Decreased levels of protein kinase C enzyme activity and protein kinase C mRNA in primary colon tumors

Abstract

We have previously reported decreased protein kinase C (PKC) enzyme activity in primary human colorectal carcinomas. The purpose of this study was to extend these findings to a larger number of cases and to also examine the levels of expression of mRNAs that encode specific isoforms of PKC in these tumors. Colorectal carcinomas and paired grossly normal adjacent mucosal samples were collected from 39 patients. Complete histopathologic analyses were performed on all samples. PKC enzyme activity in both the cytosolic and particulate fractions was quantitated by measuring the amount of 32P incorporated into histone Type III-S. Northern blot nucleic acid hybridization was performed using polyA+ RNA extracted from both the tumor and normal tissue samples and 32P-labeled probes for specific isoforms of PKC. The paired sample t-test was used to determine the statistical significance of tumor to normal ratios of both enzyme activity and mRNA levels. The mean value for cellular PKC enzyme activity in the colon tumors from 39 patients was about 60 percent of that found in the paired adjacent grossly normal mucosa samples (P < 0.001). The subcellular distribution of PKC activity was similar in normal and tumor samples (about 70 percent in the particulate fraction). The abundance of PKC alpha mRNAs varied considerably among 28 tumor/normal pairs, with a mean tumor to normal (T:N) ratio of 1.0 +/- 0.6 for the 9.9-kb mRNA band and 1.4 +/- 0.7 for the 3.5-kb band. The abundance of PKC beta mRNAs was decreased in 30 of 39 tumors, with a mean T:N ratio of 0.6 +/- 0.4 for both the 9.4- and 3.5-kb bands for all 39 samples (P < 0.001). None of the parameters measured correlated with Dukes stage or the grade of the tumor. These studies extend previous evidence that total PKC enzyme activity is frequently decreased in primary human colon tumors. Our finding that this is often associated with decreased levels of PKC beta mRNA suggest that this is not simply due to post-translational down-regulation of this enzyme system. Further studies are required to determine whether these changes in PKC alpha and PKC beta mRNAs are due to altered de novo transcription or mRNA stability. It will also be of interest to examine the expression of other isoforms of PKC in colon tumors.