Evidence that the entire Golgi apparatus cycles in interphase HeLa cells: sensitivity of Golgi matrix proteins to an ER exit block

We tested whether the entire Golgi apparatus is a dynamic structure in inte rphase mammalian cells by assessing the response of 12 different Golgi regi on proteins to an endoplasmic reticulum (ER) exit block. The proteins chose n spanned the Golgi apparatus and included both Golgi glycosyltransferases and putative matrix proteins. Protein exit from ER was blocked either by mi croinjection of a GTP-restricted Sar1p mutant protein in the presence of a protein synthesis inhibitor, or by plasmid-encoded expression of the same d ominant negative Sar1p. All Golgi region proteins examined lost juxtanuclea r Golgi apparatus-like distribution as scored by conventional and confocal fluorescence microscopy in response to an ER exit block, albeit with a diff erential dependence on Sar1p concentration. Redistribution of Ga1NAcT2 was more sensitive to low Sar1p(dn) concentrations than giantin or GM130. Redis tribution was most rapid for p27, COPI, and p115. Giantin, GM130, and GaINA cT2 relocated with approximately equal kinetics. Distinct ER accumulation c ould be demonstrated for all integral membrane proteins. ER-accumulated Gol gi region proteins were functional. Photobleaching experiments indicated th at Golgi-to-ER protein cycling occurred in the absence of any ER exit block . We conclude that the entire Golgi apparatus is a dynamic structure and su ggest that most, if not all, Golgi region-integral membrane proteins cycle through ER in interphase cells.