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Eukaryot Cell. 2014 Sep;13(9):1191-9. doi: 10.1128/EC.00104-14. Epub 2014 Jul 7.

Quality control of plasma membrane proteins by Saccharomyces cerevisiae Nedd4-like ubiquitin ligase Rsp5p under environmental stress conditions.

Author information

1
Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara, Japan.
2
Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara, Japan hiro@bs.naist.jp.

Abstract

In Saccharomyces cerevisiae, when a rich nitrogen source such as ammonium is added to the culture medium, the general amino acid permease Gap1p is ubiquitinated by the yeast Nedd4-like ubiquitin ligase Rsp5p, followed by its endocytosis to the vacuole. The arrestin-like Bul1/2p adaptors for Rsp5p specifically mediate this process. In this study, to investigate the downregulation of Gap1p in response to environmental stresses, we determined the intracellular trafficking of Gap1p under various stress conditions. An increase in the extracellular ethanol concentration induced ubiquitination and trafficking of Gap1p from the plasma membrane to the vacuole in wild-type cells, whereas Gap1p remained stable on the plasma membrane under the same conditions in rsp5(A401E) and Δend3 cells. A (14)C-labeled citrulline uptake assay using a nonubiquitinated form of Gap1p (Gap1p(K9R/K16R)) revealed that ethanol stress caused a dramatic decrease of Gap1p activity. These results suggest that Gap1p is inactivated and ubiquitinated by Rsp5p for endocytosis when S. cerevisiae cells are exposed to a high concentration of ethanol. It is noteworthy that this endocytosis occurs in a Bul1/2p-independent manner, whereas ammonium-triggered downregulation of Gap1p was almost completely inhibited in Δbul1/2 cells. We also found that other environmental stresses, such as high temperature, H₂O₂, and LiCl, also promoted endocytosis of Gap1p. Similar intracellular trafficking caused by ethanol occurred in other plasma membrane proteins (Agp1p, Tat2p, and Gnp1p). Our findings suggest that stress-induced quality control is a common process requiring Rsp5p for plasma membrane proteins in yeast.

PMID:
25001409
PMCID:
PMC4187619
DOI:
10.1128/EC.00104-14
[Indexed for MEDLINE]
Free PMC Article

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