The Exploration Of Pig New Genes And Genes With New Functional-mutations After Domestication

During the evolution of pig genome,new genetic information,including new genes and genes with new functional-mutations,are generated continuously.Exploring the new genetic information is tremendously important for breeding investigation and application.Many investigations in model species prove that new genes are related to male reproduction functions,body growth,and immune activities.Therefore,it is valuable to explore and analyze new genes in pig genome.In addition,many investigations in pig and other domesticated species have revealed that two major evolutionary forces,the artificial directional selection and the relaxation of natural purifying selection,may impose selection pressure on genes with new functional mutations after domestication,thus promoting the emergence of new traits.However,many questions are still unclear:1)whether the increasing of functional mutations is a general process in intraspecies and interspecies genomes;2)which factor,the artificial directional selection or the relaxation of natural purifying selection,drives the increasing of new functional mutations;3)the expression pattern and functional significance for genes under different selection pressure.In order to explore new genetic information within pig genome,this study used the typical new gene identification method in model species and high-quality annotations in outgroup species to identify and analyze new genes.Then,the differentially evolving genes were identified and the genes with increased functional-mutations were focused and explored.Meanwhile,the genomic data of other species were used to repeat analyses and confirm the soundness of methods in this study.The major results are on the following:1)Totally,1065 new genes in pig genome have been identified.The origination rate is 21/million year,similar with the rate of new gene evolution in primates.39%of new genes drives the rapid recent expansion of the olfactory gene family.The results suggest that new genes drive the continuous update of pig genome.In addition,based on gene annotation,new genes are related to stimulus response,growth and development,immune response,and reproduction process.The de novo assembling of a wild boar genome has revealed that there are probably no unambiguous domestication specific new genes.2)The RNA-based new genes have been majorly focused as typical examples for new genes.This study has in total identified 94 trans-traffic RNA-based new genes and36 cis-traffic RNA-based new genes,suggesting the retrotransposition events are very important forces to drive genome evolution.The gene traffic between parental gene and new genes reveal that X chromosome have received and produced excessive trans-traffic new genes with excess rate 530%and 506%.In addition,X chromosome is also the hub to receive and produce cis-traffic new genes,with excess rate 579%and 719%.3)Expression data from microarray and RNAseq have demonstrated that 50%RNA-based new genes have testis expression bias.75%X derived trans-traffic RNA-based new genes have higher expression in testis than their parental genes,suggesting the mechanism of producing trans-traffic new genes in X chromosome may be the compensation for parental gene functions during the meiotic sex chromosome inactivation(MSCI).Muscle and testis expression for RNA-based new genes are highly correlated(r~2=82.1%,p<0.001),suggesting that RNA-based new genes may have important pleiotropism for the muscle growth and male reproduction.4)The expression and gene family analyses for cis-traffic RNA-based new genes have revealed that 71.4%of X derived cis-traffic new genes have highest expression in testis,indicating the cis-traffic new genes in X chromosome may also have important function in male reproduction.This study identified 8 cis-traffic new genes in a tandem duplication family along X chromosome with the specific expression in testis.This result is consistent with the previous finding in mouse that X chromosome has a highly duplicated tandem duplication of the required spermatogenesis genes.Due to influence of MSCI,their high expression in testis may indicate their functions in post-meiotic cells.The other 12 RNA-based new genes are related to spermatogenesis and male reproduction processes.The annotation for the remaining RNA-based new genes show that these genes may have functions involving reproduction,muscle development,and immune response,et al.5)The Ka/Ks ratios analyses for differentially evolving genes between pre-and post-domestication stages have revealed that Ka/Ks ratios increase at post-domestication stage.Similar pattern has been found for other 5 small to medium-size domesticated species,suggesting that domestication process may have increased the ratios of functional mutations.In pig genome,401 differentially evolving genes have been identified,among which 74%(295)are genes with increased functional-mutations.For other 5 domesticated species,the ratios of genes with increased functional-mutations are also higher than 50%,suggesting the generality of new functional-mutations accumulation.Among the 295 genes with increased functional-mutations,68%of genes are under the artificial directional selection,indicating the selection forces to boost functional mutation accumulation is the artificial directional selection,rather than the relaxation of natural purifying selection.6)By using 214 wild and domestic resequencing public data,12 newly sequenced genomic data,and 5 outgroup species,this study has confirmed the above result that domestic pigs have accumulated more functional mutations than wild boars.In addition,European wild boars have accumulated higher ratio of new(derived)functional mutations than Asian wild boars.The effective population size computation suggests that this result may be due to the reduced natural purifying selection efficiency caused by lowered effective population size in European wild boars.7)The comparison of differentially evolving genes across different species has revealed that 8 genes are shared at least in 2 species.These shared genes may be correlated with domestication processes,such as neuron system development,male and female reproduction activities,nutrient molecular metabolism and biosynthesis,growth,and immune response.8)The expression pattern analyses using RNAseq data in 27 tissues for genes with new functional-mutations have revealed that,in comparison to genes under the relaxation of purifying selection,the genes under the artificial directional selection show higher likelihood of expression abundance and tissue-specificity.This result suggests that the artificial directional selection may be related to specifically selected traits during domestication.Only one gene with tissue specific expression is under the relaxation of natural purifying selection;while at least 90%(208/214)genes under the artificial directional selection show tissue biased expression.16 genes under the artificial directional selection show correlated biased expression in brain and endocrine related tissues.9)Kr/Kc>1 have been found in all 8 domesticates,suggesting that the percentages of radical functional mutations are higher than those of conservative functional mutations.Meanwhile,Kr/Kc ratios are higher in domestic groups than in wild groups,suggesting domestication process have promoted the accumulation of radical mutations.The Kr/Kc ratios are positively correlated with the effective population sizes.This result is consistent with the investigation of directional selection efficiency in primate species.10)The joint analyses of new genes and the differentially evolving genes show that 8 new genes are also the differentially evolving genes.The gene annotation suggests that these new genes may be related to immune response activities.In summary,this study has used many genomic and transcriptomic data,using popular and commonly used methods to explore pig new genes and genes with new functional-mutations.The new genetic information may be helpful to our understanding about pig genomic evolution and may provide some clues for pig breeding operations..