Study On The Metabolic Regulation Of GGPP In Tomato(Solanum Lycopersicum)Fruits

Carotenoids are a group of natural pigments distributed widely in plants,algae,fungi,and bacteria.Carotenoids play crucial roles in photosynthesis and photoprotection.They can serve as precursors for various kind of apocarotenoids,such as the hormones abscisic acid(ABA)and strigolactones(SL),and other volatile terpenoids.Apart from their central functions in plants,carotenoids play critical roles in human nutrition and health as essential components of human diets.They provide dietary sources of provitamin A and serve as antioxidants to reduce the onset of some chronic diseases,such as cardiovascular diseases,cancers,and age-related eye diseases.Geranylgeranyl pyrophosphate synthase(GGPP)is the precursor of carotenoids and is synthesized by GGPP synthase(GGPPS).The heterodimers consist of large subunit(LSU,aka.GGPPS)with catalytic activity and small subunit(SSU)with no activity alone.It is reported that SSU can change the kinetics of LSU and the specificity of product when it interacts with LSU.According to the amino acid sequences,SSU can be divided into two categories:SSU? and SSU?.At present,the structure and function of GGPPSs have been studied more thoroughly,but GGPPPS.SSUs are poorly understood.In order to further study the SSUs,the SlSSU? and SlSSU? genes have been retrieved from the tomato genome database and cloned.Tomato fruit is an excellent material for studying carotenoid accumulation,so we used the tomato fruit-specific E8 promoter to overexpress SlSSU?/? in the fruit of the transgenic plants.After analyzing the phenotype and metabolism of transgenic plants,we found that at the vegetative growth stage SSU?overexpression(SSU?-OE)plants grew faster than wild-type(WT)plants,and SSU?overexpression(SSU?-OE)plants grew taller.In the process of fruit development,the carotenoid content of transgenic fruits during the immature green(IM)stage was not significantly different from the WT,while the lycopene content of SSU?-OE ripe fruits was significantly higher than that of the wide type.Considering that plants that overexpress SSU? and SSU? have differences in growth and fruit carotenoid content,and that small subunits alone are not functional,we speculate that these two SSUs may interact with different SlGGPPS(s)and thus have different effects on metabolism.To verify this conjecture,we first quantitatively analyzed the genes related to carotenoid metabolism in tomato fruit.The transcription levels of GGPP downstream genes in the two over-expressed fruits were somewhat different,but the transcription level of SIGGPPSs showed a similar change:the content of SIGGPPS2 and SIGGPPS4 increased,and the content of SlGGPPS1 and SIGGPPS3 remained basically unchanged.In consideration of the inconsistency of gene level and protein level,we also explored the interaction between SlSSUs and SIGGPPSs at the protein level.By subcellular localization and color complementation experiments,we initially determined that SlGGPPS1,SlGGPPS2 and SlGGPPS4 were active.The results of the bimolecular fluorescence complementation experiments showed that SlSSU? can interact with SlGGPPS4,and SlSSU? can interact with SlGGPPS2 and SlGGPPS4.Therefore,We speculate that SlSSU? interact with SlGGPPS2 and SlGGPPS4,which may promote the increase of downstream products,so that SSU?-OE fruits accumulate more lycopene at maturity.