Chromatin 3D interaction analysis of the<i>STARD10</i>locus unveils<i>FCHSD2</i>as a new regulator of insulin secretion

Hu, Ming; Cebola, Inês; Carrat, Gaëlle; Jiang, Shuying; Nawaz, Sameena; Khamis, Amna; Canouil, Mickaël; Froguel, Philippe; Schulte, Anke; Solimena, Michele; Ibberson, Mark; Marchetti, Piero; Cardenas-Diaz, Fabian L.; Gadue, Paul; Hastoy, Benoît; Alemeida-Souza, Leonardo; McMahon, Harvey T.; Rutter, Guy A.

doi:10.1101/2020.03.31.017707

Cited by 3 publications

(8 citation statements)

References 55 publications

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“…Altogether, these data support the potential regulation of TTLL6 by NEUROG3. Interestingly, the NEUROG3 binding site assigned to the ARAP1 gene coincided with an islet active enhancer (R11, Figure 8E) belonging to a regulatory region of the STARD10 and FCHSD2 genes, highly enriched in T2D/FG variants [27; 62]. The variable region (VR) within this locus was demonstrated to be required for insulin secretion [62].…”

Section: Resultsmentioning

confidence: 99%

Extensive NEUROG3 occupancy in the human pancreatic endocrine gene regulatory network

Schreiber

Mercier

Jiménez

et al. 2021

Preprint

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Objective: Mice lacking the bHLH transcription factor (TF) Neurog3 do not form pancreatic islet cells, including insulin secreting beta cells, causing diabetes. In human, homozygous mutations of NEUROG3 manifest with neonatal or childhood diabetes. Despite this critical role in islet cell development, the precise function and downstream genetic programs regulated directly by NEUROG3 remain elusive. We therefore mapped genome-wide NEUROG3 occupancy in human induced pluripotent stem cell (iPSC)-derived endocrine progenitors and determined NEUROG3 dependency of associated genes to uncover direct targets. Methods: We generated a novel hiPSC line (NEUROG3-HA-P2A-Venus), where NEUROG3 is HA-tagged and fused to a self-cleaving fluorescent VENUS reporter. We used the CUT&RUN technique to map NEUROG3 occupancy and epigenetic marks in pancreatic endocrine progenitors (PEP) differentiated from this hiPSC line. We integrated NEUROG3 occupancy data with chromatin status and gene expression in PEPs and their NEUROG3-dependence. In addition, we searched whether NEUROG3 binds type 2 diabetes mellitus (T2DM)-associated variants at the PEP stage. Results: CUT&RUN revealed a total of 863 NEUROG3 binding sites assigned to 1268 unique genes. NEUROG3 occupancy was found at promoters as well as at distant cis-regulatory elements frequently overlapping within PEP active enhancers. De novo motif analyses defined a NEUROG3 consensus binding motif and suggested potential co-regulation of NEUROG3 target genes by FOXA, RFX or PBX transcription factors. Moreover, we found that 22% of the genes downregulated in NEUROG3−/− hESC-derived PEPs are bound by NEUROG3 and thus likely to be directly regulated. NEUROG3 targets include transcription factors known to have important roles in islet cell development or function, such as NEUROD1, PAX4, NKX2-2, SOX4, MLXIPL, LMX1B, RFX3, and NEUROG3 itself. Remarkably, we uncovered that NEUROG3 binds transcriptional regulator genes with enriched expression in human fetal pancreatic alpha (e.g., IRX1, IRX2), beta (e.g., NKX6-1, SMAD9, ISX, TFCP2L1) and delta cells (ERBB4) suggesting that NEUROG3 could control islets subtype programs. Moreover, NEUROG3 targets genes critical for insulin secretion in beta cells (e.g., GCK, ABCC8/KCNJ11, CACNA1A, CHGA, SCG2, SLC30A8 and PCSK1). In addition, we unveiled a panel of ncRNA potentially regulated by NEUROG3. Lastly, we identified several T2DM risk SNPs within NEUROG3 peaks suggesting a possible developmental role of NEUROG3 in T2DM susceptibility. Conclusion: Mapping of NEUROG3 genome occupancy in PEPs uncovers an unexpectedly broad, direct control of the endocrine gene regulatory network (GRN) and raises novel hypotheses on how this master regulator controls islet and beta cell differentiation.

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Section: Resultsmentioning

confidence: 99%

Extensive NEUROG3 occupancy in the human pancreatic endocrine gene regulatory network

Schreiber

Mercier

Jiménez

et al. 2021

Preprint

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“…These data recapitulate the pattern of improved proinsulin processing observed at the human GWAS signal. STARD10 inactivation reduces GSIS both in mice ( 116 ) and in human EndoC-βH1 cells ( 117 ) and leads to profound changes in secretory granule structure in mouse beta cells ( 118 ). Solution of the 3D structure of STARD10 and direct binding assays revealed that STARD10 binds to and may transport inositol phospholipids, contributing to the failure of normal granule biogenesis in carriers of risk alleles (where STARD10 expression is lowered) ( 116 ).…”

Section: Variants Associated With Type 2 Diabetesmentioning

confidence: 99%

“…Nevertheless, as proof of concept, Ferrer’s group have demonstrated that alteration of variants containing enhancer activity by CRISPR interference affects the expression of multiple genes in EndoC-βH1 cells ( 48 ). Our own work, focusing on an enhancer cluster at the STARD10 locus, also showed that an enhancer cluster regulates not only STARD10 but also FCHSD2 through chromatin looping ( 117 ). Detailed analyses of how these and other variants affect chromatin structure, enhancer activity and gene expression are now warranted to elucidate the molecular mechanisms of disease pathways.…”

Section: Variants Associated With Type 2 Diabetesmentioning

confidence: 99%

“…Similarly, pancreatic duodenum homeobox-1 (PDX1), an important regulator of the INS gene, was also mutated by CRISPR-Cas9 resulting in a line with defective glucose-induced Ca 2+ influx and insulin secretion ( 244 ). Our laboratory has inactivated the type 2 diabetes-related STARD10 and FCHSD2 genes in EndoC-βH1 cells using a lentiviral approach and demonstrated effects on insulin secretion (and see above) ( 117 ). Furthermore, Fang et al used CRISPR screening technology and identified several genes involved in insulin regulation in mouse MIN6 cells ( 172 ).…”

Section: Application Of Genome Editing To Disease-relevant Genetic mentioning

confidence: 99%

“…Malkon’s laboratory ( 130 ) have deleted a variant-containing enhancer region within the ADCY5 gene in INS1 cells and demonstrated reduced ADCY5 gene expression. Also using CRISPR-Cas9, we deleted an active enhancer within an enhancer cluster in the STARD10 locus in EndoC-βH1 cells which reduced the expression of both the STARD10 and FCHSD2 genes ( 117 ). Moreover, CRISPR interference techniques (CRISPRi and CRISPRa) have been used to modulate the transcriptional activities of several enhancer regions in GWAS-identified genetic loci to demonstrate that altered enhancer activity impacts the expression of multiple genes within enhancer hub ( 48 ).…”

Section: Application Of Genome Editing To Disease-relevant Genetic mentioning

confidence: 99%

See 2 more Smart Citations

Functional Genomics in Pancreatic β Cells: Recent Advances in Gene Deletion and Genome Editing Technologies for Diabetes Research

Cherkaoui

Misra

et al. 2020

Front. Endocrinol.

Self Cite

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The inheritance of variants that lead to coding changes in, or the mis-expression of, genes critical to pancreatic beta cell function can lead to alterations in insulin secretion and increase the risk of both type 1 and type 2 diabetes. Recently developed clustered regularly interspaced short palindromic repeats (CRISPR/Cas9) gene editing tools provide a powerful means of understanding the impact of identified variants on cell function, growth, and survival and might ultimately provide a means, most likely after the transplantation of genetically “corrected” cells, of treating the disease. Here, we review some of the disease-associated genes and variants whose roles have been probed up to now. Next, we survey recent exciting developments in CRISPR/Cas9 technology and their possible exploitation for β cell functional genomics. Finally, we will provide a perspective as to how CRISPR/Cas9 technology may find clinical application in patients with diabetes.

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A genome-wide CRISPR screen identifies regulators of beta cell function involved in type 2 diabetes risk

Navarro-Guerrero

Bevacqua

et al. 2021

Preprint

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Identification of the genes and processes mediating genetic association signals for complex disease represents a major challenge. Since many of the genetic signals for type 2 diabetes exert their effects through pancreatic islet-cell dysfunction, we performed a genome-wide pooled CRISPR loss-of-function screen in human pancreatic beta cells. We focused on the regulation of insulin content as a disease-relevant readout of beta cell function. We identified 580 genes influencing this phenotype: integration with genetic and genomic data provided experimental support for 20 candidate type 2 diabetes effector transcripts including the autophagy receptor CALCOCO2. Our study highlights how cellular screens can augment existing multi-omic efforts to accelerate biological and translational inference at GWAS loci.

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Chromatin 3D interaction analysis of theSTARD10locus unveilsFCHSD2as a new regulator of insulin secretion

Cited by 3 publications

References 55 publications

Extensive NEUROG3 occupancy in the human pancreatic endocrine gene regulatory network

Extensive NEUROG3 occupancy in the human pancreatic endocrine gene regulatory network

Functional Genomics in Pancreatic β Cells: Recent Advances in Gene Deletion and Genome Editing Technologies for Diabetes Research

A genome-wide CRISPR screen identifies regulators of beta cell function involved in type 2 diabetes risk

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