Functional SNPs of INCENP Affect Semen Quality by Alternative Splicing Mode and Binding Affinity with the Target Bta-miR-378 in Chinese Holstein Bulls

Abstract

Inner centromere protein (INCENP) plays an important role in mitosis and meiosis as the main member of chromosomal passenger protein complex (CPC). To investigate the functional markers of the INCENP gene associated with semen quality, the single nucleotide polymorphisms (SNPs) g.19970 A>G and g.34078 T>G were identified and analyzed. The new splice variant INCENP-TV is characterized by the deletion of exon 12. The g.19970 A>G in the exonic splicing enhancer (ESE) motif region results in an aberrant splice variant by constructing two minigene expression vectors using the pSPL3 exon capturing vector and transfecting vectors into MLTC-1 cells. INCENP-TV was more highly expressed than INCENP-reference in adult bull testes. The g.34078 T>G located in the binding region of bta-miR-378 could affect the expression of INCENP, which was verified by luciferase assay. To analyze comprehensively the correlation of SNPs with sperm quality, haplotype combinations constructed by g.19970 A>G and g.34078 T>G, as well as g.-692 C>T and g.-556 G>T reported in our previous studies, were analyzed. The bulls with H1H12 and H2H2 exhibited a higher ejaculate volume than those with H2H10 and H9H12, respectively (P < 0.05). Bulls with H11H11 and H2H10 exhibited higher initial sperm motility than those with H2H2 (P < 0.05). The expression levels of INCENP in bulls with H1H12 and H11H11 were significantly higher than those in bulls with H9H12 (P < 0.05), as determined by qRT-PCR. Findings suggest that g.19970 A>G and g.34078 T>G in INCENP both of which appear to change the molecular and biological characteristics of the mRNA transcribed from the locus may serve as a biomarkers of male bovine fertility by affecting alternative splicing mode and binding affinity with the target bta-miR-378.

Gene Polymorphisms in Boar Spermatozoa and Their Associations with Post-Thaw Semen Quality

“…Among the UTR variants at the 3'-UTR, two SNP polymorphisms, rs339379734 (OXSR1) and rs337913978 (SCLT1), showed significant associations with post-thaw semen quality. Evidence has indicated that SNPs at the 3'-UTR of the targeted mRNA have been the principal elements of microRNA adhesion, and they are associated with semen quality [19]. We suggest that the 3'-UTR SNPs, detected in this study, might play a significant role at the transcription level.…”

A RNA-Seq Analysis to Describe the Boar Sperm Transcriptome and Its Seasonal Changes

“…miR-106b has also been related to autophagy and cellular stress in intestinal epithelial HCT116 cells (Zhai et al, 2013). A study in cattle identified a single nucleotide polymorphisms in a miR-378 target site of the INCENP semen quality associated gene (Liu et al, 2016). In humans, miR-378 was found to also target the autophagy related protein 12 gene ( ATG12 ) in cervical cancer (Tan et al, 2018).…”

Relationship of cysteine‐rich secretory protein‐3 gene and protein with semen quality in stallions

“…Compared with exploring the associations of SNPs in a gene with biological traits, research on haplotype combinations between multiple SNPs may be more constructive, among which interaction among SNPs can be considered (Liu et al, ). In this investigation, it was observed that the haplotype of the CRISP‐3 gene for the four SNPs included in our study has a great influence on MOT, SV, NM and MI (Table ).…”