RNA-sequencing analysis of Intestinal stem cells from wild-type and circBtnl1-deficient mice
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE218136 | GEO | 2022/11/23
REPOSITORIES: GEO
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Project description:Background & Aims: Hierarchical organization of intestine relies on their stem cells by self-renew and producing committed progenitors. Although signals like Wnt are known to animate the continued renewal by maintaining intestinal stem cells (ISCs) activity, molecular mechanisms especially E3 ubiquitin ligases that modulate ISCs ‘stemness’ and supportive niche have not been well understood. Here, we investigated the role of Cullin 4B (Cul4b) in regulating ISC functions. Methods: We generated mice with intestinal epithelial-specific disruption of Cul4b (pVillin-cre; Cul4bfn/Y), inducible disruption of Cul4b (Lgr5-creERT2; Cul4bfn/Y, CAG-creERT2; Cul4bfn/Y) and their control (Cul4bfn/Y). Intestinal tissues were analyzed by histology, immunofluorescence, RNA sequencing and mass spectrum. Intestinal organoids deprived from mice with pVillin-Cre; Cul4bfn/Y, Lgr5-Cre; Cul4bfn/Y, Tg-Cul4b and their controls were used in assays to measure intestinal self-renewal, proliferation and differentiation. Wnt signaling and intestinal markers were analyzed by immunofluorescence and immunoblot assays. Differential proteins upon Cul4b ablation or Cul4b-interacting proteins were identified by mass spectrometry. Results: Cul4b specifically located at ISCs zone. Block of Cul4b impaired intestinal homeostasis maintenance by reduced self-renewal and proliferation. Transcriptome analysis revealed that Cul4b-null intestine lose ISC characterization and showed disturbed ISC niche. Mechanistically, reactivated Wnt pathway could recover intestinal dysfunction of Cul4b knockout mice. Analysis of differential total and ubiquitylated proteins uncovered the novel targeting substrate of Cullin-Ring ubiquitin ligase 4b (CRL4b), immunity-related GTPase family M member 1 (Irgm1) in intestine. Decreased Irgm1 rescued abnormally interferon signaling, overemphasized autophagy and downstream phosphate proteins in Cul4b knockout mice. Conclusion: We conclude that Cul4b is essential for ISC self-renewal and Paneth cell function by targeting Irgm1 and modulating Wnt signaling. Our results demonstrate that Cul4b is a novel ISC stemness and niche regulator.
2022-08-11 | PXD021528 | Pride
Project description:Intestinal stem cells (ISCs) residing in the crypts are critical for the continual self-renewal and rapid recovery of intestinal epithelium. The regulatory mechanism of ISCs is not fully understood. Here we report that CREPT, a recently identified tumor-promoting gene, is preferably expressed in the crypts, where the ISCs reside, but not in the villi. The Lgr5+ ISCs have much higher CREPT protein level than Lgr5- cells. To explore the function of CREPT in ISCs, we isolated WT and CREPT deleted Lgr5+ ISCs (Lgr5-CREPTKO) to perform Next generation sequencing.
2020-11-08 | GSE143604 | GEO
Project description:We previously demonstrated that SRCAP regulates the self-renewal of ESCs and modulates lymphoid lineage commitment. We further validated that SRCAP was mainly distributed in liver, spleen and intestine by Northern blot. SRCAP was also highly expressed in Lgr5+ ISCs. We then sought to explore the physiological role of SRCAP in the regulation of self-renewal maintenance of ISCs.We identified that Srcap deficiency impairs the self-renewal of ISCs and intestinal epithelial regeneration. Through SRCAP ChIP-sequencing, we sough to identify the key gene regulated by SRCAP in ISC self-renewal.
2020-01-28 | GSE144267 | GEO
Project description:Intestinal stem cells (ISCs) residing in the crypts are critical for the continual self-renewal and rapid recovery of intestinal epithelium. The regulatory mechanism of ISCs is not fully understood. Here we report that CREPT, a recently identified tumor-promoting gene, is preferably expressed in the crypts, where the ISCs reside, but not in the villi. The Lgr5+ ISCs have much higher CREPT protein level than Lgr5- cells. To explore the function of CREPTduring regeneration, we isolated irradiated WT and CREPT deleted intestines (Vil-CREPTKO) to perform Next generation sequencing.
2020-11-08 | GSE143605 | GEO
Project description:Purpose: circRNAs emerge as critical modulators in various biogical processes, but their roles in intestinal stem cells (ISC) reamin unclear. Here we want to identify a functional circRNA in ISCs, and firstly, we explore the expression profile of circRNA in ISCs. Methods: We isolated Lgr5+ ISCs and Lgr5- non-ISCs from the intestine tissue, followed by circRNA seq. Results: Many circRNAs are differently expressed in ISCs and non-ISCs, and we selected circRNAs highly expressed in ISCs for further investigation. Conclusions: circRNAs may play a critical role in ISC self-renewal.
2018-11-29 | GSE123051 | GEO
Project description:We previously demonstrated that SRCAP regulates the self-renewal of ESCs and modulates lymphoid lineage commitment. We further validated that SRCAP was mainly distributed in liver, spleen and intestine by Northern blot. SRCAP was also highly expressed in Lgr5+ ISCs. We then sought to explore the physiological role of SRCAP in the regulation of self-renewal maintenance of ISCs.We then generated Srcapflox/flox mice through insertion of loxP sequences flanking at the exon5 of Srcap gene locus. We established Srcapflox/flox;Lgr5GFP-CreERT2 mice through crossing Srcapflox/flox mice with Lgr5GFP-CreERT2 mice. With administration of tamoxifen (TAM), Srcap was completely deleted in Lgr5+ ISCs. We identified that Srcap deficiency impairs the self-renewal of ISCs and intestinal epithelial regeneration. Through RNA-sequencing, we sough to identify the key gene regulated by SRCAP in ISC self-renewal.
2020-01-15 | GSE143657 | GEO
Project description:Using EphB2 or the ISC marker Lgr5, we have FACS-purified and profiled intestinal stem cells (ISCs), crypt proliferative progenitors and late transient amplifying cells to define a gene expression program specific for normal ISCs. A frequent complication in colorectal cancer (CRC) is regeneration of the tumor after therapy. The intestinal stem cell signature predicts disease relapse in CRC and identifies a stem cell-like population that displays robust tumor- initiating capacity in immunodeficient mice as well as long-term self-renewal potential. We FACS purified mouse intestinal crypt cells according to their EphB2 or Lgr5 contents. We used Affymetrix chips to hybridize 2 samples from EphB2 high, 2 samples from EphB2 medium and 2 samples from EphB2 low cells (one sample from each group in a first hybridization on February 2009 plus an additional sample from each group on March 2009). Additionally, we hybridized one sample from Lgr5-EGFP high and one sample from Lgr5-EGFP low cells, obtained from Lgr5-EGFP knock-in mice (Barker et al., 2007).
2011-03-14 | E-GEOD-27605 | biostudies-arrayexpress
Project description:Perturbed intestinal epithelial homeostasis demonstrated as decreased Lgr5+ intestinal stem cells (Lgr5 ISCs) and increased secretory lineages were observed in our study where Lkb1 was specfically deleted in Lgr5 ISCs using Lgr5-EGFP-creERT2 (Tamoxifen) deletor. To gain mechanistic insight how Lkb1 maintains intestinal epithelial stem cell homeostasis, Lkb1 deficient ISCs (Lgr5-high cells) and progenitors (Lgr5-low cells) are isolated by flow cytometry and profiled by RNA sequencing to compare with controls (Lkb1 wild type ISCs and progenitors).
2020-02-29 | E-MTAB-6501 | biostudies-arrayexpress
Project description:The intestinal epithelium is continuously regenerated by highly proliferative Lgr5+ intestinal stem cells (ISCs). The existence of a population of quiescent ISCs has been suggested yet its identity and features remain controversial. Here we describe that the expression of the RNA-binding protein Mex3a labels a subpopulation of Lgr5+ cells that divide less frequently and contribute to regenerate all intestinal lineages with slow kinetics. Single cell transcriptomic analysis revealed two classes of Lgr5-high cells, one of them defined by the Mex3a-expression program and by low levels of proliferation genes. Lineage tracing experiments show that large fraction of Mex3a+ cell population is continuously recalled into the rapidly dividing self-renewing ISC pool in homeostatic conditions. Chemotherapy and radiation target preferentially rapidly dividing Lgr5+ cells but spare the Mex3a-high/Lgr5+ population, which helps sustain the renewal of the intestinal epithelium during treatment.
2017-03-14 | GSE90856 | GEO
Project description:Purpose: To examine the expression profile of WT and Stat6 KO intestinal stem cells. Methods: We isolated Lgr5+ ISCs from the intestine tissue, and generated Stat6 KO ISCs through CRISPR/Cas9 approach. Results: Many genes are changed upon Stat6 KO cells. We pay special attention on Wnt/β-catenin signaling that is the most critical pathway in ISCs, and discovered Stat6 drove the expression of Wnt target genes. Conclusions: Stat6 crosstalks with Wnt signaling to drive ISC self-renewal.
2018-12-05 | GSE123353 | GEO