Vps20p and Vta1p interact with Vps4p and function in multivesicular body sorting and endosomal transport in Saccharomyces cerevisiae

• Published in: Journal of cell science Vol. 116; no. Pt 19; pp. 3957 - 3970
• Main Authors: Yeo, Sebastian C L, Xu, Linghui, Ren, Jihui, Boulton, Victoria J, Wagle, Mahendra D, Liu, Cong, Ren, Gang, Wong, Peisze, Zahn, Regina, Sasajala, Piriya, Yang, Hongyuan, Piper, Robert C, Munn, Alan L
• Format: Journal Article
• Language: English
• Published: England 01.10.2003
• Subjects: Adenosine Triphosphatases - metabolism; Adenosine Triphosphate - metabolism; Biological Transport - physiology; Carboxypeptidases - metabolism; Carrier Proteins - genetics; Carrier Proteins - metabolism; Cathepsin A; Cloning, Molecular; Endosomal Sorting Complexes Required for Transport; Endosomes - metabolism; Membrane Proteins - genetics; Membrane Proteins - metabolism; Mutation; Protein Binding; Protein Structure, Tertiary; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Transport Vesicles - metabolism; Two-Hybrid System Techniques; Vesicular Transport Proteins
• ISSN: 0021-9533; 1477-9137
• DOI: 10.1242/jcs.00751

Vps4p (End13p) is an AAA-family ATPase that functions in membrane transport through endosomes, sorting of soluble vacuolar proteins to the vacuole, and multivesicular body (MVB) sorting of membrane proteins to the vacuole lumen. In a yeast two-hybrid screen with Vps4p as bait we isolated VPS20 (YMR077c) and the novel open reading frame YLR181c, for which the name VTA1 has recently been assigned (Saccharomyces Genome Database). Vps4p directly binds Vps20p and Vta1p in vitro and binding is not dependent on ATP - conversely, Vps4p binding to Vps20p is partially sensitive to ATP hydrolysis. Both ATP binding [Vps4p-(K179A)] and ATP hydrolysis [Vps4p-(E233Q)] mutant proteins exhibit enhanced binding to Vps20p and Vta1p in vitro. The Vps4p-Vps20p interaction involves the coiled-coil domain of each protein, whereas the Vps4p-Vta1p interaction involves the (non-coiled-coil) C-terminus of each protein. Deletion of either VPS20 (vps20Delta) or VTA1 (vta1Delta) leads to similar class E Vps- phenotypes resembling those of vps4Delta, including carboxypeptidase Y (CPY) secretion, a block in ubiquitin-dependent MVB sorting, and a delay in both post-internalisation endocytic transport and biosynthetic transport to the vacuole. The vacuole resident membrane protein Sna3p (whose MVB sorting is ubiquitin-independent) does not appear to exit the class E compartment or reach the vacuole in cells lacking Vps20p, Vta1p or Vps4p, in contrast to other proteins whose delivery to the vacuole is only delayed. We propose that Vps20p and Vta1p regulate Vps4p function in vivo.