This Article Full Text Full Text (PDF) All Versions of this Article: 1541-7786.MCR-06-0057v1 4/9/683    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Mustafi, R. Articles by Bissonnette, M. Search for Related Content PubMed PubMed Citation Articles by Mustafi, R. Articles by Bissonnette, M.

---

Signaling and Regulation

Protein Kinase- Inhibits Collagen I–Dependent and Anchorage-Independent Growth and Enhances Apoptosis of Human Caco-2 Cells

Departments of Medicine and Surgery, University of Chicago, Chicago, Illinois

Requests for reprints: Marc Bissonnette, Department of Medicine, The University of Chicago Hospitals and Clinics, MC 4076, 5841 South Maryland Avenue, Chicago, IL 60637. Phone: 773-702-8597; Fax: 773-702-2182. E-mail: mbissonn{at}medicine.bsd.uchicago.edu

Colonic carcinogenesis is accompanied by abnormalities in multiple signal transduction components, including alterations in protein kinase C (PKC). The expression level of PKC-, an atypical PKC isoform, increases from the crypt base to the luminal surface and parallels crypt cell differentiation in normal colon. In prior studies in the azoxymethane model of colon cancer, we showed that PKC- was down-regulated in rat colonic tumors. In this study, we showed that PKC- is expressed predominantly in colonic epithelial and not stromal cells, and loss of PKC- occurs as early as the adenoma stage in human colonic carcinogenesis. To assess the regulation of growth and differentiation by PKC-, we altered this isoform in human Caco-2 colon cancer cells using stable constitutive or inducible expression vectors, specific peptide inhibitors or small interfering RNA. In ecdysone-regulated transfectants grown on collagen I, ponasterone A significantly induced PKC- expression to 135% of empty vector cells, but did not alter nontargeted PKC isoforms. This up-regulation was accompanied by a 2-fold increase in basal and 4-fold increase in insulin-stimulated PKC- biochemical activity. Furthermore, PKC- up-regulation caused >50% inhibition of cell proliferation on collagen I (P < 0.05). Increased PKC- also significantly enhanced Caco-2 cell differentiation, nearly doubling alkaline phosphatase activity, while inducing a 3-fold increase in the rate of apoptosis (P < 0.05). In contrast, knockdown of this isoform by small interfering RNA or kinase inhibition by myristoylated pseudosubstrate significantly and dose-dependently increased Caco-2 cell growth on collagen I. In transformation assays, constitutively up-regulated wild-type PKC- significantly inhibited Caco-2 cell growth in soft agar, whereas a kinase-dead mutant caused a 3-fold increase in soft agar growth (P < 0.05). Taken together, these studies indicate that PKC- inhibits colon cancer cell growth and enhances differentiation and apoptosis, while inhibiting the transformed phenotype of these cells. The observed down-regulation of this growth-suppressing PKC isoform in colonic carcinogenesis would be predicted to contribute to tumorigenesis. (Mol Cancer Res 2006;4(9):683–94)