Nitrogen-regulated ubiquitination of the Gap1 permease of Saccharomyces cerevisiae.
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.
Springael
J Y
JY
Laboratoire de Physiologie Cellulaire et de Génétique des Levures, Université Libre de Bruxelles-Campus Plaine CP244, B-1050 Brussels, Belgium.
André
B
B
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
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