LsKN1 and LsOFP6 synergistically regulate the bolting time by modulating the gibberellin pathway in lettuce

Bolting time is an important agronomic trait in lettuce (Lactuca sativa) production. Premature bolting significantly reduces crop quality and marketability. Here, we report map-based cloning and characterization of a LsKN1 gene that controls bolting in lettuce. A segregating population was developed by crossing a crisphead-type cultivar with a stem-type cultivar to genetically map and clone the LsKN1 gene. In the late-bolting parent (crisphead), the LsKN1 was activated by a CACTA-like transposon which was inserted into the first exon of LsKN1. Complementation test, overexpression, and CRISPR/cas9 knockout showed that the activated LsKN1 allele (LsKN1^TP) delays bolting in lettuce. ChIP-seq and phytohormone analysis demonstrated that LsKN1 regulates gibberellin (GA) biosynthesis and response. LsKN1^TP binds to the promoter of the LsGA20ox1 and LsRGA1, and down- and upregulates their expression, respectively. Furthermore, LsRGA1 interacts with LsKN1^TP to enhance the repression of GA biosynthesis. LsOFP6 acts as a safeguard, interacting with LsKN1^TP to prevent excessive inhibition of GA biosynthesis and response during the vegetative-to-reproductive phase transition. The LsKN1-LsOFP6 module orchestrates the GA pathway to regulate bolting time in lettuce, which provides insight into the bolting development in lettuce and offers valuable genetic resources for breeding lettuce varieties resistant to premature bolting.