干露胁迫对长牡蛎基因组DNA甲基化的影响

Effects of air exposure on genomic DNA methylation in the Pacific oyster(Crassostrea gigas)

为探讨干露胁迫对海产贝类基因组DNA甲基化的影响,应用荧光标记甲基化敏感扩增多态性(fluorescencelabeled methylation sensitive amplified polymorphism,F-MSAP)技术,比较了不同干露条件下(0 d、0.5 d、1 d、3 d、5 d、7 d、9 d和11 d)长牡蛎(Crassostrea gigas)基因组DNA甲基化的变化。结果表明,对照组(干露处理0 d)闭壳肌与鳃组织的总体甲基化水平分别为29.76%和29.82%;干露处理0.5 d、1 d、3 d、5 d、7 d、9 d和11 d的长牡蛎全基因组甲基化水平呈现先增高后降低的趋势,其中,闭壳肌组织的总体甲基化水平分别为36.59%、38.86%、43.02%、39.30%、51.13%、46.79%和35.06%,鳃组织总体甲基化水平分别为39.39%、42.13%、39.36%、43.54%、56.19%、38.57%和28.99%;干露处理7 d的长牡蛎甲基化水平明显高于其他时期(P<0.05),11 d时甲基化水平基本恢复至初始状态。甲基化变异模式分析发现,闭壳肌与鳃组织DNA甲基化变异位点存在差异,甲基化升高位点变化程度较大(P<0.05)。以上结果表明,长牡蛎通过改变DNA甲基化模式来应答干露胁迫,发生了不同程度的甲基化与去甲基化反应,DNA甲基化与长牡蛎的抗逆性状密切相关。

The Pacific oyster （Crassostrea gigas） inhabits the intertidal zone and shows tolerance to air exposure conditions. Most marine invertebrates have been demonstrated to suffer large-scale mortality following sudden changes in osmolality, causing huge economic losses to commercial aquaculture. To explore the effect of air ex- posure on the genomic DNA methylation of marine shellfish, methylation polymorphisms in genomic DNA in the Pacific oyster were analyzed using the fluorescence-labeled methylation sensitive amplified polymorphism （F-MSAP） technique. The oysters were exposed to air individually for 0 d, 0.5 d, 1 d, 3 d, 5 d, 7 d, 9 d, and 11 d. The incidence of DNA methylation in adductor and gill tissue was 29.76% and 29.82%, respectively, in the control group （day 0）. Compared with the control group, the level of total methylation in various groups under air expo- sure increased initially, after which it decreased. The total level of methylation on days 0.5, 1, 3, 5, 7, 9, and 11 was 36.59%, 38.86%, 43.02%, 39.30%, 51.13%, 46.79%, and 35.06%, respectively, in adductor tissues and 39.3%, 42.13%, 39.36%, 43.54%, 56.19%, 38.57%, and 28.99%; respectively, in gill tissues. The methylation level on day 7 was higher than that following other air exposure durations （P〈0.05）; at the end of the experiment （11 d）, the methylation level nearly returned to that of the control. Moreover, data regarding genomic DNA methylation mu- tation patterns at CCGG sites were not consistent with cytosine methylation patterns and patterns of variation be- tween adductor and gill tissue; hypermethylation sites were more numerous than demethylation sites （P〈0.05）, indicating that remarkable changes occurred in the hypermethylation sites of the Pacific oyster. The results of the present study implied that the alteration of methylation patterns in C. gigas in response to air exposure might be strongly linked to stress resistance. These studies are fundamental to direct further research regarding DNA me- thylation in the aquatic shellfish epigenome, specific gene expression, and stress tolerance.