5580

PRKCD

ALPS3|CVID9|MAY1|PKCD|nPKC-delta;protein kinase C, delta;PRKCD;protein kinase C, delta

protein kinase C delta VIII|protein kinase C delta type|tyrosine-protein kinase PRKCD

3p21.31

protein-coding

Apoptotic death of vascular smooth muscle cells (SMCs) is a prominent feature of blood vessel remodeling. In the present study, we examined the novel PKC isoform protein kinase C delta (PKCdelta) and its role in vascular SMC apoptosis. In A10 SMCs, overexpression of PKCdelta was sufficient to induce apoptosis, whereas inhibition of PKCdelta diminished H2O2-induced apoptosis. Moreover, evidence is provided that the tumor suppressor p53 is an essential mediator of PKCdelta-induced apoptosis in SMCs. Activation of PKCdelta led to accumulation as well as phosphorylation of p53 in SMCs; this induction correlated with apoptosis. Furthermore, blocking p53 induction with small interference RNA or targeted gene deletion prevented PKCdelta-induced apoptosis, whereas restoring p53 expression rescued the ability of PKCdelta to induce apoptosis in p53 null SMCs. We also establish that PKCdelta regulates p53 at both transcriptional and post-translational levels. Specifically, the transcriptional regulation required p38 MAPK, whereas the post-translational modification, at least for serine 46, did not involve MAPK. Additionally, PKCdelta, p38 MAPK, and p53 co-associate in cells under conditions favoring apoptosis. Together, our data suggest that SMC apoptosis proceeds through a pathway that involves PKCdelta, the intermediary p38 MAPK, and the downstream target tumor suppressor p53.

Protein kinase C (PKC)-delta is proapoptotic in human keratinocytes, and is downregulated or inactivated in keratinocytes expressing the activated Ha-ras oncogene, making it a candidate tumor suppressor gene for squamous cell carcinoma (SCC). We evaluated the significance of PKC-delta loss in transformed human keratinocytes using tumorigenic HaCaT Ras II-4 cells that have significantly reduced PKC-delta levels. Re-expression of PKC-delta by retrovirus transduction caused an increase in apoptosis and growth inhibition in culture. The growth inhibition induced by PKC-delta could be partially reversed by Bcl-x(L) expression, indicating that apoptosis was in part responsible for PKC-delta-induced growth inhibition. PKC-delta re-expression suppressed the tumorigenicity of HaCaT Ras II-4 cells in nude mice (P<0.05), and the small tumors that did form contained elevated levels of activated caspase-3, indicating increased apoptosis. In addition, we found that 29% (12/42) of human Bowens disease (squamous carcinoma in situ) or SCC cases had absent or reduced PKC-delta when compared to the surrounding normal epidermis. These results indicate that PKC-delta inhibits transformed keratinocyte growth by inducing apoptosis, and that PKC-delta may function as a tumor suppressor in human SCCs where its loss in cells harboring activated ras could provide a growth advantage by conferring resistance to apoptosis.

Transforming growth factor-beta1 (TGF-beta1) induces apoptosis in normal hepatocytes and hepatoma cells. PDCD4 is involved in TGF-beta1-induced apoptosis via the Smad pathway. The tumor promoter 12-O-tetradecanoylphorbor-13-acetate (TPA), a protein kinase C stimulator, inhibits TGF-beta1-induced apoptosis. However, the mechanisms of TPA action on PDCD4 expression remain to be elucidated. Therefore. the regulatory mechanism of PDCD4 expression by PKC was investigated. The treatment of the human hepatoma cell line, Huh7 with TPA suppressed PDCD4 protein expression and TGF-beta1 failed to increase the PDCD4 protein expression. PKC inhibitors Ro-31-8425 or bisindolylmaleimide-1-hydrocholoride (pan-PKC inhibitors) and rottlerin (PKCdelta inhibitor), but not Go6976 (PKCalpha inhibitor), enhanced the induction of PDCD4 protein by TGF-beta1. Furthermore, siRNA-mediated knockdown of PKCdelta and epsilon, but not PKCalpha, augmented the TGF-beta1-stimulated PDCD4 protein expression. However, TPA or pan-PKC inhibitor did not alter the PDCD4 mRNA expression either under basal- and TGF-beta1-treated conditions. The down-regulation of PDCD4 by TPA was restored by treatment with the proteasome inhibitor MG132. These data suggest that two isoforms of PKCs are involved in the regulation of the PDCD4 protein expression related to the proteasomal degradation pathway.

Oncogenic activation of K-ras occurs commonly in non-small cell lung cancer (NSCLC), but strategies to therapeutically target this pathway have been challenging to develop. Information about downstream effectors of K-ras remains incomplete, and tractable targets are yet to be defined. In this study, we investigated the role of protein kinase C delta (PKCdelta) in K-ras-dependent lung tumorigenesis by using a mouse carcinogen model and human NSCLC cells. The incidence of urethane-induced lung tumors was decreased by 69% in PKCdelta-deficient knockout (deltaKO) mice compared with wild-type (deltaWT) mice. deltaKO tumors are smaller and showed reduced proliferation. DNA sequencing indicated that all deltaWT tumors had activating mutations in KRAS, whereas only 69% of deltaKO tumors did, suggesting that PKCdelta acts as a tumor promoter downstream of oncogenic K-ras while acting as a tumor suppressor in other oncogenic contexts. Similar results were obtained in a panel of NSCLC cell lines with oncogenic K-ras but which differ in their dependence on K-ras for survival. RNA interference-mediated attenuation of PKCdelta inhibited anchorage-independent growth, invasion, migration, and tumorigenesis in K-ras-dependent cells. These effects were associated with suppression of mitogen-activated protein kinase pathway activation. In contrast, PKCdelta attenuation enhanced anchorage-independent growth, invasion, and migration in NSCLC cells that were either K-ras-independent or that had WT KRAS. Unexpectedly, our studies indicate that the function of PKCdelta in tumor cells depends on a specific oncogenic context, as loss of PKCdelta in NSCLC cells suppressed transformed growth only in cells dependent on oncogenic K-ras for proliferation and survival.

The introduction of the techniques of molecular biology as tools to study skin carcinogenesis has provided more precise localization of biochemical pathways that regulate the tumor phenotype. This approach has identified genetic changes that are characteristic of each of the specific stages of squamous cancer pathogenesis: initiation, exogenous promotion, premalignant progression, and malignant conversion. Initiation can result from mutations in a single gene, and the Harvey allele of the ras gene family has been identified as a frequent site for initiating mutations. Heterozygous activating mutations in c-rasHa are dominant, and affected keratinocytes hyperproliferate and are resistant to signals for terminal differentiation. An important pathway impacted by c-rasHa activation is the protein kinase C (PKC) pathway, a major regulator of keratinocyte differentiation. Increased activity of PKC alpha and suppression of PKC delta by tyrosine phosphorylation contribute to the phenotypic consequences of rasHa gene activation in keratinocytes. Tumor promoters disturb epidermal homeostasis and cause selective clonal expansion of initiated cells to produce multiple benign squamous papillomas. Resistance to differentiation and enhanced growth rate of initiated cells impart a growth advantage when the epidermis is exposed to promoters. The frequency of premalignant progression varies among papillomas, and subpopulations at high risk for progression have been identified. These high-risk papillomas overexpress the alpha 6 beta 4 integrin and are deficient in transforming growth factor beta 1 and beta 2 peptides, two changes associated with a very high proliferation rate in this subset of tumors. The introduction of an oncogenic rasHa gene into epidermal cells derived from transgenic mice with a null mutation in the TGF beta 1 gene have an accelerated rate of malignant progression when examined in vivo. Thus members of the TGF beta gene family contribute a tumor-suppressor function in carcinogenesis. Accelerated malignant progression is also found with v-rasHa transduced keratinocytes from skin of mice with a null mutation in the p53 gene. The similarities in risk for malignant conversion by initiated keratinocytes from TG beta 1 and p53 null geneotypes suggest that a common, growth-related pathway may underly the tumor-suppressive functions of these proteins in the skin carcinogenesis model.