Amino acids regulate retrieval of the yeast general amino acid permease from the vacuolar targeting pathway.
Intracellular sorting of the general amino acid permease (Gap1p) in Saccharomyces cerevisiae depends on availability of amino acids such that at low amino acid concentrations Gap1p is sorted to the plasma membrane, whereas at high concentrations Gap1p is sorted to the vacuole. In a genome-wide screen for mutations that affect Gap1p sorting we identified deletions in a subset of components of the ESCRT (endosomal sorting complex required for transport) complex, which is required for formation of the multivesicular endosome (MVE). Gap1p-GFP is delivered to the vacuolar interior by the MVE pathway in wild-type cells, but when formation of the MVE is blocked by mutation, Gap1p-GFP efficiently cycles from this compartment to the plasma membrane, resulting in unusually high permease activity at the cell surface. Importantly, cycling of Gap1p-GFP to the plasma membrane is blocked by high amino acid concentrations, defining recycling from the endosome as a major step in Gap1p trafficking under physiological control. Mutations in LST4 and LST7 genes, previously identified for their role in Gap1p sorting, similarly block MVE to plasma membrane trafficking of Gap1p. However, mutations in other recycling complexes such as the retromer had no significant effect on the intracellular sorting of Gap1p, suggesting that Gap1p follows a genetically distinct pathway for recycling. We previously found that Gap1p sorting from the Golgi to the endosome requires ubiquitination of Gap1p by an Rsp5p ubiquitin ligase complex, but amino acid abundance does not appear to significantly alter the accumulation of polyubiquitinated Gap1p. Thus the role of ubiquitination appears to be a signal for delivery of Gap1p to the MVE, whereas amino acid abundance appears to control the cycling of Gap1p from the MVE to the plasma membrane.
Rubio-Texeira
Marta
M
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Kaiser
Chris A
CA
D026905:Amino Acid Transport Systems / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D000596:Amino Acids / Q000378:metabolism / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D009154:Mutation; D021381:Protein Transport / Q000187:drug effects; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D014617:Vacuoles / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000502:physiology*
D026905:Amino Acid Transport Systems; D000596:Amino Acids; C491145:GAP1 protein, S cerevisiae; C513355:LST4 protein, S cerevisiae; C513356:LST7 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins; D049452:Green Fluorescent Proteins