Identification of three signaling molecules required for calcineurin-dependent monopolar growth induced by the DNA replication checkpoint in fission yeast

Cell polarity is coordinately regulated with the cell cycle. Growth polarity of the fission yeast Schizosaccharomyces pombe transits from monopolar to bipolar during G2 phase, termed NETO (new end take off). Upon perturbation of DNA replication, the checkpoint kinase Cds1/CHK2 induces NETO delay through activation of Ca2+/calmodulin-dependent protein phosphatase calcineurin (CN). CN in turn regulates its downstream targets including the microtubule (MT) plus-end tracking CLIP170 homologue Tip1 and the Casein kinase 1γ Cki3. However, whether and which Ca2+ signaling molecules are involved in the NETO delay remains elusive. Here we show that 3 genes (trp1322, vcx1 and SPAC6c3.06c encoding TRP channel, antiporter and P-type ATPase, respectively) play vital roles in the NETO delay. Upon perturbation of DNA replication, these 3 genes are required for not only the NETO delay but also for the maintenance of cell viability. Trp1322 and Vcx1 act downstream of Cds1 and upstream of CN for the NETO delay, whereas SPAC6c3.06c acts downstream of CN. Consistently, Trp1322 and Vcx1, but not SPAC6c3.06c, are essential for activation of CN. Interestingly, we have found that elevated extracellular Ca2+ per se induces a NETO delay, which depends on CN and its downstream target genes. These findings imply that Ca2+-CN signaling plays a central role in cell polarity control by checkpoint activation.