The Arabidopsis dwarf1 mutant is defective in the conversion of 24-methylenecholesterol to campesterol in brassinosteroid biosynthesis

Since the isolation and characterization of dwarf1-1 (dwf1-1) from a T-DNA insertion mutant population, phenotypically similar mutants, including deet iolated2 (det2), constitutive photomorphogenesis and dwarfism (cpd), brassi nosteroid insensitive1 (bri1), and dwf4, have been reported to be defective in either the biosynthesis or the perception of brassinosteroids. We prese nt further characterization of dwf1-1 and additional dwf1 alleles. Feeding tests with brassinosteroid-biosynthetic intermediates revealed that dwf1 ca n be rescued by 22 alpha-hydroxycampesterol and downstream intermediates in the brassinosteroid pathway. Analysis of the endogenous levels of brassino steroid intermediates showed that 24-methylenecholesterol in dwf1 accumulat es to 12 times the level of the wild type, whereas the level of campesterol is greatly diminished, indicating that the defective step is in C-24 reduc tion. Furthermore, the deduced amino acid sequence of DWF1 shows significan t similarity to a flavin adenine dinucleotide-binding domain conserved in v arious oxidoreductases, suggesting an enzymatic role for DWF1. In support o f this, 7 of 10 dwf1 mutations directly affected the flavin adenine dinucle otide-binding domain. Our molecular characterization of dwf1 alleles, toget her with our biochemical data, suggest that the biosynthetic defect in dwf1 results in reduced synthesis of bioactive brassinosteroids, causing dwarfi sm.