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Protein kinase C isozyme expression and down-modulation in growing, quiescent, and transformed renal proximal tubule
L Dong1, J L Stevens, D Fabbro
1W. Alton Jones Cell Science Center, Inc., Lake Placid, New York 12946-1099.
Abstract:
Renal alpha-protein kinase C (PKC) is rapidly down-modulated modulated in animals treated with the renal toxin and tumor promoter, folic acid (Dong et al., Cancer Res., 53: 4542-4549, 1993). To further explore the role of PKC isozymes in renal growth and carcinogenesis, we compared phorbol ester receptor and PKC isozyme content, distribution, and regulation in primary and oncogene-altered rat renal proximal tubule epithelial cells (RPTE) in culture. Immunoblot analysis and RNase protection assays indicated that RPTE expressed at least four PKC isozymes, alpha, delta, epsilon, and zeta. Total phorbol ester receptors were decreased in primary proliferating, E1A-immortalized, and SV40-transformed RPTE compared to primary quiescent RPTE. The decrease in PDBu binding was largely due to a specific decrease in alpha-PKC protein content to approximately 50% of the level in quiescent RPTE. Degradation rates and message levels were compared to determine the mechanism for the decrease in alpha-PKC. Whereas alpha-PKC message levels in quiescent and proliferating primary RPTE were comparable, alpha-PKC degradation was increased in proliferating cells. These results indicate that the decreased alpha-PKC content was due largely to increased turnover. Phorbol ester stimulated the rate of degradation, thus demonstrating a link between degradation rate and PKC activation. These results suggest that the increased basal degradation rate in proliferating and oncogene-altered cells reflects an increase in activity of PKC in these cells.
Insights
Decreased alpha-protein kinase C (PKC) in kidney cells is linked to increased cell turnover and activation. This suggests a role for PKC in renal cell proliferation and cancer development.
Area of Science:
- Cell Biology
- Molecular Biology
- Oncology
Background:
- Renal alpha-protein kinase C (PKC) is downregulated by folic acid, a renal toxin and tumor promoter.
- PKC isozymes play a role in renal growth and carcinogenesis.
- Understanding PKC regulation in renal cells is crucial for cancer research.
Purpose of the Study:
- To investigate the role of PKC isozymes in renal cell proliferation and oncogenesis.
- To compare PKC isozyme content, distribution, and regulation in normal and altered rat renal proximal tubule epithelial cells (RPTE).
- To elucidate the mechanisms behind altered PKC levels in proliferating and transformed renal cells.
Main Methods:
- Primary and oncogene-altered rat renal proximal tubule epithelial cells (RPTE) were cultured.
- Immunoblot analysis and RNase protection assays were used to quantify PKC isozymes and receptors.
- Cellular degradation rates and message levels were assessed to determine PKC regulation.
Main Results:
- RPTE cells expressed PKC isozymes alpha, delta, epsilon, and zeta.
- Total phorbol ester receptors and alpha-PKC protein content were significantly decreased in proliferating and transformed RPTE compared to quiescent cells.
- Increased degradation, not altered message levels, accounted for the reduced alpha-PKC in proliferating cells.
- Phorbol ester stimulation increased alpha-PKC degradation, linking activation to turnover.
Conclusions:
- Decreased alpha-PKC content in proliferating and oncogene-altered renal cells is primarily due to increased protein degradation.
- The findings suggest that elevated basal degradation rates reflect increased PKC activity in these cells.
- This study highlights the importance of alpha-PKC regulation in renal cell growth and the potential link to carcinogenesis.