Functional Study Of A Strigolactone Receptor D14 Interacting Protein In Rice

Rice is one of the most important grain crops in China,its yield is of great significance to our food security.Tiller is the branch of rice stem,the shape and number of tillers are key agronomic traits which contribute to the crop architecture.,the number of grain-bearing tillers per plant directly determines the yield.Strigolactones(SLs),a class of plant hormones identified recently,controls many aspects of plant architecture above and under ground,such as inhibiting shoot branching.Mutants deficient in biosynthesis or signalling of SL exhibit increased tillering with reduced plant height in rice.Dwarf14(D14),the receptor of SL,could recognize SL and form a complex with D3 and D53,the F-box protein and repressor,to initiate signalling transduction.This could induce the degradation of D53 via ubiquitination and release downstream genes from transcriptional repression,then regulate plant growth and development such as tiller number,plant height and elongation of hypocotyl.In previous work,our research group identified a protein interacting with At D14 in Arabidopsis,named it as At DIP8.In this work,we identified DIP8 orthologues in rice,and carried out preliminary functional characterization.First,we analyzed the protein sequence of DIP8,which belongs to the Dna JE protein family.We constructed CRISPR/Cas9 system targeting DIP8,obtained a series of transgenic rice by Agrobacterium-mediated transformation.Then,we identified gene edited individuals with divergent base mutations,some of which are homozygous mutants.The phenotype analysis of dip8 mutants shows a reduced tiller number,which is opposite to the phenotype of d14,indicating DIP8 is a positive regulator in rice tillering.Further analysis showed that dip8 mutants possessed increased plant height while tiller number decreased,and their panicle length were shorter than wild type.The capability of regulating aboveground architecture,such as tillering,height and panicle length,demonstrates that DIP8 is a new architecture regulating gene.We quantified the expression level of strigolactone responsive gene in dip8 mutant.We found that D10,the biosynthesis gene of strigolactone,was downregulated,suggesting strigolactone signalling was enhanced in dip8 and was consistent with dip8 phenotype.Moreover,Y2 H assay showed that DIP8 interacts with Os TB1,the key tillering regulator in strigolactone signalling pathway.This interaction might be a important way through which DIP8 regulates strigolactone signalling.We then constructed overexpression plant materials of DIP8,and obtained homozygous transgene lines.Taken together,based on the research on SL signalling pathway,we investigated DIP8,the interacting protein of strigolactone receptor D14,revealed that DIP8 was a new tillering regulator in rice and could regulate strigolactone signalling.These discoveries could expand our acknowledge of the regulation on shoot branching.In the future,we could do further research on the function and mechanism of DIP8 based on this study,expecting to clarify the mechanism underlying rice tillering and provide scientific basis for crop molecular breeding.