QUANTIFICATION OF FREE CYTOSOLIC Ca2+CONCENTRATION IN Saccharomyces cerevisiae

AGUILAR, PABLO S; PONCE DAWSON, SILVINA; VILLARRUEL, CECILIA

Parana

Congreso; LIV SAIB Annual Meeting; 2018

SAIB

Saccharomyces cerevisiae haploid cells secrete mating pheromones (a & α) that bind to receptors in the plasma membrane of cells of the oppositesex. The recognition between pheromone and receptor triggers a signaling cascade, which includes a transient raise in cytosolic Ca2+ levels. Inturn, this activates the calmodulin (CM), calcineurin (CN) and Crz1 transcription factor signaling pathway (Ca2+-CN-Crz1 pathway). Activationof this pathway is necessary for maintaining cell viability during prolonged periods of time in the presence of high pheromone concentrations.Currently it is accepted that an influx from extracellular calcium is induced in response to sexual pheromone, generating an elevation of cytosolicCa2+ for 90 minutes. However, a recently published work indicates that there is not one single and continuous calcium elevation but several shortduration discrete pulses (10-90 seconds). Bursts of Ca2+ are also observed in non pheromone treated cells, but in a sporadic way. These resultssuggest that the presence of pheromone results in an increase in the frequency of cytosolic Ca2+ pulses. Having a reliable estimate of cytosolicCa2+ concentration is key to achieve a better understanding of calcium signaling during sexual pheromone response. In this work, we performFluorescence Correlation Spectroscopy (FCS) experiments in the cytosol of S. cerevisiae haploid cells using GCaMP6f as the Ca2+dye toquantify the free concentration of this ion.