A Novel c.125 T>G (p.Val42Gly) Mutation in The Human INS Gene Leads to Neonatal Diabetes Mellitus via a Decrease in Insulin Synthesis.

BACKGROUND

Neonatal diabetes mellitus is likely caused by monogenic mutations, several of which have been identified. mutations have a broad spectrum of clinical presentations, ranging from severe neonatal onset to mild adult onset, which suggests that the products of different mutant INS alleles behave differently and utilize distinct mechanisms to induce diabetes. In this study, a neonatal diabetes mellitus patient's gene was sequenced, and functional experiments were conducted.

METHODS

The neonatal diabetes mellitus patient's genomic DNA was extracted, and the patient's , and genes were sequenced. A novel mutation was identified in , and the open reading frame of this human mutant gene was inserted into the pMSCV-PIG plasmid. The constructed pMSCV-PIG plasmid was combined with VSV-g and Gag-pol and transfected into 293T cells to package the lentivirus. To stably overexpress the mutant gene, INS-1 cells were infected with the virus. The levels of insulin in the cell culture medium and cytoplasm were determined by ELISA and immunocytochemistry, respectively.

RESULTS

A heterozygous mutation, c.125T>G (p. Val42Gly), was identified in a neonatal diabetes mellitus patient's gene. The human mutant INS open reading frame was overexpressed in INS-1 cells, and the mutant insulin was undetectable in the cell culture medium and cytoplasm.

CONCLUSIONS

The novel heterozygous activating mutation c.125 T>G (p.Val42Gly) impairs the synthesis of insulin by pancreatic beta cells, resulting in diabetes.