<i>TaRIP2</i> Positively Regulates Wheat Pollen Wall Formation Through <scp>MYB80</scp> ‐Controlled Lipid Metabolism

Male-sterile genes and mutants are critical for hybrid seed production in monocotyledonous crops. Lipids are essential structural components of male reproductive organs, such as the anther and pollen. Here, we show that the pollen-preferential gene TaRIP2 is essential for wheat anther development and pollen formation. RT-qPCR analysis revealed TaRIP2 is specifically expressed during the callose and tetrad stages. Using CRISPR/Cas9, we generated TaRIP2 mutants (rip2), which displayed smaller, wilted anthers with defective cuticles and a low proportion of viable pollen grains (~5.7%). Microscopy revealed that the mutant Rip2 microspores had a smaller size, a smooth exine lacking sculptural elements and fewer organelles. RNA-seq identified differentially expressed genes (DEGs) enriched in pathways related to pollen wall formation. KEGG analysis showed these DEGs are involved in cutin, suberine and wax biosynthesis and fatty acid degradation. Fatty acids C16:0, C18:0 and C18:2 were significantly elevated in rip2 anthers. These phenotypes coincided with the downregulation of genes involved in lipid metabolism and anther development. Dual-luciferase and EMSA assays confirmed TaRIP2 is directly regulated by the transcription factor MYB80. Together, our results show TaRIP2 regulates pollen wall formation through the MYB80-control lipid metabolic pathways.