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Surface Lin28A Expression Consistent With Cellular Stress Parallels Indicators of Senescence

ABSTRACT: Single-cell RNA-seq (10X Genomics Chromium) to profile Adult cardiac interstitial sorted based on the presence of Lin28 on cell membrane surface Lin28S+/-.

ORGANISM(S): Mus musculus

PROVIDER: GSE186176 | GEO | 2021/10/22

REPOSITORIES: GEO

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Surface Lin28A expression consistent with cellular stress parallels indicators of senescence.

Project description:AimsDeclining cellular functional capacity resulting from stress or ageing is a primary contributor to impairment of myocardial performance. Molecular pathway regulation of biological processes in cardiac interstitial cells (CICs) is pivotal in stress and ageing responses. Altered localization of the RNA-binding protein Lin28A has been reported in response to environmental stress, but the role of Lin28A in response to stress in CICs has not been explored. Surface Lin28A redistribution is indicative of stress response in CIC associated with ageing and senescence.Methods and resultsLocalization of Lin28A was assessed by multiple experimental analyses and treatment conditions and correlated to oxidative stress, senescence, and ploidy in adult murine CICs. Surface Lin28A expression is present on 5% of fresh CICs and maintained through Passage 2, increasing to 21% in hyperoxic conditions but lowered to 14% in physiologic normoxia. Surface Lin28A is coincident with elevated senescence marker p16 and beta-galactosidase (β-gal) expression in CICs expanded in hyperoxia, and also increases with polyploidization and binucleation of CICs regardless of oxygen culture. Transcriptional profiling of CICs using single-cell RNA-Seq reveals up-regulation of pathways associated with oxidative stress in CICs exhibiting surface Lin28A. Induction of surface Lin28A by oxidative stress is blunted by treatment of cells with the antioxidant Trolox in a dose-dependent manner, with 300 μM Trolox exposure maintaining characteristics of freshly isolated CICs possessing low expression of surface Lin28A and β-gal with predominantly diploid content.ConclusionSurface Lin28A is a marker of environmental oxidative stress in CICs and antioxidant treatment antagonizes this phenotype. The biological significance of Lin28 surface expression and consequences for myocardial responses may provide important insights regarding mitigation of cardiac stress and ageing.

| S-EPMC10409908 | biostudies-literature

Surface Lin28A Expression Consistent With Cellular Stress Parallels Indicators of Senescence

Project description:Surface Lin28A Expression Consistent With Cellular Stress Parallels Indicators of Senescence

| PRJNA772780 | ENA

Transcriptional features of biological age maintained in cultured cardiac interstitial cells

Project description:Single-cell RNA-seq (10X Genomics Chromium) to profile cultured cardiac interstitial cells derived from neonatal and adult cardiac tissue

2020-12-31 | GSE164026 | GEO

"Youthful" Phenotype of c-Kit+ Cardiac Fibroblasts [DDR2_cKit]

Project description:Single-cell RNA-seq (10X Genomics Chromium) to profile Adult cardiac interstitial cells sorted based on the presence of cKit, DDR2 and CD45 on cell membrane surface .

2021-11-18 | GSE188804 | GEO

Single cell gene expression, immune receptor and cell surface marker expression profiling of Spike-specfic splenic T and B cells of immunized mice

Project description:Spike-specific T and B cells from splenocytes of immunized mice were profiled with 10x sequencing for gene expression (Chromium Next Automated GEM 5’ v2 kit), cell surface marker (Chromium Automated 5’ Feature Barcode kit) and immune receptor (Chromium Automated Mouse BCR/TCR Amplification and Library Construction kit)

2023-08-07 | GSE222403 | GEO

LIN28A modulates splicing and gene expression programs in breast cancer cells [RIP-Seq]

Project description:The goals of this study were to identify LIN28 downstream gene targets in breast cancer cells. We use a subclone of the MCF-7 breast cancer cell line, MCF-7M as our model system. Methods: mRNA-protein complexes (mRNP) lysates were prepared from MCF-7M cells and incubated with Protein-A Sepharose beads (Sigma-Aldrich) and either LIN28 (Abcam) or control normal rabbit serum IgG antibodies. LIN28 interacting mRNAs were identified by whole genome sequencing. Results: Using an optimized data analysis workflow, we mapped approximately 13 million sequence reads for LIN28-IP and CTL- IP (IgG), respectively to the to the human genome (build h19). Conclusions: mRNA were significantly bound by LIN28 if LIN28 RIP had 2.5 fold increase in normalized reads compared to IgG. We found that LIN28 was predominantly bound at coding exons and 3'UTRs, 38% & 45% respectively, in the 843 mRNAs within MCF-7M genome.

2015-07-27 | GSE71009 | GEO

RNA-seq analysis of mRNAs immunoprecipitated by wild-type (WT) or phospho-mimetic (S200D) FLAG-LIN28A in PA1 cells

Project description:To explore the effect of LIN28 (S200) phosphorylation on LIN28's association with mRNAs on a transcriptome-wide scale, we performed mRNA-seq on RNAs immunoprecipitated (RIP-seq) by wild-type (WT) or phospho-mimetic (S200D) LIN28 in PA1 cells.

2016-10-28 | GSE83905 | GEO

RNA-seq analysis of mRNAs immunoprecipitated by wild-type (WT) or phospho-null (S200A) FLAG-LIN28A in HeLa cells

2016-10-28 | GSE83904 | GEO

LIN28A modulates splicing and gene expression programs in breast cancer cells [RNA-Seq]

Project description:The goals of this study were to identify LIN28 downstream gene targets in breast cancer cells. We use a subclone of the MCF-7 breast cancer cell line, MCF-7M as our model system. Methods: mRNA profiles from MCF-7M breast cancer cells treated with siRNA against non-targeting control (NT), LIN28, hnRNP A1, LIN28 and hnRNPA1 (LIN28A1) for 72 hrs were generated by deep sequencing, in duplicate, using Illumina HiSeq 2000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays Results: Using an optimized data analysis workflow, we mapped over 200 million sequence reads per sample to the human genome (build h19). Each of the four groups had two biological replicates. We developed a custom method to identify alternative splicing events and identified 111 genes with significant (FDR<0.05) differential splicing for LIN28 depleted cells compared to non-targeting siRNA control, as well as 249 and 182 genes for hnRNP A1 and LIN28A1 respectively. RNA-seq data were validated with by qRT–PCR analysis of a subset of genes. Conclusions: Results reveal that LIN28 regulates alternative splicing and steady state mRNA expression of genes implicated in aspects of breast cancer biology. Notably, cells lacking LIN28 undergo significant isoform switching of the ENAH gene, resulting in a decrease in the expression of ENAH exon 11a isoform. Expression of ENAH isoform 11a has been shown to be elevated in breast cancers that express HER2. mRNA profiles of MCF-7M cells treated with siRNA for NT control, LIN28, hnRNP A1, and LIN28 plus hnRNP A1 (A1) (LIN28A1) were generated by deep sequencing, in duplicate, using Illumina HiSeq 2000

2015-07-27 | E-GEOD-71012 | biostudies-arrayexpress

Joint single-cell/single-nucleus RNA-seq on hearts from healthy or HFpEF mice

Project description:In this project we explore the cellular heterogeneity of a mouse model of heart failure with preserved ejection fraction (HFpEF) involving a two-hit model of feeding a high fat diet (HFD) along with L-NAME administration. Healthy adult male mice (C57BL/6J inbred) were fed either a normal chow diet or HFD/L-NAME for 10 weeks or 15 weeks before performing sequencing experiments. Both cardiomyocytes (CMs) and total interstitial population (TIP) were captured using a protocol to jointly capture and sequence single-nuclei (for cardiomyocytes) and single-cells (for TIP) using the 10x Genomics Chromium system.

2024-05-08 | E-MTAB-13362 | biostudies-arrayexpress

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