Loss-of-function and dominant-negative mechanisms associated with hepatocyte nuclear factor-1 beta mutations in familial type 2 diabetes mellitus

Hepatocyte nuclear factor (HNF)-1 beta, a homeodomain-containing transcript ion factor, regulates gene expression in a dimerized form in pancreas, live r, and some other tissues. Recent genetic studies have identified two HNF-1 beta mutations, R177X and A263fsinsGG, in subjects with a monogenic form o f type 2 diabetes. Despite the defects being in the same gene, diverse seve rities of disease are observed in the affected subjects. To investigate the molecular mechanism by which mutations might cause various phenotypic feat ures, wild type and mutant proteins were transiently expressed in insulin-p roducing (MIN6) and hepatic (HepG2) cells. Luciferase reporter assay showed that both mutations resulted in a marked reduction of transactivation acti vity. Because their dimerization activity was found to be intact by the yea st two-hybrid system, it was possible that they were dominant-negative to w ild type activity. When co-expressed with wild type, both of the mutants si gnificantly decreased wild type activity in HepG2 cells. In contrast, altho ugh A263fsinsGG functioned similarly in MIN6 cells, R177X failed to affect wild type activity in this cell line. Immunohistochemical analysis of the m utants suggests that this functional divergence might be generated by the m odification of nuclear localization. These results suggest that HNF-1 beta mutations may impair pancreatic beta-cell function by loss-of-function and dominant-negative mechanisms.