Silencing of DMBT1 in neuroblastoma cell lines is not due to methylation of CCWGG motifs on its promoter.
DMBT1 is one of the putative suppressor genes on 10q25-qter, which frequently lacks expression in many different kind of tumors, such as glioblastoma, and lung, esophageal and colorectal cancer. However, little is known about the reasons for this lack of expression in neoplasia. In a previous report, our group demonstrated how MC-IXC, a neuroblastoma cell line which lacked DMBT1 expression, restored it after a 5-Aza-2'-deoxycitidine treatment. So, we wondered whether DMBT1 aberrant promoter methylation could be responsible for DMBT1 silencing in several tumor cell lines, in spite of the fact that there is no CpG island near the 5' end of the gene. We studied the possibility that methylation in CCWGG sequences of the DMBT1 promoter (where "W" means "A" or "T") is able to silence the gene, as had previously been reported for TP53 in leukemia. We digested genomic DNA by the methylation sensitive restriction enzyme EcoR II (C|CWGG), and made two PCRs to amplify the three CCWGG domains placed in the 1 kb upstream DMBT1 5' end. After the PCRs, we could not find correlation between methylation in CCWGG domains and DMBT1 lack of expression. A positive regulator of DMBT1 might be silenced by aberrant methylation.
Muñoz
J
J
Molecular Neuro-Oncology Laboratory, University of Navarra, 31008 Pamplona, Spain.
Castresana
J S
JS
D001483:Base Sequence; D002135:Calcium-Binding Proteins; D045744:Cell Line, Tumor; D019175:DNA Methylation; D004268:DNA-Binding Proteins; D020868:Gene Silencing; D006801:Humans; D008969:Molecular Sequence Data; D009447:Neuroblastoma / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011956:Receptors, Cell Surface / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D025521:Tumor Suppressor Proteins
D002135:Calcium-Binding Proteins; C108174:DMBT1 protein, human; D004268:DNA-Binding Proteins; D011956:Receptors, Cell Surface; D025521:Tumor Suppressor Proteins