The functional difference of natural variations in conserved BR signalling genes and the genetic basis of rice indica-japonica differentiation are important yet unknown. Here, we discovered natural variations of the four key components (OsBRI1, OsBAK1, OsGSK3 and OsBZR1) in BR signalling pathway by GWAS using an indicator of indica-japonica differentiation in rice. Two major BR signalling haplotypes (BSHs), caused by co-selected variations of the four genetically unlinked genes, were identified to be highly differentiated between rice subspecies. The genetic contributions of BSHs to grain yield and quality were much higher than that of each component. Introducing alleles of japonica into indica employing substitution lines of OsBAK1, complementation lines of OsGSK3 and genetic populations of OsBRI1/OsBAK1/OsGSK3 confirmed their functional differences between two subspecies. The BSH differentiation led to weaker interaction between OsBRI1 and OsBAK1, stronger autophosphorylation and kinase activity of OsGSK3, less RNA/proteins and stronger phosphorylation of OsBZR1, and weaker BR sensitivity in indica than japonica rice, and regular expression trends of BR-response genes between subspecies, and then synergistically enhanced yield and superior quality of indica. Our results demonstrate that BSHs contribute to rice inter-subspecies diversity, and will provide proof-of-concept breeding strategy and useful targets in crops.