Detailed search for protein kinase(s) involved in plasma membrane H+−ATPase activity regulation of yeast cells

This study displays a screening using yeast strains deficient in protein kinases known to exist in Saccharomyces cerevisiae. From 95 viable single mutants, 20 mutants appear to be affected in the glucose-induced extracellular acidification. The mutants that are unaffected in calcium signaling were t...

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Published in:	FEMS yeast research Vol. 15; no. 2
Main Authors:	Pereira, Renata R., Castanheira, Diogo, Teixeira, Janaina A., Bouillet, Leoneide E. M., Ribeiro, Erica M. C., Trópia, Maria M. J., Alvarez, Florencia, Correa, Lygia F. M., Mota, Bruno E. F., Conceição, Luis Eduardo F. R., Castro, Ieso M., Brandão, Rogelio L.
Format:	Journal Article
Language:	English
Published:	England Oxford University Press 01-03-2015
Subjects:	Acidification Adenosine triphosphatase C-Terminus Calcium signalling Cell Membrane - enzymology Cell Membrane - metabolism Enzymes Glucose Glucose - metabolism Hydrogen Hygromycin Hygromycin B Kinases Mutants Mutation Phosphorylation Plasma membranes Protein kinase Protein Kinases - analysis Protein Kinases - genetics Proteins Proton-Translocating ATPases - metabolism Saccharomyces cerevisiae - enzymology Saccharomyces cerevisiae - genetics Saccharomyces cerevisiae - metabolism Yeast plasma membrane H yeast protein kinases ATPase glucose signaling plasma membrane H+−ATPase
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Summary:	This study displays a screening using yeast strains deficient in protein kinases known to exist in Saccharomyces cerevisiae. From 95 viable single mutants, 20 mutants appear to be affected in the glucose-induced extracellular acidification. The mutants that are unaffected in calcium signaling were tested for their sensitivity to hygromycin B. Furthermore, we verified whether the remaining mutants produced enzymes that are appropriately incorporated at plasma membrane. Finally, we measure the kinetic properties of the enzyme in purified plasma membranes from glucose-starved as well as glucose-fermenting cells. We confirmed the kinase Ptk2 involvement in H+−ATPase regulation (increase of affinity for ATP). However, the identification of the kinase(s) responsible for phosphorylation that leads to an increase in Vmax appears to be more complex. Complementary experiments were performed to check how those protein kinases could be related to the control of the plasma membrane H+−ATPase and/or the potential membrane. In summary, our results did not permit us to identify the protein kinase(s) involved in regulating the catalytic efficiency of the plasma membrane H+−ATPase. Therefore, our results indicate that the current regulatory model based on the phosphorylation of two different sites located in the C-terminus tail of the enzyme could be inappropriate. Activation of plasma membrane H+−ATPase in yeast cells probably involve different protein kinases.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	1567-1356 1567-1364
DOI:	10.1093/femsyr/fov003