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Detection of protamine 2 in bovine spermatozoa and testicles
Author(s) -
Hamilton T. R. S.,
Simões R.,
Mendes C. M.,
Goissis M. D.,
Nakajima E.,
Martins E. A. L.,
Visintin J. A.,
Assumpção M. E. O. A.
Publication year - 2019
Publication title -
andrology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.947
H-Index - 43
eISSN - 2047-2927
pISSN - 2047-2919
DOI - 10.1111/andr.12610
Subject(s) - protamine , biology , andrology , sperm , spermatogenesis , messenger rna , chromatin , microbiology and biotechnology , western blot , gene expression , gene , genetics , endocrinology , medicine , biochemistry , heparin
Background Sperm DNA integrity is crucial for transmission of genetic information to future generations and DNA damage can occur during chromatin packaging. Chromatin packaging involves the replacement of somatic nucleosomal histones by nuclear proteins called protamines. Protamine 1 ( PRM 1) is transcribed and translated in spermatids of all mammals; however, protamine 2 ( PRM 2) is transcribed in low levels in spermatids and it is not yet described in bull mature spermatozoa. Objectives The aim of this study was to assess gene and protein expression of PRM 2 and corroborate gene and protein expression of PRM 1 in bull spermatozoa and testis. Materials and methods For this purpose, absolute q‐ RT ‐ PCR was performed to calculate the number of copies of PRM 1 and PRM 2 mRNA s in bovine epididymal spermatozoa and testicular tissue. Western blot and mass spectrometry were performed to identify PRM 1 and PRM 2 in samples of bovine epididymal spermatozoa. Samples of bovine testicular tissue were collected to identify PRM 1 and PRM 2 by immunohistochemistry. Results We evaluated that the number of PRM 1 mRNA copies was about hundred times higher than PRM 2 mRNA copies in sperm and testicular samples ( p < 0.0001). In addition, we estimated the PRM 1: PRM 2 ratio based on mRNA number of copies. In spermatozoa, the ratio was 1: 0.014, and in testicle, the ratio was 1: 0.009. We also evaluated the immunolocalization for PRM 1 and PRM 2 in bovine testis, and both proteins were detected in spermatids. Western blot and mass spectrometry in bovine epididymal spermatozoa confirmed these results. Conclusion Our work identifies, for the first time, PRM 2 in bovine epididymal spermatozoa and in testis. Further studies are still needed to understand the role of PRM 2 on the chromatin of the spermatozoa and to verify how possible changes in PRM 2 levels may influence the bull fertility.