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HNF1A deficiency impairs β-cell fate, granule maturation and function (scRNA-seq of 309 hESC-derived cells: Differentiation day 25)

ABSTRACT: HNF1A deficiency impairs β-cell fate, granule maturation and function (scRNA-seq of 309 hESC-derived cells: Differentiation day 25)

PROVIDER: PRJNA532396 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR12813904.fastq.gz	Fastqsanger.gz
	SRR12813905.fastq.gz	Fastqsanger.gz
	SRR12813906.fastq.gz	Fastqsanger.gz
	SRR12813907.fastq.gz	Fastqsanger.gz
	SRR12813908.fastq.gz	Fastqsanger.gz

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HNF1A deficiency impairs β-cell fate, granule maturation and function (scRNA-seq of 309 hESC-derived cells: Differentiation day 25)

Project description:Mutations in HNF1A cause Maturity Onset Diabetes of the Young type 3, the second most frequent form of diabetes caused by single gene mutation. We generated human pancreatic stem cell-derived endocrine cells with mutations in HNF1A and show that HNF1A deficiency impairs scβ-cell fate, insulin granule maturation and the secretion of insulin in a glucose responsive manner. Single-cell RNA sequencing reveals that HNF1A orchestrates a network of genes involved in glucose metabolism, zinc transport, calcium ion binding and hormone exocytosis. Furthermore, in both patients and stem cell-derived β-cells, HNF1A deficiency altered the stoichiometry of secreted c-peptide to insulin. Sulfonylurea, used in the treatment of these patients, restored both insulin secretion and stoichiometry. Significantly, uncoupling of c-peptide and insulin secretion as described here questions the common practice in using c-peptide as a proxy to evaluate β-cell function. We also demonstrate that correction of the HNF1A locus restores function, providing a path to cell therapy.

2019-04-12 | GSE129653 | GEO

HNF1A deficiency impairs β-cell fate, granule maturation and function

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HNF1A deficiency impairs β-cell fate, granule maturation and function

Project description:HNF1A deficiency impairs β-cell fate, granule maturation and function

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Dm miR-309 cluster KO Expression Profiling

Project description:To investigate whether early zygotic miR-309 cluster miRNA expression might contribute to the down-regulation of maternal transcripts during the maternal to zygotic transition in Drosophila, we performed microarray analyses of control and miR-309 cluster mutant embryos at 0-1h and 2-3h of embryonic development. One-hour egg collections of w1118 and miR-309-6delta1 mutant flies at 25?C were either aged (2-3h sample) at 25?C or directly processed (0-1h sample). Six independent collections per genotype and time-point were performed. Using this approach, we found that maternal transcripts were stabilized in the miR-309 cluster mutant, indicating that the miR-309 cluster miRNAs are required for the down-regulation of these transcripts during the maternal to zygotic transition in Drosophila.

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An HNF1A truncation associated with maturity-onset diabetes of the young impairs pancreatic progenitor differentiation by antagonising HNF1B function

Project description:Purpose: The goal of this study is to conduct and compare NGS-derived transcriptome profiling (RNA-seq) of progenitor lines derived from 3 HNF1A-WT and 3 HNF1A-CRISPR (with p291fsinsC mutation) human induced pluripotent stem cell lines. Methods: mRNA profiles of WT/CRISPR pancreatic progenitor cells obtained after in-vitro differentiation for 14 days were generated by deep sequencing using Illumina HiSeq 2000 sequencer. The raw RNA-sequencing read files were processed using the research computing facility at King’s College London, Rosalind (https://rosalind.kcl.ac.uk). RNA-sequencing reads were trimmed from adapters using Trimmomatic/0.39 (Bolger et al., 2014) and quality control was done using FASTQC/0.11.8 (Andrews, 2010). Reads were mapped to a concatenated genome sequence of human GRCh38/hg38 using STAR/2.4.2 (Dobin and Gingeras, 2015). Transcript abundance was measured using featureCount in Subread package (Liao et al., 2013). Results: The DESeq2 package (Love et al., 2014) was used to identify differentially expressed genes. We found 919 significantly regulated genes (271 upregulated, 648 downregulated), with 16 HNF1B targets and 19 HNF1A specific targets within the significantly dysregulated genes when comparing HNF1A-CRISPR versus HNF1A-WT groups. Conclusions: HNF1A and HNF1B gene targets are disturbed by the p291fsinsC mutation engineered in the CRISPR-Cas9 lines.

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2022-05-01 | E-MTAB-11473 | biostudies-arrayexpress

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Project description:Detailed comparison of human pluripotent stem cells, to determine how hESC and iPSC differ in their protein expression profiles. The data was acquired in a TMT 10-plex SPS-MS3. 4 replicates of hESC compared to 4 replicates of HipSci human iPSCs.

2021-11-03 | PXD014502 | Pride

Expression profile of HNF1A knockdown and overexpression in 22RV1 and LNCaP cells respectively

Project description:To determine genes regulated by HNF1A in 22Rv1, we performed siRNA mediated knockdown of HNF1A using pooled siHNF1A (L-008215-00-0005 5 nmol) and pooled siSCR purchased from Dharmacon. RNA was harvested 3 days after transfection and gene expression profiling was performed. Similarly to determine genes regulated by HNF1A in LNCaP, we performed retroviral transduction of LNCaP cells in duplicate for HNF1A and empty vector control expression. cDNA for HNF1A in RC211201 vector (origene) was subcloned into a murine stem cell virus (MSCV)-based retroviral vector with hygromycin selection marker (Addgene). After 3 days of transduction, cells were selected for four days in hygromycin and later on RNA was harvested for gene expression profiling.

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Identification of HNF1A targets in WT hPSC-derived kidney organoids

Project description:To determine the transcriptional targets of HNF1A in human kidney cells, we performed chromatin immunoprecipitation sequencing on D28 kidney organoids generated from WT hPSCs using an established protocol by Morizane et al. (2015) doi: 10.1038/nbt.3392. ChIP-Seq analysis revealed that HNF1A preferentially bound onto the promoter regions of various genes that are predominantly expressed in renal proximal epithelial cells. qRT-PCR validation was performed on various human kidney cell models to confirm that HNF1A binds to promoter regions of selected target genes to regulate their gene expression.

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