Abstract：
Long noncoding RNA (lncRNA) can regulate mammary gland development and lactation physiological activities. However, the molecular genetic mechanisms of lncRNA in mammary gland involution and cell remodeling remain unclear. This work analyzed the expression characteristics and molecular functions of lncRNA in goat mammary gland tissue at the late lactation (LL), dry period (DP), and late gestation (LG) stages. Sequencing results showed that 3074 lncRNAs were identified in non-lactating goat mammary gland tissue. Statistical analysis of lncRNA length characteristics and exon number found that goat lncRNAs were shorter in length, had fewer exons, and significantly lower expression levels than those of protein-coding genes. 331 differentially expressed lncRNAs were identified in the three comparison groups (LLvsDP, DPvsLG, and LLvsLG), which indicated that the lncRNAs expression at the transcriptional level were changed during mammary involution. Interestingly, lncRNAs were more actively expressed during the dry period compared to lactation, suggesting that lncRNAs in mammary glands are developmentally specific. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses revealed that lncRNAs could regulate immune function, cell proliferation, apoptosis, hormones, substance metabolism, transport, and intercellular communication in the mammary gland through various action modes. Among them, cis-acting lncRNAs enhanced the protection of mammary gland health during the dry period and late gestation. The above reflects the particular mechanisms of lncRNA to adapt to the developmental needs of mammary involution and remodeling. Furthermore, in the lncRNA-miRNA-mRNA network associated with mammary gland development, the expression of LOC102168552 was higher in late gestation than in the dry period and late lactation. Its expression was positively correlated with PRLR and negatively correlated with chi-miR-324-3p. Overexpression of LOC102168552 in goat mammary epithelial cells cultured in vitro could up-regulate PRLR to activate the prolactin signaling pathway by competitively binding to chi-miR-324-3p, promoting cell proliferation, reducing cell cycle arrest in the G1 / S phase, and inhibiting apoptosis. However, overexpression of LOC102168552 alone did not affect mammary cell growth status and the prolactin signaling pathway. This indicates that LOC102168552 must rely on chi-miR-324-3p to inhibit mammary cell apoptosis. In conclusion, the above analysis revealed that lncRNAs in goat mammary tissue are differentially expressed at different stages of involution. As expected, lncRNAs adaptively regulate various physiological activities during mammary gland involution through multiple modes of action, in preparation for a new round of lactation. These findings provide a reference and help further understand the regulatory role of lncRNAs in mammary cell involution and remodeling.
摘要：
长链非编码RNA（lncRNA）可以调节乳腺发育和泌乳生理活动。然而,lncRNA在乳腺退化和细胞重塑中的分子遗传机制尚不清楚。本工作分析了lncRNA在泌乳晚期山羊乳腺组织中的表达特征和分子功能,干燥期（DP），和妊娠晚期（LG）阶段。测序结果显示在非泌乳山羊乳腺组织中鉴定出3074个lncRNAs。lncRNA长度特征和外显子数目的统计分析发现,山羊lncRNA的长度较短,外显子较少，和显著低于蛋白质编码基因的表达水平。在三个比较组中鉴定了331种差异表达的lncRNAs（LLvsDP，DPvsLG,和LLvsLG)，这表明lncRNAs在转录水平的表达在乳腺退化过程中发生了变化。有趣的是,与哺乳期相比，lncRNAs在干旱期表达更活跃，这表明乳腺中的lncRNAs是发育特异性的。基因本体论（GO）和京都基因和基因组百科全书（KEGG）途径富集分析显示，lncRNAs可以调节免疫功能，细胞增殖，凋亡，荷尔蒙，物质代谢，运输,以及通过各种作用方式在乳腺中的细胞间通讯。其中,顺式作用lncRNAs在干旱期和妊娠晚期增强了对乳腺健康的保护。以上反映了lncRNA适应乳腺退化和重塑的发育需求的特定机制。此外，在与乳腺发育相关的lncRNA-miRNA-mRNA网络中，LOC102168552的表达在妊娠晚期高于干旱期和泌乳晚期。其表达与PRLR呈正相关,与chi-miR-324-3p呈负相关。LOC102168552在体外培养的山羊乳腺上皮细胞中的过表达可通过竞争性结合chi-miR-324-3p上调PRLR激活催乳素信号通路,促进细胞增殖，减少细胞周期停滞在G1/S期，抑制细胞凋亡。然而,单独过表达LOC102168552并不影响乳腺细胞生长状态和催乳素信号通路.这表明LOC102168552必须依赖chi-miR-324-3p来抑制乳腺细胞凋亡。总之,以上分析显示，山羊乳腺组织中的lncRNAs在退化的不同阶段差异表达。不出所料,lncRNAs通过多种作用模式适应性调节乳腺退化过程中的各种生理活动，为新一轮哺乳做准备.这些发现提供了参考，并有助于进一步了解lncRNAs在乳腺细胞退化和重塑中的调节作用。
