Functional analysis of the human orthologue of the RSP5-encoded ubiquitin protein ligase, hNedd4, in yeast

• Published in: Current genetics Vol. 43; no. 1; pp. 1 - 10
• Main Authors: Gajewska, Beata, Shcherbik, Natalia, Oficjalska, Danuta, Haines, Dale S, Zoładek, Teresa
• Format: Journal Article
• Language: English
• Published: United States Springer-Verlag 01-04-2003; Springer Nature B.V
• Subjects: Antifungal Agents - pharmacology; Cycloheximide - pharmacology; Endosomal Sorting Complexes Required for Transport; Gene Transfer Techniques; High temperature; Hot Temperature; Humans; Hydrogen-Ion Concentration; Hypertension; Mutation; Nedd4 Ubiquitin Protein Ligases; Saccharomyces cerevisiae - drug effects; Saccharomyces cerevisiae - enzymology; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - physiology; Ubiquitin-Protein Ligase Complexes - genetics; Ubiquitin-Protein Ligase Complexes - physiology; Ubiquitin-Protein Ligases - genetics; Ubiquitin-Protein Ligases - physiology; Yeasts
• ISSN: 0172-8083; 1432-0983
• DOI: 10.1007/s00294-003-0371-x

hNedd4 and Rsp5p are orthologous ubiquitin ligases that contain a catalytic Hect domain, a C2 domain and multiple WW domains that mediate interactions with proteins. hNedd4 associates with the epithelial sodium channel and mutations disrupting this interaction lead to Liddle's syndrome, a heritable hypertension. Yeast Rsp5p ubiquitinates plasma membrane receptors and transporters and regulates their endocytosis. To determine whether the human enzyme has activity in yeast, hNEDD4 was expressed in yeast from the RSP5 or GAL1/10 promoters. Ectopic hNedd4 improved the growth and partially suppressed the endocytosis defect of rsp5 mutant cells, although it did not restore the viability of the rsp5-Δ strain. Wild-type cells harboring hNedd4 grew better at elevated temperature and on media containing cycloheximide. In contrast, hNedd4 WW domain mutants inhibited the growth of yeast when expressed at high levels. Our results show that hNedd4 affects cell growth, endocytosis and cycloheximide tolerance of yeast cells.