MOE Key Laboratory of Marine Genetics and Breeding, College of Marine Life Sciences, Ocean University of China, 5 Yushan Road, Qingdao 266003, China.
Key Laboratory of Tropical Aquatic Germplasm of Hainan Province, Sanya Oceanog Institute, Ocean University of China, Sanya 572000, China.
Int J Mol Sci. 2022 Mar 21;23(6):3392. doi: 10.3390/ijms23063392.
Black rockfish is an economically important fish in East Asia. Little mention has been paid to the sperm cryopreservation in black rockfish. In this study, the optimal cryodiluent was selected from 48 combinations by detecting various sperm parameters. Transcriptome and methylome analysis were further performed to explore the molecular mechanism of inevitable cryoinjuries. The results showed that cryopreservation had negative effects on the viability, DNA integrity, mitochondrial activity, total ATPase and LDH of sperm even with optimal cryodiluent (FBS + 15% Gly). Transcriptome and methylome analysis revealed that the expression of 179 genes and methylation of 1266 genes were affected by cryopreservation. These genes were enriched in GO terms of death, G-protein coupled receptor signaling pathway, response to external stimulus and KEGG pathways of phospholipase D signaling pathway and xenobiotic and carbohydrate metabolism pathways. The role of PIK3CA and CCNA2 were highlighted in the protein-protein interaction network, and the sperm quality-related imprinted gene was identified among the 7 overlapping genes between transcriptome and methylome. Overall, the cryodiluent for black rockfish sperm was optimized, providing a feasible method for cryopreservation. The transcriptome and methylome data further demonstrated the underlying molecular mechanisms of cryoinjuries, proving clues for improvement of cryopreservation method of black rockfish.
黑鲷是东亚地区一种具有重要经济价值的鱼类。然而，关于黑鲷精子的冷冻保存，目前研究甚少。在本研究中，通过检测各种精子参数，从 48 种组合中选择了最佳的冷冻稀释液。进一步进行转录组和甲基组分析，以探讨不可避免的冷冻损伤的分子机制。结果表明，即使使用最佳的冷冻稀释液（FBS+15%Gly），冷冻保存对精子的活力、DNA 完整性、线粒体活性、总 ATP 酶和 LDH 均有负面影响。转录组和甲基组分析显示，179 个基因的表达和 1266 个基因的甲基化受到冷冻保存的影响。这些基因富集在 GO 术语的死亡、G 蛋白偶联受体信号通路、对外界刺激的反应以及磷脂酶 D 信号通路和外来物和碳水化合物代谢途径的 KEGG 途径中。PIK3CA 和 CCNA2 在蛋白质-蛋白质相互作用网络中的作用得到了强调，在转录组和甲基组之间的 7 个重叠基因中鉴定出了与精子质量相关的印迹基因。总的来说，优化了黑鲷精子的冷冻稀释液，为冷冻保存提供了一种可行的方法。转录组和甲基组数据进一步证明了冷冻损伤的潜在分子机制，为黑鲷精子冷冻保存方法的改进提供了线索。
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