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Vol. 20, Issue 21, 4563-4574, November 1, 2009

This Article Full Text Full Text (PDF) Supplemental Materials All Versions of this Article: E09-04-0272v1 20/21/4563    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Angers, C. G. Articles by Merz, A. J. PubMed PubMed Citation Articles by Angers, C. G. Articles by Merz, A. J.

HOPS Interacts with Apl5 at the Vacuole Membrane and Is Required for Consumption of AP-3 Transport Vesicles

Department of Biochemistry, University of Washington, Seattle, WA 98195-3750

Submitted April 6, 2009; Revised August 25, 2009; Accepted August 27, 2009
Monitoring Editor: Sandra Lemmon

Adaptor protein complexes (APs) are evolutionarily conserved heterotetramers that couple cargo selection to the formation of highly curved membranes during vesicle budding. In Saccharomyces cerevisiae, AP-3 mediates vesicle traffic from the late Golgi to the vacuolar lysosome. The HOPS subunit Vps41 is one of the few proteins reported to have a specific role in AP-3 traffic, yet its function remains undefined. We now show that although the AP-3 subunit, Apl5, binds Vps41 directly, this interaction occurs preferentially within the context of the HOPS docking complex. Fluorescence microscopy indicates that Vps41 and other HOPS subunits do not detectably colocalize with AP-3 at the late Golgi or on post-Golgi (Sec7-negative) vesicles. Vps41 and HOPS do, however, transiently colocalize with AP-3 vesicles when these vesicles dock at the vacuole membrane. In cells with mutations in HOPS subunits or the vacuole SNARE Vam3, AP-3 shifts from the cytosol to a membrane fraction. Fluorescence microscopy suggests that this fraction consists of post-Golgi AP-3 vesicles that have failed to dock or fuse at the vacuole membrane. We propose that AP-3 remains associated with budded vesicles, interacts with Vps41 and HOPS upon vesicle docking at the vacuole, and finally dissociates during docking or fusion.

Address correspondence to: Alexey J. Merz (merza{at}u.washington.edu).

Abbreviations used: ALP, alkaline phosphatase; AP, adaptor protein complex; CORVET, class C core vacuole/endosomal tethering; CPY, carboxypeptidase Y; GEF, guanine nucleotide exchange factor; HOPS, homotypic fusion and vacuole protein sorting; SNARE, soluble N-ethylmaleimide–sensitive factor attachment protein receptor; VPS-C, vacuolar protein sorting class C.