Persistence of Golgi matrix distribution exhibits the same dependence on Sar1p activity as a Golgi glycosyltransferase.

We investigated the relative distributional persistence of Golgi "matrix" proteins and glycosyltransferases to an endoplasmic reticulum exit block induced by expression of a GDP-restricted Sar1p. HeLa cells were microinjected with plasmid encoding the GDP-restricted mutant (T39N) of Sar1p to block endoplasmic reticulum exit and then scored for the distribution of GM130 (Golgi matrix protein of 130 kDa), a cis located golgin; p27, a member of the p24 family of proteins; giantin, a protein that interacts indirectly with GM130; and the Golgi glycosyltransferase, N-acetylgalactosaminyltransferase-2 (GalNAcT2). All of these proteins lost their compact, juxtanuclear distribution and displayed characteristics of endoplasmic reticulum/cytoplasmic accumulation with the same dependence on plasmid concentration. The kinetics of redistribution of GM130 and GalNAcT2 were identical. Expression of Sar1pT39N displaced the COPII coat protein Sec13p from endoplasmic reticulum exit sites consistent with disruption of these sites. This occurred without disturbing the overall distribution of endoplasmic reticulum membrane. Furthermore, the reassembly of a juxtanuclear Golgi matrix as assayed by the distribution of GM130 following washout of the Golgi disrupting drug, brefeldin A, was blocked by microinjected Sar1pT39N plasmids. We conclude that the persistence, i.e. stability and maintenance, of Golgi matrix distribution and its reassembly following drug disruption are exquisitely dependent on Sar1p activity.