools is critical for cellular homeostasis, neurons are differentially vulnerable to insults to distinct tRNA anticodon families. Cell type-specific translatome analysis suggests that the balance between tRNA availability and codon demand may underlie such differential resilience. Our work provides a platform for investigating the complexities of mRNA translation and tRNA biology in the brain.

overall_design: The RiboTag mouse line (B6N.129-Rpl22tm1.1Psam/J mice) was crossed to appropriate Cre driver lines to obtain Rpl22 Fl/+, CRE + mice, expressing HA-tagged Rpl22 in the appropriate cell populations. The following populations were profiled: Cerebellar granule cells (Gabra6-cre, cerebellum), layer V cortical neurons (Rbp4-cre, cortex), dentate gyrus granule cells (Rbp4-cre, hippocamus), inhibitory interneurons (Dlx6a-cre, hippocampus), astrocytes (Aldh1-cre, cortex), and microglia (Tmem119-cre, cortex). HA-tagged RPL22 and associated mRNAs were immunoprecipitated using Pierce anti-HA magnetic beads (ThermoFisher Scientific) from the appropriate brain regions of 6-week-old mice, and mRNA libraries were made using the Kapa Stranded mRNA-Seq Kit (Roche) according to the manufacturer’s protocol.The design includes three biological replicates for each cell type. For cerebellar granule cells, astrocytes, layer V cortical neurons, and dentate gyrus granule cells, one sample each was sequenced twice to account for batch effects.

first: Susan

last: Ackerman


title: HyperTRIBE assay of VSV-GFP-infected HEK293T cells overexpressing ADAR1-CD, PTENα-N(WT)-ADAR1-CD, or PTENα-N(6R/D)-ADAR1-CD fusion protein

city: Beijing

first: Zeliang

last: Yang

address: No.38 Xueyuan Road,Haidian

institute: Peking University Health Science Center

summary: To explore the binding sites of PTENα on viral or host RNAs, we performed HyperTRIBE assay of HEK293T cells overexpressing ADAR1-CD, PTENα-N(WT)-ADAR1-CD, or PTENα-N(6R/D)-ADAR1-CD fusion protein under VSV-GFP (green fluorescent protein-expressing vesicular stomatitis virus) infection.

overall_design: Five 10cm2 dishes of HEK293T cells at 70% confluency were transfected with 10μg of pCCL-EF1a-CMV-NGFR plasmid encoding ADAR1-CD, PTENα-N(WT)-ADAR1-CD, or PTENα-N(6R/D)-ADAR1-CD fusion protein per dish. At 24 hours after transfection, NGFR-positive cells were sorted and infected with VSV-GFP (MOI = 0.01). 6 hours later, cells were collected  and total RNA was separated using TRIzol reagent.

first: Xin

last: Zhang

first: Dan

last: Lu


title: RNA-seq of wild-type and Ptenα-/- MEFs under VSV infection

summary: To explore the effect of Ptenα on antiviral immunity pathway, we conducted RNA transcriptome profiling of wild-type and Ptenα-/- MEFs (mouse embryonic fibroblast cells) in response to VSV (vesicular stomatitis virus)

overall_design: We infected wild-type and Ptenα-/- MEFs with VSV for eight hours and perform RNA sequecing

first: Yue

last: Yin

last: Lv