Related Experiment Videos
No evidence for oncogenic mutations in guanine nucleotide-binding proteins of human adrenocortical neoplasms
1Developmental Endocrinology Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892.
Abstract:
G-Proteins are membrane-bound heterotrimeric polypeptides that couple receptor signals to second messenger systems such as cAMP. Recently, point mutations at 2 codons of the highly preserved alpha-chain of Gs, the adenyl cyclase-stimulating G-protein, were found in GH-secreting pituitary tumors. These mutations resulted in constitutively activated Gs alpha and high intracellular cAMP levels. In addition, point mutations at similar codons of a different G-protein, G(i) alpha 2, were reported in adrenocortical neoplasms, suggesting a potential role of this isoform in the genesis of these tumors. We reevaluated the frequency of constitutively activating point mutations in the alpha-chain of the stimulatory (Gs alpha) and inhibitory (G(i) alpha 2) G-proteins in human adrenocortical tumors. Seven adrenocortical carcinomas, 2 human adrenocortical tumor cell lines, and 11 adrenocortical adenomas were studied. Genomic DNA was purified from either frozen tumor tissue or paraffin-embedded sections. Using specific primers and the polymerase chain reaction, DNA fragments surrounding codons 201 and 227 (Gs alpha) and 179 and 205 (G(i) alpha 2) were amplified and visualized on a 2% agarose gel. In a second asymmetric polymerase chain reaction, using nested primers, single stranded DNA was generated using 1-10 microL of the initial amplification mixture and directly sequenced using the dideoxy chain termination method of Sanger. We found no mutations at codons 201, 227 and 179, 205 of Gs alpha and G(i) alpha 2, respectively, in the tumors studied. We conclude that previously identified oncogenic point mutations in the stimulatory and inhibitory alpha-chain of G-proteins do not appear to be present at high frequency in adrenal neoplasms. Thus, the mechanism(s) of tumorigenesis in these tumors is different from that in GH-secreting adenomas and may involve oncogenic mutations of other cell constituents.
Insights
Constitutively activating G-protein mutations are common in pituitary tumors but not found in adrenal neoplasms. This suggests different genetic mechanisms drive adrenal tumor development compared to pituitary tumors.
Area of Science:
- Molecular Biology
- Oncology
- Endocrinology
Background:
- G-proteins are crucial signaling molecules linking receptors to cellular responses like cAMP production.
- Activating mutations in Gs alpha G-proteins drive GH-secreting pituitary tumors by increasing cAMP.
- Similar mutations in Gi alpha 2 G-proteins were implicated in adrenocortical neoplasms.
Purpose of the Study:
- To investigate the frequency of specific oncogenic G-protein alpha-chain mutations in human adrenal tumors.
- To determine if Gs alpha and Gi alpha 2 activating mutations contribute to adrenal neoplasm development.
Main Methods:
- Genomic DNA extraction from 7 adrenocortical carcinomas, 2 cell lines, and 11 adenomas.
- Polymerase chain reaction (PCR) amplification of Gs alpha (codons 201, 227) and Gi alpha 2 (codons 179, 205) regions.
- Direct DNA sequencing of amplified fragments using the Sanger method.
Main Results:
- No activating point mutations were detected in Gs alpha codons 201 or 227 across all studied adrenal tumors.
- No activating point mutations were detected in Gi alpha 2 codons 179 or 205 in the adrenal neoplasms.
- The frequency of these specific G-protein mutations in adrenal tumors is significantly lower than in pituitary tumors.
Conclusions:
- Oncogenic point mutations in the alpha-chains of Gs and Gi alpha 2 G-proteins are not frequent drivers of human adrenal neoplasms.
- The tumorigenesis mechanisms in adrenal neoplasms likely differ from those in GH-secreting pituitary adenomas.
- Further research is needed to identify other genetic alterations involved in adrenal tumor development.