[Roles of cytokinin oxidases in regulating rice callus regeneration]

Cytokinins, a class of plant hormones that can regulate the growth and development of plant cells, are closely related to crop yields and essential for promoting callus growth and differentiation. Cytokinin oxidases/dehydrogenases (CKXs) irreversibly catalyze the inactivation of cytokinins by side chain breakage and can dynamically regulate the cytokinin content in plants, thus participating in the photomorphogenesis, growth, and leaf senescence. However, the roles of OsCKXs in callus regeneration remain unclear. In this study, the plant tissue culture and cytokinin assay were employed to systematically investigate the changes in cytokinin content and regenerative capacity of callus from CKX family mutants in rice. We found that the mutation of OsCKX2 significantly increased the content of cytokinins and up-regulated the expression of regeneration-related genes WOX11 and LEC1 in the callus during regeneration, thereby increasing the regeneration rate. The mutation of OsCKX1 and OsCKX7 significantly reduced the content of cytokinins, induced severe browning, and down-regulated the expression of WOX11 and LEC1 in the callus during regeneration, thus reducing the regeneration rate. The treatment with cytokinins significantly mitigated the callus browning and improved the regeneration rate of osckx1 and osckx7 mutants, which indicated that cytokinin content was involved in callus browning. RT-qPCR results showed that the expression of OsCKX2 was high, while that of OsCKX1 and OsCKX7 was very low in the process of callus regeneration. These results indicate that OsCKX2, OsCKX1, and OsCKX7 are involved in the regulation of rice callus regeneration. The findings provide a theoretical basis for improving rice callus regeneration.