Bro1 coordinates deubiquitination in the multivesicular body pathway by recruiting Doa4 to endosomes.
Ubiquitination directs the sorting of cell surface receptors and other integral membrane proteins into the multivesicular body (MVB) pathway. Cargo proteins are subsequently deubiquitinated before their enclosure within MVB vesicles. In Saccharomyces cerevisiae, Bro1 functions at a late step of MVB sorting and is required for cargo protein deubiquitination. We show that the loss of Bro1 function is suppressed by the overexpression of DOA4, which encodes the ubiquitin thiolesterase required for the removal of ubiquitin from MVB cargoes. Overexpression of DOA4 restores cargo protein deubiquitination and sorting via the MVB pathway and reverses the abnormal endosomal morphology typical of bro1 mutant cells, resulting in the restoration of multivesicular endosomes. We further demonstrate that Doa4 interacts with Bro1 on endosomal membranes and that the recruitment of Doa4 to endosomes requires Bro1. Thus, our results point to a key role for Bro1 in coordinating the timing and location of deubiquitination by Doa4 in the MVB pathway.
Molecular, Cellular, and Developmental Biology, 347 UCB, University of Colorado, Boulder, CO 80309-0347, USA.
Natalie
N
Luhtala
Greg
G
Odorizzi
D002478:Cells, Cultured; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004705:Endocytosis / Q000235:genetics; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D056827:Endosomal Sorting Complexes Required for Transport; D011992:Endosomes / Q000378:metabolism* / Q000648:ultrastructure; D007425:Intracellular Membranes / Q000378:metabolism / Q000648:ultrastructure; D008854:Microscopy, Electron; D009097:Multienzyme Complexes / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D046988:Proteasome Endopeptidase Complex; D017434:Protein Structure, Tertiary / Q000235:genetics; D021381:Protein Transport / Q000235:genetics; D011956:Receptors, Cell Surface / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D022161:Transport Vesicles / Q000378:metabolism* / Q000648:ultrastructure; D025801:Ubiquitin / Q000378:metabolism*; D043222:Ubiquitin Thiolesterase; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism*
C480237:Bro1 protein, S cerevisiae; C084434:DOA4 protein, S cerevisiae; D056827:Endosomal Sorting Complexes Required for Transport; D009097:Multienzyme Complexes; D011956:Receptors, Cell Surface; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D033921:Vesicular Transport Proteins; D010450:Endopeptidases; D043222:Ubiquitin Thiolesterase; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex