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In flowering plants, ideal male reproductive development requires the systematic coordination of various processes, in which timely differentiation and degradation of the anther wall, especially the tapetum, is essential for both pollen formation and anther dehiscence. Here, we show that OsGPAT3, a conserved glycerol-3-phosphate acyltransferase gene, plays a critical role in regulating anther wall degradation and pollen exine formation. The gpat3-2 mutant had defective synthesis of Ubisch bodies, delayed programmed cell death (PCD) of the inner three anther layers, and abnormal degradation of micropores/pollen grains, resulting in failure of pollen maturation and complete male sterility. Complementation and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) experiments demonstrated that OsGPAT3 is responsible for the male sterility phenotype. Furthermore, the expression level of tapetal PCD-related and nutrient metabolism-related genes changed significantly in the gpat3-2 anthers. Based on these genetic and cytological analyses, OsGPAT3 is proposed to coordinate the differentiation and degradation of the anther wall and pollen grains in addition to regulating lipid biosynthesis. This study provides insights for understanding the function of GPATs in regulating rice male reproductive development, and also lays a theoretical basis for hybrid rice breeding. [ABSTRACT FROM AUTHOR]