Functional Analysis Of An Arabidopsis Trithorax-Related Gene OsSET7 In Rice

Grain shape,which is a determinate trait of rice quality and yield,is always the hot topics for rice genome research.As a quantitative trait,grain shape is regulated by many QTLs.These QTLs encode different types of proteins which are involved in different pathways,such as the proteasomal degradation pathway,G-protein signaling and phytohormones regulation.Current studies have shown that the epigenetic regulation plays vital roles in the process of plant growth and development,but the mechanism in epigenetic regulation on rice grain shape control remained largely unknown.In this study,we obtained a mutant with significantly increased in grain length through screene the Rice Mutant Database(RMD).The main results are as follows:1.The mutant co-segregation test.Flanking sequence showed that T-DNA was inserted in the promoter of OsSET7.Three primers PCR distinguished mutant as homozygote and heterozygote,qRT-PCR detection showed that the expression of OsSET7 in mutant was down-regulated.Co-segregation test demonstrated that the genotype is consistent with the phenotype in mutant.2.The osset7 increases the spikelet hull cell proliferation.SEM and paraffin section results showed that the average length and width of the outer and inner epidermal cells of osset7 glumes was indistinguishable from that of the wild type,but the cell numbers were significantly increased in the mutant in longitudinal direction both in outer and inner glumes.Therefore,we inferred that the increased cell number of the osset7 spikelet hull was likely due to the enhanced cell division.3.OsSET7 may participate in spikelet hull cell division regulation,we detected the expression of cell cycle-related genes in spikelet hulls,including E2F1,E2F2,CYCA2;2,CDKA1-1 CDKA1-2,CDKA2,MCM2,MCM7 and MCM10.As a result,these genes were markedly up-regulated in osset7.4.The osset7 mutant grains display opaque and wrinkled phenotypes.We analyzed starch morphology using SEM.In the wild type,starch granules were polygonal and similar in size.Tightly packed starch granules were abundant in both the peripheral and central areas of the endosperm cells.Whereas,starch granules in the mutant endosperm were round and smaller,and the packaging of starch granules was also looser than that in the wild type.5.Gene and protein structure analysis of OsSET7.OsSET7 containing 362 amino acids encoded by 1616 bp genomic DNA and shares high homology with ATXR5 and ATXR6 in Arabidopsis,they were assigned to ATXR subfamily of SET proteins.6.OsSET7 can interact with OsPCNA.Yeast two-hybrid system and bimolecular fluorescence complementation(BiFC)assay validated that OsSET7 could combine with proliferating cell nuclear antigen(PCNA)in rice.