Hypotonic stress-induced calcium signaling in Saccharomyces cerevisiae involves TRP-like transporters on the endoplasmic reticulum membrane

• Hypotonic shock induces both calcium influx and calcium release from secretory compartments in budding yeast.
• Yal053w/Flc2 is required for calcium release from intracellular stores, upon hypotonic shock in YPD growing budding yeast cells.
• Flc2 is a member of a fungal specific branch of TRP-like calcium transporters spanning calcium-related spray protein from Neurospora crassa and Pkd2 from S. pombe with significant similarity to human polycystin 2 and mucolipin.

Saccharomyces cerevisiae cells respond to hypotonic stress (HTS) by a cytosolic calcium rise, either generated by an influx of calcium from extracellular medium, when calcium is available, or by a release from intracellular stores in scarcity of extracellular calcium. Calcium release from intracellular compartments is peculiarly inhibited by external calcium in a calcineurin-independent and Cch1-, but not Mid1-, driven manner. HTS-induced calcium release is also negatively regulated by the ER protein Cls2 and involves a poorly characterized protein, FLC2/YAL053W gene product, previously proposed to be required for FAD transport in the ER, albeit, due to its molecular features, it was also previously classified as an ion transporter. A computational analysis revealed that this gene and its three homologs in S. cerevisiae, together with previously identified Schizosaccharomyces pombe pkd2 and Neurospora crassa calcium-related spray protein, belong to a fungal branch of TRP-like ion transporters related to human mucolipin and polycystin 2 calcium transporters. Moreover, disruption of FLC2 gene confers severe sensitivity to Calcofluor white and hyper-activation of the cell wall integrity MAPK cascade, suggesting a role in cell wall maintenance as previously suggested for the fission yeast homolog. Perturbation in cytosolic resting calcium concentration and hyper-activation of calcineurin in exponentially growing cells suggest a role for this transporter in calcium homeostasis in yeast.

Dependence of the initial rate of hypotonic shock-induced (HTS) cytosolic calcium concentration increase upon extracellular free calcium concentration in the wild-type (left) or in the flc2Δ strain (right). Flc2 is a TRP-like channel localized in the ER and plasma membrane with close similarity to N. crassa spray protein and far similarity to mammalian mucolipin 2 and polycystic kidney disease/polycystin 2 calcium channels. Flc2 is required for HTS-induced ER calcium release and contributes to regulation of HTS-induced calcium influx from extracellular medium.Figure optionsDownload as PowerPoint slide