Editing strigolactone biosynthesis genes in tomato reveals novel phenotypic effects and highlights D27 as a breeding target for parasitic weed resistance

Parasitic weed infestations represent an increasing threat to agriculture worldwide, especially in the Mediterranean region. Phelipanche ramosa (L.) and P. aegyptiaca (Pers.) (broomrapes) cause severe yield losses in field-grown tomato (Solanum lycopersicum L.). Strigolactones (SLs) are apocarotenoid phytohormones that play a critical role in plant physiology and development but also act as the primary germination signals for parasitic weed seeds. In this study, we generated CRISPR/Cas9 tomato knock-out (KO) lines targeting the SlD27 gene and three other key genes involved in SL biosynthesis (SlCCD7, SlCCD8, SlMAX1), all in the same genetic background. All edited lines exhibited undetectable SL levels in root exudates, leading to a strong reduction in the in vitro germination of Phelipanche spp. seeds. Consistently, reduced parasitism was also observed in vivo when Sld27 lines were tested. A comprehensive evaluation of morphological, reproductive, and fruit-related traits revealed gene-specific phenotypic effects, including changes in vegetative growth, fruit set, fruit development, and volatilome. Specifically, knock-out of two CCDs and the SlMAX1 affected shoot architecture, fruit development, and the production of volatile organic compounds during fruit ripening. In contrast, the Sld27 lines, newly developed in this study, displayed a mild phenotype generally comparable to non-edited control plants and likely due to the expression of SlD27 paralogues. Overall, our results indicate that SlD27 represents a promising breeding target for enhancing resistance to parasitic weeds in tomato while minimizing negative impacts on plant development and fruit quality.