Shi Haixia, Yan Zunqiang, Du Hong, Zhang Bo, Gun Shuangbao
College of Animal Science and Technology, Gansu Agricultural University, Lanzhou, 730030, China.
Institute of Livestock, Grass and Green Agriculture, Gansu Academy of Agricultural Sciences, Lanzhou, 730030, China.
BMC Genomics. 2024 Dec 30;25(1):1258. doi: 10.1186/s12864-024-11170-9.
The Hezuo (HZ) pig, a famous indigenous breed in China, is characterized by precocious puberty compared with foreign-introduced pig breeds. Sexual maturation is a complex physiological process, and in recent years, circular RNAs (circRNAs), a new class of noncoding RNAs with endogenous regulatory functions, have been shown to play important roles in regulating sexual maturation. However, the dynamic expression and regulatory mechanism of circRNAs during sexual maturation in HZ pigs remain unclear. In this study, we performed RNA sequencing and bioinformatics analysis to reveal circRNA expression patterns in the testes of HZ boars at 30 days (sexual immaturity; Ha) and 120 days (sexual maturity; Hb), with Landrace (LC) boars of the same age (La and Lb) as controls. Subsequently, an abundant circ_005678 (circPAN3) transcribed from the PAN3 gene, was functionally investigated by RT-qPCR, Western Blot, CCK-8, and flow cytometry.
We identified 31,134 circRNAs in 12 samples, and 2,562, 2,401, 749, and 831 differentially expressed (DE) circRNAs were identified in the Ha-vs-Hb, La-vs-Lb, Ha-vs-La, and Hb-vs-Lb groups, respectively. The results of functional enrichment analyses indicated that these source genes of the DE circRNAs were involved mainly in testicular development and spermatogenesis. Furthermore, we constructed a circRNA-miRNA-mRNA interaction network and functionally analyzed the target genes. GO functional annotation of the target genes suggested that they were mainly involved in biological processes such as gland development, cell proliferation, and reproduction. KEGG pathway analysis further revealed that these genes were enriched mainly in signaling pathways involved in testicular development and spermatogenesis, including the PI3K-Akt and MAPK signaling pathways. Cellular assays revealed that circPAN3 promoted proliferation and inhibited apoptosis in immature Sertoli cells, whereas opposite changes were observed by circPAN3 knockdown.
This study revealed the dynamic expression profiles and regulatory mechanisms of circRNAs during sexual maturation in HZ pigs. Further functional studies demonstrated that circPAN3 promoted the proliferation and inhibited the apoptosis of immature Sertoli cells, suggesting that circPAN3 may be closely related to the characteristics of precocious puberty in HZ boars. These findings provide a new perspective for exploring the regulatory mechanism of circRNAs in precocious puberty in HZ pigs.
合作猪（HZ猪）是中国著名的地方猪种，与引进的国外猪种相比，其特点是性早熟。性成熟是一个复杂的生理过程，近年来，环状RNA（circRNA）作为一类具有内源性调节功能的新型非编码RNA，已被证明在调节性成熟中发挥重要作用。然而，HZ猪性成熟过程中circRNA的动态表达及调控机制尚不清楚。在本研究中，我们进行了RNA测序和生物信息学分析，以揭示30日龄（性未成熟；Ha）和120日龄（性成熟；Hb）的HZ公猪睾丸中circRNA的表达模式，并以相同年龄的长白猪（LC）公猪（La和Lb）作为对照。随后，通过RT-qPCR、蛋白质免疫印迹、CCK-8和流式细胞术对从PAN3基因转录的丰富circ_005678（circPAN3）进行了功能研究。
我们在12个样本中鉴定出31,134个circRNA，在Ha与Hb、La与Lb、Ha与La、Hb与Lb组中分别鉴定出2,562、2,401、749和831个差异表达（DE）circRNA。功能富集分析结果表明，这些DE circRNA的来源基因主要参与睾丸发育和精子发生。此外，我们构建了circRNA- miRNA -mRNA相互作用网络，并对靶基因进行了功能分析。靶基因的GO功能注释表明，它们主要参与腺体发育、细胞增殖和生殖等生物学过程。KEGG通路分析进一步揭示，这些基因主要富集在参与睾丸发育和精子发生的信号通路中，包括PI3K-Akt和MAPK信号通路。细胞实验表明，circPAN3促进未成熟支持细胞的增殖并抑制其凋亡，而circPAN3敲低则观察到相反的变化。
本研究揭示了HZ猪性成熟过程中circRNA的动态表达谱和调控机制。进一步的功能研究表明，circPAN3促进未成熟支持细胞的增殖并抑制其凋亡，提示circPAN3可能与HZ公猪的早熟特性密切相关。这些发现为探索circRNA在HZ猪早熟调控机制方面提供了新的视角。
