Functional Analysis Of OsHOX1 And OsHOX28 In Regulating Plant Architecture And Application Exploration Of Heading Date Gene Ehd1

Plant architecture is an important agronomic trait in rice,which is determined by tiller angle,tiller number and plant height.Spreading tiller may affect planting density,but too compact culms are more susceptible to insects and pathogens.Too many tillers may lead to thin and fragile culms which are not resistant to logging;but few number of tiller is detrimental to yield production.Hence,in order to obtain the optimized tiller angle and number,it is of significance to investigating the mechanism of regulation of plant architecture.In this study,we performed explorative study of five HD-ZIP Ⅱ transcription factors.Overexpression of OsHOX1 and OsHOX28 outspreaded the tiller angle and also reduced the tiller number.Further investigation revealed that OsHOX1 and OsHOX28 could be induced by auxin treatment and the proteins encoded by both genes possessed transcriptional suppressing activity.Complicated transcriptional interaction between OsHOX1 and other HD-ZIP members were detected.Both suppression and induction of HD-ZIP Ⅰ members were existed,but only suppression were found for that of HD-ZIP Ⅱ,and overexpression of OsHOX1 had little expressional impact on HD-ZIP Ⅲ members.Meanwhile,OsHOX1 and OsHOX28 could reduce the expression of themselves and each other,which suggested a negative feedback regulation mechanism were involving in the regulation.Results from yeast one hybrid showed that both OsHOX1 and OsHOX28 could bind a DNA fragment from upstream of OsHOX1.Biosynthesis of not only auxin but also strigolactone were affected by OsHOX1 and OsHOX28.d17 CRISPR in the background of overexpressing of OsHOX1 or OsHOX28 showed similar tiller number to that of d17 single mutant,but displayed no difference in tiller angle from plants overexpressing OsHOX1 or OsHOX28.These results suggested the regulation of tiller number of OsHOX1 or OsHOX28 require a functional D17.Furthermore,OsHOX1 and OsHOX28 down regulated the expression of Os HSFA2 D and LA1,thus through Os HSFA2D-LA1 pathway to regulate tiller angle.And further investigation indicated the existence of interactions between genes involving in auxin and strigolactone biosynthesis.Reduction of strigolactone causing by mutation of D17 induced the synthesis of auxin,and enhanced auxin synthesis in turn promoted the synthesis of strigolactone.But the regulations of these plant hormones by OsHOX1 and OsHOX28 were independent of each other.Overall,this study first reported the regulation of plant architecture involving HD-ZIP Ⅱ transcription factors,OsHOX1 and OsHOX28.Through independent modulation of biosynthesis of auxin and strigolactone,these genes redundantly control not only tiller angel but also tiller number in rice,which may provide theoretical basis for breeding of new variety with better architecture in the future.The yield of rice is the main concern in both research study and agriculture production.Heading date is one of the most important agronomic characters determining the cultivation region and season of a specific variety.In this study,we mapped a quantitative locus(QTL),q Ehd10,controlling heading date,plant height and grain number per panicle on chromosome 10 in rice genome.Further investigation confirmed that q Ehd10 is a cis-variant allele of Ehd1.Then we continued our study by focusing on the yield contribution of Ehd1.Variations in both the coding and regulating region of Ehd1 between the parents,Zhensan97 and Zhongzao18 were detected by comparing sequencing.Differences in the coding region were not the causal variations since no phenotype change was found in the complementary test.And significant differences of Ehd1 expression were found between near isogenic lines in both long-day and short-day conditions.Overexpression of Ehd1 and Ghd7 together resembled the phenotype of overexpression of Ehd1 alone,and double mutant ghd7 ehd1 showed similar phenotype of single mutant ghd7.These results demonstrated that Ehd1 acted down stream of Ghd7 as a negative regulator of grain yield.Meanwhile,we found that as the expression of Ehd1 declined in these genetic materials,their heading date delayed,but plant height and grain yield of which increased accordingly.This negative correlation was also found in the 45 varieties randomly selected from a core germplasm collection.According to these results,we hypothesized that beneficial variations of heading date and yield could be artificially induced by simply modulating Ehd1 expression.To test this hypothesis,a continuum variations of heading date was created by differentially suppressing of Ehd1 in ghd7 mutant.Further analysis showed that not only the expression of Ehd1 but also that of Hd3 a and RFT1 were negatively correlated with heading date and grain yield.Moreover,we performed the same experiment in another variety Nipponbare which possessed a functional allele of Ghd7.Similar negative correlation between heading date and the expression of Ehd1 was detected,which proved the generality of our approach.Hence,we propose an effective and flexible breeding strategy: regardless of genetic background,continuum variations of heading date and grain yield could be artificially induced by differentially suppressing of Ehd1.And for a specific cultivation region,we could screen and select of heading date best suited for the local environmental conditions to maximize the grain yield.