title: DNA methylation-mediated silencing of nonsteroidal anti-inflammatory drug-activated gene (NAG-1/GDF15) in glioma cell lines.
abstract: Nonsteroidal anti-inflammatory drug-activated gene, NAG-1, a transforming growth factor-β member, is involved in tumor progression and development. The association between NAG-1 expression and development and progression of glioma has not been well defined. Glioblastoma cell lines have lower basal expression of NAG-1 than other gliomas and normal astrocytes. Most primary human gliomas have very low levels of NAG-1 expression. NAG-1 basal expression appeared to inversely correlate with tumor grade in glioma. Aberrant promoter hypermethylation is a common mechanism for silencing of tumor suppressor genes in cancer cells. In glioblastoma cell lines, NAG-1 expression was increased by the demethylating agent, 5-aza-2'-deoxycytidine. To investigate whether the NAG-1 gene was silenced by hypermethylation in glioblastoma, we examined DNA methylation status using genomic bisulfite sequencing. The NAG-1 promoter was densely methylated in several glioblastoma cell lines as well as in primary oligodendroglioma tumor samples, which have low basal expression of NAG-1. DNA methylation at two specific sites (-53 and +55 CpG sites) in the NAG-1 promoter was strongly associated with low NAG-1 expression. The methylation of the NAG-1 promoter at the -53 site blocks Egr-1 binding and thereby suppresses Nag-1 induction. Treatment of cells with low basal NAG-1 expression with NAG-1 inducer also did not increase NAG-1. Incubation with a demethylation chemical increased Nag-1 basal expression and subsequent incubation with a NAG-1 inducer increased NAG-1 expression. We concluded from these data that methylation of specific promoter sequences causes transcriptional silencing of the NAG-1 locus in glioma and may ultimately contribute to tumor progression.
authors: Laboratory of Molecular Carcinogenesis, NIEHS, National Institutes of Health, Research Triangle Park, NC 27709, USA.
authors: Mitsutoshi
authors: M
authors: Kadowaki
authors: Hiroki
authors: H
authors: Yoshioka
authors: Hideki
authors: H
authors: Kamitani
authors: Takashi
authors: T
authors: Watanabe
authors: Paul A
authors: PA
authors: Wade
authors: Thomas E
authors: TE
authors: Eling
mesh_terms: D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology; D017209:Apoptosis / Q000235:genetics; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D001932:Brain Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D048708:Cell Growth Processes / Q000235:genetics; D045744:Cell Line, Tumor; D019175:DNA Methylation; D000077209:Decitabine; D051766:Early Growth Response Protein 1 / Q000235:genetics / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D005909:Glioblastoma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D055436:Growth Differentiation Factor 15 / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D013467:Sulindac / Q000031:analogs & derivatives / Q000494:pharmacology; D014162:Transfection
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; C526140:GDF15 protein, human; D055436:Growth Differentiation Factor 15; D006877:Hydroxamic Acids; D013467:Sulindac; C012589:trichostatin A; C025462:sulindac sulfide; D000077209:Decitabine; D001374:Azacitidine