Ablation of the epithelial-specific splicing factor Esrp1 results in ureteric branching defects and reduced nephron number
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE81716 | GEO | 2016/05/21
SECONDARY ACCESSION(S): PRJNA322348
REPOSITORIES: GEO
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Project description:BackgroundAbnormalities in ureteric bud (UB) branching morphogenesis lead to congenital anomalies of the kidney and reduced nephron numbers associated with chronic kidney disease (CKD) and hypertension. Previous studies showed that the epithelial fibroblast growth factor receptor 2 (Fgfr2) IIIb splice variant supports ureteric morphogenesis in response to ligands from the metanephric mesenchyme during renal organogenesis. The epithelial-specific splicing regulator Esrp1 is required for expression of Fgfr2-IIIb and other epithelial-specific splice variants. Our objective was to determine whether Esrp1 is required for normal kidney development.ResultsAblation of Esrp1 in mice, alone or together with its paralog Esrp2, was associated with reduced kidney size and increased incidence of renal aplasia. Three-dimensional imaging showed that embryonic Esrp1 knockout (KO) kidneys had fewer ureteric tips and reduced nephron numbers. Analysis of alternative splicing in Esrp-null ureteric epithelial cells by RNA-Seq confirmed a splicing switch in Fgfr2 as well as numerous other transcripts.ConclusionsOur findings reveal that Esrp1-regulated splicing in ureteric epithelial cells plays an important role in renal development. Defects in Esrp1 KO kidneys likely reflect reduced and/or absent ureteric branching, leading to decreased nephron induction secondary to incorrect Fgfr2 splicing and other splicing alterations. Developmental Dynamics 245:991-1000, 2016. © 2016 The Authors. Developmental Dynamics published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists.
| S-EPMC5096029 | biostudies-literature
Project description:The epithelial splicing regulatory proteins, ESRP1 and ESRP2 are essential for mammalian development through regulation of a global program of alternative splicing of genes involved in maintenance of epithelial cell function. To further inform our understanding of the molecular functions of ESRP1 we performed enhanced crosslinking immunoprecipitation coupled with high throughput sequencing (eCLIP) in epithelial cells of mouse epidermis. The genome-wide binding sites of ESRP1 were integrated with RNA-Seq analysis of alterations in splicing and total gene expression that result from epidermal ablation of Esrp1 and Esrp2. These studies demonstrated that ESRP1 functions in splicing regulation occur primarily through direct binding in a position-dependent manner to either promote exon inclusion or skipping. In addition, we also identified widespread binding of ESRP1 in 3’ and 5’ untranslated regions (UTRs) of genes involved in epithelial cell function, suggesting that its post-transcriptional functions extend beyond splicing regulation.
2022-09-12 | GSE212304 | GEO
Project description:Epithelial Splicing Regulatory Proteins 1 and 2 (ESRP1 and ESRP2) are recently discovered epithelial-specific RNA-binding proteins that promote splicing of the epithelial variant of the FGFR2, ENAH, CD44, and CTNND1 transcripts. To catalogue a larger set of splicing events under the regulation of the ESRPs, we profiled splicing changes induced by RNA interference-mediated knockdown of ESRP1 and ESRP2 expression in a human epithelial cell line using the splicing-sensitive Affymetrix Exon ST1.0 Arrays. Analysis of the microarray data using the previously described MADS tool resulted in the identification of over a hundred candidate ESRP-regulated splicing events. We were able to independently validate 37 of these targets by RT-PCR. The ESRP-regulated events encompass all known types of alternative splicing events. Importantly, a number of these regulated splicing events occur in gene transcripts that encode proteins with well-described roles in the regulation of actin cytoskeleton organization, cell-cell adhesion, cell polarity, and cell migration. In sum, this work reveals a novel list of transcripts differentially spliced in epithelial and mesenchymal cells, implying that coordinated alternative splicing plays a critical role in determination of cell type identity. Keywords: control / knockdown comparison Short interfering knockdown of ESRP1 and ESRP2 in human PNT2 prostatic epithelium cells was performed as described before (Warzecha et al., 2009, Molecular Cell 33:591-601). The efficiency of ESRP1 and ESRP2 knockdown was monitored by quantitative RT-PCR as described before (Warzecha et al., 2009, Molecular Cell 33:591-601). In all cases the efficiency of the knockdown was close to 80%. We conducted Exon array profiling on RNAs from four siESRP1/2-treated samples and four siGFP controls.
2010-08-18 | E-GEOD-17468 | biostudies-arrayexpress
Project description:Nephrons, the functional units of the kidney, develop from progenitor cells (cap mesenchyme [CM]) surrounding the epithelial ureteric bud (UB) tips. Reciprocal signaling between UB and CM induces nephrogenesis and UB branching. Although low nephron number is implicated in hypertension and renal disease, the mechanisms that determine nephron number are obscure. To test the importance of nephron progenitor cell number, we genetically ablated 40% of these cells, asking whether this would limit kidney size and nephron number or whether compensatory mechanisms would allow the developing organ to recover. The reduction in CM cell number decreased the rate of branching, which in turn allowed the number of CM cells per UB tip to normalize, revealing a self-correction mechanism. However, the retarded UB branching impaired kidney growth, leaving a permanent nephron deficit. Thus, the number of fetal nephron progenitor cells is an important determinant of nephron endowment, largely via its effect on UB branching.
| S-EPMC4049224 | biostudies-literature
Project description:Tissue- and cell-type specific regulators of alternative splicing (AS) are an essential layer of posttranscriptional gene regulation necessary for normal cellular function, patterning, and development. Here we report the Epithelial splicing regulatory proteins (Esrps) are required for patterning of multiple organs, with loss of both paralogs, Esrp1 and Esrp2, resulting in increasingly severe phenotypes. Global profiling of the Esrp splicing regulatory network from total epidermis revealed varied splicing sensitivity of Esrp targets upon loss of Esrp1 or double knockout. This may explain the progressive phenotypes seen in Esrp knockout mice, and these mice provide a unique genetic tool to evaluate functional consequences of epithelial splicing events in vivo. RNA from purified total epidermis (basal keratinocyte layer to cornified layer) of E18.5 mouse embryos were harvested by Trizol extraction. (Esrp1+/+, Esrp2-/- (n=2), Esrp1-/-, Esrp2+/+ (n=3), Esrp1-/-, Esrp2+/- (n=2), and Esrp1-/-, Esrp2-/- (n=2). 1 ug of total RNA was used for for RNA-seq library preparation using the TruSeq™ Stranded mRNA LT Sample Prep Kit (Illumina). 100x2 bp paired-end RNA-seq reads were generated on a HiSeq 2000 sequencer.
2015-09-08 | E-GEOD-64357 | biostudies-arrayexpress
Project description:Epithelial specific splicing regulatory protein 1 and 2 (ESRP1 and ESRP2) are important regulators of alternative splicing during EMT. To study the alternative splicing events regulated by ESRP1/2 at a genome wide scale, we used lentiviral shRNAs to knockdown ESRP1/2 in H358 cells and performed RNA-seq in biological triplicates. We used lentiviral based shRNAs targeting ESRP1 and ESRP2 to knockdown both regulators in human H358 cells. We harvested total RNA and protein from ESRP1/2 knockdown and control knockdown in biological triplicates. We made cDNA libraries for each replicate and subjected them to RNA-seq.
2016-04-05 | E-GEOD-75489 | biostudies-arrayexpress
Project description:Epithelial Splicing Regulatory Proteins 1 and 2 (ESRP1 and ESRP2) are recently discovered epithelial-specific RNA-binding proteins that promote splicing of the epithelial variant of the FGFR2, ENAH, CD44, and CTNND1 transcripts. To catalogue a larger set of splicing events under the regulation of the ESRPs, we profiled splicing changes induced by RNA interference-mediated knockdown of ESRP1 and ESRP2 expression in a human epithelial cell line using the splicing-sensitive Affymetrix Exon ST1.0 Arrays. Analysis of the microarray data using the previously described MADS tool resulted in the identification of over a hundred candidate ESRP-regulated splicing events. We were able to independently validate 37 of these targets by RT-PCR. The ESRP-regulated events encompass all known types of alternative splicing events. Importantly, a number of these regulated splicing events occur in gene transcripts that encode proteins with well-described roles in the regulation of actin cytoskeleton organization, cell-cell adhesion, cell polarity, and cell migration. In sum, this work reveals a novel list of transcripts differentially spliced in epithelial and mesenchymal cells, implying that coordinated alternative splicing plays a critical role in determination of cell type identity. Keywords: control / knockdown comparison
2009-08-15 | GSE17468 | GEO
Project description:Alternative splicing contributes to gene expression dynamics in many tissues, yet its role in auditory development remains unclear. We performed whole exome sequencing in individuals with sensorineural hearing loss (SNHL) and identified pathogenic mutations in Epithelial Splicing Regulatory Protein 1 (ESRP1). Patient derived induced pluripotent stem cells showed alternative splicing defects consistent with impaired ESRP1 function. To determine how mutations in ESRP1 cause hearing loss we evaluated Esrp1-/- mouse embryos and uncovered alterations in cochlear morphogenesis, auditory hair cell differentiation and cell fate specification. Transcriptome analysis revealed impaired expression and splicing of genes with essential roles in inner ear development and auditory function. In particular, aberrant splicing of Fgfr2 blocked stria vascularis formation due to erroneous ligand usage, which was corrected by reducing Fgf9 gene dosage. These findings implicate mutations in ESRP1 as a novel cause of SNHL and demonstrate the complex interplay between alternative splicing, inner ear development, and auditory function.
2017-09-01 | GSE90821 | GEO
Project description:Kidney development depends critically on proper ureteric bud branching giving rise to the entire collecting duct system. The transcription factor HNF1B is required for the early steps of ureteric bud branching. Yet, the molecular and cellular events regulated by HNF1B are poorly understood. We report that specific removal of Hnf1b from the ureteric bud leads to defective cell-cell contacts and apico-basal polarity during the early branching events. High resolution ex vivo imaging combined with a membranous fluorescent reporter strategy show decreased mutant cell-rearrangements during mitosis-associated cell dispersal and severe epithelial disorganisation. Molecular analysis reveals downregulation of Gdnf-Ret pathway components and suggests that HNF1B acts both upstream and downstream of Ret-signaling by directly regulating Gfrα1 and Etv5. Subsequently, Hnf1b-deletion leads to massively mispatterned ureteric tree network, defective collecting duct differentiation and disrupted tissue architecture leading to cystogenesis. Consistently, mRNA-seq analysis performed from E15.5 control and mutant kidneys shows that the most impacted genes encode intrinsic cell-membrane components with transporter activity. Our study uncovers a fundamental and recurrring role of HNF1B in epithelial organization during early ureteric bud branching and further patterning and differentiation of the collecting duct system. Article submitted to Development. Authors: Audrey Desgrange, Claire Heliot,Ilya Skovorodkin, Saad U. Akram, Janne Heikkilä, Veli-Pekka Ronkainen, Ilkka Miinalainen I., Seppo J. Vainio and Silvia Cereghini
2017-12-15 | GSE106085 | GEO
Project description:During development, distinct progenitors contribute to the nephrons versus the ureteric epithelium of the kidney. Indeed, previous pluripotent stem cell-derived models of kidney tissue either contain nephrons or pattern specifically to the ureteric epithelium. By reanalysing the transcriptional distinction between distal nephron and ureteric epithelium in human fetal kidney, we show here that while existing nephron-containing kidney organoids contain distal nephron epithelium and no ureteric epithelium, this distal nephron segment alone displays significant in vitro plasticity and can adopt a ureteric epithelial tip identity when isolated and cultured in defined conditions. “Induced” ureteric epithelium cultures can be cryopreserved, serially passaged without loss of identity and transitioned towards a collecting duct fate. Indeed, cultures harbouring loss-of-function mutations in PKHD1 recapitulate the cystic phenotype associated with autosomal recessive polycystic kidney disease.
2021-01-15 | GSE161255 | GEO