Plant fructokinase (FRK) guarantees the growth and development of higher plants by participating in carbohydrate metabolism. In this study, a new fructokinase, OsFRK3, was identified using bioinformatics analysis, enzyme assay, bacterial growth assay, and yeast complementation test. Then, we created OsFRK3 knockout transgenic lines (osfrk3-1 and osfrk3-2) by the CRISPR/Cas9 technology. We found that the 1000-grain weight decreased notably (approximately -3.6% and -6.1%, respectively) in osfrk3-1 and osfrk3-2. Evidently decreased grain width, grain thickness, and endosperm filling rate were detected in the osfrk3 mutants (osfrk3-1 and osfrk3-2) compared with those of the WT. In addition, the content of seed total starch was significantly decreased by 3.42 and 4.80% in osfrk3 lines, compared with that in the WT. The level of maltose was significantly reduced in the mutants, while that of sucrose and fructose was obviously increased in the mutants. The transcript levels of OsGBSS1, OsBEIIb, OsPFP1β, and OsAGPL1 were significantly decreased in the osfrk3 mutants. These results suggest that OsFRK3 may positively regulate the accumulation of starch through influencing the sugar metabolism.
Keywords: Oryza sativa; OsFRK3; fructokinase; grain filling; starch.