Analysis Of Evolution And Function Of C2 Domain Proteins In Arabidopsis

C2 domain is a calcium(Ca+)binding domain of approximately 130 residues in length that is widely present in eukaryotes and is evolutionarily conserved.So far,studies have shown that a large number of proteins containing the C2 domain play a role in signal transduction and membrane transport,so the biological processes involved are very extensive.However,most of the current researches on proteins containing C2 domain were concentrated in humans and animals,and the understanding in plants is still very limited.In the process of studying the temporal regulation of Arabidopsis flowering time,we found a gene C2-1 encoding the C2 domain,and its function has not been reported.We are interested in: What role does the gene encoding C2 domain protein play in the process of plant vegetative growth to reproductive growth? Does it affect the response of plants to stress?Does it have an effect on seed germination and dormancy? To this end,we use bioinformatic and molecular methods to investgate the biological function of the proteins containing the C2 domain in Arabidopsis thaliana in plant flowering time control and stress.The relationship between the two has been systematically studied and the following findings have been obtained:1)Bioinformatic analysis of C2-1 gene in Arabidopsis thaliana,and clustering and functional prediction analysis of proteins containing C2 domain in Arabidopsis thaliana,and analysis of epigenetic information distribution characteristics.The results indicated that,Arabidopsis contains a total of 90 C2 domain proteins which clusted into five subfamilies.C2-1 gene was likely to be involved in some important biological processes,such as seed germination and stress in Arabidopsis.We further analyzed the quantitative and structural characteristics of C2 proteins in B.rapa and rice.The results showed that,there were 161 C2 domain proteins in B.rapa and 91 C2 proteins in rice.Cluster analysis indicated that the C2 domain proteins in Arabidopsis,B.rapa and rice belonged to six subfamilies.2)The flowering time of WT(Col),c2-1 single mutant,oe216 and oe114 overexpression transgenic lines was observed respectively,and the results showed that the flowering time of oe216 and oe114 overexpression lines were significantly delayed comparing with wild type,while the flowering time of the c2-1 single mutant was similar to that of the wild type.This result indicated that C2-1 gene played a negative role in theflowering time control of Arabidopsis thaliana.3)After treatment with different concentrations of NaCl,we found that the seed germination rates of oe216 and oe114 overexpression lines on 150 mM NaCl MS medium were significantly higher than wild type,while those of c2-1 single mutant were significantly lower than wild type.This result indicated that C2-1 gene plays a positive regulation in Arabidopsis seed germination.4)Quantitative PCR was used to detect the expression of flowering time-regulated genes(FLC,SOC1,FT,CO and MAFs)in WT(Col),c2-1 single mutant,oe216 and oe114 overexpressing transgenic plants.The results showed: The expression level of the FLC gene was significantly higher in the oe216 and oe114 overexpression lines than in the wild type,while the FLC expression in the c2-1 single mutant was similar to the wild type.The expression patterns of the MAF2,MAF4 and MAF5 genes(homologous to FLC)were similar to the Col wild type.In contrast to the FLC gene,the expression levels of SOC1 and FT in oe216 and oe114 overexpression lines were significantly reduced.There was no significant change in the expression level of the CO gene in the four materials.There was no significant change in the basic expression levels of COR15 a,COR15b and RD29 a in overexpression lines;whereas there was a significant decrease in c2-1 mutants.When subjected to salt stress,the expression levels of COR15 a,COR15b and RD29 a in overexpressing plants were nearly doubled,while in c2-1 single mutants decreased by nearly 90%.Our results explained why at the transcriptional level the c2-1 single mutant,oe216 and oe114 overexpression lines exhibited various phenotypes.In summary,the results showed that the C2-1 gene plays a negative role in the flowering time control of Arabidopsis,while it plays a positive role in stress resistance and seed germination.