Vacuolar-type H⁺-ATPase (V-ATPase) is a highly conserved proton pump responsible for acidification of intracellular organelles and potential drug target. It is a multisubunit complex comprising a cytoplasmic V₁ domain responsible for ATP hydrolysis and a membrane-embedded V_o domain that contributes to proton translocation across the membrane. Saccharomyces cerevisiae V-ATPase is composed of 14 subunits, deletion of any one of which results in well-defined growth defects. As the structure of V-ATPase and the function of each subunit have been well-characterized in yeast, this organism has been recognized as a preferred model for studies of V-ATPases. In this study, to assess the functional relatedness of the yeast and human V-ATPase subunits, we investigated whether human V-ATPase subunits can complement calcium- or pH-sensitive growth, acidification of the vacuolar lumen, assembly of the V-ATPase complex, and protein sorting in yeast mutants lacking the equivalent yeast genes. These assessments revealed that 9 of the 13 human V-ATPase subunits can partially or fully complement the function of the corresponding yeast subunits. Importantly, sequence similarity was not necessarily correlated with functional complementation. We also found that besides all V_o domain subunits, the V₁ F subunit is required for proper assembly of the V_o domain at the endoplasmic reticulum. Furthermore, the human H subunit fully restored the level of vacuolar acidification, but only partially rescued calcium-sensitive growth, suggesting a specific role of the H subunit in V-ATPase activity. These findings provide important insights into functional homologies between yeast and human V-ATPases.

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Journal Article | Research Support, Non-U.S. Gov't

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Abe M, Saito M, Tsukahara A, Shiokawa S, Ueno K, Shimamura H, Nagano M, Toshima JY, Toshima J

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