miR-7081-5p靶向SOX6对羊驼黑色素细胞色素的调控  

MiR-7081-5p Regulates Melanogenesis by Targeting SOX6 in Melanocytes of Alpaca

作　　者：王飞 钟圣伟 李勇[1] 胡小芬[1] 周作红[1] 杨姗姗 WANG Fei;ZHONG Shengwei;LI Yong;HU Xiaofen;ZHOU Zuohong;YANG Shanshan(College of Animal Science and Technology,Jiangxi Agricultural University,Nanchang 330045,China)

机构地区：[1]江西农业大学动物科学技术学院,江西南昌330045

出　　处：《江西农业大学学报》2021年第5期1116-1123,共8页Acta Agriculturae Universitatis Jiangxiensis

基　　金：国家自然科学基金地区基金项目(31960688);江西省教育厅科技计划项目(GJJ180181);江西省自然科学基金项目(20181BAB204016)。

摘　　要：【目的】miR-7081-5p在白色和棕色羊驼皮肤中呈差异表达,推测miR-7081-5p可能参与色素生成。【方法】运用生物信息学和双荧光素酶报告实验对miR-7081-5p的靶基因进行预测和验证;通过脂质体转染的方法,运用qRT-PCR和Western blotting检测miR-7081-5p过表达对靶基因和黑色素生成相关基因表达水平的影响;使用碱溶法检测miR-7081-5p过表达对黑色素含量的影响;使用分光光度计检测miR-7081-5p过表达对酪氨酸酶活性的影响。【结果】生物信息学预测SOX6是miR-7081-5p靶基因之一,双荧光素酶报告实验证实SOX6是其靶基因之一。qRT-PCR和Western blotting结果显示:在羊驼黑色素细胞中,过表达miR-7081-5p显著降低了SOX6、MITF、TYR、TYRP1和TYRP2的表达,其中mRNA表达水平分别降低了40.51%、63.63%、59.22%、58.84%和63.80%,蛋白质表达水平分别降低了71.69%、65.23%、63.23%、58.52%和62.95%。黑色素含量检测结果显示:过表达miR-7081-5p显著降低了总黑色素、真黑素和褐黑素的相对含量,分别下降了49.03%,62.4%和63.8%。酪氨酸酶活性检测结果显示:过表达miR-7081-5p,黑色素细胞内酪氨酸酶活性显著降低了63.06%。【结论】miR-7081-5p靶向SOX6抑制了羊驼黑色素细胞的色素生成。[Objective]miR-7081-5p is differentially expressed in white and brown skins of alpaca,suggesting that it may be involved in the melanogenesis.[Method]Using bioinformatics and dual luciferase report experiments the target genes of miR-7081-5p were predicted and verifyied;by liposome transfection,using qRT-PCR and Western blotting the effect of miR-7081-5p overexpression on the expression levels of target genes and melanogenesis-related genes was detected;using alkaline solution method the effect of miR-7081-5p overexpression on melanin content was detected;using spectrophotometer the effect of miR-7081-5p overexpression on tyrosinase activity was detected.[Result]Predicting the sex-determining region Y-box 6(SOX6)with bioinformatics is one of the target genes of miR-7081-5p.Luciferase activity assays confirmed that SOX6 was targeted by miR-7081-5p.qRT-PCR and Western blotting results showed that overexpression of miR-7081-5p in melanocytes,the mRNA levels of SOX6,MITF,TYR,TYRP1 and TYRP2 were significantly decreased by 40.51%,63.63%,59.22%,58.84% and 63.80%,respectively;the protein levels of SOX6,MITF,TYR,TYRP1 and TYRP2 were significantly decreased by 71.69%,65.23%,63.23%,58.52% and 62.95%,respectively.The total alkali melanin,pheomelanin and eumelanin were significantly decreased by 49.03%,62.4% and 63.8%in melanocytes of overexpressed miR-7081-5p,respectively.The tyrosinase activity was significantly decreased by 63.06% in melanocytes of overexpressed miR-7081-5p.[Conclusion]These results indicate that miR-7081-5p inhibits melanogenesis in meanocytes of alpaca by targeting SOX6.

关 键 词：miR-7081-5p SOX6 MITF 黑色素细胞 色素生成 

分 类 号：S852.2[农业科学—基础兽医学]