Sarabi Mostafa Moradi, Naghibalhossaini Fakhraddin
Department of Biochemistry, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.
Autoimmune Diseases Research Center, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.
Cell Biochem Funct. 2015 Oct;33(7):427-33. doi: 10.1002/cbf.3126. Epub 2015 Sep 28.
There are conflicting reports regarding the association between DNA methyltransferases (DNMTs) expression and global or gene-specific DNA methylation in colorectal cancer (CRC) cells. To correlate DNMTs expression with DNA methylation, we quantified DNMT1, DNMT3A and DNMT3B mRNA levels in five CRC cell lines (HCT116, LS180, HT29/219, Caco2 and SW742) by real-time reverse-transcriptase polymerase chain reaction (PCR) assay. In addition, we examined the global 5-methyl cytosine levels and the methylation patterns of 12 CpG islands in these CRC cells by enzyme-linked immunosorbent assay and methylation-specific PCR methods, respectively. The average expression levels of three DNMTs in HCT116, Caco2, HT29/219 and SW742, relative to the expression level in LS180 (taken to be 1), were 90.1, 31.6, 2.66 and 1.86. Our data indicated that overall about 1.45%, 1.03%, 0.98%, 0.86% and 0.85% of the cytosines were methylated in the genome of HCT116, Caco2, HT29/219, SW742 and LS180 cells, respectively. The 5-mC percentages were positively correlated with the relative cellular DNMTs expression in five CRC cell lines as verified by Pearson correlation test. However, we found no positive correlation between mRNA expression of DNMTs and gene promoter hypermethylation in these cells. Our results suggest that cellular DNMT expression is positively correlated with global DNA methylation level but not with regional DNA hypermethylation at each locus.
关于DNA甲基转移酶（DNMTs）表达与结直肠癌（CRC）细胞中整体或基因特异性DNA甲基化之间的关联，存在相互矛盾的报道。为了将DNMTs表达与DNA甲基化相关联，我们通过实时逆转录聚合酶链反应（PCR）测定法，对五种CRC细胞系（HCT116、LS180、HT29/219、Caco2和SW742）中的DNMT1、DNMT3A和DNMT3B mRNA水平进行了定量。此外，我们分别通过酶联免疫吸附测定法和甲基化特异性PCR方法，检测了这些CRC细胞中整体5-甲基胞嘧啶水平以及12个CpG岛的甲基化模式。相对于LS180中的表达水平（设为1），HCT116、Caco2、HT29/219和SW742中三种DNMTs的平均表达水平分别为90.1、31.6、2.66和1.86。我们的数据表明，HCT116、Caco2、HT29/219、SW742和LS180细胞基因组中分别约有1.45%、1.03%、0.98%、0.86%和0.85%的胞嘧啶发生了甲基化。经Pearson相关性检验证实，五种CRC细胞系中5-mC百分比与相对细胞DNMTs表达呈正相关。然而，我们发现这些细胞中DNMTs的mRNA表达与基因启动子高甲基化之间没有正相关。我们的结果表明，细胞DNMT表达与整体DNA甲基化水平呈正相关，但与每个位点的区域DNA高甲基化无关。
