Electron Tomography Reveals Rab6 Is Essential to the Trafficking of trans-Golgi Clathrin and COPI-Coated Vesicles and the Maintenance of Golgi Cisternal Number

We have shown previously thatRab6, a small, trans-Golgi-localizedGTPase, acts upstream of the conserved oligomericGolgi complex (COG) andZW10/RINT1 retrograde tether complexes to maintainGolgi homeostasis. In this article, we present evidence from the unbiased and high-resolution approach of electron microscopy and electron tomography thatRab6 is essential to the trans-Golgi trafficking of two morphological classes of coated vesicles; the larger corresponds to clathrin-coated vesicles and the smaller to coat proteinI(COPI)-coated vesicles. On the basis of the site of coated vesicle accumulation, cisternal dilation and the normal kinetics of cargo transport from the endoplasmic reticulum (ER) toGolgi followed by delayedGolgi to cell surface transport, we suggest thatGolgi function in cargo transport is preferentially inhibited at the trans-Golgi/trans-Golgi network (TGN). The >50% increase inGolgi cisternae number inRab6-depletedHeLa cells that we observed may well be coupled to the trans-Golgi accumulation ofCOPI-coated vesicles; depletion of the individualRab6 effector, myosinIIA, produced an accumulation of uncoated vesicles with if anything a decrease in cisternal number. These results are the first evidence for aRab6-dependent protein machine affectingGolgi-proximal, coated vesicle accumulation and probably transport at the trans-Golgi and the first example of concomitant cisternal proliferation and increasedGolgi stack organization under inhibited transport conditions.