Human disease phenotypes associated with haploinsufficient gene requirements are often not recapitulated well in animal models. Here, we have investigated the association between human GATA6 haploinsufficiency and a wide range of clinical phenotypes that include neonatal and adult-onset diabetes using CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9-mediated genome editing coupled with human pluripotent stem cell (hPSC) directed differentiation. We found that loss of one GATA6 allele specifically affects the differentiation of human pancreatic progenitors from the early PDX1+ stage to the more mature PDX1+NKX6.1+ stage, leading to impaired formation of glucose-responsive β-like cells. In addition to this GATA6 haploinsufficiency, we also identified dosage-sensitive requirements for GATA6 and GATA4 in the formation of both definitive endoderm and pancreatic progenitor cells. Our work expands the application of hPSCs from studying the impact of individual gene loci to investigation of multigenic human traits, and it establishes an approach for identifying genetic modifiers of human disease.
Bibliographical noteFunding Information:
We thank Drs. Licia Selleri and Wenjun Guo and members of the D.H. laboratory for insightful discussions and critical reading of the manuscript. The mouse anti-NKX6.1 and rat anti-C-Peptide antibodies were obtained from the Developmental Studies Hybridoma Bank, created by the NICHD of the NIH and maintained at The University of Iowa, Department of Biology, Iowa City, IA 52242, USA. This study was funded in part by the New York State Stem Cell Science Program (NYSTEM) (C029567 and C029156), the NIH/NIDDK (R01DK096239, DP3DK111907 and DP2DK098093-01), the Tri-Institutional Stem Cell Initiative (#2016-004), and a MSKCC Cancer Center Support grant (P30 CA008748). K.L. was supported by the Korean Government Scholarship Program for Study Overseas and Mogam Science Scholarship Foundation. N.V. was supported by the Howard Hughes Medical Institute (HHMI) Medical Research Fellowship. Z.Z. was supported by a NYSTEM postdoctoral fellowship (C026879) from the Center for Stem Cell Biology of the Sloan Kettering Institute.
© 2017 Elsevier Inc.
- CRISPR/Cas9 genome editing
- GATA6 and GATA4
- definitive endoderm
- genetic modifier
- human embryonic stem cells
- human pluripotent stem cells disease modeling
- insulin producing pancreatic beta cells
- pancreatic agenesis and neonatal diabetes
- pancreatic progenitor