CALCINEURIN IS ESSENTIAL FOR DNA-SYNTHESIS IN SWISS 3T3 FIBROBLASTS

DNA synthesis was measured 16 h after stimulation of Swiss 3T3 fibrobl asts in the resting phase with various growth factors (platelet-derive d growth factor, fibroblast growth factor, lysophosphatidic acid and t hrombin). When extracellular Ca2+ was cheated by EGTA, or when the inf lux of Ca2+ from outside to inside the cell was blocked by cobalt, DNA synthesis was completely inhibited. As there was no effect whatsover on DNA synthesis when Ca2+ was chelated, or when the influx of Ca2+ wa s blocked up to the first 4h after growth stimulation, it was conclude d that, at an early stage, Ca2+ influx from outside to inside the cell is not related to the transition from the G(1) to the S phase. A Ca2/calmodulin-dependent protein kinase II inhibitor (KN-62) had no effec t on DNA synthesis. However, cyclosporin A and FK-506, which are inhib itors of Ca2+/calmodulin-dependent protein phosphatase 2B (calcineurin ), markedly inhibited DNA synthesis stimulated by all of the growth fa ctors. These results indicate that calcineurin plays a role, not only in activation of T-cells of the immune system in the initial phase, bu t also in DNA synthesis in fibroblasts. It was concluded that Ca2+ inf lux from outside to inside the cell during the mid-to-late G(1) phase, followed by calcineurin activation, is essential as a mechanism of gr owth signal transduction.