TAL, encoding a transaldolase, increases grain size and promotes grain filling in rice.

INTRODUCTION: Grain size and grain filling are critical determinants of rice yield and grain quality. Although transaldolase (TAL) is a key enzyme of the pentose phosphate pathway and is known to be essential for vegetative growth, whether it contributes to the regulation of grain size and filling has yet to be determined. OBJECTIVE: This study aims to systematically characterize the function of TAL, with particular emphasis on its effects on grain size and grain filling in rice. METHODS: We applied CRISPR/Cas9-mediated knockout and RNA interference (RNAi)-mediated knockdown to assess loss-of-function phenotypes. Concurrently, we established stable TAL overexpression lines to evaluate gain-of-function phenotypes. To identify transcriptional regulators upstream of TAL, we integrated multiple molecular approaches: yeast one-hybrid (Y1H) assays for protein-DNA interaction detection, electrophoretic mobility shift assays (EMSAs) for in vitro binding validation, and dual-luciferase (dual-LUC) transient expression assays for in vivo transcriptional regulatory activity quantification. RESULTS: CRISPR/Cas9-mediated knockout of the TAL gene and RNAi-based silencing of TAL affect multiple aspects of rice growth and development, including decreased grain weight and poor grain filling, accompanied by additional pleiotropic effects such as delayed flowering and a marked yield penalty. In contrast, TAL overexpression not only promoted earlier flowering but, more importantly, substantially enhanced grain size and promoted filling. Furthermore, TAL was associated with increased net photosynthetic performance and elevated starch accumulation in leaves and mature grains. Biochemical analysis revealed that TAL possesses transaldolase activity and modulates S7P and E4P homeostasis within the pentose phosphate pathway. We confirmed that TAL is localized to chloroplasts in rice protoplasts, and is preferentially expressed in the pericarp and aleurone layer, as well as in the vascular tissues, suggesting a role in assimilate transport during grain filling. Mechanistically, we identified that OsRA2 functions as a potential transcriptional repressor of TAL by binding to its promoter. CONCLUSION: This study reveals that TAL, as an important regulatory factor in the pentose phosphate pathway, not only affects multiple aspects of rice growth and development, but more importantly, plays a prominent role in regulating grain size and grain filling.