Abstract
The endosomal sorting complexes required for transport (ESCRTs) impact multiple cellular processes including multivesicular body sorting, abscission, and viral budding. The AAA-ATPase Vps4 is required for ESCRT function, and its full activity is dependent upon the co-factor Vta1. The Vta1 carboxyl-terminal Vta1 SBP1 Lip5 (VSL) domain stimulates Vps4 function by facilitating oligomerization of Vps4 into its active state. Here we report the identification of the Vps4 stimulatory element (VSE) within Vta1 that is required for additional stimulation of Vps4 activity in vitro and in vivo. VSE activity is autoinhibited in a manner dependent upon the unstructured linker region joining the amino-terminal microtubule interacting and trafficking domains and the carboxyl-terminal VSL domain. The VSE is also required for Vta1-mediated Vps4 stimulation by ESCRT-III subunits Vps60 and Did2. These results suggest that ESCRT-III binding to the Vta1 microtubule interacting and trafficking domains relieves linker region autoinhibition of the VSE to produce maximal activation of Vps4 during ESCRT function.
Highlights
Vta1 promotes Vps4 ATPase activity and facilitates endosomal sorting complexes required for transport (ESCRTs)-III stimulation of Vps4
Stimulation by the Vta1 SBP1 Lip5 (VSL) domain is less robust than the activity of full-length Vta1, suggesting that additional Vta1-Vps4 interactions are relevant for enhancement of Vps4 activity
We have demonstrated that yeast Vta1 stimulates Vps4 through an additional surface (VSE) proximal to the VSL domain
Summary
Vta promotes Vps ATPase activity and facilitates ESCRT-III stimulation of Vps. Results: The Vta VSE (Vps stimulatory element) mediates ESCRT-III-enhanced activation of Vps and contributes to Vta function in vivo. The VSE is required for Vta1-mediated Vps stimulation by ESCRT-III subunits Vps and Did2 These results suggest that ESCRT-III binding to the Vta microtubule interacting and trafficking domains relieves linker region autoinhibition of the VSE to produce maximal activation of Vps during ESCRT function. Vta promotes Vps function in vivo and can enhance Vps ATP hydrolysis and Vps disassembly of ESCRT-III in vitro [20, 32,33,34,35,36] Both Vps and Vta harbor amino-terminal microtubule interacting and trafficking (MIT) domains, which recognize elements within ESCRT-III subunits (MIT-interacting motifs, or MIMs) and facilitate recruitment of Vps and Vta to ESCRT-III [29, 37,38,39,40,41,42]. These observations suggest that the binding of ESCRT-III subunits Vps and Did to the Vta MIT domains induces conformational changes in the Vta linker region that promote VSE activation of Vps in addition to the stimulation afforded by Vta VSLVps association
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