Methylation and mutation of the inhibinα gene in human melanoma cells and regulation of PTEN expression and AKT/PI3K signaling by a demethylating agent.
Oncol Rep
; 47(2)2022 02.
Article
en En
| MEDLINE
| ID: mdl-34958114
ABSTRACT
Inhibin suppresses the pituitary secretion of folliclestimulating hormone and has been reported to act as a tumor suppressor gene in the gonad in mice. Epigenetic modifications, mutations, changes in the loss of heterozygosity (LOH) of the inhibinα gene and regulation of gene expression in response to a demethylating agent [5aza2'deoxycytidine (5AzadC)] in human melanoma cells were assessed. In addition, the association between a mutation in the 5'untranslated region (5'UTR) of the inhibinα subunit and the expression of phosphatidylinositol 3,4,5trisphosphatedependent Rac exchanger 2 (PREX2) and phosphatase and tensin homolog (PTEN) as well as AKT/PI3K signaling was determined. The methylation status of the CpG sites of the inhibinα promoter was analyzed by methylationspecific PCR in bisulfitetreated DNA. Cell viability was counted using the trypan blue assay, mRNA expression was examined via reverse transcriptionquantitative PCR, and protein expression was examined via western blot analysis. The inhibinα promoter was hypermethylated in G361, SKMEL3, SKMEL24 and SKMEL28 cells and moderately methylated in SKMEL5 cells. Inhibinα gene mutations were observed in the 5'UTR exon 1 of G361, SKMEL5, SKMEL24 and SKMEL28 cells as well as in exon 2 of SKMEL3 cells. Allelic imbalance, including LOH, in the inhibinα gene was detected in human melanoma cells. Treatment with 5AzadC increased inhibinα mRNA and protein levels, inhibited cell proliferation, and delayed the doubling times of surviving melanoma cells. In 5AzadCtreated cells, PREX2 protein expression was slightly increased in G361 and SKMEL24 cells and decreased in SKMEL3, SKMEL5 and SKMEL28 cells. However, the protein expression of PTEN was decreased in melanoma cells. In addition, AKT and PI3K protein phosphorylation levels increased in all melanoma cells, except of G361 cells, demonstrating decreased PI3K protein phosphorylation. These data provided evidence that methylation, mutation and LOH are observed in the inhibin αsubunit gene and gene locus in human melanoma cells. Furthermore, the demethylating agent reactivated inhibinα gene expression and regulated PREX2 expression. AKT/PI3K signaling increased as PTEN expression decreased. In addition, mutations in the tumor suppressor inhibinα, PTEN and p53 genes were not associated with transcriptional silencing, gene expression and cell growth as analyzed through experiments and literature reviews. These data demonstrated that methylation and mutations were associated with the inhibinα gene in human melanoma cells and indicated the regulation of PTEN expression and AKT/PI3K signaling by a demethylating agent.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Metilación de ADN
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Fosfatidilinositol 3-Quinasas
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Fosfohidrolasa PTEN
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Proteínas Proto-Oncogénicas c-akt
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Inhibinas
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Melanoma
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Mutación
Límite:
Humans
Idioma:
En
Revista:
Oncol Rep
Asunto de la revista:
NEOPLASIAS
Año:
2022
Tipo del documento:
Article