RASSF1A, BLU, NORE1A, PTEN and MGMT expression and promoter methylation in gliomas and glioma cell lines and evidence of deregulated expression of de novo DNMTs.
Methylation of CpG islands in gene promoters can lead to gene silencing. Together with deletion or mutation, it may cause a loss of function of tumor suppressor genes. RASSF1A (3p21.3), NORE1A (1q32.1) and BLU (3p21.3) have been shown to be downregulated by methylation in cancer, and PTEN (10q23.3) and MGMT (10q26.1) are located in areas commonly deleted in astrocytomas. MGMT methylation predicts a better response and a longer overall survival in patients with glioblastomas treated with temozolomide. We analyzed 53 astrocytoma samples and 10 high-grade glioma cell lines. Gene expression was assessed by RT-PCR. Bisulfite sequencing, MSP and a melting curve analysis-based real-time PCR were performed to detect promoter methylation. Treatments with 5'-aza-2'-deoxicitidine were applied to restore gene expression in cell lines. Ninety-two percent of tumor samples were methylated for RASSF1A, 30%-57% for BLU and 47% for MGMT, suggesting promoter methylation of these genes to be a common event in glioma tumorigenesis. Only 4% of the tumors revealed a methylated promoter for NORE1A. No association between methylation and loss of expression could be established for PTEN. We identified de novo DNMTs overexpression in a subset of tumors which may explain the methylation phenotype of individual gliomas.
Lorente
Aiala
A
Brain Tumor Biology Unit, University of Navarra, Irunlarrea 1, Pamplona, Spain.
Mueller
Wolf
W
Urdangarín
Edurne
E
Lázcoz
Paula
P
Lass
Ulrike
U
von Deimling
Andreas
A
Castresana
Javier S
JS
D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D001254:Astrocytoma / Q000235:genetics; D045744:Cell Line, Tumor; D003598:Cytoskeletal Proteins; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics / Q000378:metabolism; D019175:DNA Methylation; D015254:DNA Modification Methylases / Q000235:genetics / Q000378:metabolism; D045643:DNA Repair Enzymes / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D016147:Genes, Tumor Suppressor / Q000502:physiology; D005910:Glioma / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D051059:PTEN Phosphohydrolase / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D003598:Cytoskeletal Proteins; C411762:RASSF1 protein, human; C454042:RASSF5 protein, human; D025521:Tumor Suppressor Proteins; C472720:ZMYND10 protein, human; D015254:DNA Modification Methylases; D004248:DNA (Cytosine-5-)-Methyltransferases; C515491:MGMT protein, human; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human; D020559:Monomeric GTP-Binding Proteins; D045643:DNA Repair Enzymes