基于转录组测序分析猪子宫内膜妊娠中的关键非编码RNA及其调控通路

Key non-coding RNAs and regulatory pathways in porcine endometrial pregnancy based on transcriptome sequencing

【目的】探究调控具有不同产仔数的川乡黑猪和藏猪的猪子宫内膜妊娠过程的相关miRNA、lncRNA及靶基因,进一步揭示影响不同品种猪子宫内膜妊娠相关非编码RNA富集的信号通路。【方法】对平均产仔数为11.35和6.70的川乡黑猪和藏猪第2、5胎次的子宫内膜进行分离,提取RNA,质检合格后进行转录组测序,测序数据经过拼接、比对到miRNA和lncRNA并预测其靶基因,进行GO和KEGG富集分析筛选到调控川乡黑猪和藏猪不同产仔数的候选基因以及信号富集通路。【结果】不同胎次的川乡黑猪和藏猪之间存在多个miRNA、lncRNA及靶基因的差异表达,其中第2胎次具有1571个miRNA和2042个lncRNA差异表达,第5胎次具有1042个miRNA和2096个lncRNA差异表达。进一步根据靶基因的GO和KEGG富集分析发现川乡黑猪在促性腺激素释放激素(GnRH)分泌、ECM受体互作以及Wnt信号通路等的富集程度高于藏猪。同时发现川乡黑猪相比藏猪有醛固酮调节钠重吸收、谷胱甘肽代谢、果糖和甘露糖代谢等过程的富集。此外,已鉴定的关键候选调控基因包括FSHR(Follicle stimulating hormone receptor)、DBX1(Developing brain homeobox 1)、ARID2(AT-rich interaction domain 2)、TNC(Tenascin C)、NT5C2(Neuropilin 2)、LAMC3(Laminin subunit gamma 3)、MADD(MAP kinase activating death domain)等。【结论】高产仔数的川乡黑猪相比低产仔数的藏猪在子宫内膜妊娠过程中有更高的促性腺激素释放激素(GnRH)分泌、ECM受体互作以及Wnt信号通路富集(GO分析结果),同时也有代谢途径的参与(KEGG分析结果)。涉及到的关键调控基因主要有FSHR、DBX1等。本研究为探究非编码RNA在猪子宫内膜妊娠过程中的功能及其分子调控机制提供了新的思路。

[Objective]The study aims to explore miRNA,IncRNA and target genes regulating endometrial pregnancy in Chuanxiang Black pigs and Tibetan pigs with different liter sizes,and to further reveal the signal pathways affecting the enrichment of non-coding RNA related to endometrial pregnancy in different breeds of pigs.[Method]Transcriptome sequencing was performed on the endometrium of the 2nd and 5th litters Chuanxiang Black pigs and Tibetan pigs with an average litter size of 11.35 and 6.70,respectively.RNA was extracted,tran-scriptome sequencing was carried out after quality inspection,sequencing data were spliced,miRNA and lncRNA were aligned,and target genes were predicted.GO and KEGG enrichment analysis were performed to screen candidate regulatory genes and signaling pathways regula-ting different liter sizes of Chuanxiang Black pigs and Tibetan pigs.[Result]Multiple miRNAs,IncRNAs and target genes were differential-ly expressed between Chuanxiang Black pigs and Tibetan pigs.The second parity had 1571 miRNAs and 2042 IncRNAs differentially ex-pressed,and the fth parity had 1042 miRNAs and 2096 IncRNAs differentially expressed.Further GO and KEGG enrichment analysis re-vealed that Chuanxiang Black pigs had higher enrichment in gonadotropin-releasing hormone(GnRH)secretion,ECM-receptor interaction and Wnt signaling pathway than Tibetan pigs.Functional enrichment was also found in some metabolic pathways,including aldosterone regu-lation of sodium reabsorption,glutathione metabolism,fructose and mannose metabolism,etc.Besides,key candidate regulatory genes iden-tified included FSHR(Follicle stimulating hormone receptor),DBXI(Developing brain homeobox 1),ARID2(AT-rich interaction domain 2),TNC(Tenascin C),NT5C2(Neuropilin 2),LAMC3(laminin subunit gamma 3),MADD(MAP kinase activating death domain),etc.[Conclusion]Chuanxiang Black pigs with high litter size have higher gonadotropin-releasing hormone(GnRH)secretion,ECM-receptor in-teraction,Wnt signaling pathway enrichment(CO analysis result),and metabolic pathways(KEGG analysis result)during endometrial preg-nancy compared with Tibetan pigs.The key regulatory genes involved mainly include FSHR,DBXI,etc..Overall,the study provides new insights for exploring the function and molecular regulation mechanism of non-coding RNA in the process of porcine endometrial pregnancy.