Conservation of function between mammalian and plant steroid 5α-reductases

Abstract

Arabidopsis det2 mutants are small darkgreen dwarfs displaying pleiotropic defects in light-regulated development during multiple stages of the plant life cycle. The DET2 gene encodes a protein that shares Ϸ40% sequence identity with mammalian steroid 5␣-reductases and is implicated in the synthesis of a class of plant steroids, the brassinosteroids. Here we show that the DET2 protein, when expressed in human embryonic kidney 293 cells, catalyzes the 5␣-reduction of several animal steroid substrates and has similar kinetic properties to the mammalian steroid 5␣-reductase enzymes. Moreover, human steroid 5␣-reductases expressed in det2 mutant plants can substitute for DET2 in brassinosteroid biosynthesis. These data indicate that DET2 is an ortholog of the mammalian steroid 5␣-reductases and provide further evidence that brassinosteroids play an essential role in light-regulated plant development. The structural and functional conservation between DET2 and human steroid 5␣-reductases raise interesting issues concerning the evolutionary origin of the steroid hormone signaling system. Steroid hormones play important roles in mammalian development and homeostasis (1). While many steroids have been identified in plants (2), to date only the brassinosteroids (BRs) are widely distributed throughout the plant kingdom and elicit unique growth promoting activity when applied exogenously (3). Despite these properties, the physiological role of endogenous BRs in plant development has been unclear. We recently isolated the Arabidopsis DET2 gene by positional cloning (4) and found that DET2 encodes a 262-amino acid protein that shares significant sequence identity with mammalian steroid 5␣-reductases, enzymes that play crucial roles in steroid hormone metabolism and action by catalyzing an NADPHdependent reduction of the ⌬ 4,5 double bond in a variety of steroids (Fig. 1A) (5). Mutations in DET2, including a missense mutation at residue Glu-204, which is absolutely required for human steroid 5␣-reductase (hS5R) activity (5), lead to many defects in light-dependent Arabidopsis development that can be ameliorated by application of brassinolide, an active BR (4, 6). Based on these results, we proposed that DET2 encodes a 5␣-reductase in a BR biosynthetic pathway (Fig. 1B) and that BRs constitute an important class of plant hormones (4).Support for the postulated importance of BRs in plant development came from the identification of a second Arabidopsis gene, CPD (constitutive photomorphogenesis and dwarfism) (7). The CPD gene encodes a protein that shares sequence identity with several mammalian cytochrome P450 proteins including steroid hydroxylases. Mutations in CPD cause phenotypic defects that are similar to those of det2 mutations. Moreover, brassinolide treatment also restores a wild-type phenotype to cpd mutants. Rescue studies with intermediates in the BR biosynthetic pathway suggest that CPD may encode a steroid 23-hydroxylase (7).In mammals, steroid hormones are synthesized from cholesterol via pregnenolone thr...

CYP90A1/CPD, a Brassinosteroid Biosynthetic Cytochrome P450 of Arabidopsis, Catalyzes C-3 Oxidation

“…Substrate Specificity of DET2-Previous in vitro assays demonstrated that DET2 catalyzes 5␣ reduction of 4-en-3-one to 3-one (7). In vivo feeding of labeled CR has also suggested a role for this enzyme in the conversion of 22-OH-4-en-3-one to 22-OH-3-one, a reaction in the early C-22 oxidation branch of BR biosynthesis (14).…”

Arabidopsis Actin-Depolymerizing Factor AtADF4 Mediates Defense Signal Transduction Triggered by thePseudomonas syringaeEffector AvrPphB

“…To construct pMD1-gAtADF4, a genomic DNA fragment starting from 665 bp upstream of the AtADF4 start codon (e.g. ATG) to the stop codon TAA, was cloned into the binary vector pMD1 (Li et al, 1997), which was digested with HindIII and XbaI to remove 35S promoter. Primers 5#-GCGAAGCTTacatcttgtcttcacataatgaaaac-3# (forward) and 5#-GCGTCTAGAttaACCCATTTGTTGACCACCTGTCATTGAAGCCATgttgacgcggcttttcaaaacatcaagatcc-3# (reverse) were used to amplify this fragment.…”

Caenorhabditis elegans LET-767 is able to metabolize androgens and estrogens and likely shares common ancestor with human types 3 and 12 17β-hydroxysteroid dehydrogenases

“…It is noteworthy that the human type 2 5a-reductase is also able to convert campesterol into campestanol and it is identified as ortholog of det2 in plant (Li et al 1997). The similarity between the two cases, LET-767/17b-HSD12 and det2/5a-reductase, suggests that LET-767 is involved in the biosynthesis of the active steroid in C. elegans that has a structure different from sex steroids in mammals but it could play an equivalent role.…”

“…Similar to LET-767 that is able to metabolize vertebrate hormone androgen and estrogen, it has been shown that the plant enzyme responsive for det2 mutation, which causes small dark green dwarfs to display pleiotropic defects in lightregulated development during multiple stages of the Arabidopsis plant life cycle, is also able to convert vertebrate steroid hormone testosterone into dihydrotestosterone although its natural plant steroid hormone is campestanol, a brassinosteroid (Li et al 1997). It is noteworthy that the human type 2 5a-reductase is also able to convert campesterol into campestanol and it is identified as ortholog of det2 in plant (Li et al 1997).…”