This study was designed to evaluate the seasonal expression of seminal plasma proteins from two bovine breeds adapted to a subtropical climate and their associations with post-thawing sperm and environmental characteristics. Semen samples were obtained three times in summer and three times in winter from four Crioulo Lageano and four Angus bulls. Seminal plasma was obtained by centrifugation, and the other portion of the semen was cryopreserved. Seminal plasma proteins were identified by 2D-nanoUPLC-MS. Post-thawing assessments of sperm kinetics, morphology and membrane integrity were performed. Environmental data such as air temperature, air humidity and black globe temperature (BGT) were recorded, and the temperature-humidity index (THI) was calculated in summer and winter. Results showed that the climate varied significantly between seasons. Although no statistical differences were observed in semen quality between breeds, the protein profiles varied within and between seasons. We suggest that the most critical proteins in summer affecting sperm characteristics were TIMP-2, DNase, Clusterin, CFAH and GPx6. TIMP-2 and DNase showed a higher abundance in Crioulo Lageano in comparison with Angus, while Clusterin, CFAH and GPx6 presented a lower abundance. To the best of our knowledge, this is the first report of a recently evolved type of glutathione peroxidase, GPx6, in seminal plasma of bovines. In winter, five proteins were considered to be more critical: BSP1, BSP3, CCL2, Sulfhydryl oxidase and TIMP-2. BSP1 and TIMP-2 showed a lower abundance while BSP3, CCL2 and Sulfhydryl oxidase presented a higher abundance in this season in Crioulo Lageano in comparison with Angus.

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