Systematic mutational analysis of the intracellular regions of yeast Gap1 permease.

<h4>Background</h4>The yeast general amino acid permease Gap1 is a convenient model for studying the intracellular trafficking of membrane proteins. Present at the plasma membrane when the nitrogen source is poor, it undergoes ubiquitin-dependent endocytosis and degradation upon addition...

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Autores principales:	Ahmad Merhi, Nicolas Gérard, Elsa Lauwers, Martine Prévost, Bruno André
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2011
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/d07e65b3c57f471c89683091c8cbe24b
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spelling	oai:doaj.org-article:d07e65b3c57f471c89683091c8cbe24b2021-11-18T06:55:34ZSystematic mutational analysis of the intracellular regions of yeast Gap1 permease.1932-620310.1371/journal.pone.0018457https://doaj.org/article/d07e65b3c57f471c89683091c8cbe24b2011-04-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/21526172/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203<h4>Background</h4>The yeast general amino acid permease Gap1 is a convenient model for studying the intracellular trafficking of membrane proteins. Present at the plasma membrane when the nitrogen source is poor, it undergoes ubiquitin-dependent endocytosis and degradation upon addition of a good nitrogen source, e.g., ammonium. It comprises 12 transmembrane domains (TM) flanked by cytosol-facing N- and C-terminal tails (NT, CT). The NT of Gap1 contains the acceptor lysines for ubiquitylation and its CT includes a sequence essential to exit from the endoplasmic reticulum (ER).<h4>Principal findings</h4>We used alanine-scanning mutagenesis to isolate 64 mutant Gap1 proteins altered in the NT, the CT, or one of the five TM-connecting intracellular loops (L2, -4, -6, -8 and -10). We found 17 mutations (in L2, L8, L10 and CT) impairing Gap1 exit from the ER. Of the 47 mutant proteins reaching the plasma membrane normally, two are unstable and rapidly down-regulated even when the nitrogen source is poor. Six others are totally inactive and another four, altered in a 16-amino-acid sequence in the NT, are resistant to ammonium-induced down-regulation. Finally, a mutation in L6 causes missorting of Gap1 from the secretory pathway to the vacuole. Interestingly, this direct vacuolar sorting seems to be independent of Gap1 ubiquitylation.<h4>Conclusions</h4>This study illustrates the importance of multiple intracellular regions of Gap1 in its secretion, transport activity, and down-regulation.Ahmad MerhiNicolas GérardElsa LauwersMartine PrévostBruno AndréPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 6, Iss 4, p e18457 (2011)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Ahmad Merhi Nicolas Gérard Elsa Lauwers Martine Prévost Bruno André Systematic mutational analysis of the intracellular regions of yeast Gap1 permease.
description	<h4>Background</h4>The yeast general amino acid permease Gap1 is a convenient model for studying the intracellular trafficking of membrane proteins. Present at the plasma membrane when the nitrogen source is poor, it undergoes ubiquitin-dependent endocytosis and degradation upon addition of a good nitrogen source, e.g., ammonium. It comprises 12 transmembrane domains (TM) flanked by cytosol-facing N- and C-terminal tails (NT, CT). The NT of Gap1 contains the acceptor lysines for ubiquitylation and its CT includes a sequence essential to exit from the endoplasmic reticulum (ER).<h4>Principal findings</h4>We used alanine-scanning mutagenesis to isolate 64 mutant Gap1 proteins altered in the NT, the CT, or one of the five TM-connecting intracellular loops (L2, -4, -6, -8 and -10). We found 17 mutations (in L2, L8, L10 and CT) impairing Gap1 exit from the ER. Of the 47 mutant proteins reaching the plasma membrane normally, two are unstable and rapidly down-regulated even when the nitrogen source is poor. Six others are totally inactive and another four, altered in a 16-amino-acid sequence in the NT, are resistant to ammonium-induced down-regulation. Finally, a mutation in L6 causes missorting of Gap1 from the secretory pathway to the vacuole. Interestingly, this direct vacuolar sorting seems to be independent of Gap1 ubiquitylation.<h4>Conclusions</h4>This study illustrates the importance of multiple intracellular regions of Gap1 in its secretion, transport activity, and down-regulation.
format	article
author	Ahmad Merhi Nicolas Gérard Elsa Lauwers Martine Prévost Bruno André
author_facet	Ahmad Merhi Nicolas Gérard Elsa Lauwers Martine Prévost Bruno André
author_sort	Ahmad Merhi
title	Systematic mutational analysis of the intracellular regions of yeast Gap1 permease.
title_short	Systematic mutational analysis of the intracellular regions of yeast Gap1 permease.
title_full	Systematic mutational analysis of the intracellular regions of yeast Gap1 permease.
title_fullStr	Systematic mutational analysis of the intracellular regions of yeast Gap1 permease.
title_full_unstemmed	Systematic mutational analysis of the intracellular regions of yeast Gap1 permease.
title_sort	systematic mutational analysis of the intracellular regions of yeast gap1 permease.
publisher	Public Library of Science (PLoS)
publishDate	2011
url	https://doaj.org/article/d07e65b3c57f471c89683091c8cbe24b
work_keys_str_mv	AT ahmadmerhi systematicmutationalanalysisoftheintracellularregionsofyeastgap1permease AT nicolasgerard systematicmutationalanalysisoftheintracellularregionsofyeastgap1permease AT elsalauwers systematicmutationalanalysisoftheintracellularregionsofyeastgap1permease AT martineprevost systematicmutationalanalysisoftheintracellularregionsofyeastgap1permease AT brunoandre systematicmutationalanalysisoftheintracellularregionsofyeastgap1permease
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Systematic mutational analysis of the intracellular regions of yeast Gap1 permease.

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