【摘要】：microRNA-411a-3p(miR-411a-3p)在不同毛色羊駝皮膚中差異表達(dá),提示其可能在毛色形成中起調(diào)控作用。為證明miR-411a-3p在羊駝皮毛黑色素形成中的作用,本研究通過生物信息學(xué)預(yù)測及靶基因搜索,發(fā)現(xiàn)胰島素樣生長因子1受體(IGF1R)是miR-411a-3p的靶基因,繼而構(gòu)建了含Igf1r mRNA 3'-UTR的熒光素酶報告基因載體。報告基因轉(zhuǎn)染結(jié)合報告酶活性測定證明,Igf1r是miR-411a-3p的靶基因。原位雜交顯示,miR-411a-3p主要在羊駝黑色素細(xì)胞胞質(zhì)中表達(dá),提示其可能在黑色素細(xì)胞中具有重要作用。qRT-PCR揭示,棕色和黑色羊駝皮膚中miR-411a-3p表達(dá)量明顯低于白色羊駝。qRT-PCR聯(lián)合蛋白質(zhì)印跡分析顯示,在羊駝黑色素細(xì)胞過表達(dá)miR-411a-3p,伴隨Igf1r下調(diào),小眼畸形相關(guān)轉(zhuǎn)錄因子(Mitf)依賴的毛色基因酪氨酸酶(Tyr)、酪氨酸酶相關(guān)蛋白-2(Tyrp2)及黑色素表達(dá)水平明顯下調(diào)。上述結(jié)果提示,miR-411a-3p可通過靶向抑制Igf1r負(fù)調(diào)控羊駝黑色素黑色素合成。該結(jié)果將加深我們對羊駝毛色形成機制的認(rèn)識。
[Abstract]:The differential expression of microRNA-411a-3p (miR-411a-3p) in the skin of alpaca with different color suggests that miR-411a-3p may play a regulatory role in the formation of wool color. In order to demonstrate the role of miR-411a-3p in the formation of melanin in alpaca fur, we found that insulin like growth factor 1 receptor (IGF1R) is the target gene of miR-411a-3p by bioinformatics prediction and target gene search. Then the luciferase reporter gene vector containing Igf1r mRNA 3'-UTR was constructed. The reporter gene transfection combined with reporter enzyme activity showed that Igf1r was the target gene of miR-411a-3p. In situ hybridization showed that miR-411a-3p was mainly expressed in the cytoplasm of melanocytes of alpaca, suggesting that miR-411a-3p may play an important role in melanocytes. The expression of miR-411a-3p in brown and black alpaca skin was significantly lower than that in white alpaca. QRT-PCR combined with Western blot analysis showed that the overexpression of miR-411a-3p, was accompanied by down-regulation of Igf1r in the melanocytes of alpaca. The expression of tyrosinase (Tyr), tyrosinase protein-2 (Tyrp2) and melanin were down-regulated by (Mitf) dependent transcription factor. These results suggest that miR-411a-3p can inhibit the negative regulation of melanin synthesis in alpaca by targeting Igf1r. This result will deepen our understanding of the formation mechanism of alpaca coat color.
【作者單位】： 山西農(nóng)業(yè)大學(xué)動物科技學(xué)院;
【基金】：國家大學(xué)生創(chuàng)新創(chuàng)業(yè)訓(xùn)練項目(No.201710113025) 山西省“三晉學(xué)者”人才專項資助~~
【分類號】：S829.9

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