The Agronomic Traits And Brassinosteroid Response Of The Transgenic Rice Overexpressing OsOFP5

Brassinosterol(BR)is a class of steroid phytohormones,which regulates cell elongation,division and differentiation,and plays an important role in regulating almost all plant growth and development processes.BR in rice plays a crucial role in regulating agronomic traits such as plant height,leaf angle,tiller number,seed development,grain filling and grain shape,and more and more studies have found that using BR-related genes can significantly improve rice yield.The Ovate Family Proteins(OFPs)are plant-specifc transcription factors that regulate plant growth and development and abiotic stress.The rice genome encodes 33 OFP proteins,of which OsOFP1,OsOFP3,OsOFP8,OsOFP19 and OsOFP22 have been reported to be involved in BR signaling and regulate agronomic traits such as the plant architecture and grain shape.However,the functions of most OsOFP family members are unknown.In this study,we identified the agronomic traits of OsOFP5 overexpressing transgenic rice(OsOFP5-OE)and its brassinosterol responses,analyzed the expression pattern of BR-related genes in OsOFP5-OE,and identified the protein interactions between OsOFP5 and other OsOFP family members or BR signaling proteins,in order to clarify the functions of OsOFP5 involved in regulating BR responses,thereby regulating the agronomic traits such as the plant architecture and grain shape of rice.The main results are presented as follows:1.The OsOFP5 protein contains the conserved OVATE domain and the promoter region of OsOFP5 contains hormone-responsive cis-acting elements such as BRRE.The expression pattern of OsOFP5 gene was analyzed by fluorescence quantitative PCR(Real-time quantitative polymerase chain reaction,q RT-PCR),and the results showed that its expressions in germination and seedling stages were much higher than those in other periods.The expression of OsOFP5 was in response to the induction of BR or abscisic acid(Abscisic Acid,ABA),and the cold and salt stresses also promoted the expression of this gene.In rice protoplasts,OsOFP5 is localized in both the nucleus and the cytoplasm.2.In the mature stage of OsOFP5-OE,the internode length was changed,whereas the flag leaf angle and tiller number showed no significantly difference from the wild type(WT).The ear length,the number of secondary branches and the number of spikelet were significantly increased,however the seed setting rate was decreased.The grain shape of OsOFP5-OE appeared narrower as the width was decreased,and the length-width ratio was increaseed.The grain size and1000-grain weight were decreased significantly.OsOFP5-OE rice showed some potential for yield increase,such as increased plant height,which contributes to improve biological yield;the unchanged plant width has little effect on plantingdensity;the number of secondary branch and spikelet were increased.Meanwhile,it is accompanied by the unfavorable traits including decreased seed set and grain weight.3.OsOFP5-OE seedlings showed reduced plant height and root length,and larger second leaf inclination compared with the WT,which is consistent with the phenotype of high concentration BR inhibiting seedling growth.The elongation of the coleoptile and the second leaf inclination were more sensitive to exogenous BR treatment,indicating that OsOFP5 overexpression promoted BR signaling in the transgenic rice.The down-regulated transcript levels of BR biosynthesis enzyme coding genes and the up-regulated expression of BR catabolic enzyme coding genes in OsOFP5-OE indicated that the enhanced BR signaling feedback suppressed the biosynthesis of BR.The expression of genes encoding the BR signaling member BZR1 and DLT were up-regulated in OsOFP5-OE.The interaction between OsOFP5 and OsOFP8 was confirmed by yeast two-hybrid and immunoprecipitation assays.The above results indicate that OsOFP5 is involved in BR signaling at least in part through its interaction with OsOFP8,thereby regulating the development of plant height,panicle shape,and grain size of the transgenic rice.The results of ranscriptome sequencing of OsOFP5-OE seedlings showed that more than half of the differentially expressed genes(DEGs)had no functional annotation,and the functional annotated DEGs were enriched in catalytic activity,oxidation-reduction process and other functions and biological processes.OsOFP5 Overexpression regulated the expression of multiple cytochrome P450 encoding genes,with two extremely significantly DEGs in response to BR induction.