Functional analyses of three different mutations in the AVP-NPII gene causing familial neurohypophyseal diabetes...

Familial neurohypophyseal diabetes insipidus (FNDI), a rare disorder, which is clinically characterized by polyuria and polydipsia, results from mutations in the arginine vasopressin-neurophysin II (AVP-NPII) gene. The aim of this study was to perform functional analyses of three different mutations (p.G45C, 207_209delGGC, and p.G88V) defined in the AVP-NPII gene of patients diagnosed with FNDI, which are not included in the literature.

For functional analysis studies, the relevant mutations were created using PCR-based site-directed mutagenesis and restriction fragment replacement strategy and expressed in Neuro2A cells. AVP secretion into the cell culture medium was determined by radioimmunoassay (RIA) analysis. Fluorescence imaging studies were conducted to determine the differences in the intracellular trafficking of wild-type (WT) and mutant AVP-NPII precursors. Molecular dynamics (MD) simulations were performed to determine the changing of the conformational properties of domains for both WT and 207-209delGGC mutant structures and dynamics behavior of residues.

Reduced levels of AVP in the supernatant culture medium of p.G45C and p.G88V transfected cells compared to 207_209delGGC and WT cells were found. Fluorescence imaging studies showed that a substantial portion of the mutant p.G45C and p.G88V AVP-NPII precursors appeared to be located in the endoplasmic reticulum (ER), whereas 207_209delGGC and WT AVP-NPII precursors were distributed throughout the cytoplasm.

The mutations p.G45C and p.G88V cause a failure in the intracellular trafficking of mutant AVP-NPII precursors. However, 207_209delGGC mutation does not result in impaired cellular trafficking, probably due to not having any significant effect in processes such as the proper folding, gain of three-dimensional structure, or processing. These results will provide valuable information for understanding the influence of mutations on the function of the AVP precursor hormone and cellular trafficking. Therefore, this study will contribute to elucidate the mechanisms of the molecular pathology of AVP-NPII mutations.