0 avis
Ubiquitin Is Required for Sorting to the Vacuole of the Yeast General Amino Acid Permease, Gap1
Archive ouverte
Edité par CCSD ; American Society for Biochemistry and Molecular Biology -
International audience. In yeast, ubiquitin plays a central role in proteolysis of a multitude of proteins and serves also as a signal for endocytosis of many plasma membrane proteins. We showed previously that ubiquitination of the general amino acid permease (Gap1) is essential to its endocyto-sis followed by vacuolar degradation. These processes occur when NH 4 , a preferential source of nitrogen, is added to cells growing on proline or urea, i.e. less favored nitrogen sources. In this study, we show that Gap1 is ubiquitinated on two lysine residues in the cytosolic N terminus (positions 9 and 16). A mutant Gap1 in which both lysines are mutated (Gap1 K9K16) remains fully stable at the plasma membrane after NH 4 addition. Furthermore , each of the two lysines harbors a poly-ubiquitin chain in which ubiquitin is linked to the lysine 63 of the preceding ubiquitin. The Gap1 K9 and Gap1 K16 mutants, in which a single lysine is mutated, are down-regulated in response to NH 4 although more slowly. In proline-grown cells lacking Npr1, a protein kinase involved in the control of Gap1 trafficking, newly synthesized Gap1 is sorted from the Golgi to the vacuole without passing through the plasma membrane (accompanying article, De Craene, J.-O., Soetens, O., and André , B. (2001) J. Biol. Chem. 276, 43939 – 43948). We show here that ubiquitination of Gap1 is also required for this direct sorting to the vacuole. In an npr1 mutant, neo-synthesized Gap1 K9K16 is rerouted to and accumulates at the plasma membrane. Finally, Bul1 and Bul2, two proteins interacting with Npi1/Rsp5, are essential to ubiq-uitination and down-regulation of cell-surface Gap1, as well as to sorting of neosynthesized Gap1 to the vacuole, as occurs in an npr1 mutant. Our results reveal a novel role of ubiquitin in the control of Gap1 trafficking, i.e. direct sorting from the late secretory pathway to the vacuole. This result reinforces the growing evidence that ubiquitin plays an important role not only in inter-nalization of plasma membrane proteins but also in their sorting in the endosomes and/or trans-Golgi.
Consulter en ligne
Suggestions
Du même auteur
