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The natural protein kinase C alpha mutant is present in human thyroid neoplasms
C Prévostel1, V Alvaro, F de Boisvilliers
1INSERM U401, Montpellier, France.
Abstract:
An altered protein expression of Ca(2+)-dependent protein kinase C (PKC) isoforms and a point mutation in the PKC alpha cDNA (position 908 of the nucleotide sequence, position 294 of the amino acid sequence, substitution of an aspartic acid by a glycine) have been previously described in a subpopulation of human pituitary tumors. In this work, we screened 16 thyroid tissue samples (four follicular adenomas, five colloid adenomas, three papillary carcinomas, one follicular carcinoma and three normal tissues adjacent to the tumors) for the presence of the PKC alpha point mutation and for PKC alpha, beta 1, beta 2, epsilon and delta protein expression. Screening for the presence of the PKC alpha mutant was performed by a subcloning technic. The polymerase chain reaction products were generated using reverse-transcribed cDNAs, subcloned and sequenced (10 clones were routinely sequenced). The PKC alpha point mutation at position 908 of the cDNA sequence was found in four out of the nine adenomas and in the follicular carcinoma. It was neither detected in the papillary carcinomas nor in the adjacent normal tissues (one was the adjacent normal tissue of the follicular carcinoma; in this sample, genomic DNA and cDNA were used to look for the presence of the mutant), demonstrating the somatic nature of this mutant. Western blot analysis of PKC isoforms showed that the expression of all isoforms was higher in the thyroid neoplasms as compared with their adjacent normal tissue (n = 3). It was also higher in the samples containing the PKC mutant (two follicular adenomas, two colloid adenomas and the follicular carcinoma) as compared with the tumors where it was not detected (three papillary carcinomas and five adenomas). Samples could be ordered according to their increasing PKC expression as follows: normal adjacent tissue < follicular adenomas without PKC alpha mutant < or = papillary carcinoma < follicular adenomas with PKC mutant < follicular carcinoma with PKC mutant. In conclusion, the discovery of the PKC alpha mutant in thyroid neoplasms demonstrates that this mutant is not particular to human pituitary tumors where it was originally detected. It is a somatic mutation and its presence is concomitant with high levels of all of the PKC isoforms analysed. The presence of the PKC mutant in thyroid neoplasms raises the question of its importance in thyroid tumorigenesis.
Insights
A specific protein kinase C alpha (PKC) gene mutation, previously found in pituitary tumors, was also detected in thyroid neoplasms. This mutation correlates with increased expression of various PKC isoforms, suggesting a role in thyroid cancer development.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Altered protein expression of Ca(2+)-dependent protein kinase C (PKC) isoforms and a specific point mutation in PKC alpha cDNA have been previously identified in human pituitary tumors.
- The role and prevalence of this PKC alpha mutation and its associated protein expression in thyroid neoplasms remain largely unexplored.
Purpose of the Study:
- To investigate the presence of the PKC alpha point mutation in various thyroid tissue samples, including follicular adenomas, colloid adenomas, papillary carcinomas, follicular carcinoma, and adjacent normal tissues.
- To analyze the protein expression levels of PKC alpha, beta 1, beta 2, epsilon, and delta isoforms in these thyroid tissues.
- To determine the correlation between the PKC alpha mutation and the expression of PKC isoforms in thyroid neoplasms.
Main Methods:
- Screening for the PKC alpha point mutation involved subcloning polymerase chain reaction products from reverse-transcribed cDNAs, followed by sequencing.
- Protein expression of PKC isoforms was assessed using Western blot analysis.
- Thyroid tissue samples included follicular adenomas (n=4), colloid adenomas (n=5), papillary carcinomas (n=3), follicular carcinoma (n=1), and adjacent normal tissues (n=3).
Main Results:
- The PKC alpha point mutation was identified in four out of nine adenomas and in the follicular carcinoma, but not in papillary carcinomas or adjacent normal tissues, confirming its somatic nature.
- Western blot analysis revealed higher expression of all analyzed PKC isoforms in thyroid neoplasms compared to adjacent normal tissues.
- Tumors harboring the PKC alpha mutant exhibited significantly higher PKC isoform expression than those without the mutation, with a clear order of increasing PKC expression observed.
Conclusions:
- The PKC alpha mutant, initially discovered in pituitary tumors, is also present in thyroid neoplasms, indicating it is not tumor-specific.
- The presence of the PKC alpha mutant is associated with elevated levels of multiple PKC isoforms in thyroid tumors.
- This discovery raises important questions about the potential role of the PKC alpha mutant in the development and progression of thyroid cancer.