Evidence for Metabolic Conversions of Rat Neurophysins Within Neurosecretory Granules of the Hypothalamo-Neurohypophysial System

Abstract

Neural lobes of rats contain 3 major neurophysins, known as I, II and III (RNP I, RNP II, RNP III). RNP I appears to be associated with vasopressin, and RNP II and III with oxytocin. Burford and Pickering suggested, on the basis of indirect evidence, that RNP III may be a breakdown product of RNP II. The present work provides direct support for this concept. When purified RNP II was incubated in vitro with extract of rat neural lobe, RNP II was converted to RNP III and then converted to another protein product, RNP III''. The time-dependent loss of RNP II and concurrent gain of RNP III, and then the appearance of RNP III'' possibly from the conversion of RNP III, could be demonstrated at an incubation pH of 4.5 (close to what is thought to be the pH within neurosecretory granules), but not at an incubation pH of 7.0. Proteins electrophoretically similar to RNP III and III'' can be demonstrated in the neural lobes of rats. The products of incubation do not appear to be experimental artifacts, and the further facts that the conversions occur at pH 4.5 rather than at pH 7.0, and that RNP III has been extracted from neurosecretory granules, strongly suggest that the in vitro findings reflect physiological conversions taking place within neurosecretory granules in vivo. Wholly analogous evidence was obtained for metabolic conversion of RNP I to RNP I'': conversion in vitro occurs at an incubation pH of 4.5 but not at 7.0; time-dependent loss of RNP I is accompanied by a concurrent and proportional gain of RNP I''; and a protein electrophoretically similar to RNP I'' can be demonstrated in neural lobes of rats. Considering the available data on the structure of rat neurophysins, it seems likely that RNP II is converted to RNP III by proteolytic cleavage on the carboxyl side of the Phe residue located near the C-terminal end of the molecule.