Molecular Brake SlKNU Regulates Flower Formation and Fruit Development in Tomato

Floral meristems (FMs) are small clusters of stem cells that orchestrate flower formation and ultimately determine the size and structure of the fruit. Precise timing in FM termination is essential: too early and flowers are incomplete, too late and fruits become malformed. In Arabidopsis, the CLV3–WUS feedback loop and KNUCKLES (KNU) gene ensure meristem homeostasis and proper organ number. Tomatoes, however, have a more complex, sympodial flowering system, and how the KNU homolog fine-tunes this process has remained a mystery. Due to these unresolved mechanisms, deeper research was needed to uncover how SlKNU coordinates floral meristem determinacy and fruit size in tomato.

A research team from Nanjing University and Huaiyin Normal University has discovered that the tomato gene SlKNU acts as a molecular brake that stops excessive meristem growth to define fruit size. The study (DOI: 10.1093/hr/uhae331), published in Horticulture Research on March 1, 2025, demonstrates that SlKNU directly represses the stem cell genes SlWUS and SlCLV3 to maintain floral meristem balance. By combining CRISPR/Cas9 knockout and overexpression experiments, the team revealed that this single gene determines the boundary between flower formation and fruit development-opening new paths for precision fruit breeding.

Through CRISPR/Cas9 gene editing, the researchers created SlKNU knockout mutants (CR-slknu-15 and CR-slknu-17). These plants produced flowers with more petals, stamens, and carpels than wild-type plants, leading to fruits with significantly more locules and greater diameters. Microscopic analyses showed that their floral meristems were markedly enlarged, confirming that SlKNU restrains meristem activity to control fruit size. Conversely, when SlKNU was overexpressed, plants displayed stunted growth and reduced meristem vigor, while still maintaining normal carpel development.

At the molecular level, dual-luciferase, EMSA, and yeast one-hybrid assays confirmed that SlKNU binds directly to promoter regions of SlCLV3 and SlCLV1, suppressing their transcription. The protein's EAR repression domain was shown to be crucial for this activity. Strikingly, when SlKNU was expressed in Arabidopsis knu-2 mutants, it completely restored normal flower formation, revealing a conserved function across species. Together, these results identify SlKNU as a master regulator that bridges floral meristem determinacy and fruit development.

Our work shows that SlKNU serves as the critical 'off switch' for floral stem cells, shaping the number of carpels and, consequently, the size of tomato fruits. By directly repressing SlWUS, SlCLV3, and SlCLV1, SlKNU orchestrates a delicate balance between growth and termination within the floral meristem. Understanding this molecular logic not only explains why some tomatoes grow bigger but also provides a new handle for precision breeding in crop improvement."

Professor Bo Sun, corresponding author of the study

This discovery provides a genetic framework for manipulating tomato fruit size through precise control of meristem activity. By fine-tuning SlKNU expression, breeders may one day optimize fruit yield and shape without sacrificing plant fertility. The conserved function of SlKNU across species also hints at a universal mechanism that could be leveraged in other fruit-bearing crops. Integrating SlKNU-based regulation with CRISPR gene-editing technologies may pave the way for high-efficiency, high-yield cultivars, aligning with global goals for sustainable food production and agricultural innovation.