Studies On Physiological Mechanism Of Tillering Of A Dwarf-multi-tiller Rice Mutant D12W191

Tillering is one of important factors which affect the plant architecture and yield of rice.Regulating the occurrence of tillers in rice is very important to shape a high quality population and increase the yield.A dwarf-multi-tiller rice mutant(designed D12W191)and its wild-type Wuyujing 3 were used as materials in this study.The basal nodes and buds were research subjects.We investigated levels of IAA,Z,ZR and GA.The expression of genes regulating tiller outgrowth and proteins was respectively analyzed using RT-qPCR and iTRAQ.The study were to explore mechanisms of tiller formation of the dwarf-multi-tiller rice mutant,which aimed to enrich the theory of rice tiller formation,and to provide theoretical support for the chemical regulation of rice tille.The main results are as follows:(1)The tiller number of mutant D12W191 was much higher than that of Wuyujing 3,but plant height was lower than Wuyujing 3.It was observed that the growth rate of tiller buds of D12W191 was slower than that of Wuyujing 3 at tillering stage.After jointing stage no new tillers of Wuyujing 3 came out,but D12W191 continued to form new tillers.The panicle length of D12W191 was shorter,but grain length and width has no difference between D12W191 and Wuyujing 3.It also showed that number of grain per panicle,filled grain rate and 1000 grain weight of D12W191 were lower than that of Wuyujing 3,which led to a lower grain yield of D12W191.(2)The tillering phenotype of D12W191 may be due to the higher content of the endogenous cytokinin.The contents of Z and ZR in tiller buds of D12W191 were higher than Wuyujing 3,but no significant difference was determined in tiller nodes.Furthermore,a lower level of IAA in tiller nodes of D12W191 than Wuyujing 3 were observed.In tiller buds the content of IAA between Wuyujing 3 and D12W191 was at the same level.The content of GA in tiller buds and nodes of D12W191 was no different with Wuyujing 3.The expression of OsIPT4,OsIPT5,OsIPT7,OsIPT8 was up-regulated both in tiller buds and tiller nodes of D12W191,while a completely contrary trend was seen for OsCKiXl,OsCKiX4,OsCKX5,OsCKX9,OsRR6,OsRR7 and OsRR11.The change trend of IAA synthesis genes was different between tiller buds and tiller nodes of Wuyujing 3 and D12W191.In tiller buds of D12W191 YUCCA3 and YUCCA4 did not change significantly,but the expression of YUCCA6 was much greater than Wuyujing 3.In tiller nodes of D12W191 YUCCA3 and YUCCA6 showed significant decline in expression,but YUCCA4 showed obvious rise.The expression of OsD3 and OsD27 in tiller buds and tiller nodes of D12W191 had no significant difference with Wuyujing 3,but in tiler buds of D12W191 the expression level of OsD10 was higher,which was in stark contrast to the tiller nodes.And the expression of OsD14 in tiller buds of D12W191 showed no difference with Wuyujing 3,but was down-regulated in tiller nodes.OsTB1,a gene specially expressed in buds,was down-regulated in D12W191.The treatment with exogenous cytokinin synthesis inhibitor Lovastatin was able to fully inhibit the outgrowth of tiller buds of two materials.(3)In total,174 differentially expressed proteins between tiller buds of Wuyujing 3 and D12W191 were identified by using iTRAQ of which 110 proteins were up-regulated and 60 protein were down-regulated.These proteins were involved in biosynthesis of secondary metabolites(14%),protein synthesis,folding and degradation(13%),stress response(12%),DNA replication(7%),amino acid synthesis(6%),energy metabolism(5%)lipid transport(5%),hormone synthesis and signal transduction(3%)and so on.Several differentially expressed proteins involved in energy metabolism were up-regulated in the mutant tiller buds.Hormone synthesis and signal transduction were changed.The member of GH3 family ARP1 was up-regulated in the mutant tiller buds,causeing a decrease of free-standing IAA in buds.But cytokinins activated protein LOG4 was down-regulated.This might exist a negative feedback regulation mechanism,because the contents of free state zeatin were much higher than that of wild type,and then negatively regulating the expression of LOG4 protein.ACC oxidase 1,a key enzyme in ethylene biosynthesis,was up-regulated in mutant tiller buds,which indicated that ethylene might promote the occurrence of tiller buds.At the same time,it was also found that the expression abscisic acid responsive factor ABH1 and 4-coumarate--CoA ligase-like 1 related of jasmonic acid synthesis between Wuyujing 3 and D12W191reached significant level,indicating that the two hormones may be involved in the regulation of rice tillering.