Protein kinase C ␦ (PKC ␦) is tyrosine-phosphorylated and catalytically inactive in mouse keratinocytes transformed by a ras oncogene. In several other model systems, Src kinases are upstream regulators of PKC ␦. To examine this relationship in epidermal carcinogenesis, v-ras transformed mouse keratinocytes were treated with a selective Src kinase inhibitor (PD 173958). PD 173958 decreased autophosphorylation of Src, Fyn, and Lyn kinases and prevented tyrosine phosphorylation of the Src kinase

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... ubstrate p120. PD 173958 also prevented PKC ␦ tyrosine phosphorylation and activated PKC ␦ as detected by membrane translocation. Expression of keratinocyte differentiation markers increased in PD 173958-treated v-ras-keratinocytes, and fluid-filled domes emerged, indicative of tight junction formation. Antisense PKC ␦ or bryostatin 1 inhibited dome formation, while overexpression of PKC ␦ in the presence of PD 173958 enhanced the formation of domes. Plasmids encoding phenylalanine mutants of PKC ␦ tyrosine residues 64 and 565 induced domes in the absence of PD 173958, while phenylalanine mutants of tyrosine residues 52, 155, and 187 were inactive. Thus, Src kinase mediated post-translational modification of PKC ␦ on specific tyrosine residues in ras-transformed mouse keratinocytes inactivates PKC ␦ and contributes to alterations in the differentiated phenotype and tight junction formation associated with neoplasia.