Reference: Ren J, et al. (2008) DOA1/UFD3 Plays a Role in Sorting Ubiquitinated Membrane Proteins into Multivesicular Bodies. J Biol Chem 283(31):21599-611

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Abstract

Ubiquitin (Ub) is a sorting signal that targets integral membrane proteins to the interior of the vacuole/lysosome by directing them into lumenl vesicles of multivesicular bodies (MVBs). The Vps27-Hse1 complex, which is homologous to the Hrs-STAM complex in mammalian cells, serves as an Ub-sorting receptor at the surface of early endosomes. We have found that Hse1 interacts with Doa1/Ufd3. Doa1 is known to interact with Cdc48/p97 and Ub, and is required for maintaining Ub levels. We find that the Hse1 SH3 domain binds directly to the central PFU domain of Doa1. Mutations in Doa1 that block Hse1 binding but not Ub binding do not alter Ub levels but do result in the mis-sorting of the MVB sorting of GFP-Cps1. Loss of Doa1 also causes a synthetic growth defect when combined with loss of Vps27. Unlike loss of Doa1 alone, the doa1 vps27 double mutant phenotype is not suppressed by Ub overexpression, demonstrating the effect is not due to indirect consequence of lowered Ub levels. Loss of Doa1 results in a defect in the accumulation of GFP-Ub within yeast vacuoles, implying that there is a reduction in the flux of ubiquitinated membrane proteins through the MVB pathway. This defect was also reflected by an inability to properly sort Vph1-GFP-Ub, a modified subunit of the multiprotein Vacuolar ATPase complex, which carries an in-frame fusion of Ub as an MVB sorting signal. These results reveal novel roles for Doa1 in helping process ubiquitinated membrane proteins for sorting into MVBs.

Reference Type
Journal Article
Authors
Ren J, Pashkova N, Winistorfer S, Piper RC
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