Phospholipase C is required for glucose-induced calcium influx in budding yeast.
Intracellular calcium is a second messenger involved in several processes in yeast, such as mating, nutrient sensing, stress response and cell cycle events. It was reported that glucose addition stimulates a rapid increase in free calcium level in yeast. To investigate the calcium level variations induced by different stimuli we used a reporter system based on the photoprotein aequorin. Glucose addition (50 mM) to nutrient-starved cells induced an increase in free intracellular calcium concentration, mainly due to an influx from external medium. The increase of calcium reached its maximum 100-120 s after the stimulus. A concentration of about 20 mM glucose was required for a 50% increase in intracellular calcium. This response was completely abolished in strain plc1 Delta and in the isogenic wild-type strain treated with 3-nitrocoumarin, a phosphatidylinositol-specific phospholipase C inhibitor, suggesting that Plc1p is essential for glucose-induced calcium increase. This suggests that Plc1p should have a significant role in transducing glucose signal. The calcium influx induced by addition of high glucose on cells previously stimulated with low glucose levels was inhibited in strains with a deletion in the GPR1 or GPA2 genes, which suggests that glucose would be detected through the Gpr1p/Gpa2p receptor/G protein-coupled (GPCR) complex. Moreover, the signal was completely abolished in a strain unable to phosphorylate glucose, which is consistent with the reported requirement of glucose phosphorylation for GPCR complex activation.
Tisi
Renata
R
Dipartimento di Biotecnologie e Bioscienze, Università degli Studi di Milano-Bicocca, Piazza della Scienza 2, 20126 Milan, Italy.
Baldassa
Simona
S
Belotti
Fiorella
F
Martegani
Enzo
E
D000331:Aequorin / Q000235:genetics / Q000378:metabolism; D001059:Apoproteins / Q000235:genetics / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D003470:Culture Media / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005656:Fungal Proteins / Q000378:metabolism; D044385:GTP-Binding Protein alpha Subunits; D005947:Glucose / Q000494:pharmacology*; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D004718:Saccharomycetales / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D010738:Type C Phospholipases / Q000378:metabolism*; D014867:Water / Q000494:pharmacology
D001059:Apoproteins; D003470:Culture Media; D005656:Fungal Proteins; C109334:GPR1 protein, S cerevisiae; D044385:GTP-Binding Protein alpha Subunits; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C060014:apoaequorin; D014867:Water; D000331:Aequorin; D010738:Type C Phospholipases; C083907:Gpa2 protein, S cerevisiae; D020962:Heterotrimeric GTP-Binding Proteins; D005947:Glucose; D002118:Calcium