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| Status |
Public on Mar 29, 2024 |
| Title |
CRISPR Screening Uncovers a Long-Range Enhancer for ONECUT1 in Pancreatic Differentiation and Links a Diabetes Risk Variant [RNA-seq] |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
|
| Summary |
Functional enhancer annotation is a valuable first step for understanding tissue-specific transcriptional regulation and prioritizing disease-associated non-coding variants for investigation. However, unbiased enhancer discovery in physiologically relevant contexts remains a major challenge. To discover regulatory elements pertinent to diabetes, we conducted a CRISPR interference (CRISPRi) screen in the human pluripotent stem cell (hPSC) pancreatic differentiation system. Among the enhancers uncovered, we focused on a long-range enhancer ~664 kb from the ONECUT1 promoter, as coding mutations in ONECUT1 cause pancreatic hypoplasia and neonatal diabetes. Homozygous enhancer deletion in hESCs was associated with a near-complete loss of ONECUT1 gene expression and compromised pancreatic differentiation. We then identified a type 2 diabetes (T2D) associated variant (rs528350911) in the enhancer which disrupts a GATA motif. Introduction of the risk variant into hESCs revealed substantially reduced binding of key pancreatic transcription factors (GATA4, GATA6 and FOXA2) on the edited allele, accompanied by a subtle reduction of ONECUT1 transcription, supporting a causal role for this risk variant in metabolic disease. This work expands our knowledge about transcriptional regulation in pancreatic development through the characterization of a long-range enhancer and highlights the utility of enhancer discovery in disease-relevant settings for understanding monogenic and complex disease.
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| Overall design |
RNA-sequencing (RNA-seq) in WT and ONECUT1e-664kb deletion hPSC cells (heterozygous and homozygous) differentiated to the pancreatic lineage.
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| |
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| Contributor(s) |
Huangfu D, Kaplan SJ |
| Citation(s) |
39163202 |
| |
| Submission date |
Mar 12, 2024 |
| Last update date |
Aug 23, 2024 |
| Contact name |
Samuel Joseph Kaplan |
| Organization name |
Sloan Kettering Institute
|
| Department |
Developmental Biology Program
|
| Street address |
430 E 67th St
|
| City |
New York |
| State/province |
New York |
| ZIP/Postal code |
10065 |
| Country |
USA |
| |
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| Platforms (1) |
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
|
| Samples (18)
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| This SubSeries is part of SuperSeries: |
| GSE267330 |
CRISPR Screening Uncovers a Long-Range Enhancer for ONECUT1 in Pancreatic Differentiation and Links a Diabetes Risk Variant |
|
| Relations |
| BioProject |
PRJNA1086827 |
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