Study On The Cis-trans Effects Of Gene Transcriptional Regulation In Rats And Mice

Transcription regulation of genes plays a very important role in the evolution of species.More than 40 years ago,the famous evolutionary biologists King and Wilson found that the difference between humans and chimpanzees at the macromolecular level is very small,and this cannot explain the huge difference between the two species in terms of morphology,physiology and behavior.Thus,we suggest that changes in gene transcriptional regulation may be responsible for the differences between humans and chimpanzees.However,direct experimental evidence on the evolution of gene transcriptional regulation is markedly lacking.In recent years,with the development of experimental methods and bioinformatics tools,domestic and foreign researchers can carry out research in this area.Among them,the hybrid model is a classic model that can be used to distinguish the effect size of cis-elements and trans-factors in gene transcription.Therefore,studying the roles of cis-and trans-regulation through hybrid models is of great significance for understanding the evolution of species at the molecular phenotype level.Regarding the above research background,this thesis uses rat and mouse fusion cells and hybrid mice as research models,and uses RNA-seq,ATAC-seq and Ch IP-seq to profile the transcriptome,chromatin open regions and DNA-protein interactions.Combining the three methods,we carried out three levels of research,namely,the research on the interaction of cis-trans regulation,the research on cis-regulation alone,and the research on trans-regulation alone.At the research level of cis-trans regulatory interactions,by analyzing the differences in cis-trans regulation at the level of gene expression and chromatin openness between rats and mice,as well as changes in inheritance patterns,it was found that at the level of gene expression regulation,more gene s are co-regulated by cis-elements and trans-factors.At the level of chromatin regulation,more genes are conserved.In addition,in terms of inheritance patterns,gene expression was found to show more incompatibility,while chromatin openness is mainly conservative.At the level of cis-elements,by analyzing the distribution of monoallelic genes in different tissues,it was found that monoallelic genes are regulated specifically by tissues and cells at the level of gene expression and chromatin openness.It was further found that chromatin openness in the gene promoter region is not sufficient for the formation of monoallelic expression.At the level of trans-factors,through the alignment analysis of the amino acid sequences of transcription factors in the whole genome of multiple species,it was found that the mouse CDX2 protein has 3 specific amino acid mutations in the DNA binding region.We further studied the CDX2 protein through functional analysis of the differences in transcriptional regulation between rats and mice and the functional changes caused by mutations in three specific amino acids of mouse CDX2 protein.It was found that there was no obvious functional difference between mouse CDX2 and rat CDX2,but the three amino acids alone and combinatorial mutations can result in partial loss of function.In summary,this thesis systematically explores the evolutionary patterns of gene transcriptional regulation in rats and mice and studies the roles of cis-elements and trans-factors in the evolution of rats and mice in detail.In the meantime,this thesis explores the application scope of multi-omics and found that the evolutionary regulation mode between mammalian species rats and mice,is different from that of lower animals,that is,the influence of cis-regulation does not increase significantly.At the same time,it is also found that monoallelic genes are regulated specifically by tissues.In addition,evolutionarily conserved transcription factors such as CDX2 also have amino acid mutations that are subject to such strong positive selection but do not result in functional changes in the species.In conclusion,this thesis has laid a good foundation for the study of cis-trans evolutionary regulation and provided ideas and methods that can be used fo r reference.Among them,the research on fusion cells of rats and mice has provided insights into the study of the cis-trans evolutionary regulation among mammalian species.Future investigation could use similar research methods to overcome the defects of reproductive isolation,which in turn makes it possible to explore the differences in the evolutionary regulation of humans and other species.The study of hybrid mouse monoallelic genes provides an effective research method and the possibility to seek new autosomal monoallelic and imprinted genes.The study of transcription factor CDX2 provides a method reference for the study of evolutionary differences caused by protein amino acid sequence mutations in trans-factors.