Ras oncogene mediated transformation of mouse keratinocytes to a benign squamous papilloma phenotype is associated with activation of PKCa. Tumors produced by ras transformation of keratinocytes derived from skin targeted PKCa transgenic mice produced in our laboratory grow faster and achieve a larger size, but do not convert to malignancy at a higher rate. Activation of a PKCa transgene in mouse epidermis induced a marked intraepidermal neutrophilic infiltrate and epidermal and follicular apoptosis that is blocked by inhibition of AP-1 activity. Multiple chemokines are upregulated and supernatants cultured from transgenic keratinocytes contain CCR1 mediated chemotactic properties. Microarray analysis has revealed a spectrum of PKCa regulated genes, including differentiation markers, chemokines/cytokines and proteases. The AP-1 and NFkB pathways interact in this model. This laboratory has shown that both PKCa and PKCd activate independent keratinocyte death pathways. Activation of PKC also inactivates the AKT kinase in mouse keratinocytes through prevention or reversal of AKT phosphorylation. Inhibition of PKC activity enhances IGF-1- mediated protection from UV induced apoptosis, presumably by enhancing AKT activity as detected by AKT phosphorylation. The specific PKC isoforms targeting AKT appear to be d and/or e. The genomic structures of C. elegans, mouse, rat and human PKCd were analyzed after sequencing a genomic fragment of mouse PKC. Overall exon-intron genomic structure was highly conserved among mammals while significantly diverged in C. elegans. A 1.7 kb region of the PKCd promoter revealed a hitherto unknown NFkB site, and TNFa was shown to upregulate the expression of the gene. A conditional floxed targeting vector has been constructed to specifically delete PKCd and is being sequence verified. These mice will be crossed with the K5 CRE mice to delete PKCd in the epidermis. Disruption of the nuclear tumor suppressor gene PML function in skin by targeting the PML-RARa fusion oncogenic transgene in transgenic mice cause spontaneous skin tumor induction. This is associated with cutaneous retinoid deficiency. When PML is targeted to the epidermis in transgenic mice, transgenic keratinocytes senesce more rapidly and produce smaller tumors when transformed by a ras oncogene.
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