MAEL 基因啟動子目標甲基化會激發跳躍子單元活性導致去氧核醣核酸不穩定
鄭裕生、黃詩凱、林永明
國立成功大學醫學院附設醫院泌尿部
Targeted methylation of MAEL promoter results in DNA instability by re-activating transposable elements
Yu-Sheng Cheng, Shi-Kae Wee, Yung-Ming Lin
Department of Urology, National Cheng Kung University College of Medicine and Hospital, Tainan, Taiwan
Purpose:
In mouse model, MAEL plays an important role in the piRNA-mediated defense of the germ line from retrotransposons. In our previous study, human dysregulated methylation of MAEL promoter may lead to de-repression of LINE-1, which may contribute to one of the causes of spermatogenic failure in infertile men. This study is focused on investigating whether the targeted methylation of MAEL promoter has the direct impact on genome instability.
Materials and methods:
The targeted DNA methylation(TDM) in MAEL promoter region(from -188 to +294) is established in H358 human cell line. MAEL and LINE-1expression levels in TDM cells were determined and compared with un-methylated cells. To investigate the impact of dysregulated methylation in MAEL promoter, we determined the Double-Strand Breaks (DSBs) by immunofluorescence with histone γ-H2AX in TDM cells compared with un-methylated cells. In patients with hypospermatogenesis and non-obstructive azoospermia, the mRNA transcript levels were determined by quantitative real-time RT-PCR compared to normal spermatogenesis. The methylation levels of MAEL promoter region were investigated by pyrosequencing technology. Moreover, LINE-1 expression was determined afterward. The significance was set at P < 0.05.
Results:
In H358 human cell line, MAEL expression levels of TDM cells decreased significantly compared to un-methylated cells. Nevertheless, LINE-1 transcripts were significantly higher in TDM cells compared to un-methylated cells. There were more Double-Strand Breaks (DSB) in TDM cells than in un-methylated cells.
In human testis, the mRNA transcript levels of MAEL were significantly lower in patients with hypospermatogenesis (P = 0.0131). There are 26 CpGs in our predicted promoter region (-131 to +177, TSS=1) of the MAEL gene. Total 26 CpGs showed significantly higher methylation levels in HS group. Moreover, LINE-1 transcript levels were significantly higher in HS versus NS (P = 0.0303).
Conclusions:
Dysregulated methylation of MAEL promoter might lead to DNA instability by reactivating transposable elements in human cells. We believed this mechanism responsible for one of the causes in human male infertility.