Let-7d在牙鲆变态发育期间的表达及靶基因预测

MicroRNAs(miRNAs)通过抑制编码基因的转录调控关键的生物学进程。研究发现,let-7是时序发育过程中的重要调控因子,在促进幼体向成体转变的发育过程中起着极其重要的作用。利用Solexa测序技术从牙鲆中成功鉴定出let-7家族的10个成员((let-7a,let-7b,let-7c,let-7d,let-7e,let-7f,let-7g,let-7h,let-7i,let-7j)。相对于let-7家族其他成员,let-7d的拷贝数最多,推测其在牙鲆发育过程中扮演着更重要的角色。运用实时荧光定量PCR方法分析let-7d在牙鲆变态发育不同时期的表达,结果发现,在对照组和甲状腺激素处理组,let-7d的表达模式类似:前5个时期(孵化后17 d,17dph;变态Ⅰ期,20 dph;变态Ⅱ期,23 dph;变态Ⅲ期,29 dph;变态Ⅳ期,36dph)表达量逐渐上升;变态晚期(36 dph)达到高峰;变态结束(41 dph)表达量呈现下降的趋势。定量结果显示,甲状腺激素下调let-7d的表达,与对照组呈显著性差异(P<0.05)。对牙鲆不同组织中let-7d的表达情况分析表明,let-7d主要存在于牙鲆的头部(脑,鳃,心脏)。靶基因预测发现,在目前已公布的482条牙鲆mRNA序列中,其中有30条mRNA的3’非翻译区(3’-UTRs)存在let-7d结合位点。这些预测的靶基因涉及到发育过程的许多方面:蛋白折叠,细胞分化,信号转导,病毒免疫反应和信号通路等。这些结果说明,let-7d在调控牙鲆从仔鱼向稚鱼转变的变态发育过程中起着极其重要的作用。

Expression character and target predictions of let-7dduring metamorphosis in Japanese flounder(Paralichthys olivaceus)

Recent work has demonstrated that microRNAs(miRNAs) are involved in critical biological processes by suppressing the translation of coding genes.let-7 is a critical regulator of developmental timing events at the larval-to-adult transition.MiRNAs in Paralichthys olivaceus were identified by Solexa sequencing.Ten miRNAs belonged to the let-7 family(from let-7a to let-7j).The copy numbers of let-7d were significantly higher than the other members of the let-7 family.Quantitative analysis revealed that the expression levels of let-7d in the normal control group and the thyroid hormones(THs) treatment group had a common characteristic: expression gradually increased to a peak level,and then decreased to a low level.Let-7d levels in the THs treatment group were 50% less than those of the normal control group at each stage of development(P < 0.05).Let-7d was detected in all tissues examined,whereas at relatively higher levels in gill,heart and brain.Data of target predictions indicated that the 3′-untranslated regions(3′-UTRs) of 30 registered mRNAs had target sites matched with let-7d.These genes were grouped into families: protein fold,cell proliferation,signal transduction,virus immune response and signaling path-ways.The results of the present study suggested that let-7d played an essential role in regulating the developmental events during metamorphosis in Paralichthys olivaceus.