摘要Amylose content(AC)is the main factor de-termining the palatability,viscosity,trans-parency,and digestibility of rice(Oryza sativa)grains.AC in rice grains is mainly controlled by different alleles of the Waxy(Wx)gene.The AP2/EREBP transcription factor OsEBP89 in-teracts with the MYC-like protein OsBP5 to synergistically regulate the expression of Wx.Here,we determined that the GLYCOGEN SYNTHASE KINASE 5(OsGSK5,also named SHAGGY-like kinase 41[OsSK41])inhibits the transcriptional activation activity of OsEBP89 in rice grains during amylose biosynthesis.The loss of OsSK41 function enhanced Wx ex-pression and increased AC in rice grains.By contrast,the loss of function of OsEBP89 re-duced Wx expression and decreased AC in rice grains.OsSK41 interacts with OsEBP89 and phosphorylates four of its sites(Thr-28,Thr-30,Ser-238,and Thr-257),which makes OsEBP89 unstable and attenuates its inter-action with OsBP5.Wx promoter activity was relatively weak when regulated by the phos-phomimic variant OsEBP89E-OsBP5 but relatively strong when regulated by the non-phosphorylatable variant OsEBP89A-OsBP5.Therefore,OsSK41-mediated phosphorylation of OsEBP89 represents an additional layer of complexity in the regulation of amylose bio-synthesis during rice grain development.In addition,our findings provide four possible sites for regulating rice grain AC via precise gene editing.