?Study of ionic homeostasis regulation mechanisms: Physiological and transcriptomic analysis of the hal4hal5 mutant of Saccharomyces cerevisiae.

         The regulation of intracellular ion concentrations is a fundamental property of living cells. Many important physiological parameters like cellular volume, turgor, intracellular pH, ionic strength or internal cation concentrations depend on the regulation of the uptake and efflux systems for the main monovalent cations: protons, sodium and potassium. Potassium, the major cellular cation, is actively retained so that it is present at very high concentrations internally, and its concentrations are the principal determinant of the physiological parameters formerly commented on. The threshold for the toxicity of other monovalent cations such as sodium and lithium is much lower than that for potassium. Sodium and lithium ions must be prevented from accumulating in the cytosol to protect essential and sensitive enzymes. Eukaryotic cells employ primary active transport, mediated by P-type ATPases, and secondary transport,mediated by channels and cotransporters, to keep high potassium concentrations, essential for the cells, and low sodium concentrations, that could have toxic effects. In Saccharomyces cerevisiae the main potassium uptake system is encoded by the TRK1 and TRK2 genes.

         The present work focuses on the study of two proteins kinases, Hal4 and Hal5. The over-expression of the HAL4 and HAL5 genes improves the tolerance of the cells to toxic cations like sodium or lithium, whereas, its disruption produces hypersensitivity to these cations. It has been checked out that these effects depend on the Trk1-Trk2 potassium transport system, and everything seems to indicate that these two kinases act like activators of this system, increasing the entry of potassium in the cells, and therefore, making the membrane potential decrease. This decrease of the membrane potential also reduces the uptake of toxic cations, improving salt tolerance.

         Along this work, different phenotypes that of cells with the mutation of the HAL4 and HAL5 genes have been studied. We have done measurements of the internal levels of potassium, of the intracellular pH, of amino acids uptake, we have carried out a transcriptomic analysis of the hal4hal5 mutant, and we have tried to find out how the Hal4 and Hal5 kinases regulate the Trk1-Trk2 potassium transport system. We have also found a new relation between these two protein kinases and a set of different transporters of the plasma membrane, bringing to light that, probably, Hal4 and Hal5 exert other functions in the cells independent of the ones related with the potassium transport. Precisely, it has been observed that these two protein kinases are necessary for the stability at the plasma membrane of some proteins dedicated to ion and nutrient transport.

         In conclusion, a new mechanism of regulation of transporter proteins of the plasma membrane has been identified, and that it affects ionic homeostasis and the nutrient uptake from the media.