Coalescence of microsomal vesicles from rat liver: a phenomenon occurring in parallel with enhancement of the glycosylation activity during incubation of stripped rough microsomes with GTP.

Abstract

Rough microsomes from rat liver were subjected to various treatments and incubated afterwards with UDP-N-acetyl-[14C]glucosamine and GDP-mannose in the presence of GTP (0.5 mM), or of other nucleotides. In agreement with earlier results, the preparations previously treated to strip off the ribosomes and incubated in the presence of GTP assembled dolichol-linked oligosaccharides and transferred these oligosaccharides to endogenous protein acceptors much more actively than untreated preparations, or stripped preparations incubated in the absence of GTP. Thin-section and freeze-fracture electron microscopy revealed that pyrophosphate-treated preparations incubated with GTP are aggregated and contain numerous vesicles as large as 1-4 .mu.m, or more. Such large vesicles were not present before incubation and thus were considered to have been formed through coalescence of regular-sized ones. Like glycosylation, the coalescence phenomenon depends on the removal of ribosomes, because it occurred whether ribosomes were stripped, at least partly, with pyrophosphate, KCl, or puromycin, but not when rough microsomes had been washed with 0.25 M sucrose or with KCl and MgCl2. Like glycosylation, it also depends on the addition of GTP and was not induced by ATP, UTP, CTP and nonhydrolyzable analogs of GTP. Rough microsomes coalesced when pyrophosphate-treated preparations were incubated with GTP in the absence of nucleotide sugars or in the presence of tunicamycin, indicating that the coalescence phenomenon does not result from the glycosylation of some membrane constituents.