MODY1 mutation Q268X in hepatocyte nuclear factor 4alpha allows for dimerization in solution but causes abnormal...

Area of Science:

• Molecular endocrinology
• Genetics of diabetes

Background:

• Mutations in hepatocyte nuclear factor (HNF)-4alpha are linked to maturity-onset diabetes of the young, type 1 (MODY1).
• HNF-4, a nuclear receptor, has independent DNA-binding and ligand-binding domains.

Purpose of the Study:

• To investigate the functional properties of the HNF4.Q268X mutant, the first identified MODY1 mutation.
• To assess DNA binding, dimerization, and transactivation of the HNF4.Q268X mutant in vitro and in vivo.

Main Methods:

• Electrophoretic mobility shift assay (EMSA) for DNA binding.
• Co-immunoprecipitation assay for protein dimerization.
• Transient transfection assays for transactivation activity.

Main Results:

• HNF4.Q268X mutant protein failed to bind DNA, either alone or with wild-type HNF-4 (HNF4.wt).
• HNF4.Q268X demonstrated the ability to dimerize with HNF4.wt in solution.
• The mutant did not exhibit a dominant negative effect on HNF4.wt transactivation in vivo.
• Differential subcellular localization of HNF4.Q268X was observed, requiring SDS for extraction.

Conclusions:

• The MODY1 phenotype results from a loss of functional HNF-4 protein.
• The loss of function is exacerbated in tissues with low HNF-4 expression, such as the pancreas.