UC San Diego

Maturity-onset diabetes of the young type 1 (MODY1) is a form of monogenic diabetes caused by single base pair mutation in the hepatocyte nuclear factor-4-alpha (HNF4) gene that results in neonatal hyperinsulinemic hypoglycemia, macrosomia, and early onset diabetes. Although many HNF4 mutations have been identified in patients and their families, the molecular mechanism of MODY1 progression remains unknown. This study aims to understand the mechanism underlying HNF4 R141X mutations in pancreatic beta cells using human embryonic stem cell (hESC)-derived beta cells and genome editing CRISPR/Cas9 technology. By differentiating hESCs into insulin-producing beta cells using a stepwise protocol, we found that in vitro generated beta cells carrying the R141X mutation show phenotypes similar to clinical presentation of MODY1. At the immature stage, which is likely equivalent to the neonatal stage in vivo, mutant beta cells hypersecrete insulin at low glucose, which can be reduced by treatment of the potassium channel activator diaxozide. At the stage when wild-type (WT) beta cells become functionally mature and secrete insulin in response to glucose levels, HNF4 mutants hypersecrete insulin in both low and high glucose conditions. At both stages, mutant beta cells consistently secrete higher insulin in response to KCl-induced depolarization than WT. Furthermore, RNA sequencing of immature beta cells revealed upregulation of genes involved in cell cytotoxicity in R141X mutants. This is validated by an increase in cleaved Caspase-3 in mutants, suggesting that HNF4-mutated cells might be prone to cell death. Overall, our results show that hESC-generated R141X mutant beta cells behave similarly to beta cells from MODY1 patients and this mutation may induce apoptosis in beta cells.