Response Of Several Dehydrogenases In Mung Bean To Cadmium Stress

Based on the annotation file of mung bean genome,we identified alcohol dehydrogenase gene(ADHs),succinate dehydrogenase gene(SDHs),cytochrome c oxidase gene(COXs),Mannitol dehydrogenase gene(MTDs)and aldo-keto reductase gene(AKRs)using HMMER and Pfam softwares,and further analyzed the gene structure and phylogenetic.Furthermore,ADH expression patterns and the activity of ADH enzyme in root,stems,and leaves of mung bean seedlings treated with or without cadmium(Cd)were detected using transcriptome and enzymology.The main results are as follows.1.A total of 15 ADHs harbored in mung bean genome,which had similar gene structure and common motifs.Phylogenetic analysis clustered the proteins,i.e.,alcohol dehydrogenase(Vr ADHs),into three clades,which were more closely related to soybean Gm ADHs.Besides Vr ADH10,which have no ADH＿zinc＿N domain,other Vr ADHs contain the typical ADH＿N and ADH＿zinc＿N domains of the ADH family.Transcriptome analysis showed that 12 Vr ADHs were expressed in roots,stems,and leaves of mung bean seedlings in the control and Cd treatment groups,but the expression levels were different.In the control seedlings,the genes with highest expression level were Vr ADH1,Vr ADH3,and Vr ADH14 in roots,stems,and leaves,respectively,while the genes with highest expression level were Vr ADH4,Vr ADH14,and Vr ADH3 in roots,stems,and leaves under Cd treatment.After 1 day of Cd treatment,the expression of Vr ADH4 in roots was significantly increased by90.39-fold.After 9 days of Cd treatment,the expression level of Vr ADH14 in stems and leaves was significantly increased by 12.04-and 76.20-fold,respectively.The analysis of enzyme activity showed that Cd stress increased the activity of ADH enzyme in roots,stems and leaves as a whole,and decreased compared with the control at 1 day in stems and leaves.2.A total of 12 SDHs harbored in mung bean genome,except some SDHs which do not contain motif,and other SDHs have similar structure and motif.Phylogenetic analysis classified 12 Vr SDHs encoding succinate dehydrogenase protein(Vr SDHs),into two branches,which were more closely related to Arabidopsis At SDHs.It contains many types of domain,but the same evolutionary clade contains the same domain.Transcriptome analysis showed that 10 Vr SDHs were expressed in roots,stems and leaves of mung bean seedlings in control and Cd treatment groups,but the expression levels were different.The highest expression levels in roots,stems and leaves of the control group were Vr SDH2,Vr SDH2 and Vr SDH1.2,respectively,while the highest expression levels in roots,stems and leaves of the control group were Vr SDH1.1.After 5 days of Cd treatment,the expression levels of Vr SDH1.1 in roots and leaves were significantly increased by 17.97-and 3.33-fold.After 9 days of Cd treatment,the expression of Vr SDH1.1 in stem was significantly increased by15.01-fold.The results of enzyme activity analysis showed that Cd stress increased the SDH enzyme activity in roots,stems and leaves as a whole.3.A total of 19 COXs harbored in mung bean genome,except some of which do not contain motif,and other COXs have some differences in structure and motif.Phylogenetic analysis classified 19 Vr COXs encoded cytochrome C oxidase proteins(Vr COXs)into two branches,which were closely related to soybean Gm COXs.Most Vr COXs contain cytochrome c oxidase subunit domain.Transcriptome analysis showed that 15 Vr COXs were expressed in roots,stems and leaves of mung bean seedlings in Cd treatment groups,but the expression levels were different.In the control group,the highest expression level was Vr COX5 C in root,stem and leaf,while the highest effect of Cd treatment on root,stem and leaf were Vr COX5 B,Vr COX5 B and Vr COX19.1,respectively.After 1 day of Cd treatment,the expression level of Vr COX5 B in roots was significantly increased by 2.51-fold.After 5 days of Cd treatment,the expression of Vr COX5 B in stem and leaf was significantly increased by 1.21-and 1.02-fold,respectively.The results of enzyme activity analysis showed that Cd stress increased the activity of COX enzyme in roots,stems and leaves as a whole,and decreased in roots and leaves at 5 and 7 days.4.A total of 10 MTDs harbored in mung bean genome,which have similar structures and common motifs.Phylogenetic analysis showed that 10 Vr MTDs encoded by mannitol dehydrogenase proteins(Vr MTDs)were divided into two branches,which were more closely related to soybean Gm MTDs.All Vr MTDs have the same two domains,ADH＿N and ADH＿zinc＿N.Transcriptome analysis showed that 8 Vr MTDs were expressed in roots,stems and leaves of mung bean seedlings in control and Cd treatment groups,but the expression levels were different.The highest expression levels in roots,stems and leaves of the control group were Vr MTD2.3,Vr MTD2.2 and Vr MTD2.2,respectively,while the highest expression levels in roots,stems and leaves of the control group were Vr MTD2.5.After 9 days of Cd treatment,the expression of Vr MTD2.5 in roots,stems and leaves was significantly increased by250.83-,621.40-and 6.64-fold,respectively.The results of enzyme activity analysis showed that Cd stress increased the MTD enzyme activity in roots,stems and leaves as a whole.5.A total of 9 AKRs harbored in mung bean genome,which have similar structures and common motifs.Phylogenetic analysis showed that 9 Vr AKRs encoded aldo-keto reductase proteins(Vr AKRs)were classified into two branches,which were more closely related to soybean Gm AKRs.All AKRs contain the same one domain,the Aldo＿ket＿red domain.Transcriptome analysis showed that 6 Vr AKRs were expressed in roots,stems and leaves of mung bean seedlings in control and Cd treatment groups,but the expression levels were different.In the control group,the highest expression levels were Vr AKR1.5,Vr AKR1.5 and Vr AKR2 in roots,stems and leaves,respectively,while the highest effect of Cd treatment on root,stem and leaves was Vr AKR1.3,Vr AKR1.2 and Vr AKR2,respectively.After 1 day of Cd treatment,the expression of Vr AKR1.3 in roots was significantly increased by 14.06-fold.After 5days of Cd treatment,the expression of Vr AKR1.2 in stem and leaves was significantly increased by 6.90-and 1.87-fold,respectively.The results of enzyme activity analysis showed that Cd stress increased the AKR enzyme activity in roots,stems and leaves as a whole.In conclusion,the results showed that Cd stress significantly increased the expression of Vr ADHs,Vr SDHs,Vr COXs,Vr MTDs and Vr AKRs,but also decreased the expression of some genes.There were significant differences in the expression characteristics of genes in the roots,stems and leaves of mung bean seedlings,and the expression of some genes showed tissue specificity.The results of enzyme activity analysis also showed that Cd stress increased or decreased the activities of ADH,SDH,MTD,COX and AKR enzymes in roots,stems and leaves,which was confirmed by the results of gene expression,indicating that these Vr ADHs,Vr SDHs,Vr COXs,Vr MTDs and Vr AKRs were involved in the response process of mung bean to Cd stress.