Altered calcium handling following the recombinant overexpression of protein kinase C isoforms in HaCaT cells

Both changes in intracellular calcium concentration ([Ca 2+ ] i ) and activation of certain protein kinase C (PKC) isoforms play a crucial role in keratinocyte functions. To better understand the interaction between these two signalling pathways we investigated the resting [Ca 2+ ] i and the extracellular ATP-induced changes in [Ca 2+ ] i on HaCaT cell clones overexpressing either the classical α or the β PKC isoform. These PKC isoenzymes were previously shown to decrease (a) or increase (β) cell proliferation and augment (a) or suppress (β) cell differentiation. Keratinocyte clones with decreased proliferation rate were found to have unaltered resting [Ca 2+ [ i , but responded with greater calcium transients to the application of 180 μM of ATP. In contrast, clones with increased proliferation rate had elevated resting [Ca 2+ ] i and suppressed calcium responses to ATP. Calcium transients on PKCβ clones displayed a faster falling phase. Each clone had a distinct purinergic receptor expression pattern, some of which paralleled the altered proliferation rate and calcium handling. Keratinocytes overexpressing PKCβ revealed decreased P2X1 and increased P2Y1 receptor expression as compared with the control or PKCa clones. The expression level of P2X7 was significantly increased in keratinocytes overexpressing PKCa. On the other hand neither the P2X2 nor the P2Y2 expression was altered significantly in the cell types investigated. These data indicate that a modified proliferation and differentiation pattern is associated with altered calcium handling in keratinocytes. The observations also suggest that different PKC isoenzymes have different effects on the phosphatidyl-inositol signalling pathway.