College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang, P. R. China.
Sichuan Animal Science Academy, Chengdu, Sichuan, China.
Anim Biotechnol. 2022 Jun;33(3):493-507. doi: 10.1080/10495398.2020.1806860. Epub 2020 Aug 18.
The lncRNA-599547 (619-nt in length) is identified in secondary hair follicle (SHF) of cashmere goat, but its functional roles in regulating the inductive property of dermal papilla cells (DPCs) remains unknown. We found that lncRNA-599547 had significantly higher expression in dermal papilla of cashmere goat SHF at anagen than its counterpart at telogen. The overexpression of lncRNA-599547 led to a significant increase of ALP and LEF1 expression in DPCs ( < 0.05), whereas, the siLncRNA-1 mediated silencing of lncRNA-599547 significantly down-regulated the expression of ALP and LEF1 in DPCs ( < 0.05). Based on biotin-labeled RNA pull-down assay, we found that lncRNA-599547 directly interacted with chi-miR-15b-5p in DPCs. Based on both overexpression and silencing analysis of lncRNA-599547, our results indicate that lncRNA-599547 promotes the expression of Wnt10b in DPCs but without modulating its promoter methylation level. Using the mRNA-3'UTR fragments of goat Wnt10b containing the predicted binding sites of chi-miR-15b-5p in Dual-luciferase Reporter Assays, we show that lncRNA-599547 modulates the expression of Wnt10b at the chi-miR-15b-5p mediated post-transcriptional level. Taken together, our results indicate that lncRNA-599547 sponges miR-15b-5p to positively regulate the expression of Wnt10 gene, and thereby contributes the inductive property of DPCs in cashmere goat.
lncRNA-599547（长度为 619 个核苷酸）在羊绒山羊次生毛囊（SHF）中被鉴定出来，但它在调节真皮乳头细胞（DPC）的诱导特性中的功能作用尚不清楚。我们发现，lncRNA-599547 在羊绒山羊 SHF 的生长期真皮乳头中的表达明显高于休止期。lncRNA-599547 的过表达导致 DPC 中 ALP 和 LEF1 的表达显著增加（<0.05），而 siLncRNA-1 介导的 lncRNA-599547 沉默则显著下调 DPC 中 ALP 和 LEF1 的表达（<0.05）。基于生物素标记的 RNA 下拉实验，我们发现 lncRNA-599547 与 DPC 中的 chi-miR-15b-5p 直接相互作用。基于 lncRNA-599547 的过表达和沉默分析，我们的结果表明 lncRNA-599547 促进了 DPC 中 Wnt10b 的表达，但不调节其启动子甲基化水平。通过 Dual-luciferase Reporter 测定，我们使用含有 chi-miR-15b-5p 预测结合位点的山羊 Wnt10b 的 mRNA-3'UTR 片段，证明 lncRNA-599547 在 chi-miR-15b-5p 介导的转录后水平调节 Wnt10b 的表达。总之，我们的结果表明，lncRNA-599547 作为 miR-15b-5p 的海绵，正向调节 Wnt10 基因的表达，从而促进羊绒山羊 DPC 的诱导特性。
