Effects of various mutations in the neurophysin/glycopeptide portion of the vasopressin gene on vasopressin expression in vitro

The vasopressin gene encodes three polypeptides besides the signal peptide: vasopressin, neurophysin II (neurophysin), and the carboxy-terminal glycop eptide (glycopeptide). Although the function of vasopressin is well charact erized, those of the latter two are not completely understood. In the prese nt study, we investigated the effects of various mutations within the neuro physin/glycopeptide portion of the vasopressin gene on vasopressin secretio n in vitro, to clarify the role of each peptide in vasopressin biosynthesis . Expression vectors containing the vasopressin gene, either wild-type or v arious mutants, mere transiently transfected into AtT20 cells, which are kn own to have the enzymes necessary for the proper processing of the vasopres sin precursor protein. The amount of vasopressin secreted into the culture medium was estimated by specific radioimmunoassay. Variable degrees of decr eased vasopressin secretion were observed with mutant vasopressin genes har boring deletions or amino acid substitutions in neurophysin. The naturally- occurring frame-shift mutation in the hereditary diabetes insipidus (Brattl eboro) rat completely eliminated vasopressin expression. In contrast, a mis sense mutation found in patients with familial neurogenic diabetes insipidu s only partially decreased vasopressin secretion. Finally, the mutant vasop ressin gene lacking the N-linked glycosylation site in glycopeptide had no effect on vasopressin expression. Our data suggest that 1) intact neurophys in is not indispensable for vasopressin expression, although an altered str ucture of neurophysin significantly affects the secretion of the hormone; 2 ) the pathogenesis of diabetes insipidus with the two naturally-occurring m utations found in the rat (Brattleboro rat) and human (familial central dia betes insipidus) seem to be different; and 3) glycosylation of the carboxy- terminal glycopeptide is not essential for the expression of vasopressin.