Genome-edited zebrafish model of ABCC8 loss-of-function disease

KATP channel gain- (GOF) and loss-of-function (LOF) mutations in SUR1 or Kir6.2 underlie human neonatal diabetes mellitus (NDM) and congenital hyperinsulinism (CHI), respectively. Genetically modified mice with transgenic overexpression of GOF mutations recapitulate many features of human NDM and transgenic mice expressing incomplete KATP LOF do reiterate mild hyperinsulinism, although complete KATP knockout mice do not exhibit persistent hyperinsulinism. We have shown that islet excitability and glucose homeostasis in zebrafish are regulated by identical KATP channels to those in mammals, and to explore the possibility of using zebrafish for modeling CHI, we have examined SUR1 truncation mutation (K499X) introduced into the abcc8 gene. Patch-clamp analysis confirmed complete absence of channel activity in {beta}-cells from K499X (SUR1-/-) fish. No difference in random blood glucose was detected in heterozygous SUR1+/- fish, nor in homozygous SUR1-/- fish, mimicking findings in SUR1 knockout mice. Mutant fish also demonstrated impaired glucose tolerance, similar to LOF mouse models. In paralleling features of mammalian hyperinsulinism resulting from loss-of-function mutations, this gene-edited animal provides a valid zebrafish model of KATP LOF driven-dependent pancreatic disease. Key PointsO_LILoss-of function in the Kir6.2 (KCNJ11) and SUR1 (ABCC8)-encoded pancreatic islet {beta}-cell KATP channels underlie congenital hyperinsulinism. Mouse models reiterate key features, but zebrafish models could provide a powerful model for further analysis and therapy testing. C_LIO_LIAn early nonsense mutation in exon 10 of SUR1 was generated by ENU mutagenesis. C_LIO_LIPatch-clamp analysis revealed an absence of {beta}-cells of SUR1 truncation mutants. C_LIO_LICa imaging demonstrated elevated basal [Ca]i in {beta}-cells with SUR1 truncation. C_LIO_LIHomozygous SUR1 truncation mutants had normal fasting glucose but impaired glucose tolerance as adults, mimicking findings in mouse SUR1 knockouts. C_LIO_LIIn paralleling features of mammalian diabetes and hyperinsulinism resulting from equivalent loss-of-function mutations, this gene-edited animal provides a valid zebrafish model of KATP -LOF dependent pancreatic diseases. C_LI