College of Animal Science and Technology, Shanxi Agricultural University, Taigu 030801, China.
Domest Anim Endocrinol. 2010 Apr;38(3):200-9. doi: 10.1016/j.domaniend.2009.10.004. Epub 2009 Nov 25.
Although the influence of endocrine factors is well established, the molecular and cellular mechanisms controlling coat color are not completely understood. A major mechanism for post-transcriptional regulation of gene expression is through the action of microRNAs (miRNAs), which anneal to the 3' untranslated region of mRNAs in a sequence-specific fashion and either block translation or promote transcript degradation. In this study, we investigated the expression of miRNAs in the skin of alpacas with brown vs white coat color using a microarray screen; identified potential mRNA targets for identified miRNAs among coat color genes; and subsequently determined the ability of a specific, differentially expressed miRNA (miR-25) to suppress expression of micropthalmia-associated transcription factor (MITF), a predicted miR-25 target gene that regulates genes linked to coat color. Expression of 10 different miRNA species in the skin of alpacas with brown vs white coat color was identified from microarray screens. Of the 10 alpaca skin miRNAs identified, predicted binding sites in the 3' untranslated region of RNAs encoding for known genes linked to coat color were primarily for miR-25, but sites were also identified for miR-129 and miR-377. Potential miR-25 binding sites were present in transcripts encoding for 11 coat color genes, including MITF. An inverse relationship between transcript abundance for MITF and miR-25 was observed in skin samples collected from alpacas with white vs brown coat color. Furthermore, overexpression of miR-25 in cultured melanocytes reduced MITF mRNA and protein abundance and corresponding mRNA abundance for the MITF-regulated enzymes tyrosinase and tyrosinase-related protein 1. Results support a novel functional role for miRNA-25 in the regulation of gene expression linked to coat color.
虽然内分泌因素的影响已得到证实，但控制毛色的分子和细胞机制尚不完全清楚。转录后调控基因表达的主要机制是通过 microRNAs（miRNAs）的作用，miRNAs 以序列特异性的方式与 mRNA 的 3'非翻译区结合，要么阻止翻译，要么促进转录本降解。在这项研究中，我们使用微阵列筛选研究了棕色和白色被毛的羊驼皮肤中 miRNAs 的表达；鉴定了毛色基因中潜在的 miRNA 靶mRNA；随后确定了特定的、差异表达的 miRNA（miR-25）抑制小眼相关转录因子（MITF）表达的能力，MITF 是一个预测的 miR-25 靶基因，调节与毛色相关的基因。从微阵列筛选中鉴定出棕色和白色被毛的羊驼皮肤中 10 种不同 miRNA 物种的表达。在鉴定的 10 种羊驼皮肤 miRNA 中，编码与毛色相关的已知基因的 RNA 的 3'非翻译区的预测结合位点主要是 miR-25，但也鉴定出 miR-129 和 miR-377 的结合位点。潜在的 miR-25 结合位点存在于编码 11 个毛色基因的转录本中，包括 MITF。在白色和棕色被毛的羊驼皮肤样本中观察到 MITF 转录本丰度与 miR-25 呈负相关。此外，在培养的黑色素细胞中过表达 miR-25 会降低 MITF mRNA 和蛋白丰度以及 MITF 调节的酶酪氨酸酶和酪氨酸酶相关蛋白 1 的相应 mRNA 丰度。结果支持 miRNA-25 在调节毛色相关基因表达中的新的功能作用。
