title: Antigen Specific VNAR Screening in Whitespotted Bamboo Shark (Chiloscyllium plagiosum) with Next Generation Sequencing

status: Public on Jan 01 2024

city: hangzhou

first: bo

last: zhong

state: zhejiang

address: No. 928, 2nd Street, Xiasha Higher Education Park, Qiantang District, Hangzhou City, Zhejiang Province

institute: Zhejiang Sci-Tech University

summary: IgNAR exhibits significant promise in the fields of cancer and anti-virus biotherapies. Notably, the variable regions of IgNAR (VNAR) possess comparable antigen binding affinity with much smaller molecular weight (~12 kDa) compared to IgNAR. Antigen specific VNAR screening is a changeling work, which limits its application in medicine and therapy fields. Though phage display is a powerful tool for VNAR screening, it has a lot of drawbacks, such as small library coverage, low expression levels, unstable target protein, complicating and time-consuming procedures.  Here we report VNAR screening with next generation sequencing (NGS) could effectively overcome the limitations of phage display, and we successfully identified approximately 3000 BAFF-specific VNARs in Chiloscyllium plagiosum vaccinated with the BAFF antigen. The results of modelling and molecular dynamics simulation and ELISA assay demonstrated that one out of the top five abundant specific VNARs exhibited higher binding affinity to the BAFF antigen than those obtained through phage display screening. Our data indicates NGS would be an alternative way for VNAR screening with plenty of advantages.

overall_design:
Next Generation Sequencing results of VNAR in immune BAFF antigen and untreated  Whitespotted Bamboo Shark (Chiloscyllium plagiosum)
Different primer pairs were used for VNAR libraries' preparations. NR means the library was prepared with primers IgNAR_NF&IgNAR_NR. OR means the library was prepared with primers IgNAR_NF&IgNAR_OR.

first: Bo

last: Zhong

first: Shengjie

last: Sun

first: Zhan

last: Luo

first: Guodong

last: Chen


title: Rescue of blood coagulation Factor VIII exon-16 mis-splicing by antisense oligonucleotides

status: Public on Jan 17 2024

city: Santa Cruz

first: Guillermo

middle: A

last: Chacaltana

state: CA

address: 1156 High Street

department: Chemistry

country: USA

institute: University of California, Santa Cruz

summary: The human Factor VIII (F8) protein is essential for the blood coagulation cascade and specific F8 mutations cause the rare bleeding disorder Hemophilia A (HA). Here, we investigated the impact of HA-causing single-nucleotide mutations on F8 pre-mRNA splicing. We found that 14/97 (∼14.4%) coding sequence mutations tested in our study induced exon skipping. Splicing patterns of 4/11 (∼36.4%) F8 exons tested were especially sensitive to the presence of common disease-causing mutations. RNA-chemical probing analyses revealed a three-way junction structure at the 3′ end of intron 15 (TWJ-3-15). TWJ-3-15 sequesters the polypyrimidine tract, a key determinant of 3′ splice site strength. Using exon-16 of the F8 gene as a model, we designed specific antisense oligonucleotides (ASOs) that target TWJ-3-15 and identified three that promote the splicing of F8 exon-16. Interaction of TWJ-3-15 with ASOs increases accessibility of the polypyrimidine tract and inhibits the binding of hnRNPA1-dependent splicing silencing factors. Moreover, ASOs targeting TWJ-3-15 rescue diverse splicing-sensitive HA-causing mutations, most of which are distal to the 3’ splice site being impacted. The TWJ-3-15 structure and its effect on mRNA splicing provide a model for HA etiology in patients harboring specific F8 mutations and provide a framework for precision RNA-based HA therapies.

overall_design: F8 Exon16 RNAs (WT & MTs of interest) were chemically probed using a SHAPE reagent, 2-aminopyridine-3-carboxylic acid imidazolide (2A3 which creates chemical adducts in the backbone of the RNA that can be readout as mismatches at the DNA level at positions where these adducts were made, or DMSO a non-modifying agent as a control. Replicates of the same experimental condition in these experiments are labeled as "A" or "B" (32 different experiments, 64 total samples sequenced), ex: F8Exon16_A_DMSO > F8Exon16 Replicate A, DMSO control; F8Exon16_A_2A3 > F8Exon16 Replicate A, 2A3 treated.

first: Victor

last: Tse

first: Martin

last: Gutierrez

first: Nicholas

last: Forino

first: Arcelia

last: Jimenez

first: Michael

last: Stone

first: Jeremy

last: Sanford


title: Periostin Upregulated in Cancer-Associated Fibroblasts upon Direct Contact with Cancer Cells Promotes Esophageal Squamous Cell Carcinoma Progression by Enhancing the Migration of Cancer Cells and Stromal Cells

status: Public on Jan 08 2024

city: Kamakura

first: Satoshi

last: Kondo

state: Kanagawa

address: 10-1 tebiro 6-chome

country: Japan

institute: Toray Industries,Inc.

summary:
Cancer-associated fibroblasts (CAFs) in the tumor microenvironment are involved in the progression of esophageal squamous cell carcinoma (ESCC). We generated CAF-like cells by direct co-culture of human bone marrow-derived mesenchymal stem cells (MSCs), one of the origins of CAFs, with ESCC cell lines and found that periostin is highly expressed in CAF-like cells. Periostin activated Akt and Erk signaling pathways in ESCC cells, enhancing survival and migration via integrin β4, one of the receptors for periostin. Periostin also enhanced migration of MSCs and macrophages and caused macrophages to acquire tumor-associated macrophage (TAM)-like properties.
High periostin expression in cancer stroma was associated with several clinicopathological factors and expressions of CAF markers and numbers of infiltrating TAMs. Moreover, ESCC patients with high periostin expression exhibited significantly poor postoperative outcomes. Thus, periostin secreted from CAFs enhanced the migration of ESCC cells, MSCs, and macrophages and contributed to developing the tumor microenvironment. These results indicate that periostin may be a novel therapeutic target for ESCC.

overall_design: To induce the CAF-like cells, MSCs and TE cells were seeded in the same dish. Direct co-culture was performed for 4 days in DMEM high-glucose (Wako) with 10% FBS and 1% antibiotic-antimycotic. MSCs and TE cells were seeded and cultured similarly without co-culture as controls. Cell suspensions were mixed with CD326 (EpCAM) microbeads (#130-061-101; Miltenyi Biotec) and incubated at 4°C for 30 minutes. Prepared cell suspensions were separated from EpCAM-positive and -negative cells using the autoMACS Pro Separator. Mono-cultured TE cells and mono-cultured MSCs were collected using autoMACS Pro Separator as well as cells after direct co-culture. We performed cDNA microarray analysis between mono-cultured MSCs and CAF9 (MSC co-cultured with TE-9 cells) cells.

first: S

last: Miyako

first: Y

last: Koma


title: Liver transcriptome of an LPS infusion model for porcine sepsis

status: Public on Dec 31 2023

city: Gent

first: Steven

last: Timmermans

address: Technologiepark 71

country: Belgium

institute: VIB-UGent

type: Expression profiling by high throughput sequencing

summary: Sepsis is a life-threatening organ dysfunction resulting from a dysregulated host response to infection. This is best studied in humans and various mouse models, however finding sof these models to not always translate easy to the clinic. In order to improve this transfer, and because sepsis also plays a significant role in vetrenairy medicine, we use the pig as a model organism in sepsis research. We compare two modes of porcine sepsis iduction: fecal infusion and LPS infusion and also compare thse o wha we can find in mice

overall_design: 2 conditions LPS and PBS (controls) each with 3 biological replicates

first: Fëllanza

last: Halimi

first: Tineke

last: Vanderhaeghen

first: Bruno

last: Garcia

first: Jolien

last: Vandewalle

first: Antoine

last: Herpain

first: Laura

last: Dhont

first: Siska

last: Croubels

first: Claude

last: Libert


title: Integrative multi-omics analysis of intratumoral and intertumoral heterogeneity in malignant pleural mesothelioma

status: Public on Jan 04 2024

city: Houston

first: Sung Wook

last: Kang

state: TX

address: one plaza

department: Surgery

institute: Baylor college of medicine

summary: Malignant pleural mesothelioma (MPM) is a deadly pleural malignancy with over 90% mortality within five years and a median survival of approximately 7 months without treatment. The current first-line chemotherapy, a combination of pemetrexed and cisplatin, modestly extends survival, with the addition of bevacizumab offering a further 2-month benefit. Radiation therapy, while not effective as a primary treatment due to the extensive field required, plays a role in multimodal approaches or as localized therapy.  The diversity in patient outcomes and responses to treatments indicates significant tumor heterogeneity in MPM, underscoring the need for therapies that consider this variability and target the molecular specifics of individual MPM cases. Tumor heterogeneity is observed at both intertumor and intratumor levels, encompassing tumor cells and their microenvironments. Molecular profiling studies have further highlighted this heterogeneity, complicating the identification of a unique biomarker for MPM.

overall_design: To address these challenges and improve clinical outcomes, an enhanced understanding of MPM's biological heterogeneity is crucial. This can inform prognostic assessments and the development of precise therapeutic strategies, including immunotherapy and targeted therapy. Our approach to dissecting MPM heterogeneity involves comprehensive testing using whole exome sequencing (WES), total RNA sequencing, mass cytometry, and mass spectrometry on tissue samples collected from three distinct sites during macroscopic complete resection. This multi-faceted analysis promises to deepen our understanding of MPM and pave the way for more personalized treatment modalities.

first: Hyun-Sung

last: Lee

first: Christopher

last: Amos

first: Bryan

last: Burt

first: SungWook


title: Identification of GLIS3 and HNF1b target genes in kidney

city: Research Triangle Park

first: Hong Soon

state: NC

address: 111 T.W. Alexander Dr.

department: IIDL

institute: NIH/NIEHS

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Comparison of cistromes from GLIS3 and HNF1b ChIP-Seq analysis using mouse kidney was performed to examine whether there was a significant overlap in target genes between GLIS3 and HNF1b.

overall_design: GLIS3 ChIP-Seq was performed with kidneys from Glis3-eGFP mice using GFP antibody, and HNF1b ChIP-Seq was performed with WT kidneys using HNF1b antibody (Invitrogen #720259).

first: Anton

middle: M

last: Jetten

first: Hong

middle: S

first: Justin

last: Collier

first: Yun-Gil

last: Roh

first: Sara

last: Grimm


title: Borrelia burgdorferi fitness is constrained during Peromyscus leucopus infection

status: Public on Jan 03 2024

city: Boston

first: Jeffrey

last: Bourgeois

address: 136 Harrison Ave

institute: Tufts University

summary: The Lyme disease spirochete Borrelia burgdorferi drives a range of acute and chronic maladies in humans and other incidental hosts infected with the pathogen. However, the primary vertebrate reservoir, Peromyscus leucopus appears spared from any symptomology following infection. This has led to a common hypothesis that P. leucopus and B. burgdorferi exist symbiotically: P. leucopus restrain their immune response against the microbe and enable the enzootic cycle while B. burgdorferi avoids causing damage to the host. While aspects of this hypothesis have been tested, the exact interactions that occur between P. leucopus and B. burgdorferi during infection remain largely unknown. Here we utilized an inbred colony of P. leucopus in order to compare B. burgdorferi (B31) fitness in these rodents to the traditional B. burgdorferi murine models—C57BL/6J and C3H/HeN Mus musculus, which develop signs of inflammation akin to human disease. We find that in contrast to our expectations, B. burgdorferi were able to reach much higher burdens in M. musculus, and that the overall kinetics of infection differed between the two rodent species. Surprisingly, we also found that P. leucopus remained infectious to larval Ixodes scapularis for a far shorter period than either M. musculus strain. In line with these observations, we found that P. leucopus does launch a modest but sustained inflammatory response against B. burgdorferi in the skin, which we hypothesize leads to reduced bacterial viability and infectivity in these hosts. These observations provide new insight into the nature of reservoir species and the B. burgdorferi enzootic cycle.

overall_design: Ear punches were collected from P. leucopus following infection with B. burgdorferi strain B31 at 1 week, 2 weeks, 4 weeks, and 8 weeks post infected nymph placement and compared to ear punches collected from rodents one week following uninfected nymph placement. Ear punches were placed directly in RNA-later reagent overnight. Samples were frozen at -80C before beadbeating in QIAzol and RNA isolation with the miRNeasy kit by Qiagen with an additional RWT wash. TURBO Dnase was used to degrade DNA following isolation and samples were cleaned with RNeasy MinElute Cleanup Kit by Qiagen. Purified RNA was sent to Azenta/Genewiz for library preparation and sequencing (Illumina HiSeq 2x150bp)

first: Stephanie

last: You

first: Luke

middle: H

last: Clendenen

first: Muskan

last: Shrestha

first: Tanja

last: Petnicki-Ocwieja

first: Sam

middle: R

last: Telford III

first: Linden

middle: T

last: Hu


title: RNA-Seq data from V321L mouse dentate gyrus (DG)

city: Bethesda

first: David

last: Talmage

state: MD

address: 35 Convent Dr, Room 3B-101

department: NINDS

institute: National Institutes of Health

summary: Schizophrenia is a complex genetic neurodevelopmental disorder. eQTLs form bulk of the genetic variation contributing to schizophrenia. However, there is no known mechanism explaining regulation of expression of eQTL associated risk genes and underlying neurodevelopmental processes that are part of the cellular pathology. Neuregulin1-ErbB4 signaling plays a crucial role in synaptogenesis and has been implicated in schizophrenia. Gamma secretase mediated NRG1 nuclear back-signaling is thought to be involved in transcriptional regulation. A psychosis-associated missense mutation in NRG1 (rs74942016) has been predicted to impair nuclear back-signaling thereby implicating this mode of signaling in the underlying cellular pathology.

overall_design: 12 samples - 6 VV control, 6 LL mutants (3Male/3Female per genotype)

first: Prithviraj

last: Rajebhosale


title: Oncogenic PIK3CA corrupts growth factor signalling specificity

status: Public on Dec 27 2023

city: Dundee

first: Ralitsa

middle: Radostinova

last: Madsen

address: Dow Street

department: Medical Research Council Protein Phosphorylation and Ubiquitylation Unit

country: United Kingdom

institute: School of Life Sciences, University of Dundee

summary: Pathological activation of the PI3K/AKT pathway is among the most frequent defects in human cancer and is also the cause of rare overgrowth disorders. Yet, unlike the related oncogenic RAS/MAPK pathway, there is currently no systematic understanding of the quantitative flow of information within PI3K/AKT signalling and how it is perturbed by disease-causing mutations. Here, we develop scalable, single-cell approaches for systematic studies of signal processing within the PI3K pathway, enabling precise calculations of its information transfer for different growth factors. Using genetically engineered human cell models with allele dose-dependent expression of PIK3CAH1047R, we show that this oncogene is not a simple, constitutive pathway activator but a context-dependent modulator of extracellular signal transfer. PIK3CAH1047R reduces information transfer downstream of IGF1 while selectively enhancing EGF-induced signalling and transcriptional responses. This leads to a gross reduction in signalling specificity, akin to “blurred” signal perception. The associated increase in signalling heterogeneity increases phenotypic diversity in a cervical cancer cell model and in human induced pluripotent stem cells. Collectively, these findings and the accompanying methodological advances lay the foundations for a systematic mapping of the quantitative mechanisms of PI3K/AKT-dependent signal processing and phenotypic control in health and disease.

overall_design: Total RNA sequencing of CRISPR/Cas9 HeLa clones either expressing wildtype PIK3CA or one or two copies of PIK3CA-H1047R alongside a C-terminal frameshift variant on the remaining alleles. One clone, RM12, has a C-terminal frameshift on all three PIK3CA alleles and therefore no catalytically active PIK3CA protein.


title: Direct RNA sequencing experiment of humanized  liver mice with metabolic treatments

status: Public on Jan 23 2024

first: Haiming

last: Cao

address: 9000 Rockville Pike

department: NHLBI

institute: NIH

summary:
To make the human liver accessible to metabolic treatments, we employed a liver-specific humanized mouse model in which approximately 50% of the mouse hepatocytes were replaced by human ones.
To capture transcriptomes reflecting pathophysiology and therapeutic development of metabolic diseases, we subjected the humanized mice to  dietary intervention and the key metabolic transcriptional factor agonist treatments.  We then performed rna exoression profiling analysis using data obtained from nanopore direct RNA sequencing of these humanized mice.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
For the dietary treatment, the humanized mice were allowed free access to food (AL, n=4 for donor1, n=3 for donor2) or subjected to a twenty-four hours food withdrawal (Fast, n=4 for donor1, n=3 for donor2). For the transcription factor agonist treatments, the humanized mice were injected with DMSO (n=4), fenofibrate (n=4, 50mg/kg, Sigma-Aldrich, Cat. F6020), rosiglitazone (n=4,10mg/kg, Sigma-Aldrich, Cat. R2408) and GW4064 (n=4, 30mg/kg, Sigma-Aldrich, Cat. G5172) by i.p. injection. The livers were collected after 6 hours fasting and stored in liquid nitrogen immediately after mice sacrificed.The 75 ug of fresh Trizol isolated RNAs were adjusted to a volume of 100 μL with nuclease-free water. The polyadenylated RNAs were selected using the Dynabeads™ mRNA Purification Kit (Invitrogen, Cat. 61006) following the manufacturer's instructions. The 500 ng of fresh isolated polyadenylated RNAs were used for nanopore direct RNA sequencing (DRS) generally following the Oxford Nanopore Technologies (ONT) SQK-RNA002 kit protocol, including the optional reverse transcription step recommended by ONT. Libraries were loaded onto ONT R9.4 flow cells (Oxford Nanopore Technologies) and were sequenced using the GridION platform, with the standard MinKNOW/19.12.5 protocol script being used.

UPDATE: [12-13-2024] The following files were updated:
GSE224278_all.human.isoforms.donor1.bed.gz
GSE224278_all.mouse.isoforms.donor1.bed.gz
GSE224278_human.isoforms.donor1.gtf.gz
GSE224278_mouse.isoforms.donor1.gtf.gz
GSE224278_nanopore_human_transcripts_tpm_donor1.txt.gz

first: Chengfei

last: Jiang


title: GLIS3

first: Hong Soon

last: Kang

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

overall_design: Refer to individual Series


title: Measuring genetics, 5-mC and 5-hmC at single-base resolution

status: Public on Dec 21 2023

city: Cambridge

first: Páidí

last: Creed

address: B400, The Trinity Building Chesterford Business Park, Saffron Walden

institute: biomodal limited

type: Methylation profiling by high throughput sequencing

summary:
DNA comprises molecular information stored in genetic and epigenetic bases, both of which are vital to our understanding of biology. In human genomes, an epigenetic modification at the fifth carbon of cytosine bases comprises one fundamental pathway by which genes can be silenced or activated . Methods widely used to detect epigenetic modifications at cytosine bases rely on either deamination of unmodified cytosines to read as thymine or borane reduction of the modified oxidised cytosine bases to read as thymine. As a result, such methods fail to capture common C-to-T mutations, and importantly also fail to distinguish 5-methylcytosine (5mC - mainly found in silenced parts of the genome) from 5-hydroxymethylcytosine (5hmC - enriched in active gene bodies and enhancers). Hence, existing methods are unable to  read the complete information stored in our genomes in a single workflow.

Here, we build on biomodal's duet multiomics solution +modC, which has the unique ability to read genetic and epigenetic cytosine modifications (modC) in a single workflow. By introducing a high-fidelity methyltransferase during the workflow, we are able to construct an expanded information table that can be used to deconvolute A,T,C,G, 5-mC and 5-hmC simultaneously in a single read within a DNA molecule . We use a mouse embryonic stem cell-line, ES-E14TG2A, to map for the first time a simultaneous reading of the genome and epigenome at high depth and show how these epigenetic modifications are segregated across the genome.   In summary, using our modified approach we demonstrate simultaneous, phased reading of all six genetic and epigenetic bases. This tool provides a more complete picture of the information stored in genomes and will have applications throughout biology and medicine.

overall_design: 4 technical replicates, each 80ng input extracted gDNA material run in 6-base sequencing workflow as provided by biomodal

first: Robert

last: Crawford

first: Walraj

last: Gosal

first: Jack

last: Hardwick

first: Nicholas

middle: J

last: Harding

first: Ermira

last: Lleshi

first: Jean

last: Teyssandier

first: Michael

middle: W

last: Wilson

first: Sabri

last: Jamal

last: Monahan

first: Harry

last: Pink


title: Statistical algorithm enabled high precision tumor biomarker discovery for circulating extracellular vesicle-based cancer liquid biopsy

status: Public on Jan 31 2024

city: Gainesville

first: Mei

last: He

state: FL

address: 2033 Mowry Rd, Cancer and Genetics Research Complex

department: Pharmaceutics

institute: University of Florida

summary: Circulating extracellular vesicles (EVs) have gained significant attention for discovering tumor biomarkers. However, isolating EVs with well-defined homogeneous populations from complex biological samples is challenging. Different methods have been found to derive different EV populations carrying different biomolecules, which significantly confound biomarker discovery for developing clinical diagnostics. Building a rigorous EV isolation and standardizing assessment platform associated with -omics is essential to overcome this challenge. We introduced a novel isolation approach using a pH-responsive peptide conjugated with NanoPom magnetic beads (ExCy) for homogeneous EV isolation. Additionally, we introduced the first statistical algorithm for EV quality assessment (ExoQuality Index, EQI), which enables multi-assay quantification to provide a consistent and accurate definition of EV purity and quality; ExoQuality’s algorithm intakes multi-assay information to deconvolute complex EV heterogeneity. We analyzed our next generation sequencing on EV RNAs from pancreatic cancer patient plasma using four isolation methods; results highlighting ExCy’s isolation and EQI assessment improved biomarker identification. We identified a novel EV tumor biomarker, ATP6V0B, validated with Quantitative PCR (qPCR) by screening a pilot cohort of 23 individuals. With random forest modeling the ATP6V0B cycling threshold, we reported an AUC of 0.91, showcasing an enabling and clinically translatable liquid biopsy approach using circulating EVs.

overall_design: To benchmark our novel EV isolation method, we compared EVs isolated by different methods on the same sample sets (plasma from patients) using RNA-seq

first: Zachary

middle: F

last: Greenberg

first: Samantha

last: Ali

first: Andrew

last: Brock

first: Jinmai

first: Thomas

middle: D

last: Schmittgen

first: Song

last: Han

first: Steven

last: Hughes

first: Kiley

last: Graim


title: SMPD4 mediated sphingolipid metabolism regulates brain and primary cilia development

status: Public on Dec 24 2023

city: Columbus

first: Rolf

last: Stottmann

state: OHIO

address: 575 Children's Crossroads

department: Pediatrics

institute: Nationwide Children's Hospital

summary: SMPD4 (neutral sphingomyelinase-3/nSMase3) has recently been shown to be a new cause of microcephaly in a cohort of twenty-three pediatric patients. The function of nSMases in brain development and how SMPD4 variants cause human microcephaly and cerebellar hypoplasia was previously unknown.We developed an iPSC model to complement our mouse study. We found iPSC models from human SMPD4 patient and CRISPR/Cas9-induced SMPD4 knockout lines demonstrate a proliferation defect, increased cell death, loss of neural progenitors, and shortened primary cilia. Treatment with exogenous ceramide significantly rescues the cilia defect. SMPD4 patient and knockout cells have altered WNT signaling. We provide evidence that SMPD4 controls brain development by providing ceramide for primary ciliogenesis, suggesting a novel therapeutic strategy for SMPD4 mediated disease.

overall_design: We generated an SMPD4 knockout iPSC line (SMPD4 KO). We generated neural rosettes (NR) containing neural progenitor cells from control, KO, and patient iPSCs. We extracted RNA from these three lines for neural rosettes  at eight days in culture and stem cells (iPSCs) and compared RNA expression patterns.

first: Katherine

last: Inskeep

first: Bryan

last: Crase


title: RNA-Sequencing of HepG2 cells and rats treated with Chinese herbal medicine Pulsatilla Chinensis

status: Public on Dec 01 2023

city: beijing

first: ruijun

last: wang

address: college of life science

country: China

institute: Tsinghua university

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus


title: Single-cell analysis reveals the stromal dynamics and tumor-specific characteristics in the microenvironment of ovarian cancer

status: Public on Jan 07 2024

city: Pittsburgh

first: Ronald

last: Buckanovich

state: Pennsylvania

address: 204 Craft Avenue

department: Department of Medicine

institute: University of Pittsburgh

summary:
We characterized high-grade ovarian serous carcinoma TIME based on integrative single-cell transcriptomics analysis of public and in-house datasets. We identified a distinct transcriptomic landscape of both immune and non-immune cells between the dense and more sparse stromal tumors. High stromal tumors contain a lower fraction of infiltrating activated CXCR3+ GRZB+ IFN+ IL2R+ CD8+ T and NCR3+ KLRD1+ natural killer (NK) cells and CXCL1+ macrophages.
Next, we determined the potential interplays between non-immune and immune cells. High stromal tumors showed increased expression of CXCL12 in epithelial cancer cells and CA-MSCs. Cell-cell communication analyses suggested that epithelial cancer cells and CA-MSCs secreted CXCL12 interacts with the CXCR4 receptor on NK and CD8+ T cells. Using CXCL12 and/or CXCR4 neutralizing antibodies, we confirmed the immunosuppressive role of the CXCL12-CXCR4 axis in CA-MSC-induced immune suppression.

overall_design: OVCA mRNA profiles of the following two groups: (1) high-stromal samples, and (2) low stromal samples.

first: John

last: Mellors

first: Cynthia

last: Adams

first: Linan

last: Zhang

first: Hatice

middle: U

last: Osmanbeyoglu

first: Sandra

last: Cascio


title: RNA-Sequencing of rats treated with Chinese herbal medicine Pulsatilla Chinensis

summary: Pulsatilla chinensis (PC) is a traditional Chinese medicinewith detoxification activities. It has been used for dysentery, vaginal trichomoniasis, bacterial infections and malignant tumor treatment. Gavage administration of PC in hyperlipidemia rats for 11 weeks resulted in obviously reduced serum total cholesterol, LDL-cholesterol and ameliorated fatty liver. So we wanted to further investigate the relationship between PC and lipid metabolism related pathways or genes

overall_design: Analysis used normal diet treated rats as control samples for comparison to the model samples treated with high cholesterol-high fat diet ,and the treatment group with high cholesterol-high fat diet plus Pulsatilla Chinensis for comparison to the model samples treated with high cholesterol-high fat diet  , genes expression changes were determined by RNA-Seq to determine Pulsatilla Chinensis  responsive genes

first: Rui

last: Wang

first: Lan

last: Xie

first: Juan

last: Feng

first: Xiao

middle: L


title: Regulation of gene expression by GLIS3 during neonatal kidney development

first: Hong Soon

last: Kang

summary: Loss of transcription factor GLIS3 function in humans and mice leads to the development of neonatal polycystic kidney disease (PKD). To investigate how loss of GLIS3 function in kidney affects postnatal kidney development and PKD, we analyzed the gene expression profiles of kidneys from WT and Glis3-null mice at P7, P14, and P28 by RNA-Seq analysis using Glis3 global KO model Glis3-mCherry and kidney specific knockout model Glis3-PAX8Cre.

overall_design: Total kidney RNAs were isolated from WT and Glis3-null mice and gene expression profiles were analyzed by RNA-Seq.

first: Anton

last: Jetten

first: Chitrangda

last: Srivastava

first: Hong

first: Justin

last: Collier

first: Sara

last: Grimm


title: Single-cell RNA-seq of BHT induced lung injury

status: Public on Nov 02 2023

city: Durham

first: Purushothama Rao

last: Tata

email: pt93@duke.edu, tata.lab@yahoo.com

address: 303 Research Dr., Alexander Sands Building, Room- 451, BOX 3709

department: Department of Cell Biology

institute: Duke University School of Medicine

summary: We performed single-cell RNA-seq analysis  (10X genomics 3'-GEM scRNA-seq kit) of BHT induced lung injury

overall_design: Single-cell RNA-seq on day 6 post BHT induced lung injury

first: Aleksandra

first: Yoshihiko

last: Kobayashi

first: Pankaj

last: Agarwal


title: TMED10 regulates cell-surface expression of PD-1

status: Public on Nov 10 2023

city: Amsterdam

first: Oscar

last: Krijgsman

state: Outside the US or Canada

address: Plesmanlaan 121

department: Molecular Oncology and Immunology

country: Netherlands

institute: Netherlands Cancer Institute

summary: We found that Tmed10 regulates the cell-surface expression of PD-1 in T cells. We additionally show that treatment with AGN192403 can phenocopy Tmed10 KO by lysosomally degrading the Tmed10-PD-1 complex.

overall_design: Here, we treated T cells (WT or Tmed10 KO) with AGN (100uM) or Veh (H2O) for 24 hours. The T cells were additionally activated with anti-CD3 antibodies (2.5ug/mL in PBS) for 24 hours (Act) or not (Rest).

first: David

last: Vredevoogd

first: Daniel

last: Peeper


title: In vitro and in vivo CRISPR-Cas9 screens reveal drivers of aging in neural stem cells of the brain.

status: Public on Nov 01 2023

city: Stanford

first: Tyson

middle: Joel

last: Ruetz

address: 290 Jane Stranford Way

department: Genetics

institute: Stanford University

summary: Here we develop in vitro and in vivo high-throughput CRISPR-Cas9 screening platforms to systematically uncover genes whose knockout boosts NSC activation in old mice. Our genome-wide screening pipeline in primary cultures of young and old NSCs identifies over 300 gene knockouts that specifically restore old NSC activation. Interestingly, the top gene knockouts are involved in glucose import, cilium organization and ribonucleoprotein structures. To determine which gene knockouts have a rejuvenating effect for the aging brain, we establish a scalable CRISPR-Cas9 screening platform in vivo in old mice. Of the 50 gene knockouts we tested in vivo, 23 boost old NSC activation and production of new neurons in old brains. Notably, the knockout of Slc2a4, which encodes for the GLUT4 glucose transporter, is a top rejuvenating intervention for old NSCs. GLUT4 protein expression increases in the stem cell niche during aging, and we show that old NSCs indeed uptake ~2-fold more glucose than their young counterparts. Transient glucose starvation increases the ability of old NSCs to activate, which is not further improved by knockout of Slc2a4/GLUT4

overall_design: Sequenced guide counts from genome wide in vitro screen and in vivo sub screens

first: Anne

last: Brunet


title: Effects of siERRα on gene expression in human skin fibroblast TIG113 cell.

status: Public on Oct 17 2023

city: Aomori

first: Naoki

last: Nanashima

state: Aomori

address: 58-1 Mase, Hamadate

department: Department of Nutrition

institute: Aomori University of Health and Welfare

summary: We analyzed function of  ERRα in human skin fibroblasts which silenced ERRα gene expression by microarray analysis.

overall_design: TIG113 was seeded in a culture dish, cultured until subconfluent, and then transfected with siRNA. After 48 h, total RNA was extracted from siRNA transfected or control TIG113 cells.


title: Attenuated retinoic acid signaling is among the early responses in mouse uterus approaching embryo attachment

status: Public on Nov 15 2023

city: Athens

first: Xiaoqin

last: Ye

state: Georgia

address: 501 D.W. Brooks Drive

department: Physiology and Pharmacology

institute: The University of Georgia

summary: The uterus is transiently receptive for embryo implantation. It remains to be understood why the uterus does not reject semi-allogeneic or allogeneic embryos for implantation. Uterine early response genes at the time approaching embryo attachment may provide insights. Uteri from C57BL/6 pseudo-pregnant (PP) mice and pregnant (P) mice approaching embryo attachment on day 3 post-coitum (D3) @22 h were analyzed for early response genes by microarray. SAM algorithm retained 21,858 unique probesets. Principal component analysis (PCA) indicated a clear separation between the PP and P groups. There were 105 upregulated and 5 downregulated protein-coding genes in the pregnant uterus (fc>1.5 & q-value<5%). Gene ontology (GO) analysis of these upregulated genes revealed that 84% of the GO terms were related to immune responses, with a dominant natural killer (NK) cell activation signature. Among the top 8 upregulated protein-coding genes, Cyp26a1 inactivates retinoic acid (RA) while Lrat promotes vitamin A storage, both of which attenuate RA bioavailability; Atp6v0d2 and Gjb2 play roles in ion transport and transmembrane transport; Gzmb, Gzmc, and Il2rb are involved in immune responses; and Tdo2 is important for kynurenine pathway. Most of these genes or their related pathways have been implicated in immune regulations.  RA signaling has been implicated in tolerance and immunity and uterine NK cells have been implicated in immunotolerance at the maternal-fetal interface in the placenta. The mechanisms of immune responses approaching embryo attachment remain to be elucidated. The coordinated effects of the early response genes may hold the keys to the question of why the uterus does not reject an implanting embryo.

overall_design: Embryo attachment can be detected by faint blue dye reaction in mice and it normally occurs ~midnight 4 days post-coitum (D4 @0h or D3 @24h). The earliest time for faint blue dye reaction to be detectable in some C57BL/6 mice is D3 @22h (DOI: 10.1016/j.fertnstert.2011.01.160). Young virgin C57BL/6 mice were purchased from The Jackson Laboratory. They were randomly splitted into two sets. One set of mice were mated with stud males. Mating night was defined as D0 and mating was verified by a vaginal plug the next morning. The pregnant uterine tissues were collected on D3 @22h. Only the ones without blue dye reaction (to verify no embryo attachment yet) but with healthy-looking blastocysts (to verify pregnancy status) flushed from the uterine horns were included. The other set of mice were mated with vasectomized males to produce pseudopregnant mice. The pseudopregnant uterine tissues were also collected on D3 @22h. The uterine tissues were processed for total RNA isolation.  Four samples each from the pregnant and pseudopregnant groups with the best RNA qualities (260/280 ratio: 1.84~1.99; RIN: 6.9~8.7) were selected from microarry.


title: Type I IFN signaling pathway promotes osteoblast differentiation through targeting Sox9 [scRNA-seq]

status: Public on Nov 30 2023

first: lei

last: zhang

address: xueyuan road 38th

department: heath science center

institute: peking university

summary: Ossification of the posterior longitudinal ligament (OPLL) is a progressive ectopic bone formation of ligaments and may lead to severe neurological symptoms. However, in addition to conservative therapy for neurological symptoms or surgical management, effective pharmacotherapy and preventive intervention for OPLL remains absent. Here, to reveal the potential therapeutic target we generate single-cell maps of T-OPLL patients. We found that type I IFN signaling pathway is significantly activated during the progression of ossification. Further, stimulation of IFN β obviously promoted the calcium deposition of murine MC3T3-E1 preosteoblasts. We profiled the transcriptome in MC3T3-E1 cells after osteogenic induction, and found that IFN β promoted the expression of Sox9 and SPP1. STAT1 positively regulated the expression of Sox9 and phosphorylated STAT1 occupied on the promoter of Sox9. We also observed the localization of Sox9 in the ossificated region of ligament from OPLL patients. Thus, Sox9 is a target gene of type I IFN pathway during osteogenic induction. Furthermore, blockage of IFNAR1 by anti-IFNAR1 neutralizing antibody significantly impeded osteoblast differentiation induced by IFN β and display efficacy against ossification. Together, these results demonstrated that type I IFN signaling pathway promotes osteoblast differentiation. Anti-IFNAR1 neutralizing antibody could be used as a potential therapy for OPLL by blocking type I IFN signaling pathway.

overall_design: To investigate the mechanisms of OPLL with single-cell RNA-seq.

first: Xiao

last: Liu

first: Lei

last: Zhang

first: Ge

last: Wang

first: Wei

last: Zhao

first: Chen

last: Liang

first: Youzhi

last: Tang

first: Yenan

last: Fu

first: Bo

first: Jing

first: Xiaoguang

first: Hongquan

first: Yu

last: Yu


title: Dendritic Cell Vaccination in Conjunction with TLR Agonist Polarizes Interferon Immune Responses Associated with Long-term Survival in Malignant Glioma Patients [scRNA-seq]

status: Public on Oct 11 2023

city: Los Angeles

first: Willy

last: Hugo

state: California

address: 10833 Le Conte Ave

department: Medicine

institute: University of California, Los Angeles

summary: Autologous tumor lysate-pulsed dendritic cell (ATL-DC) vaccination is a promising immunotherapy for patients with high grade gliomas, but responses have not been demonstrated in all patients. Pre-clinical studies demonstrate that toll-like receptor (TLR) agonists can enhance the anti-tumor immune response from cancer vaccines.  To determine the most effective combination of autologous tumor lysate-pulsed DC vaccination, with or without the adjuvant toll-like receptor (TLR) agonists poly-ICLC or resiquimod, we conducted a randomized, open-label multi-arm Phase 2 clinical trial to evaluate immune responses and survival in 23 patients with newly diagnosed or recurrent WHO Grade III-IV malignant gliomas. Patients were randomized to receive ATL-DC vaccination with either a placebo, a TLR-7/8 agonist (resiquimod, topical 0.2%), or a TLR-3 agonist (poly-ICLC, 20 mcg/kg intramuscular).  Peripheral blood was obtained at baseline and following the vaccination cycle for immune monitoring. Mass cytometry, bulk and single-cell RNA sequencing analyses were performed. Analysis of the bulk RNAseq data (pre-treatment vs. post-treatment) demonstrated highly significant upregulation of type 1 and type 2 interferon gene expression selectively in patients who received adjuvant TLR agonist (either poly-ICLC or resiquimod) together with ATL-DC.  CyTOF analysis of patient peripheral blood mononuclear cells (PBMCs) showed increased levels of memory PD-1+ T-cell and monocyte populations after ATL-DC + TLR agonist administration.  In addition, scRNA-seq demonstrated a higher expression fold change of IFN-induced genes with poly-ICLC treatment in both peripheral blood monocytes and T lymphocytes. Median progression-free survival (PFS) was 8.1 months (5.5 placebo vs. 8.1 resiquimod vs. 31.4 poly-ICLC), and median overall survival (OS) was 26.6 months (7.7 placebo vs. 16.7 resiquimod vs. 52.5 poly-ICLC).   Patients who had higher expression of interferon response genes lived significantly longer and had longer time to progression compared to those with lower expression.  The results suggest that ATL-DC in conjunction with adjuvant poly-ICLCTL induces a polarized interferon response in circulating monocytes and specific activation of a CD8+ T cell population. This combination was associated with extended survival in this patient population. This interferon response signature may represent an important blood biomarker for immunotherapy in patients with malignant glioma.

overall_design: The cells for scRNAseq analysis were resuspended in PBS at a concentration of 1,000 cells/µl. We only selected representative patients from each treatment group whose PBMC quality were sufficient for single cell RNAseq processing (Placebo: 2 pairs, Poly-ICLC: 3 pairs (1 sample failed and is not included), Resiquimod (2 pairs); PBMC of two normal donors are added to provide a normal reference). Cell preparation, library preparation, and sequencing were carried out according to Chromium product-based manufacturer protocols (10X Genomics), targeting for a total of 10,000 cells sequenced. Cells from post-treatment samples (Placebo, Poly-ICLC, or Resiquimod), were compared to the pre-treatment samples to identify differentially expressed genes associated with each treatment.

first: Richard

middle: G

last: Everson

first: Lu

last: Sun

first: Joseph

last: Antonios

first: Alexander

last: Lee

first: Lizhong

last: Ding

first: Melissa

last: Bu

first: Brian

last: Lu

first: Sarah

last: Khattab

first: Carolina

last: Chavez

first: Emma

last: Billingslea-Yoon

first: Andres

last: Salazar

first: Benjamin

last: Ellingson

first: Timothy

last: Cloughesy

first: Linda

last: Liau

first: Robert

last: Prins


title: Transcriptional Reference Map of Human Natural Killler Cells

status: Public on Oct 27 2023

city: Oslo

first: Karl-Johan

last: Malmberg

address: Ullernchausseen 70

country: Norway

institute: Oslo University Hospital

summary: The functional diversity of NK cell repertoires stems from differentiation, homeostatic receptor-ligand interactions, and adaptive-like responses to viral infections. Here we performed single-cell RNA sequencing of both bulk NK and sorted NK subsets to establish a transcriptional reference map of human natural killer cells based on single-cell RNA sequencing profiles from 72,129 cells obtained from bulk (12 donors) and sorted phenotypically-defined NK cell subsets (2 donors) and tissue-resident NK cells (TrNK, 136 donors). Our own data is included here.

overall_design:
Here we performed single-cell RNA sequencing of both bulk NK and sorted NK subsets to establish a transcriptional reference map of human natural killer cells.
The raw sequencing data has been uploaded to EGA under accession EGAS50000000014.

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Submitter states that missing raw data are being made available for controlled access in EGA.
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first: Herman

last: Netskar

first: Aline

last: Pfefferle

first: Amir

last: Horowitz


title: Energy stress-induced circEPB41(2) regulates lipogenesis through histone modifications

city: HEFEI

first: Yang

last: Yang

state: ANHUI

address: NO. 96, JinZhai Road

institute: University of Science and Technology of China


title: Energy stress-induced circEPB41(2) regulates lipogenesis through histone modifications [RNA-seq]

summary: The list of circular RNAs (circRNAs) involved in tumor metabolic reprogramming is rapidly increasing, but whether circRNAs have a role in linking epigenetic modification with lipid metabolism remains elusive. Here, we demonstrate that the circRNA circEPB41(2), which expression is driven by HNRNPA1 in response to glucose deprivation, is important for de novo lipogenesis. Moreover, we uncovered a genome-wide decrease in specific histone acetylation after circEPB41(2) depletion. These changes are associated with altered gene expression and observed cellular phenotypes, suggesting their functional significance. Mechanistically, we showed that circEPB41(2) regulates histone acetylation in part by modulating the stability and polyadenylation of SIRT6 through binding and maintaining the demethylase activity of FTO. Accordingly, circEPB41(2) silencing inhibited cellular lipid accumulation, suppressed cell proliferation, and inhibited cancer xenograft growth. Thus, circEPB41(2) may consider a target for cancer treatment.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for control  and circEPB41(2) shRNA-knockdown cells (sh#1, sh#2).

first: Jingjing

last: Luo

first: Yide

last: Mei


title: Energy stress-induced circEPB41(2) regulates lipogenesis through histone modifications [H3K9ac ChIP-seq]

summary:
The list of circular RNAs (circRNAs) involved in tumor metabolic reprogramming is rapidly increasing, but whether circRNAs have a role in linking epigenetic modification with lipid metabolism remains elusive. Here, we demonstrate that the circRNA circEPB41(2), which expression is driven by HNRNPA1 in response to glucose deprivation, is important for de novo lipogenesis. Moreover, we uncovered a genome-wide decrease in specific histone acetylation after circEPB41(2) depletion. These changes are associated with altered gene expression and observed cellular phenotypes, suggesting their functional significance.
Mechanistically, we showed that circEPB41(2) regulates histone acetylation in part by modulating the stability and polyadenylation of SIRT6 through binding and maintaining the demethylase activity of FTO. Accordingly, circEPB41(2) silencing inhibited cellular lipid accumulation, suppressed cell proliferation, and inhibited cancer xenograft growth. Thus, circEPB41(2) may consider a target for cancer treatment.

overall_design: Comparative occupancy profiling by high throughput sequencing analysis for control  and circEPB41(2) shRNA-knockdown cells (shcircEPB41(2)).


title: Spatial transcriptomics of gastric cancers (Visium 10X platform)

status: Public on Dec 26 2023

city: Seocho-gu

first: Kim

last: Tae-Min

address: Banpo-daero

country: South Korea

institute: The Catholic University of Korea

summary: A total of 9 primary gastric cancers (1 pair of primary-metastasis, GC6) analyzed using Visium 10X platform-based spatial transcriptomics.

overall_design: Spatial transcriptomics of human gastric cancers were profiled.

first: Tae

last: Kim

first: Hoon

last: Hur

first: Sung

last: Lee


title: Nucleotide Excision Repair of Aflatoxin-Induced DNA Damage within the 3D Human Genome Organization

status: Public on Oct 19 2023

first: Wentao

last: Li

state: GA

address: 500 D. W Brooks Drive

department: Environmental Health Science

institute: University of Georgia

summary: Aflatoxin B1 (AFB1), a potent carcinogenic mycotoxin, is known to contribute to liver cancer development. Within the cell, bioactivated AFB1 intercalates into the DNA double helix, forming bulky DNA adducts that lead to mutagenesis if left unrepaired. Here, we adapted the tXR-seq method to map the nucleotide excision repair of AFB1-induced DNA lesions at genome-wide level. Our research uncovered that transcription-coupled repair plays a major role in repairing the AFB1-induced DNA lesions. We identified a distinctive length distribution pattern for the excision products released during repair, suggesting a unique dual incision mechanism for AFB1-induced DNA lesions. Notably, we revealed that repair activity is more pronounced on chromosomes closer to the nuclear center and A compartments undergo faster repair compared with B compartments. Additionally, we observed higher repair activity within regions encompassing TAD boundaries and loop anchors. This study provides insights into the interplay between repair, transcription, and 3D genome organization, shedding light on the mechanisms behind AFB1-associated liver cancer development.

overall_design: To investigate the impact of 3D genome organization on nucleotide excision repair, we adapted the tXR-seq method to map the genome-wide repair of AFB1-induced DNA lesions and determined the repair signals at multiple levels of genomic organizations.

first: Yiran

last: Wu

first: Muhammad

last: Muzammal Adeel

first: Aziz

last: Sancar


title: O-GlcNAcylation promotes Palbociclib resistance in breast cancer by regulating MITF nuclear translocation [RNA-seq]

status: Public on Oct 10 2023

city: WASHINGTON

first: Wenge

last: Zhu

state: DC

address: 800 22ND ST NW, Room 8480, Science & Engineering Hall

department: Biochemistry and Molecular Medicine

institute: George Washington University

summary: Cyclin-dependent kinases 4 and 6 (CDK4/6) are essential drivers of the cell cycle and are also critical for the initiation and progression of diverse malignancies. Pharmacological inhibitors targeting CDK4/6 have demonstrated significant activity against various tumor types such as breast cancer. However, resistance to  CDK4/6 inhibitors (CDK4/6i) (such as palbociclib) remains an immense obstacle in clinical and the underlying mechanisms have not been fully understood.  Using quantitative high-throughput combinational screen (qHTCS) and genomic sequencing, we report that the Microphthalmia-associated transcription factor (MITF), was significantly elevated in palbociclib-resistance cells. Inhibition of MITF can enhance the therapeutic efficacy of Palbociclib and surmount Palbociclib resistance both in vitro and in vivo. Mechanistically, we found that O-GlcNAc transferase (OGT) modifies MITF with O-GlcNAcylation at Serine 49 (Ser49) within its nuclear localization signal (NLS), thereby promoting MITF binding to importin α/ β and facilitating its nuclear transportation, which is crucial in regulating senescence. Significantly, clinical studies also confirm that MITF was elevated in palbociclib-resistance patients. Collectively, these results reveal a previously unrecognized mechanism by which MITF-mediated palbociclib resistance, and provide valuable insights for the development of innovative therapeutic strategies in future clinical contexts.

overall_design: To investigate the mechanism of Palbociclib resistance in breast cancer cell lines, we first generated Palbociclib-resistance (PR) cell line MCF-7 PR. Then we compare different genes expression between MCF-7 wild-type (WT) and MCF-7 PR cells by RNA seq. Then we generated MITF-depleted cell line by shRNA (MCF-7 PR shMITF), and compare different genes expression between MCF-7 PR and MCF-7 PR shMITF

first: Yi

last: Zhang


title: TRIBAL/TRIBA1AL suppression in human hepatocytes

city: Ottawa

first: Ruth

last: McPherson

state: Ontario

address: 40 Ruskin

country: Canada

institute: UOHI

summary: Cultured primary human hepatocytes treated with antisense oligonucleotides

overall_design: Primary human hepatocytes (2 lots from 2 distinct donors, commercially obtained from ThermoFisher Scientific ) were treated for 72 h with 1 of 2 antisense olignucleotides (and a matching control)

first: S

last: Soubeyrand

first: R


title: Transcriptional profiling of male PAB rats with compensated and decompensated right ventricule

status: Public on Sep 29 2023

city: Bad Nauheim

first: Fatemeh

last: Khassafi

address: Parkstrasse

country: Germany

institute: Max Planck Institute

summary: We performed molecular phenotyping of adaptive to maladaptive RV hypertrophy remodeling, using transcriptome analysis of RV tissue obtained from PAB rats. This data along with other datasets linked within this study, provides a resource to comprehensively compare human RV remodeling to the animal RV dysfunction models (PAB) and led to the validation of state-specific biomarkers and potential therapeutic targets for RV dysfunction.

overall_design: RNA sequencing was performed for RV tissues obtained from PAB rats which were characterized as compensated RV, decompensated RV by hemodynamics, as well as normal RV from control rats, including only male samples as follows: control (n=5), compensated RV (n=5), decompensated RV (n=4)

first: Prakash

last: Chelladurai

first: Sébastien

last: Bonnet

first: Soni

last: Pullamsetti


title: Effect of PCIF1 knockdown on gene exppression in HeLa S3 cells

status: Public on Sep 30 2023

city: Toyama

first: Yutaka

last: Hirose

state: Toyama

address: 2630 Sugitani

department: Graduate School of Medicine and Pharmaceutical Sciences

institute: University of Toyama

summary:
We previously identified PCIF1 (Phosphorylated CTD Interacting Factor 1) as a novel phosphorylated C-terminal domain (CTD) of RNA polymerase II. We also recently identified PCIF1 as a new cap-specific adenine N6 methyltransferase (CAPAM) responsible for creating N6, 2’-O-dimethyladenosine (m6Am) at the 5’-end of mRNAs. However, it remains unclear how PCIF1 regulates gene expression.
To identify genes whose expression levels are affected by PCIF1 knockdown in human cultured cells, we performed gene expression profiling by microarray analysis.

overall_design: HeLa S3 cells were seeded on 6-well plates at 1.0 × 105 cell/well and cultured at 37°C 24 h before transfection. Cells were transfected with either negative control siRNA or siRNA specific to human PCIF1 gene using Lipofectamine RNAi MAX (ThermoFisher) and further cultured for 72 h at 37°C.

first: Hiroyasu

last: Ishiguro

first: Ai

last: Sugita

first: Natsuki

last: Yanagisawa

first: Yoshiaki

last: Ohkuma

last: Tabuchi


title: RNA-Sequencing of HepG2 cells treated with Chinese herbal medicine Pulsatilla Chinensis

first: ruijun

last: wang

address: college of life science

institute: Tsinghua university

summary: Pulsatilla chinensis (PC) is a traditional Chinese medicinewith detoxification activities. It has been used for dysentery, vaginal trichomoniasis, bacterial infections and malignant tumor treatment. Gavage administration of PC in hyperlipidemia rats for 11 weeks resulted in obviously reduced serum total cholesterol, LDL-cholesterol and ameliorated fatty liver. So we wanted to further investigate the relationship between PC and lipid metabolism related pathways or genes

overall_design: Analysis used DMSO treated cell as control samples for comparison to the experimental samples treated with extracts of  Chinese herbal medicine Pulsatilla Chinensis, genes expression changes were determined by RNA-Seq to determine Pulsatilla Chinensis  responsive genes

first: Rui

last: Wang

first: Lan

last: Xie

first: Juan

last: Feng

first: Xiao


title: NFATc1 drives allergic contact dermatitis reponses by controlling the induction of IL-17-producing CD8 cells

status: Public on Nov 27 2023

city: Wuerzburg

first: Muhammad

last: Azeem

state: Bavaria

address: Josef-Schneider Str. 2

department: Dermatology, Allergology, and Venereology

institute: University Hospital Wuerzburg

summary:
A plethora of data supports a major role of CD4+ and CD8+ T lymphocytes for the initiation, progression and maintenance of allergic contact dermatitis (ACD). However, in-depth understanding of the underlying molecular mechanisms is still limited. NFATc1 , a central component of the Ca++-Calcineurin-NFAT-signalling network, plays an essential role in T cell activation. We therefore investigated its impact on contact hypersensitivity (CHS), the mouse model for ACD.
The CHS response to 2,4,6-trinitrochlorobenzene (TNCB) was diminished in Nfatc1fl/flxCd4-cre mice (Nfatc1-/-) as compared to wild-type (WT) animals and associated with a lower percentage of interleukin (IL)17-producing CD8+T (Tc17) cells in both inflamed skin and draining lymph nodes (dLN). In vitro Tc17 polarization assays revealed that Nfatc1-/- CD8+ T cells have a reduced capacity to polarize into Tc17 cells. Applying single-cell RNA sequencing, we realized that NFATc1 controls the T cell differentiation fate. In the absence of NFATc1, CD8+ T cells favour the development of Interferon (IFN)-g-secreting CD8+ T (Tc1) lymphocytes while in its presence they turn into Tc17 cells. Finally, we showed the adoptive transfer of TNCB-sensitized WT CD8+T cells rescued tThe CHS response could be rescued in naïve Nfatc1-/-mice by adoptive transfer of TNCB-sensitized WT CD8+T cells.
Our data demonstrate that NFATc1 acts as a molecular switch controllcontrolsing the development of Tc17 cells and can be used as a target to alleviate surveilling CD8+T cell-mediated CHS responses.

overall_design: C57/B6N WT mice and mice carrying NFATc1 deficiency in T cells through Cre-Lox system (Nfatc1f/f×Cd4-Cre) were used in the experiment. CD8+ T cells from both of these mice were used for the Tc17 (CD8+IL17+) polarization which followed by single cell RNA sequencing of these Tc17 cells. WT mice were the control group and NFATc1 deficient mice were the experimental group.

first: Moutaz

last: Helal

first: Stefan

last: Klein-Hessling

first: Edgar

last: serfling

first: Matthias

last: Goebeler

first: Khalid

last: Muhammad

first: Andreas

last: Kerstan


title: The genetic architecture of protein stability

city: Barcelona

first: cristina

last: Hidalgo-Carcedo

address: Dr. Aiguader 88

country: Spain

institute: CRG

summary: There are more ways to synthesize a 100 amino acid protein (20^100) than atoms in the universe.  Only a miniscule fraction of such a vast sequence space can ever be experimentally or computationally surveyed. Deep neural networks are increasingly being used to navigate high-dimensional sequence spaces. However, these models are extremely complicated and provide little insight into the fundamental genetic architecture of proteins. Here, by experimentally exploring sequence spaces >10^10, we show that the genetic architecture of at least some proteins is remarkably simple, allowing accurate genetic prediction in high-dimensional sequence spaces with fully interpretable biophysical models. These models capture the non-linear relationships between free energies and phenotypes but otherwise consist of additive free energy changes with a small contribution from pairwise energetic couplings. These energetic couplings are sparse and caused by structural contacts and backbone propagations. Our results suggest that artificial intelligence models may be vastly more complicated than the proteins that they are modeling and that protein genetics is actually both simple and intelligible.

overall_design: DNA-seq of PCR amplicon; 3 biological replicates from Saccharmocyes cerevisiae

first: Andre

last: Faure

first: Aina

last: Martí-Aranda

first: Cristina

first: Jörn

last: Schmiedel

first: Ben

last: Lehner


title: Transcriptional profiling of male and female MCT-induced rats with compensated and decompensated right ventricle

first: Fatemeh

last: Khassafi

summary: We performed molecular phenotyping of RV remodeling, using transcriptome analysis of RV tissue obtained from MCT-induced rats (of both male and female animals). The clustering approach along with precise hemodynamics assessments identified “early" and "late" subgroups of decompensated state in rat RV. Further molecular characterization of these subgroups identified distinct molecular genes and pathways within different stages of RV remodeling. This study provided a resource to comprehensively compare human RV remodeling with the current animal model of PH (MCT), and led to validation of state-specific biomarkers and potential therapeutic targets for RV dysfunction.

overall_design: RNA sequencing was performed for two batches of RV tissues obtained from MCT-induced rats (n=53) which were characterized as compensated RV, decompensated RV, as well as normal RV from control rats. Batch 1 include only male samples as follows: control (n=10), compensated RV (n=7), early decompensated RV (n=3), late decompensated RV (n=6). Second batch includes animals from both sexes as follows: control female RV (n=4), control male RV (n=4), compensated female RV (n=6), compensated male RV (n=5), decompensated female RV (n=3), decompensated male RV (n=5).

first: Prakash

last: Chelladurai

first: Sébastien

last: Bonnet

first: Soni

last: Pullamsetti


title: Transcriptional profiling of human adaptive and maladaptive right ventricular remodeling

summary: In this study, we performed molecular phenotyping of RV remodeling, by transcriptome analysis of RV tissue obtained from 40 patients. The unsupervised clustering analysis identified “early" and "late" subgroups within compensated and decompensated states, characterized by the expression of distinct signaling pathways, and different circulating levels of several ECM proteins in decompensated RV subgroups. Our study provides a resource for the subphenotyping of RV states, the identification of state-specific biomarkers, and potential therapeutic targets for RV dysfunction.

overall_design: RNA sequencing was performed for RV tissues obtained from human patients clinically categorized as compensated RV (n=14), decompensated RV (n=13) and donor controls (n=13). Followed by additional clustering labels, A to E. Cluster A represent Normal group (n=24), Cluster B represent compensated RV group (n=4), Cluster C represent severe hypertrophic RV group (n=3), Cluster D represent early decompensated group (n=5), and Cluster E represent late decompensated group (n=4).

first: Olivier

last: Boucherat


title: Dendritic Cell Vaccination in Conjunction with TLR Agonist Polarizes Interferon Immune Responses Associated with Long-term Survival in Malignant Glioma Patients [Bulk RNA-seq]

first: Willy

last: Hugo

overall_design: Total RNA was isolated from frozen PBMC of the patients isolated at baseline and after receiving vaccines with ATL-DC plus adjuvant (placebo: 5 pairs, resiquimod: 9 pairs and poly-ICLC: 9 pairs). The RNA were sent for RNA sequencing. For each pair, we compute gene expression changes pre- vs. post- ATL-DC+adjuvant treatment and compare the changes among the patients treated with placebo vs. resiquimod or poly-ICLC.

first: Richard

last: Everson

first: Lu

last: Sun

first: Joseph

last: Antonios

first: Alexander

last: Lee

first: Lizhong

last: Ding

first: Melissa

last: Bu

first: Brian

last: Lu

first: Sarah

last: Khattab

first: Carolina

last: Chavez

first: Emma

last: Billingslea-Yoon

first: Andres

last: Salazar

first: Benjamin

last: Ellingson

first: Timothy

last: Cloughesy

first: Linda

last: Liau

first: Robert

last: Prins


title: Gene expression profile of DCs of mLN and SI-LP of mice lacking RelB expression in DCs at steady-state and after helminth infection

status: Public on Sep 22 2023

city: Neuherberg

first: Caspar

last: Ohnmacht

address: Ingolstädter Landstraße 1

institute: Helmholtz Munich

summary:
Gene expression profile of dendritic cells (DC) of mesenteric lymph node (mLN) and lamina propria of small intestine (SI-LP) of control mice and mice lacking RelB expression in DCs at steady state, four days and 14 days after infection with Heligmosomoides polygyrus bakeri (Hpb)
Dendritic cells (DCs) are crucial for initiating protective immune responses and have also been implicated in the generation and regulation of Foxp3+ regulatory T cells (Tregs). Specific DC subsets or DC-intrinsic pathways regulate immunity against pathogens but also tolerance to harmless antigens derived from food or microbiota at barrier sites, but underlying mechanisms in the intestinal tract remain poorly defined. Here, we provide evidence that the alternative NF-B family member RelB controls a defined transcriptional program in migratory DC subsets of mesenteric lymph nodes and the small intestinal lamina propria. Functionally, ablation of RelB in dendritic cells result in increased Foxp3+ Treg cell numbers but decreased RORt peripheral Treg cell numbers maintained even under inflammatory conditions. Single-cell RNA-sequencing revealed a complete RelB dependency for the differentiation of cryptopatches and isolated lymphoid follicles-associated DCs (CIA-DCs) in the lamina propria of the small intestine. In addition, we show a RelB-dependent signature of migratory DCs in mesenteric lymph nodes favoring DC-Treg cell interaction by affecting the expression of chemokines (Ccl22, Ccl17), migration behavior (Cd63), co-stimulatory molecule (Cd80, Cd40, Cd200, Tnfsf4), and tolerance-related integrin (Itgb5, Itgb8) expression. Functionally, increased Treg cell numbers in DC-specific RelB knockout animals did not show any risk of increased reactions in a model of food allergy but instead prevented protective Th2 immune responses in the intestines after infection with Heligmosomoides polygyrus bakeri despite their slight steady-state type 2 immune bias. This protection was dependent on elevated Treg cell frequencies during primary infection as a result of bystander immune tolerance. Thus, RelB expression in conventional DCs acts as a rheostat to establish a tolerogenic set point that is maintained even during infection and strong type 2 immune conditions and thereby is a key regulator of intestinal homeostasis.

overall_design: Dendritic cells (CD45+, live dead-, B220-, CD64-, CD11c-high, MHCII-high) from SI-LP and mLN of control and RelBdDC mice were isolated by Fluorescence activated cell sorting (FACS) at steady state, day 4 and day 14 after Hpb infection and subjected to scRNA-Sequenzing using the 10x Genomics pipeline

first: Anna-Lena

last: Geiselhöringer


title: Next Generation Sequencing Quantitative Analysis of altered expression of circRNAs in PDAC tissues

status: Public on Sep 20 2023

city: Guangzhou

first: Yao

last: Kong

address: Zhongshan Second Road

institute: Guangdong Provincial People's Hospital

summary: Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest digestive system malignancies; its incidence is increasing and it is predicted to be the second most common cause of death within the next decade. Recent studies demonstrated the increased tendency of LN metastasis in PDAC. Yet the biological role and underlying regulatory mechanism of hsa_circ_0060665 in lymph node (LN) metastasis of PDAC remain unclear. Our study aims to explore the target genes of hsa_circ_0060665 and the underlying mechanism in the LN metastasis of PDAC.

overall_design: To detect the precise mechanism of hsa_circ_0060665 in LN metastasis of PDAC, the total RNA of PDAC tissues and paired normal adjacent tissues was extracted and the next generation sequencing was conducted to explore the expressing profile difference between PDAC tissues and paired normal adjacent tissues. The result was obtained from three independent experiments.

first: Changhao

last: Chen


title: Dendritic Cell Vaccination in Conjunction with TLR Agonist Polarizes Interferon Immune Responses Associated with Long-term Survival in Malignant Glioma Patients

first: Willy

last: Hugo

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

overall_design: Refer to individual Series


title: Bulk RNA-sequencing of normal and Kaposi sarcoma tissues from skin and gastrointestinal tract

status: Public on Sep 25 2023

city: Bethesda

first: Joseph

last: Ziegelbauer

address: 10 Center Dr. 6N110

institute: NIH / National Cancer Institute

summary: Matched tissue samples from 19 patients were harvested for total RNA and analyzed by bulk RNA-sequencing.

overall_design: Tissues were stored in RNAlater and lysed with Trizol. Tissues were homogenized and extracted for total RNA with Direct-Zol Miniprep kit (Zymo). Ribosomal RNA was removed, and sequencing libraries were prepared using NEBnext Ultra Low Input Total RNA with rRNA Depletion Prep (NEB). The reads from the FASTQ files (reads are generated by HiSeq Illumina 4000 and NovaSeq 6000) were trimmed of Illumina adapters usingcutadapt  with default parameter except for --q 10 and--minimum-length 25. We generated a ‘combined genome reference’ by catenating the human reference genome (GRCh38) and KSHV reference genome (NC009333).   The reads (average 45.9 million reads per sample) were aligned to the combined genome reference using STAR (version 2.7.8a). The transcriptome bam files created by STAR was used to estimate the read counts and Transcript per million (TPM) values at the gene level using RSEM v1.3.2. The GENCODE v30 reference gene annotation (GTF) combined with the KSHV gene annotation were used for annotation using STAR and RSEM.

first: Takanobu

last: Tagawa

first: Guruswamy

last: Mahesh


title: C/EBP⍺ Aggravated Ferroptosis and Apoptosis in Renal Tubular Epithelial Cell

status: Public on Sep 14 2023

city: Shanghai

first: Ziru

last: Xia

address: 2nd Ruijin Road

institute: Shanghai Jiaotong University

summary: Acute Kidney Injury (AKI) is widely recognized for its rapid deterioration in renal function, often resulting in unfavorable clinical outcomes. An increase in the expression of CCAAT/enhancer binding protein alpha (C/EBP⍺) has been observed in both human and mouse kidneys affected by AKI. We unraveled the downstream pathway of C/EBP⍺ through RNA-sequencing, confirming our findings with qRT-PCR and luciferase reporter assays. Furthermore, we identified a potential target for AKI reversal in a murine AKI model.

overall_design: Comparative expression of gene in HK2 cell after treatment of ICCB280 (and inducer of CEBPA) or DMSO control for 24hours

first: Xin

last: Li

first: Zhaonan

last: Wei

first: Weiming

last: Wang


title: RNA-binding proteins hnRNPM and ELAVL1 promote type-I interferon induction downstream of the nucleic acid sensors cGAS and RIG-I

status: Public on Sep 16 2023

city: Bonn

first: Thais

middle: Marina

last: Schlee-Guimarães

address: Venusberg-Campus 1

department: Institute of Clinical Chemistry and Clinical Pharmacology

institute: University Hospital, University of Bonn

summary: RIG-I like helicases (RLHs) and cGAS are crucial cytosolic sensors of viral RNA and DNA, respectively, and induce type I IFNs via TBK1/IKKε. hnRNPM was described to possess antimicrobial activities. Here, we show that hnRNPM promotes IRF3 phosphorylation and type I IFN induction downstream of cGAS and RIG-I in THP-1 cells and primary human fibroblasts. Interactome analysis revealed that hnRNPM forms a multiprotein complex with ELAVL1, TBK1, IKKε, IKKβ, and NF-κB p65. hnRNPM, ELAVL1, and TBK1 predominantly interacted in the cytosol. To our knowledge, hnRNPM and ELAVL1 represent the first non-redundant signaling components merging the cGAS-STING and RIG-I–MAVS pathways. Therefore, our findings may have implications for host defense and auto-inflammatory diseases.

overall_design: 3'-mRNA expression profiles of THP-1 dual wildtype (WT) and ELAVL1 knockout (KO) cells 6 h after stimulation of the interferon-α/β receptor (IFNAR) with 1000 U/ml interferon-α (IFNα)

first: Martin

last: Schlee

first: Alexander

last: Kirchhoff


title: Transcriptome analysis of the three equine tissue

status: Public on Aug 31 2023

city: Balice

first: Ewelina

last: Semik-Gurgul

address: Krakowska 1

department: Department of Animal Molecular Biology

country: Poland

institute: National Research Institute of Animal Production

summary: The aim of the project was to identify the tissue-specific patterns of gene expression of selected horse tissues, derived from two germ layers, endodermal (liver and lung) and mesenchymal (cardiac striated muscle) origin.

overall_design: One-condition experiment, heart vs. lung, heart vs. liver, liver vs. lung. Biological replicates: 4 heart replicates, 4 lung replicates, 4 liver replicates.

last: Semik-Guegul

first: Tomasz

last: Szmatoła

first: Artur

last: Gurgul


title: Experimental upregulation of developmentally downregulated ribosomal protein large subunits 7 and 7A promotes axon regeneration after injury in vivo

status: Public on Aug 28 2023

city: Farmington

first: Ephraim

last: Trakhtenberg

state: CT

address: 263 Farmington Ave. RM L4005

department: Neuroscience

institute: University of Connecticut School of Medicine


title: Epigenetic regulation of cell state by H2AFY governs immunogenicity in high-risk neuroblastoma

status: Public on Sep 01 2023

city: Uppsala

first: Xingqi

last: Chen

address: BMC, Husargatan 3

department: Department of Immunology, Genetics and Pathology

country: Sweden

institute: Uppsala university

summary: Childhood neuroblastoma with MYCN-amplification is classified as high-risk and often relapses after intensive treatment regimen. Immune checkpoint blockade therapy shows limited efficacy in neuroblastoma patients and the cancer intrinsic immune regulatory network is poorly understood. Here, we leveraged genome-wide CRISPR/Cas9 screens in a human co-culture system and identified H2AFY as a resistance gene to nivolumab. Genetic deletion of H2afy in MYCN-driven neuroblastoma cells reverted in vivo resistance to PD-1 blockade by eliciting concurrent activation of the adaptive and innate immunity. Analysis of single-cell RNA sequencing datasets revealed that H2AFY mRNA was enriched in adrenergic cancer cells and was associated with patient survival. Mapping of the epigenetic and translational landscape demonstrated that H2afy deletion promoted cell transition to a malignant mesenchymal-like state. With a multi-omics approach, we uncovered H2AFY-associated genes that were functionally relevant and prognostic in patients. Altogether, our study elucidates the role of H2AFY as an epigenetic gatekeeper for cell state and immunogenicity in high-risk human neuroblastoma.

overall_design: H2AFY may serve as an epigenetic gatekeeper for cell state transition and malignant behavior in human NB. To validate this hypothesis, we mapped the epigenetic landscape of control or H2afy CRISPR KO NB cells using ATAC-seq. ATAC-seq in control or H2afy CRISPR KO 9464D cells

first: Divya

last: Nagarajan

first: Minglu

last: Xie

first: Maede

last: Salehi

first: Yumeng

last: Mao


title: Experimental upregulation of developmentally downregulated ribosomal protein large subunits 7 and 7A promotes axon regeneration after injury in vivo [bulk RNA-Seq]

summary: Ribosomal proteins are involved in neurodevelopment and central nervous system (CNS) disease and injury. However, the roles of specific ribosomal protein subunits in developmental axon growth, and their potential as therapeutic targets for treating CNS injuries, are still poorly understood. Here, we show that ribosomal protein large (Rpl) and small (Rps) subunit genes are substantially (56-fold) enriched amongst the genes, which are downregulated during maturation of retinal ganglion cell (RGC) CNS projection neuron. We also show that RGC subtype-specific Rpl and Rps subunits are highly co-regulated, and partially re-upregulated after optic nerve crush (ONC). Because developmental downregulation of ribosomal proteins coincides with developmental decline in neuronal intrinsic axon growth capacity, we hypothesized that Rpl/Rps incomplete re-upregulation after injury may be a part of the cellular response which attempts to reactivate intrinsic axon growth mechanism. We found that experimentally upregulating Rpl7 and Rpl7A promoted axon regeneration after ONC in vivo. Finally, we characterized gene networks associated with Rpl/Rps, and showed that Rpl7 and Rpl7A belong to the cluster of genes, which are shared between translational and neurodevelopmental biological processes (based on gene-ontology) that are co-downregulated in maturing RGC during the decline in intrinsic axon growth capacity.

overall_design: Bulk-mRNA-seq samples from different age RGCs were purified by immunopanning for Thy1 (after immunopanning depletion of macrophages), cDNA libraries prepared using polyA-selected RNA (TruSeq RNA Library Prep Kit, Illumina), paired reads sequenced 100 bp from each end on HiSeq 2000 Sequencer (Illumina), passed QC filters, mapped to the mm10 genome and transcriptome by Hisat2, and analyzed by Cufflinks/CuffDiff, as we previously described.

first: William

middle: C

last: Theune

first: Jian

last: Xing


title: Lineage tracing of newly accrued nuclei in skeletal myofibers uncovers distinct transcripts and interplay between nuclear populations

status: Public on Aug 23 2023

city: Cincinnati

first: Douglas

last: Millay

state: OH

address: 3333 Burnet Ave Cincinnati, OH 45229

department: Molecular Cardiovascular Biology

institute: Cincinnati Children's Hospital Medical Center

summary: Multinucleated skeletal muscle cells have an obligatory need to acquire additional nuclei through fusion with activated skeletal muscle stem cells when responding to both developmental and adaptive growth stimuli. A fundamental question in skeletal muscle biology has been the reason underlying this need for new nuclei in syncytial cells that already harbor hundreds of nuclei. To begin to answer this long-standing question, we utilized nuclear RNA-sequencing approaches and developed a lineage tracing strategy capable of defining the transcriptional state of recently fused nuclei and distinguishing this state from that of pre-existing nuclei. Our findings reveal the presence of conserved markers of newly fused nuclei both during development and after a hypertrophic stimulus in the adult. However, newly fused nuclei also exhibit divergent gene expression that is determined by the syncytial environment they fuse into. Moreover, accrual of additional nuclei through fusion is required for nuclei already resident in adult myofibers to mount a normal transcriptional response to a load-induced stimulus. We propose a model of mutual regulation in the control of skeletal muscle development and adaptations, where newly fused and pre-existing myonuclei influence each other to maintain optimal functional growth.

overall_design:
For bulk nuclear sequencing, to obtain sufficient material for sequencing, skeletal muscles were pooled together for this analysis. For the early development stage, 4 × Gastrocnemius and 4 × tibialis anterior muscles per replicate were used, while for adulthood muscle overload, 4 × Plantaris muscles per replicate were used. Nuclei were extracted using the protocol described in the methods section above. The washed nuclei pellets were resuspended in sorting buffer (2% BSA/RNase-free PBS) and labeled with anti-PCM1 (1:500, HPA023374, Sigma-Aldrich) on ice for 45 minutes. Nuclei were washed twice in sorting buffer and then labeled with an Alexa Fluor 647-conjugated secondary antibody (1:100, Invitrogen) at 4°C for 30 minutes, followed by labeling with Hoechst dye. The nuclei were washed again and resuspended in sorting buffer with 0.2 U/μl Protector Rnase inhibitor prior to cell sorting to separate the PCM1-labeled fractions using a FACS cell sorter (BD Aria). The sorted nuclei were then pelleted at 3000 × g for 10 minutes before mRNA isolation using the Direct-zol RNA Microprep kit (Zymo research R2060).

For snRNA-Seq datasets, plantaris muscles from HSArtTA;TREH2B-GFP mice were used for snRNA-seq in this study. Four pieces of plantaris muscle, pooled from two mice that either received or did not receive synergistic ablation, were used for nuclei isolation using the protocol described above. After filtration via a 40 μm strainer, uuclei were labeled with Hoechst dye and 0.2 U/μl Protector RNase inhibitor (Roche). All GFP+ Hoechst stained myonuclei were purified using FACS (BD Aria, 70 μm nozzle) and gathered in sorting buffer containing Protector RNase inhibitor (0.2 U/μl). The nuclei were counted with a hemocytometer, and the concentration was fine-tuned as needed to reach the optimal range for the 10X Chromium chip. The 10X Chromium system was then used to load the nuclei using the Single Cell 3′ Reagent Kit v3.1, following the guidelines provided by the manufacturer. Around 12,000 nuclei were loaded for each operation. Sequencing was performed on an Illumina NovaSeq 6000 System.

first: Chengyi

last: Sun

first: Casey

middle: O

last: Swoboda

first: Michael

last: Petrany

first: Sreeja

last: Parmeswaran

first: Andrew

last: VonHandorf

first: Matthew

last: Weirauch

first: Christoph

last: Lepper

middle: P


title: Non-Electrophilic NRF2 ActivatorsPromote Wound Healingin Human KeratinocytesandDiabetic Miceand DemonstrateSelectiveDownstreamGene Targeting

city: Chicago

first: Terry

last: Moore

state: IL

address: 833 S Wood St

department: Pharmaceutical Sciences

institute: University of Illinois at Chicago College of Pharmacy

summary: The transcription factor NRF2 plays an important role in many biological processes and is a promising therapeutic target. NRF2 is highly expressed in the skin and is known to play a critical role in diabetic wound healing, a disease state with limited treatment options. However, many existing NRF2 activators display off-target effects due to their electrophilic mechanism, underscoring the need for alternative approaches. In this work, we investigated two recently described, non-electrophilic NRF2 activators, ADJ-310 and PRL-295, and demonstrated their efficacy in vitro and in vivo. Both ADJ-310 and PRL-295 maintained human keratinocyte cell viability at increasing concentrations and maintained or improved cell proliferation over time. Both compounds also increased cell migration, improving in vitro wound closure. ADJ-310 and PRL-295 enhanced the oxidative stress response in vitro, and RNA-sequencing data showed that PRL-295 activated NRF2 with a narrower transcriptomic effect than the widely used electrophilic NRF2 activator, CDDO-Me. In vivo, both ADJ-310 and PRL-295 improved wound healing in Leprdb/db diabetic mice. The non-electrophilic compounds ADJ-310 and PRL-295 are effective, innovative tools for the investigation of the function of NRF2 that directly address the need for alternative NRF2 activators. They offer a new approach to studying the role of NRF2 in human diseases and its potential as a therapeutic. Both non-electrophilic NRF2 activators promoted wound healing functions in human keratinocytes and improved wound healing in diabetic mice, demonstrating their efficacy both in vitro and in vivo. RNA sequencing showed that PRL-295 upregulated downstream target genes that favorably compared with those of CDDO-Me.

overall_design:
Gene expression profiling and comparative analysis of RNASeq data from human keratinocyes cells treated with 250 nM of CDDO-Me or 10 μM of PRL-295.
*** This submission does not contain raw data due to corruption of files. ***

first: May

last: Barakat

first: Luisa

middle: A

last: DiPietro

first: Lin

first: Sekhar

last: Reddy

first: Aparna

last: Ankireddy

first: Kornelia

last: Skowron

first: Tatum

last: Johnson

first: Brian

last: David

first: Junhe

last: Shi

first: Reginald

last: Woods

first: Angela

last: Xu


title: Transcriptional profiling unveils molecular subgroups of adaptive and maladaptive right ventricular remodeling in pulmonary hypertension

first: Fatemeh

last: Khassafi


title: Functional study of a small transcriptional factor CdsR

status: Public on Aug 22 2023

city: Beijing

first: zhang

last: xin

address: No.2 West Yuanmingyuan Road

department: Institute of Plant Protection

institute: Chinese Academy of Agricultural Sciences

summary: We then performed gene expression profiling using data obtained from RNA-seq of 2 different cells at three time points.

sample_organism: Bacillus thuringiensis serovar kurstaki str. HD73

overall_design: Comparative gene expression profiling of RNA-seq data for HD73 wild-type and cdsR gene deletion mutation.

first: Xin

last: Zhang

first: Hengjie

last: Wang

first: Yuhan

last: Chen

first: Qi

last: Peng

first: Fuping

last: Song


title: Effects of PARP inhibition on the transcriptome in BRCA1 wild-type and BRCA1 deficient ovarian cancer cell lines

status: Public on Aug 01 2023

city: Innsbruck

first: Leonie

last: Madersbacher

address: Innrain 80/82

country: Austria

institute: Medical University of Innsbruck

summary: Despite numerous therapeutic advances over the years, ovarian cancer, especially high grade serous ovarian carcinoma remains the deadliest gynecological malignancy. Although PARP inhibition has been shown to be an effective (maintenance) therapy for homologous recombination repair deficient or BRCA1 mutated ovarian cancer, there may be further potential for combination therapy with other drugs such as immune checkpoint inhibitors. BRCA1 mutation as well as PARP inhibitor (Olaparib) treatment influenced the activation of immune response pathways such as cGAS-STING signaling and Interferon-alpha response in ovarian cancer cell lines. Bioinformatics functional analyses uncovered further immune related cellular responses and signaling pathways such as JAK-STAT signaling.

overall_design:
To investigate the effect of the PARP inhibitor Olaparib in both BRCA1 wildype (OVCAR3) and BRCA1 mutated (UWB1.289) cells, the IC50 value for 96h treatment with Olaparib in both cell lines was determined.
Both cell lines were either treated with Olaparib (determined IC50 value) or vehicle control (DMSO) and harvested for RNA extraction after 96h. This was repeated four times.
RNA-seq was done for all samples, raw data was processed and comparative gene expression analyses were conducted.

first: Raphael

last: Gronauer

first: Hubert

last: Hackl


title: Identification of two subsets of subcompartment A1 associated with high transcriptional activity and frequent loop extrusion

status: Public on Jul 31 2023

first: Ya

last: Guo

address: 800 Dongchuan Road

department: School of Life Sciences and Biotechnology

institute: Shanghai Jiao Tong University

summary: Three-dimensional genome organization has been increasingly recognized as an important de-terminant of the precise regulation of gene expression in mammalian cells, yet the relationship between gene transcriptional activity and spatial subcompartment positioning is still not fully comprehended. Here, we first utilized genome-wide Hi-C data to infer eight types of subcom-partment (labeled A1, A2, A3, A4, B1, B2, B3, and B4) in mouse embryonic stem cells and four primary differentiated cell types, including thymocytes, macrophages, neural progenitor cells, and cortical neurons. Transitions of subcompartments may confer gene expression changes in different cell types. Intriguingly, we identified two subsets of subcompartments defined by higher gene density and characterized by strongly looped contact domains, named common A1 and variable A1, respectively. We revealed that common A1, which includes highly expressed genes and abundant housekeeping genes, shows a ~2-fold higher gene density than the variable A1, where cell type-specific genes are significantly enriched. Thus, our study supports a model in which both types of genomic loci with constitutive and regulatory high transcriptional activity can drive the subcompartment A1 formation. Special chromatin subcompartment arrangement and intradomain interactions may, in turn, contribute to maintaining proper levels of gene expression, especially for regulatory non-housekeeping genes.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for H3K27ac in K562 cells. Cleavage Under Targets and Tagmentation (CUT&Tag) for Nipbl or H3K27ac in K562 or Hela cells.

first: Shuang

last: Cui

first: Zihang

last: Yin


title: Effect of common mutations in clonal hematopoiesis of indeterminate potential (CHIP) on the transcriptome in bone marrow derived macrophages (BMDM).

status: Public on Jul 26 2023

city: Boston

first: Philipp

last: Rauch

state: MA

address: 450 Brookline Ave

institute: DFCI

summary: Clonal hematopoiesis of indeterminate potential (CHIP) is associated with an increased risk of inflammatory conditions, including cardiovascular disease, in humans. The goal of the present data set was to examine the molecular mechanism underlying the increase in inflammation seen with the two most common CHIP mutations, namely loss-of-function mutations in Dnmt3a and Tet2, using murine bone-marrow derived macrophages (BMDM) as a model system.

overall_design: We performed transcriptome analysis by RNAseq in bone-marrow derived macrophages from WT (Vav-Cre), Dnmt3aKO (Dnmt3afl/fl;Vav-Cre), Tet2KO (Tet2fl/fl;Vav-Cre) and Jak2VF (Jak2V617F/WT;Vav-Cre) mice, stimulated for 24h with either 200 mg/dL low-density lipoprotein (LDL) or vehicle control (ctl).

first: Jayakrishnan

last: Gopakumar

first: Alexander

last: Silver

first: Benjamin

last: Ebert

first: Siddhartha

last: Jaiswal


title: Genome-wide Translational Response of Candida albicans to Fluconazole Treatment

status: Public on Aug 05 2023

city: San Antonio

first: David

last: Kadosh

state: Texas

address: 7703 Floyd Curl Dr., MC: 7758

department: Department of Microbiology, Immunology & Molecular Genetics

institute: University of Texas Health Science Center at San Antonio

summary: Azoles are commonly used for the treatment of fungal infections and the ability of human fungal pathogens to rapidly respond to azole treatment is critical for the development of antifungal resistance. While the role of genetic mutations, chromosomal rearrangements and transcriptional mechanisms in azole resistance has been well-characterized, very little is known about post-transcriptional and translation mechanisms that drive this process. In addition, most previous genome-wide studies have focused on transcriptional responses to azole treatment, and likely serve as an inaccurate proxies due to extensive post-transcriptional and translational regulation. In this study we use ribosome profiling to provide the first picture of the global translational response of a major human fungal pathogen, Candida albicans, to treatment with fluconazole, one of the most widely used azole drugs. We identify sets of genes showing significantly altered translational efficiency (TE), including genes associated with a variety of biological processes such as the cell cycle, DNA repair, cell wall/cell membrane biosynthesis, transport, signaling, DNA- and RNA-binding activities and protein synthesis. Importantly, while there are similarities and differences among gene categories that are regulated by fluconazole at the translational vs. transcriptional levels, we observe very little overlap among individual genes controlled by these mechanisms. Our findings suggest that C. albicans possesses distinct translational mechanisms that are important for the response to antifungal treatment, which could eventually be targeted by novel antifungal therapies.

overall_design: Examination of gene expression by RNA-seq and Ribo-seq in 1 strain of Candida albicans grown under 2 conditions (YEPD + 1 ug/mL fluconazole at  30°C  (Fluconazole)) and (YEPD + DMSO vehicle only control at 30°C (No drug)). There are four biological replicates for each strain grown under each condition (16 samples total).

first: Saket

last: Choudhary


title: Loss-of-Function Mutations in Dnmt3a and Tet2 Lead to Accelerated Atherosclerosis and Concordant Macrophage Phenotypes

status: Public on Jul 24 2023

city: Palo Alto

first: Jayakrishnan (Jk)

state: CA

address: 240 Pasteur Dr Rm 4600

institute: Stanford

summary: We used single cell RNA sequencing (scRNA-seq) to analyze the different immune cell populations present in atheromatous plaques of mice fed an atherogenic diet with or without CHIP mutations

overall_design: CD45.1 or CD45.2 immune cells were sorted from atheromas before scRNAseq was performed. There is a 30 week cohort and a 24 week cohort, and for each timepoint we have WT, Dnmt3a KO, and Tet2 KO cells

first: Phillip

first: J

middle: K


title: ATF3- and ZIKV-dependent transcriptome of A549 cells

status: Public on Jun 30 2023

city: Albany

first: Morgan

last: Sammons

state: NY

address: 1400 Washington Ave, Life Sciences 2075

department: Biological Sciences

institute: University at Albany - SUNY

summary: We examined the transcriptome (polyA+ RNAseq) of parental or ATF3 knock-out A549 lung epithelial adenocarcinoma cell lines in response to either a mock infection or infection with ZIKV.

overall_design: Comparative gene expression profiling (polyA+ selected RNAseq) on three biological replicates

first: Cara

last: Pager


title: Restoring Tumor Immunogenicity with Dendritic Cell Reprogramming

status: Public on Jul 07 2023

city: Lund

first: Ilia

last: Kurochkin

state: Skane

address: BMC A12, Sölvegatan 17

institute: Lund University


title: Experimental upregulation of developmentally downregulated ribosomal protein large subunits 7 and 7A promotes axon regeneration after injury in vivo [scRNA-Seq]

first: Ephraim

last: Trakhtenberg

relation:
Reanalysis of: GSM4148508
Reanalysis of: GSM3466902
Reanalysis of: GSM3466903
Reanalysis of: GSM3177677
Reanalysis of: GSM3177678
Reanalysis of: GSM3907320
Reanalysis of: GSM3907321
Reanalysis of: GSM3907322
Reanalysis of: GSM3907323
Reanalysis of: GSM3907324
Reanalysis of: GSM3907325
Reanalysis of: GSM3907326
Reanalysis of: GSM3907327
Reanalysis of: GSM3907328
Reanalysis of: GSM3907329
Reanalysis of: GSM4078555
Reanalysis of: GSM4078556
Reanalysis of: GSM4078557
Reanalysis of: GSM4078558
SubSeries of: GSE240317

overall_design:
Normalized matrices and cell metadata (e.g., type assignment) for embryonic, adult uninjured, and adult injured RGCs, are available from the GEO accession numbers GSE122466 and GSE137400. Postnatal RGC scRNA-seq dataset we previously generated is available under the Gene Expression Omnibus (GEO) accession number GSE115404.
Cell-counts matrices were obtained from the GEO submissions from the previously published data. Matrices were loaded as seurat objects in R.
Seurat objects were merged and expression was normalized. A cell counts matrix for the merged dataset was then generating by extracting counts matrix from seurat object.
The condition each cell belongs to was appended to the cell ID in the columns of the matrix
mm10
One CSV files: Containg RGC embyonic, postnatal, adult and injured cells' normalized expression values for each gene.

first: William

last: Theune

first: Xing

last: Jian


title: Detection of plant microRNAs in edible-plant based products

status: Public on Sep 05 2023

city: Pamplona

first: Fermín

middle: Ignacio

last: Milagro Yoldi

state: Navarra

department: Department of Nutrition, Food Science and Physiology/Center for Nutrition Research

institute: University of Navarra

type: Non-coding RNA profiling by high throughput sequencing

summary: Plant-based diets could be a key source of microRNAs in animals. Plant microRNAs are cross-kingdom gene expression regulators that could modulate mammalian gene expression, influencing their physiology. Therefore, it is important to identify the microRNA expression profile of plant foods in order to identify potential target genes and biological functions in the mammalian host. Next-generation sequencing was applied to identify microRNAs in RNA samples derived from nuts (walnut and almond), vegetables (spinach) and fruits (orange, apple, olive, pear, and tomato). Our data revealed that edible plant contain a large number and diverse type of microRNAs.

sample_organism:
Citrus sinensis
Spinacia oleracea
Malus domestica
Prunus dulcis
Solanum lycopersicum
Olea europaea
Pyrus communis
Juglans regia

overall_design: To identify microRNAs from edible plants, next-generation sequencing was performed with total RNA isolated from 0.1 g of fruits and vegetables (orange, apple, olive, pear, tomato, spinach) and 0.5 g of nuts (walnut and almond).

first: E

last: Díez-Sainz

first: S

last: Lorente-Cebrián

first: P

last: Aranaz

middle: I

last: Riezu-Boj

first: F

last: Milagro


title: A tracheal aspirate-derived airway basal cell model reveals a proinflammatory epithelial defect in congenital diaphragmatic hernia

first: Xingbin

last: Ai

address: 55 Fruit St

department: Pediatrics

institute: MASSACHUSETTS GENERAL HOSPITAL

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

overall_design: Refer to individual Series


title: A tracheal aspirate-derived airway basal cell model reveals a proinflammatory epithelial defect in congenital diaphragmatic hernia (RNA-Seq)

summary: Congenital diaphragmatic hernia (CDH) is characterized by incomplete closure of the diaphragm and lung hypoplasia. The pathophysiology of lung defects in CDH is poorly understood. To establish a translational model of human airway epithelium in CDH for pathogenic investigation and therapeutic testing, we developed a robust methodology of epithelial progenitor derivation from tracheal aspirates of newborns. Basal stem cells from CDH patients and preterm and term, non-CDH controls were derived and analyzed by bulk RNA-sequencing, ATAC-sequencing, and air-liquid-interface differentiation. Transcriptomic and epigenetic profiling of CDH and non-CDH basal stem cells reveals a disease-specific, proinflammatory signature independent of severity or hernia size. In addition, CDH basal stem cells exhibit defective epithelial differentiation in vitro that recapitulates epithelial phenotypes found in fetal human CDH lung samples and fetal tracheas of the nitrofen rat model of CDH. Furthermore, steroid treatment normalizes epithelial differentiation phenotypes of human CDH basal stem cells in vitro and in nitrofen rat tracheas in vivo. Our findings have identified an interplay between a proinflammatory signature and BSC differentiation as a potential therapeutic target for airway epithelial defects in CDH.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for basal stem cells from CDH patients and term and preterm controls.

first: Richard

last: Wagner

first: Gaurang

middle: M

last: Amonkar

first: Wei

last: Wang

first: Paul

middle: H

last: Lerou