Influence of the SMT2 knock-out on hypocotyl elongation in Arabidopsis thaliana

Abstract

Inhibitors are very important in the study of hormone function. Brasinazole (Brz) is a specific inhibitor of brassinosteroids (BRs) biosynthesis. To expand our knowledge of the molecular mechanisms of plant steroid signaling, we performed genetic screening using medium containing Brz under dark conditions. Mutants insensitive to Brz develop longer hypocotyls than their wild type counterparts. We isolated abz453 as a Brz insensitive mutant. TAIL-PCR and the segregation ratio of T2 plants indicated a single T-DNA insertion at the 24-Sterol C-methyltransferase (SMT2) gene in the abz453 mutant. Recapitulation for putative FCP serine phosphatase (FSP), the gene neighboring SMT2, indicated no significant phenotypes, but the SMT2 anti-sense (SMT2-AS) line developed longer hypocotyls than the wild type in medium containing Brz. Additionally, the SMT2-AS line displayed similar phenotypes to the abz453 line in soil including enhanced growth and smaller silique. The abz453 and SMT2-AS mutants showed phenotypes similar to those of wild type in medium containing benzylaminopurine, pacrobutrazol and ACC (precursor for ethylene) under dark conditions. However, when brassinolide (BL) dose response was observed, the abz453 and SMT2-AS lines showed higher sensitivity than wild type. The abz453/det2 and abz453/bri1-119 double mutants showed enhanced growth compared to the det2 and bri1-119 line under both dark and light conditions. Specially, in dark conditions double mutants displayed nearly 2- and 1.5-fold longer hypocotyls than det2 and bri1-119 plants. Brz insensitivity to the SMT2 knock-out mutant and phenotypes of double mutants indicate that not only do BRI1 and DET2 influence the BRs response, as evidenced by hypocotyl elongation, but another sterol derived signals may also be affected in mutants, suggesting that another pathway is involved in hypocotyl elongation due to SMT2. (c) KSBB.