Control of Bro1-domain protein Rim20 localization by external pH, ESCRT machinery, and the Saccharomyces cerevisiae Rim101 pathway.
Bro1-domain proteins such as yeast Bro1 and mammalian AIP1/Alix are well-established participants in endosome metabolism. The Bro1-domain interacts with endosomal surface protein Snf7/Vps32 in yeast, a subunit of the ESCRT complex. Yeast Bro1-domain protein Rim20 has no role in endosome function, but is required for alkaline pH-stimulated cleavage of transcription factor Rim101. Rim20-GFP is cytoplasmic under acidic conditions but concentrated in punctate foci under alkaline conditions. Bro1-GFP also accumulates in foci, but they are more numerous under acidic than alkaline conditions. Colocalization experiments indicate that some Rim20-GFP foci correspond to Bro1-RFP foci, whereas others do not. Rim8, Rim9, Rim21, Dfg16, Snf7, Vps20, Vps23, and Vps25, which are required for Rim101 cleavage, are required for appearance of Rim20-GFP foci. ESCRT complexes accumulate on endosome-derived compartments in cells that lack the AAA-ATPase Vps4. We find that Rim20-GFP foci accumulate in a vps4 mutant background independently of external pH, Rim101 pathway-specific genes, and most ESCRT subunit genes except for SNF7. Rim20-GFP foci seem to represent endosomes, because they colocalize with Snf7-RFP and because they correspond to a perivacuolar compartment in the vps4 strain. We propose that alkaline growth conditions alter the endosomal surface to favor Rim20-Snf7 interaction and Rim101 cleavage. Our findings raise the possibility that Bro1-domain proteins may be differentially regulated in the same cell, thereby coupling endosome metabolism to signaling.
Boysen
Jacob H
JH
Integrated Program in Cellular, Molecular, and Biophysical Studies, Columbia University, New York, NY 10032, USA.
Mitchell
Aaron P
AP
D027682:Cation Transport Proteins / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004705:Endocytosis / Q000502:physiology*; D056827:Endosomal Sorting Complexes Required for Transport; D006863:Hydrogen-Ion Concentration; D008954:Models, Biological; D009154:Mutation / Q000235:genetics; D009687:Nuclear Proteins / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism; D022161:Transport Vesicles / Q000378:metabolism; D014617:Vacuoles / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000737:chemistry*
C480237:Bro1 protein, S cerevisiae; D027682:Cation Transport Proteins; D004268:DNA-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; C492183:NHX1 protein, S cerevisiae; D009687:Nuclear Proteins; C489601:RIM101 protein, S cerevisiae; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C510979:Rim20 protein, S cerevisiae; C083890:SNF7 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D017923:Sodium-Hydrogen Exchangers; D033921:Vesicular Transport Proteins