Cloning And Functional Analysis Of GmZTL3 And GmZTL4 In Soybean

ZEITLUPE(ZTL)plays an important role in the control of circadian clock and photoperiod flowering in Arabidopsis.In this study,we acquired two ZTL homologs in soybean based on homological cloning,and named them as Gm ZTL3 and Gm ZTT4,respectively.Then,their biochemical characteristics and physiology functions are studied,and the results indicate that they may possess the similar functions as At ZTL does.The main evidence are as follows:(1)Analysis of bioinformatics indicate that the protein sequence similarity between Gm ZTL3/4 and At ZTL are up to 80%,and they all possess the same three domains: LOV domain,F-box domain,and kelch repeats domain,in which the Critical amino acids are highly conserved.The LOV domain of Gm ZTL3 and Gm ZTL4 proteins possess the similar 3D structures of Phototropin2 LOV domain.(2)Inducible-expression of recombinant proteins Gm ZTL3 and Gm ZTL4 in yeast,displays a similar absorption spectra to blue light photoreceptor in Arabidopsis,in which two distinct absorption peaks are in blue light region(400~500 nm)and UV-A region(350 nm).(3)Quantitative analysis of Gm ZTL3 and Gm ZTL4 m RNA accumulation by real-time PCR reveal that their expressions are independent of diurnal cycles and concentrate in roots and hypocotyls before the unifoliolates open fully,in all kinds of leaves before flowering,in stems and petioles at flowering,and in maturation seeds and pods.(4)The subcellular localization of Gm ZTL3 and Gm ZTL4 proteins is examined by Arabidopsis protoplast transformation,and the results show that both Gm ZTL3 and Gm ZTL4 are localize in both cytoplasm and nuclear,that is similar to At ZTL proteins.(5)The ectopic expression of Gm ZTL3 in Arabidopsis results in more rosette leaves and flowering later than their parents,that is also similar to the overexpression of At ZTL in Arabidopsis.In conclusion,both Gm ZTL3 and Gm ZTL4 encode blue photoreceptors as At ZTL,and play a role in different organs and at different developmental stages,and particularly Gm ZTL3 may function as inhibitor of flowering induction in the photoperiod flowering pathway.