调控山羊绒生长mRNA和lncRNA基因簇及关键信号通路的生物信息学分析

doi:10.16431/j.cnki.1671-7236.2023.11.001

摘要/Abstract

摘要： 【目的】分析绒山羊皮肤中受外源褪黑素诱导的调控山羊绒生长mRNA和lncRNA基因簇及关键信号通路，为从基因簇角度解析外源褪黑素促山羊绒生长的分子机制提供参考。【方法】选取6只罕山白绒山羊，随机分为对照组和埋植组（褪黑素处理），每组3个重复。以自然年为试验周期，每月采集皮肤样本进行RNA测序，使用Bowtie、TopHat、Cufflinks、Cuffmerge、Cuffdiff、Cuffcompare、CPAT和CPC软件分析差异表达mRNA（differentially expressed mRNA，DE-mRNA）和差异表达lncRNA（differentially expressed lncRNA，DE-lncRNA），利用Mfuzz程序包挖掘调控山羊绒生长的基因簇，利用GSVA和GGally程序包分析调控山羊绒生长的信号通路。【结果】筛选得到组间DE-mRNAs 2 024个和DE-lncRNAs 329个，获得对照组和褪黑素处理组各8组不同动力学模式的基因簇。对照组基因簇通过Hippo信号通路、对白细胞介素-1的应答、角蛋白纤维、中间纤维、黑色素生成、细胞周期、FoxO信号通路、Wnt信号通路、Hh信号通路和p53信号通路等参与调控山羊绒生长；褪黑素处理组基因簇通过中间纤维、PI3K-Akt信号通路、聚合细胞骨架纤维、TGF-beta信号通路、生长因子活性、Wnt信号通路、Hippo信号通路、ECM与受体相互作用、p53信号通路和间隙连接等参与调控山羊绒生长。DE-mRNA和DE-lncRNA的相关性研究结果显示，外源褪黑素诱导下55个mRNAs和10个lncRNAs协调互作（PCC>0.960），调控山羊绒生长的共表达网络关系。基因集变异分析（gene set variation analysis，GSVA）可视化了FoxO、p53、PI3K-Akt、Wnt、Hh、Hippo和TGF-beta信号通路在绒山羊皮肤次级毛囊发育不同时期调控山羊绒生长的动态矩阵。外源褪黑素诱导下p53信号通路与FoxO信号通路呈极显著正相关（r=0.843，P<0.01）；Hh信号通路与Hippo（r=0.836，P<0.01）、TGF-beta（r=0.868，P<0.01）、Wnt（r=0.790，P<0.01）、PI3K-Akt（r=0.630，P<0.05）等信号通路均呈极显著或显著正相关；Hippo信号通路与Wnt（r=0.846，P<0.01）、TGF-beta（r=0.806，P<0.05）信号通路均呈极显著或显著正相关；Wnt信号通路与TGF-beta信号通路呈极显著正相关（r=0.866，P<0.01）。【结论】本研究在组学水平从基因簇角度揭示了外源褪黑素诱导下绒山羊皮肤中调控山羊绒生长的55个mRNAs和10个lncRNAs的共表达调控关系，确定了调控山羊绒生长的FoxO、p53、PI3K-Akt、Wnt、Hh、Hippo和TGF-beta共7条关键信号通路，为进一步研究外源褪黑素促山羊绒生长的机制提供参考。

关键词: 绒山羊; 皮肤; 褪黑素; RNA测序; 基因簇; 信号通路

Abstract: 【Objective】 The mRNA and lncRNA gene clusters and the key signal pathways that regulated cashmere growth induced by exogenous melatonin of skin in cashmere goats were analyzed, so as to provide a reference for resolving on the molecular mechanism of exogenous melatonin promoting cashmere growth from the perspective of gene clusters.【Method】 Six Hanshan White cashmere goats were selected and randomly assigned to control group and implanted group (melatonin treatment) with three replicates in each group.Take the natural year as the experiment period, skin samples were collected monthly for RNA-Seq analysis.Using Bowtie, TopHat, Cufflinks, Cuffmerge, Cuffdiff, Cuffcompare, CPAT and CPC software to analyze differentially expressed mRNA (DE-mRNA) and differentially expressed lncRNA (DE-lncRNA), Mfuzz package to mine the gene clusters regulating cashmere growth, GSVA and GGally package to analyze the signal pathways regulating cashmere growth.【Result】 A total of 2 024 DE-mRNAs and 329 DE-lncRNAs were identified, which were divided into 8 gene clusters with different dynamic modes in control and melatonin treatment group, respectively.The gene clusters in control group participated in regulating cashmere growth through Hippo signaling pathway, response to interleukin-1, keratin filament, intermediate filament, melanogenesis, cell cycle, FoxO signaling pathway, Wnt signaling pathway, Hh signaling pathway and p53 signaling pathway. The gene clusters in melatonin treatment group participated in regulating cashmere growth through intermediate filament, PI3K-Akt signaling pathway, polymeric cytoskeletal fiber, TGF-beta signaling pathway, growth factor activity, Wnt signaling pathway, Hippo signaling pathway, ECM-receptor interaction, p53 signaling pathway and gap junction.The correlation between DE-mRNA and DE-lncRNA showed that the co-expression network of 55 mRNAs and 10 lncRNAs (PCC>0.960) induced by exogenous melatonin regulating cashmere growth in goats.Gene set variation analysis (GSVA) visualized the dynamic matrix of FoxO, p53, PI3K-Akt, Wnt, Hh, Hippo and TGF-beta signaling pathways regulating cashmere growth at different stages of secondary hair follicle development in skin of cashmere goat.After exogenous melatonin induction, p53 signaling pathway was extremely significantly positively correlated with FoxO signaling pathway (r=0.843, P<0.01).Hh signaling pathway was extremely significantly or significantly positively correlated with Hippo (r=0.836, P<0.01), TGF-beta (r=0.868, P<0.01), Wnt (r=0.790, P<0.01) and PI3K-Akt (r=0.630, P<0.05) signaling pathways.Hippo signaling pathway was extremely significantly or significantly positively correlated with Wnt (r=0.846, P<0.01) and TGF-beta (r=0.806, P<0.05) signaling pathways.Wnt signaling pathway was extremely significantly positively correlated with the TGF-beta signaling pathway (r=0.866, P<0.01).【Conclusion】 The coexpression regulation relationship between 55 mRNAs and 10 lncRNAs regulating cashmere growth in skin of cashmere goats induced by exogenous melatonin had been revealed from the perspective of gene cluster at the omics level, the seven key signaling pathways of FoxO, p53, PI3K-Akt, Wnt, Hh, Hippo and TGF-beta regulating cashmere growth had been identified, which provided a reference for further investigations on the mechanism of melatonin promoting cashmere growth in goats.

Key words: cashmere goat; skin; melatonin; RNA-Seq; gene cluster; signal pathway

中图分类号:

S827

贾纯琰, 孙燕勇, 包永红, 张文广. 调控山羊绒生长mRNA和lncRNA基因簇及关键信号通路的生物信息学分析[J]. 中国畜牧兽医, 2023, 50(11): 4311-4324.

JIA Chunyan, SUN Yanyong, BAO Yonghong, ZHANG Wenguang. Bioinformatics Analysis of mRNA and lncRNA Gene Clusters and Key Signal Pathways Regulating Cashmere Growth[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(11): 4311-4324.

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