The Differential Expression Of Protamine 2 And Gene Polymorphism In Human Sperm Associated With Male Infertility

Protamine was discovered a century ago, it has many functions, the most important functions are: protamine will enable the male gene group concentrated in a more dense and fluidity sperm nucleus; protamine can inhibit nucleic acid or other mutant gene entering the sperm to ensure that the transmission of male genetic information is correct; Protamine can remove the transcription factor and other proteins from sperm cells so that the sperm cells lack of genetic information and epigenetic information, thus sperm genetic information can be rearranged by oocytes; protamine is related to the identification of male genetic group in the process of spermiogenesis. Protamine may be as a detecting index in the process of spermiogenesis. Protamine may play a role in the fertilized eggs. People have found that the abnormal expression of P2 is related to male infertility. Sliverst roni have firstly reported the mature sperms of 15 cases of oligospermatism infertile patients are lack of protamine, that is because of protamine abnormal or missing, histone was not replaced by protamine in the process of spermiogenesis, that appeared nucleoprotein obstacles in the transformation, leading to primary infertility, which first proposed t the relationship between the protamine and male infertility. At present, it is believed that the nucleoprotein obstacles in the transformation can affect the normal cohesion and stability of sperm chromatin, can cause the sperms are vulnerable to injury or abnormal shape of sperm head that will form abnormal sperms to weaken the sperm fertilization ability. Even after fertilization, the sperm nuclear can not depolymerize because the nucleoprotein is abnormal, thus affecting the formation of original male pronucleus and the fusion of sperm nucleus and estrogen. Some scholars have proposed that all the infertility patients with abnormal protamine are companied with the reduction or loss of protamine, there is no change or little change of P1, it suggesting that the P2 family in protamine are more closely related to infertility. The sperm nucleus containing protamine 2 is more depolymerization than the sperm nuclear without P2, and was positively correlated to the content of P2, it suggesting that the content of P2 may be related to the depolymerization of sperm nucleus after fertilized or the formation of protocaryon. The content of P2 declined in some patients, in addition, appearing a higher content of histone, which may block the combination of protamine and DNA, influence or interfere the transformation of nucleoprotein types, because the histone which constructed of sperm chromatin-binding protein was not replaced by protamine, which can lead to nucleoprotein obstacles in the transformation.It also found that protamine gene have taken place mutant in some infertile patients. The transgenic mice with protamine expression defects have shown a lot of defects in the structure of sperm nucleus and have different levels of infertility. There is evidence that dangerous levels of protamine may trigger sperm DNA is vulnerable to damage that would cause infertility or assisted reproductive defects. The genome sequence of PRM1 and PRM2 combined with the transitional 2 gene in the form of ring domain. Not all the DNA in essence are involved in the nuclear component of protamine structure, some regions still maintain nuclear body structure. Documents have shown that nearly 85% of DNA in sperm nucleus is related to protamine, and the remaining 15% of DNA is related to histone or other protein. After fertilization, the protamine genome may be removed, the protamine disulfide reduced firstly, and then DNA is formed in a nuclear body structure. The change and segmentation of chromatin after fertilization is potentially relevant to the function of protamine. Different sperm genome DNA region have differential marked with P1,P2 or histone. There are very few cases of mutant protamine gene leading to infertility, pending further study.Objective: To study the relationship between the abnormal expression of P2 and male infertility and between the mutant polymorphism sites of PRM1 gene G197T and PRM2 gene C248T and male infertility.Methods: In this study, we collected semen samples firstly, using TRIzol reagent to extract the total protein and genomic DNA of sperm, to detect the differential expression of P2 in different subjects through Western Blotting technology. First quantitative sperm protein, and then separated by SDS-PAGE and Western blot test, results showed that: the expression content of P2 in oligospermatism and lepto-sperm groups has declined, the expression content has not changed in P2 normal sperm count. Sperm extracted from the nuclear genome DNA, the gene sequences of PRM1 and PRM2 gene was acquired through the GenBank database search, specific primers was designed by Primer5.0 software; PCR product contains PRM1 gene SNP197 site and PRM2 gene SNP248 site. We use REBase database query that if BseRâ… restriction sites disappear after G197T site base mutation, the Eaeâ… restriction s will disappear too after C248T mutation site base, using sequence changes, we have adopted polymerase chain reaction - restriction fragment length State analysis methods to screen PRM1 gene G197T and PRM2 gene C248T polymorphism.Results: The expression content of P2 in infertility patients with oligospermatism and lepto-sperm has declined; the content in sperm count normal group has not changed. PRM1 gene G197T polymorphism and PRM2 gene C248T polymorphism have not found in 36 cases of male infertility patients.Conclusion: The abnormal expression of P2 may be an important reason that caused oligospermatism and lepto-sperm patients infertile. PRM1 gene G197T polymorphism and PRM2 gene C248T polymorphism has nothing to do with male infertility.