Characterization of testis-specific serine/threonine kinase 1-like (TSSK1-like) gene and expression patterns in diploid and triploid Pacific abalone (<i>Haliotis discus hannai</i>; Gastropoda; Mollusca) males

Testis-specific serine/threonine kinase 1-like (TSSK1-like), which plays important roles in late-phase spermatogenesis and male fertility, was characterized in Pacific abalone Haliotis discus hannai, an important commercial marine gastropod. Further, its expression patterns were assessed in diploid and induced triploid males showing differential degrees of testis maturation. Abalone TSSK1-like shared conserved structural features with mammalian TSSK1s and other potential metazoan orthologs, especially regarding the catalytic STKc domain. Phylogenetically, abalone TSSK1-like displayed a genetic affiliation with its molluscan TSSK1-like orthologs and human TSSK1. Additionally, abalone TSSK1-like gene showed a tetrapartite exon-intron organization, unlike the intronless structure of most amniotic tetrapodian TSSK1s. Molecular phylogenetic analysis in the metazoan lineage suggested a possible revision in the origin of the earliest ancestral TSSK1. Further, abalone TSSK1-like showed testis-predominant expression, which was significantly influenced by both age and seasonal reproductive cycles. Comparative expression analyses between diploid and triploid abalone males suggested that robust TSSK1-like expression occurred primarily at the post-meiotic stage. Additionally, RT-PCR assay indicates that mature abalone sperms retain TSSK1-like transcripts after release. Taken together, this study provides useful insights for further studies to assess male reproduction and sterility and/or partial fertility of induced male triploidy in abalone species.睾丸特异性丝氨酸/苏氨酸激酶1样基因（Testis-specific serine/threonine kinase 1-like, TSSK1-like）在精子发生后期与雄性生育调控中发挥关键作用，本研究针对重要的商业海洋腹足类物种——太平洋鲍（*Haliotis discus hannai*，皱纹盘鲍）中的该基因开展了特征分析。研究进一步检测了二倍体与诱导型三倍体雄性个体的表达模式，这些个体的睾丸成熟程度存在显著差异。鲍TSSK1样基因与哺乳动物TSSK1及其他后生动物潜在同源物共享保守的结构特征，尤以催化型STKc结构域最为突出。系统发育分析显示，鲍TSSK1样基因与软体动物类群的TSSK1样同源物及人类TSSK1具有遗传亲缘关系。此外，鲍TSSK1样基因具有四分区的外显子-内含子组织结构，而大多数羊膜四足类的TSSK1基因均为无内含子结构。针对后生动物谱系的分子系统发育分析提示，最早的祖先TSSK1基因的起源或需重新修订。研究还发现，鲍TSSK1样基因呈现睾丸优势表达模式，其表达水平显著受年龄与季节性生殖周期的调控。对二倍体与三倍体鲍雄性个体的比较表达分析表明，TSSK1样基因的高水平表达主要发生在减数分裂后阶段。此外，逆转录聚合酶链式反应（reverse transcription polymerase chain reaction, RT-PCR）实验证实，成熟鲍精子在释放后仍保留有TSSK1样基因的转录本。综上，本研究为后续探究鲍类雄性生殖机制以及诱导型雄性三倍体的不育性与/或部分生育能力提供了重要的参考依据。