Molecular characteristics and transcriptional regulatory of spermatogenesis-related gene in adult Banna mini-pig inbred line (BMI).

plays critical roles in mammalian spermatogenesis and cilium maturation. Here, the testes of 12-month-old adult boars of Banna mini-pig inbred line (BMI) were subjected to whole-transcriptome sequencing. The results indicated that the average expression (raw count) of gene in BMI testes was 16138.25, and the average expression value of the corresponding transcript ENSSSCT00000043271.2 was 123.1898. The CDS of obtained from BMI testes was 2,817 bp (GenBank accession number: OL362242). Gene structure analysis showed that was located on chromosome 2 of the pig genome with 19 exons. Protein structure analysis indicated that RFX2 contains 728 amino acids with two conserved domains. Phylogenetic analysis revealed that RFX2 was highly conserved with evolutionary homologies among mammalian species. Other analyses, including PPI networks, KEGG, and GO, indicated that BMI RFX2 had interactions with 43 proteins involving various functions, such as in cell cycle, spermatid development, spermatid differentiation, cilium assembly, and cilium organization, etc. Correlation analysis between these proteins and the transcriptome data implied that RFX2 was significantly associated with FOXJ1, DNAH9, TMEM138, E2F7, and ATR, and particularly showed the highest correlation with ATR, demonstrating the importance of RFX2 and ART in spermatogenesis. Functional annotation implied that RFX2 was involved in 17 GO terms, including three cellular components (CC), six molecular functions (MF), and eight biological processes (BP). The analysis of miRNA-gene targeting indicated that BMI was mainly regulated by two miRNAs, among which four lncRNAs and five lncRNAs competitively bound ssc-miR-365-5p and ssc-miR-744 with , respectively. Further, the dual-luciferase report assay indicated that the ssc-miR-365-5p and ssc-miR-744 significantly reduced luciferase activity of 3'UTR in the 293T cells, suggesting that these two miRNAs regulated the expression of . Our results revealed the important role of in BMI spermatogenesis, making it an intriguing candidate for follow-up studies.