Activation of a TRP-like channel and intracellular calcium dynamics during phospholipase C-mediated cell death

Abstract

The model organism Neurospora crassa undergoes programmed cell death when exposed to staurosporine. Here we show that staurosporine causes defined changes in cytosolic free Ca2+ ([Ca2+]c) dynamics and a distinct Ca2+-signature that involves Ca2+ influx from the external medium and internal Ca2+ stores. We investigated the molecular basis of this Ca2+-response with [Ca2+]c measurements combined with pharmacological and genetic approaches. Phospholipase C was identified as a pivotal player during cell death since modulation of the phospholipase C signaling pathway and deletion of PLC-2, that we show to be involved in hyphal development, results in inability to trigger the characteristic staurosporine-induced Ca2+-signature. Using Δcch-1, Δfig-1 and Δyvc-1 mutants and a range of inhibitors, we show that extracellular Ca2+ entry does not occur through the hitherto described high- and low-affinity Ca2+ uptake systems, but through the opening of plasma membrane channels with properties resembling the transient receptor potential (TRP) family. Partial blockage of the response to staurosporine after inhibition of a putative inositol-1,4,5-trisphosphate (IP3) receptor suggests that Ca2+ release from internal stores following IP3 formation combines with the extracellular Ca2+ influx.