title: Nitrogen-regulated ubiquitination of the Gap1 permease of Saccharomyces cerevisiae.
abstract: Addition of ammonium ions to yeast cells growing on proline as the sole nitrogen source induces rapid inactivation and degradation of the general amino acid permease Gap1 through a process requiring the Npi1/Rsp5 ubiquitin (Ub) ligase. In this study, we show that NH4+ induces endocytosis of Gap1, which is then delivered into the vacuole where it is degraded. This down-regulation is accompanied by increased conversion of Gap1 to ubiquitinated forms. Ubiquitination and subsequent degradation of Gap1 are impaired in the npi1 strain. In this mutant, the amount of Npi1/Rsp5 Ub ligase is reduced >10-fold compared with wild-type cells. The C-terminal tail of Gap1 contains sequences, including a di-leucine motif, which are required for NH4+-induced internalization and degradation of the permease. We show here that mutant Gap1 permeases affected in these sequences still bind Ub. Furthermore, we provide evidence that only a small fraction of Gap1 is modified by Ub after addition of NH4+ to mutants defective in endocytosis.
authors.affiliation: Laboratoire de Physiologie Cellulaire et de Génétique des Levures, Université Libre de Bruxelles-Campus Plaine CP244, B-1050 Brussels, Belgium.
authors.forename: J Y
authors.initials: JY
authors.lastname: Springael
authors.forename: B
authors.initials: B
authors.lastname: André
mesh_terms: D000595:Amino Acid Sequence; D026905:Amino Acid Transport Systems; D000818:Animals; D004705:Endocytosis; D056827:Endosomal Sorting Complexes Required for Transport; D005656:Fungal Proteins / Q000378:metabolism; D008025:Ligases / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009584:Nitrogen / Q000378:metabolism*; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology; D011817:Rabbits; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D043743:Ubiquitin-Protein Ligase Complexes; D044767:Ubiquitin-Protein Ligases; D014452:Ubiquitins / Q000378:metabolism*; D014617:Vacuoles
chemical_list: D026905:Amino Acid Transport Systems; D056827:Endosomal Sorting Complexes Required for Transport; D005656:Fungal Proteins; D026901:Membrane Transport Proteins; D000644:Quaternary Ammonium Compounds; D029701:Saccharomyces cerevisiae Proteins; D014452:Ubiquitins; D043743:Ubiquitin-Protein Ligase Complexes; D044767:Ubiquitin-Protein Ligases; D008025:Ligases; C469287:RSP5 protein, S cerevisiae; D009584:Nitrogen