The Mechanism Of Crotonylation In Human Mature Sperm

Current infertility is already a global problem,with male factors accounting for about half.Low sperm motility is one of the main causes of male infertility.Its pathogenesis is complex,involving environmental factors,genetic factors,pathogen infection,endocrine factors and other reasons,but the specific molecular pathological mechanism is unknown.Human mature sperm is a cell that is silenced by transcription,and its gene expression regulation has a weak effect on sperm function,so its functional regulation is more likely to depend on post-translational modification of proteins.At present,protein phosphorylation is the most intensive study in mature sperm,which is involved in the regulation of human sperm movement,capacitation and acrosome reaction.Although lysine is a post-translational modification hotspot,only lysine methylation and acetylation modifications have been reported in mature sperm,and other lysine post-translational modifications have rarely been studied.Lysine crotonylation(Kcr)is a novel lysine modification discovered in recent years.It regulates protein function by altering protein structure in charge and physical properties by adding a crotonyl group to the lysine residue of the substrate protein to neutralize the positive charge on lysine.Kcr is a conserved post-translational modification of proteins that is widely found in prokaryotic and eukaryotic cells.Kcr is also present in mouse testicular sperm cells and is enriched in sperm cells in the late spermatogenesis,but it has not been reported in human mature sperm.In this paper,the location,regulation system,function and possible regulation mechanism of Kcr were systematically studied in human mature sperm by immunoblotting,immunofluorescence,sperm function analysis and large sample screening.The results showed that: 1)Kcr can be detected in human mature sperm,which is present in 20-150 k Da protein,and is different from Hela cells;Kcr protein is mainly located in the tail of sperm;2)inhibition of histone acylase P300 Agent(C646)and histone deacylation enzyme Sirtuins inhibitor nicotinamide(NAM)reduce and increase Kcr content in human mature sperm,respectively,suggesting that P300 and Sirtuins are involved in the regulation of Kcr in human mature sperm;sodium crotonate(Na Cr)It can specifically increase the Kcr content of human mature sperm,and prove that crotonyl-Co A is also an acyl donor of crotonylation in human mature sperm;3)It can increase the forward direction of human sperm by using Na Cr specifically to increase human mature sperm Kcr.Exercise rate and exercise capacity such as mucus;4)1836 Kcr sites identified by crotonylation proteomics were distributed on 645 proteins;GO and KEGG enrichment analysis showed that crotonylation modified protein was mainly involved in human sperm Pathways such as energy metabolism;5)Large sample screening results showed that Kcr content in spermatozoa of humans with weak spermatozoa was significantly lower than that in normal human sperm(P < 0.001),whereas Y-chromodomain protein in normal human sperm(C)The content of DYL(with crotonyl-Co A dehydrogenase activity)was significantly lower than that in normal human sperm(P < 0.001);these can be restored by increasing the Kcr content in spermatozoa of weak sperm with a decrease in Kcr content by Na Cr specificity.The patient’s sperm forward movement rate and ATP content.In summary,Kcr is also present in human sperm,and its content is regulated by regulatory enzymes(Sirtuins and P300)and cofactors(crotonyl-Co A).Kcr is involved in the regulation of human sperm motility,and its content may be one of the causes of weak sperm disease,and the pathogenesis of weak sperm disease with decreased Kcr content is related to the increase of CDYL content and the decrease of ATP content in sperm.