Brassinosteroid Overaccumulation Differentially Modulates Arabidopsis Hypocotyl Growth in the Light and in the Dark

Background: Brassinosteroids (BRs) are plant hormones that are critical for seedling growth. In Arabidopsis thaliana (L.) Heynh., BRs stimulate hypocotyl elongation in the light but inhibit it in darkness, whereas the BR biosynthesis inhibitor brassinazole (BRZ) suppresses hypocotyl growth under both conditions. Our earlier work using BR-deficient mutants suggested that endogenous BR levels are suboptimal for hypocotyl elongation in light-grown seedlings but nearly optimal in darkness. However, this model has not been validated using BR-overaccumulating lines. Methods: To increase endogenous BR levels, we used previously generated single, double, and triple knockout mutants of three well-characterized BR-inactivating genes (BEN1, BAS1, and SOB7). These mutants accumulate BRs in an additive manner. We examined their hypocotyl responses to exogenous brassinolide (BL) and BRZ under light and dark conditions, and analyzed the expression of selected BR- and light-responsive genes. Results: BR overaccumulation led to longer hypocotyls in the light but shorter hypocotyls in the dark. In the light, BL consistently promoted hypocotyl elongation, whereas, BRZ consistently inhibited it, in both wild-type and BR-overaccumulating seedlings. In darkness, both BL and BRZ suppressed hypocotyl growth across all genotypes at moderate and high concentrations. Notably, at high BRZ concentrations, BR-overaccumulating mutants developed longer hypocotyls than wild-type seedlings under the same dark conditions. Comparisons of relative hypocotyl length (% of untreated control) showed that BR overaccumulation increased hypocotyl responsiveness to high-dose BRZ in the light but conferred partial resistance to BRZ in the dark. Conclusions: These results support and refine our previous model of BR–light/dark interactions. BR overaccumulation enhances hypocotyl growth in the light, whereas excess BRs inhibit elongation in darkness. Gene expression analyses of wild-type and BR-overaccumulating seedlings identified GATA Transcription Factor 2 (GATA2) as a candidate integrator of BR and light signaling during hypocotyl development.