Functional Dissection Of E3 Ubiquitin Ligases SINA1 In Tomato

As an important commodity character of tomato,fruit size was an important domain direction of breeding.Molecular biological function identification of the genes of the fruit size,and the molecular mechanisms regulating fruit size of tomato were not only playing a significant role in tomato breeding,but also providing an important scientific basis in the aspects of identification of new varieties.Little is known about SINA(Seven in absentia)in tomato,SIN A family contains highly conserved Sina and RING function domain,and its RING function domain can exercise the E3 ubiquitin ligase activity.In this study we found that the tomato fruit size reduced by overexpression of SlSINA1 in tomato.And the results as follows:1.Through tissue expression profile analysis,SlSINA1 were expressed in different tissues,but the highest expression was in flower,then in the bud,and minimum expression was in the root and red ripe period fruit(RR).With the development of the fruits,at 17DPA(Days post-anthesis)SlSINA1 has the highest expression level compared with other fruit periods,then relative expression level to decreased,and minimum in RR.2.We got over-expressing transgenic plants of SlSINA1,compared with control(AC,Ailsa Craig),T1 and T3 fruits of longitudinal diameter,transverse diameter and fruit weight are reduced.3.Observation of cell biology,in 0DPA,7DPA,24 DPA and 35 DPA we took fruits of the transgenic plants and AC for paraffin sections.we found pericarp thickness and cell layer numbers of pericarp decreased by comparing with AC at 0DPA,7DPA,24 DPA and 35 DPA,and the average area of pericarp cell reduced by comparing with AC at 7DPA,24 DPA and 35 DPA.As time goes on,pericarp thickness,cell layer numbers of pericarp and average area of pericarp cell were increased,it can improve 10-15 fold between 7DPA and 24 DPA,and weakly increased in other periods.Therefore,SlSINA1 may participated in regulating fruit size by influencing cell layer numbers of pericarp and the average area of pericarp cell.4.The result of yeast two-hybrid demonstrated that SlSIN A1 interacted with RAP30,SPG and FW2.2,indicating that SlSINA1 may form a complex with them.5.By detection of SlSINA1 with QPCR in transgenic plants,relative expression level of SlSINA1 distinctly increased,but relative expression level of RAP30,SPG and FW2.2 distinctly declined.6.We couldn't get SPG and FW2.2 by Western-blot,SlSINA1 might not interact with RAP30 based on Co-IP results in vivo.