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Internal Amino Acids Promote Gap1 Permease Ubiquitylation via TORC1/Npr1/14-3-3-Dependent Control of the Bul Arrestin-Like Adaptors

Molecular Physiology of the Cell, Université Libre de Bruxelles, IBMM, Gosselies, Belgium.
Molecular and Cellular Biology (Impact Factor: 4.78). 09/2012; 32(22). DOI: 10.1128/MCB.00463-12
Source: PubMed

ABSTRACT

Ubiquitylation of many plasma membrane proteins promotes their endocytosis followed by degradation in the lysosome. The yeast general amino acid permease, Gap1, is ubiquitylated and down-regulated when a good nitrogen source like ammonium is provided to cells growing on a poor nitrogen source. This ubiquitylation requires the Rsp5 ubiquitin ligase and the redundant arrestin-like Bul1 and Bul2 adaptors. Previous studies have shown that Gap1 ubiquitylation involves the TORC1 kinase complex, which inhibits the Sit4 phosphatase. This causes inactivation of the protein kinase Npr1, which protects Gap1 against ubiquitylation. However, the mechanisms inducing Gap1 ubiquitylation after Npr1 inactivation remain unknown. We here show that on a poor nitrogen source, the Bul adaptors are phosphorylated in an Npr1-dependent manner and bound to 14-3-3 proteins that protect Gap1 against down-regulation. After ammonium is added and converted to amino acids, the Bul proteins are dephosphorylated, dissociate from the 14-3-3 proteins, and undergo ubiquitylation. Furthermore, dephosphorylation of Bul requires the Sit4 phosphatase, which is essential to Gap1 down-regulation. The data support the emerging concept that permease ubiquitylation results from activation of the arrestin-like adaptors of the Rsp5 ubiquitin ligase, this coinciding with their dephosphorylation, dissociation from the inhibitory 14-3-3 proteins, and ubiquitylation.

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Available from: Bruno André, Jan 29, 2014
    • "Several potential novel targets of Rsp5 were also identified. Bmh2, a 14-3-3 protein, which has previously been shown to regulate the Rsp5 adaptor Bul1 [74], was isolated, and Ino1 (inositol 3-phosphate synthase) and Pdi1 (a protein disulfide isomerase) were also previously isolated in large-scale proteomic analyses of Rsp5- associated proteins [75,76]. Sul2 (a sulfur permease), two plasma membrane-associated t-SNAREs (Sso1 and Sso2), and Rvs161, a lipid raft protein, were also isolated, consistent with the role of Rsp5 in membrane protein trafficking. "
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