The FW2.2-like (FWL) genes encode cysteine-rich proteins with a placenta-specific 8 domain. They play roles in cell division and organ size control, response to rhizobium infection, and metal ion homeostasis in plants. Here, we target eight rice FWL genes using the CRISPR/Cas9 system delivered by Agrobacterium-mediated transformation. We successfully generate transgenic T₀ lines for 15 of the 16 targets. The targeted mutations are detected in the T₀ lines of all 15 targets and the average mutation rate is found to be 81.6%. Transfer DNA (T-DNA) truncation is a major reason for the failure of mutagenesis in T₀ plants. T-DNA segregation analysis reveals that the T-DNA inserts in transgenic plants can be easily eliminated in the T₁ generation. Of the 30 putative off-target sites examined, unintended mutations are detected in 13 sites. Phenotypic analysis reveals that tiller number and plant yield of OsFWL4 gene mutants are significantly greater than those of the wild type. Flag leaves of OsFWL4 gene mutants are wider than those of the wild type. The increase in leaf width of the mutants is caused by an increase in cell number. Additionally, grain length of OsFWL1 gene mutants is higher than that of the wild type. Our results suggest that transgene-free rice plants with targeted mutations can be produced in the T₁ generation using the Agrobacterium-mediated CRISPR/Cas9 system and that the OsFWL4 gene is a negative regulator of tiller number and plant yield.

In plants, auxin/indoleacetic acid (Aux/IAA) proteins are transcriptional regulators, which regulate developmental process and responses to phytohormones and stress treatments. A previous study has shown that the rice Aux/IAA transcription fac...

Grain size is one of the major factors determining rice grain yield. Nevertheless, our knowledge of the molecular mechanisms underlying the control rice grain size remains limited. Trihelix proteins are plant-specific transcription factors that regulate plant growth and development. However, their roles in modulating grain size in cereal crops are largely unknown. Here, we report the rice trihelix family gene Oryza sativa trihelix transcription factor related to grain size 1 (OsTGS1) as a novel regulator of grain size and weight. Mutation of OsTGS1 leads to large and heavy grains, whereas overexpression of OsTGS1 results in small and light grains. OsTGS1 regulates grain size by influencing cell division and cell expansion in spikelet hulls. OsTGS1 is expressed in various tissues, and its expression level increases during panicle development. The OsTGS1 protein is localized to the nucleus and exhibits transcriptional repressor activity. The screening of interacting proteins via a yeast two-hybrid assay revealed that OsTGS1 interacted with GSK3/SHAGGY-LIKE KINASE2 (GSK2), an important regulator of various agronomic traits, including grain size, in rice. Moreover, ostgs1 mutants are hypersensitive to exogenous brassinosteroid treatment, indicating that OsTGS1 may be involved in brassinosteroid signaling. Our study reveals the role of OsTGS1 in controlling grain size and provides a new gene resource for improving grain weight in rice.

Starch serves as a crucial storage substance in both cereal crops and root/tuber crops, with its composition and properties determining the quality of storage organs. The Waxy (Wx) gene, encoding a key enzyme in starch biosynthesis, plays a pivotal role in this metabolic pathway. However, existing reviews seldom systematically elaborate on Wx gene regulatory mechanisms from the perspective of intrinsic molecular networks. Focusing on the model crop rice, this article synthesizes research advances in Wx-mediated starch biosynthesis regulation over the past decade. We analyze the structural features of the Wx gene and factors influencing its regulatory function during starch synthesis. In conclusion, future research directions are proposed to provide references for Wx gene studies in other crops, as well as theoretical foundations for rice varietal improvement and molecular design breeding.

Grain size is a critical agronomic trait that influences grain yield in rice. However, the molecular mechanisms underlying grain size regulation in rice have not been fully elucidated. Here, we report that the GATA family transcription factor OsGATA1 is a novel regulator of grain size and weight. Loss-of-function of OsGATA1 leads to wide and heavy grains, whereas overexpression of OsGATA1 causes narrow and light grains. OsGATA1 regulates grain size by modulating cell proliferation in spikelet hulls. OsGATA1 is ubiquitously expressed in rice tissues, and its expression level increases with panicle development. The OsGATA1 protein localizes to both the nucleus and the cytoplasm and exhibits transcriptional activation activity. The screening of interacting proteins revealed that OsGATA1 interacts with GS9, a transcriptional activator involved in grain size regulation. Genetic analyses confirmed that OsGATA1 and GS9 function in a common genetic pathway to control grain size. In addition, haplotype analyses of OsGATA1 uncovered clear genetic divergence between indica and japonica subpopulations, and Hap2 and Hap3 may be superior haplotypes for grain weight improvement. Furthermore, loss-of-function of OsGATA1 results in increased expression of brassinosteroid (BR) biosynthetic genes and elevated levels of endogenous BRs, suggesting that OsGATA1 plays a role in regulating BR biosynthesis. Taken together, our findings reveal the roles of OsGATA1 in the regulation of grain size and BR biosynthesis and provide a new candidate gene for improving grain weight in rice.