Comparative Analysis Of Genetic Variation Of Mitochondrial Regulatory Genes In Polyploid Cyprinids

Mitochondria are not only important cellular energy plants,but also participate in a variety of important life metabolic activities,such as apoptosis,cell differentiation and cell information transmission.At the same time,mitochondrial dysfunction will lead to the entire cell dysfunction,thus leading to body disease.Although mitochondria is a semi-autonomous organelle,it can encode proteins by itself.It is mainly subject to nuclear gene regulation and their interaction.Mitochondrial replication and transcription are also regulated by the nuclear gene,the main regulatory genes include mitochondrial RNA polymerase(polrmt),mitochondrial transcription factor A,tfam,mitochondrial transcription Factor B1,tfb1m or mitochondrial transcription factor B2,tfb2m and so on.Thus,the variation of these regulated genes is directly related to the transcription and replication of mitochondrial DNA,which affect the life activities of cells and tissues.This study was about comparative analysis of genetic variation of mitochondrial regulatory genes in polyploid barley.Taking allotetraploid(4nAT),crucian carp(Cyprinus carpio L,CC,2n=100),red crucian carp(Carassius auratus red var,RCC,2n=100)and allotriploid crucian carp(3nRC)as research object,the evolution of mitochondrial transcription factors tfam,tfb1m and tfb2m in several different physiologic fish were investigated.The coding sequences were analyzed and protein structure were predicted.At the same time,the expression of tfam,tfb1m and tfb2m in liver,kidney,brain,testis,ovary and other tissues in several fishes were studied.Laying a good foundation for the follow study on the effect of mitochondrial regulatory factors to the mtDNA and the evolution of mtDNA.The results are as follows:1.Four kinds of fish had measured the cDNA coding sequences and the sequences were analyzed by BioEdit.The tfam protein structure of various fish were predicted by Editseq and Protean in DNASTAR software.The results showed that polyploid fish inherited the genetic characteristics of parents and the genetic variation has occurred at the same time.2.Real-time quantitative PCR(QPCR)was used to detect the number of mtDNA copies in several different tissues.The results showed that in comparison of the testis and the ovary,the copy number of mtDNA in ovary is RCC>3nRC>4nAT,while in testis the copy number of mtDNA is 4nAT>3nRC>RCC.The mtDNA copy number was the highest in the ovary of RCC,and the highest in testis of 4nAT.The copy number of mtDNA in testis of 3nRC and 4nAT was greater than in the ovary,whereas in the ovary of RCC,it was larger than in testis.The mtDNA copy number shown a certain trend with the ploidy change in ovary,testis,and the brain.The results show that it may be related to the ploidy of fish and also has tissue specificity.3.The expression of tfam,tfb1m and tfb2m mitochondrial transcription factors in RCC,4nAT and 3nRC were analyzed by RT-PCR and QPCR.We found that tfam has major regulation of mitochondrial copy number in different tissues effect.In the study of tfb2m,the expression of tfb2m in the liver,brain and kindye was almost the same as that of tfam.tfb2m and mitochondrial copy number have some effect on the transcription of mitochondrial gene cox3.In the study of tfb1m,it was found that the expression of tfblm rises with the increase of ploidy of fish,indicating that tfb1m was stable in different ploidy tissues.Summary:each regulatory factor has its focus:mitochondrial copy number was mainly regulated by the transcription factor tfam,but also by other factors,tfb2m mainly focuses on the regulation of mtDNA transcription,The expression of tfb1m in a variety of tissues of several ploidy fish was relatively stable.