Molecular Genetic Mechanism Of The Phase Transition From Growth To Spermatogenesis In Goat Testis

The development of goat testis after birth is the process of gradual transition from organ growth to spermatogenesis.Studying the transition process of developmental stages of"growth-spermatogenesis"of testis is the key to analyzing the development mechanism of goat reproductive system.In this study,goat was used to analyze the gene expression process of the testicular development stage,identify the transition-related genes and lincRNAs of the testicular development stage,detect the whole genome selection signal,screen the testicular development-related mutations,and conduct experimental verification.（1）The growth model fitting of testicular weight,the observation of tissue section and transcriptome analysis all showed that 60～90 days of age and 90～120 days of age may be the periods of major changes in testicular gene expression.WGCNA was performed on the up-and down-regulated genes in these two periods,and 9 gene modules（M1～M9）and 84 hub genes were obtained.Through the analysis of gene time series expression patterns,it was found that most of the spermatogenesis-related genes were only up-regulated at 90～120 days of age,while the growth-related genes were down-regulated at 90 and 120 days of age.During 0 to 180 days of age,the development of goat testes has gone through the following three stages:0 to 60 days is the stage of organ growth,with high expression of the genes related to neurodevelopment,angiogenesis,and cell junction adhesion;60 to 120 days of age is the transitional stage when the organ growth-related genes were down-regulated,while the spermatogenesis-related genes were up-regulated;120 to 180 days of age is the spermatogenesis stage when spermatogenesis-related genes were highly expressed,with the low expression of organ growth and immune-related genes.Compared with related studies,19 possible candidate genes of testicular precocious development in mammal（such as ODF2,STRA8 and SOX9）were found.（2）A total of 501 positive selection genes related to the transitional stage of testicular development were detected.Among them,the SPAM1 gene is up-regulated at120 days of age.The positively selected locus,SPAM1 c.716 A>T,caused an exchange of the 239_th amino acid of SPAM1 protein from glutamate into valine.The frequency of the T allele at this locus in the domestic goat population is 0.725 higher than that in the wild goat population.This locus was found under purification selection among species.According to the prediction,this mutation caused a prolonging of the corresponding alpha helix,which improved the hydrophilicity of adjacent amino acid residues.Experimental validation showed that the mutation can significantly improve the hyaluronidase activity of SPAM1 protein.（3）A total of 20269 goat testicular lincRNAs were identified,including 16931 novel lincRNAs.90～120 days of age is the stage when the expression of lincRNAs in the goat testis underwent a major change after birth.Testicular growth-related lincRNAs were mainly down-regulated at 90 days of age,while spermatogenesis-related lincRNAs were mainly up-regulated at 120 days of age.By constructing a co-expression network of lincRNAs and m RNAs,potential key lincRNAs in the process of testicular growth and spermatogenesis were found,such as ENSCHIT00000000777,ENSCHIT00000002069and ENSCHIT00000005076.The target genes of 2645 lincRNAs were predicted.In conclusion,the transition from growth to spermatogenesis in goat testes is controlled by multiple genes.There are 9,098 related genes that meet the screening criteria.Among them,the SPAM1 gene is under positive selection in domestication,and the SPAM1 c.716 A>T mutation in its coding region increases the hyaluronidase activity of SPAM1 protein.The increase in the frequency of the T allele at this locus in the goat population is the common result of the relaxation of constraints in the domestic environment and the positive selection pressure imposed by domestication on reproductive traits.This research helps to deepen the understanding of the developmental mechanism of mammalian reproductive system and provides a theoretical basis for the study of animal reproduction mechanism.