YCZ-18 Is a New Brassinosteroid Biosynthesis Inhibitor

Abstract

Plant hormone brassinosteroids (BRs) are a group of polyhydroxylated steroids that play critical roles in regulating broad aspects of plant growth and development. The structural diversity of BRs is generated by the action of several groups of P450s. Brassinazole is a specific inhibitor of C-22 hydroxylase (CYP90B1) in BR biosynthesis, and the application use of brassinazole has emerged as an effective way of complementing BR-deficient mutants to elucidate the functions of BRs. In this article, we report a new triazole-type BR biosynthesis inhibitor, YCZ-18. Quantitative analysis the endogenous levels of BRs in Arabidopsis indicated that YCZ-18 significantly decreased the BR contents in plant tissues. Assessment of the binding affinity of YCZ-18to purified recombinant CYP90D1 indicated that YCZ-18 induced a typical type II binding spectrum with a Kd value of approximately 0.79 μM. Analysis of the mechanisms underlying the dwarf phenotype associated with YCZ-18 treatment of Arabidopsis indicated that the chemically induced dwarf phenotype was caused by a failure of cell elongation. Moreover, dissecting the effect of YCZ-18 on the induction or down regulation of genes responsive to BRs indicated that YCZ-18 regulated the expression of genes responsible for BRs deficiency in Arabidopsis. These findings indicate that YCZ-18 is a potent BR biosynthesis inhibitor and has a new target site, C23-hydroxylation in BR biosynthesis. Application of YCZ-18 will be a good starting point for further elucidation of the detailed mechanism of BR biosynthesis and its regulation.

In vitro and in vivo evidence for the inhibition of brassinosteroid synthesis by propiconazole through interference with side chain hydroxylation

“…33 Protein measurements were performed using a Protein Assay Kit (Bio-Rad, Hercules, CA, USA) and bovine serum albumin as a standard. The relative purity of recombinant CYP90D1 was estimated by SDS-polyacrylamide gel electrophoresis (12% polyacrylamide) and gel staining was performed using Coomassie brilliant blue R250.…”

“…33 The data obtained were used to calculate binding constants based on linear regression analysis. Spectral determinations were performed at least twice for each experiment, confirming the reproducibility with respect to the spectral profile and position of λmax and λmin.…”

“…Another assay system, which is based on the use of BR biosynthesis intermediators, can be used to determine the inhibitory effects of small molecules in BR biosynthesis in vivo. 33 With these experimental system, we resently demonstrated that fenarimol, a pyrimiding type funjicide, inhibits BR biosynthesis. 34 Currently, numerous triazole compounds have been shown to inhibit P450s.…”

“…1), demonstrated that yucaizol is a specific inhibitor of BR biosynthesis that binds to purified recombinant CYP90D1. 33 In the course of these works, we established several assay systems that can be used to determine the target site of small molecules in BR biosynthesis. One experiment system which is based on using purified recombinant protein of CYP90D1 allows us to conduct in vitro evaluation the inhibitory activity of small molecules against BR biosynthesis.…”

Recent advances in brassinosteroid biosynthetic pathway: insight into novel brassinosteroid shortcut pathway

“…It has been recently reported that new triazole-type inhibitors based on a ketoconazole scaffold (YCZ18) inhibit a new target, CYP90D1, in Arabidopsis. 38) Aside from BR synthesis inhibitors, BR antagonists have recently been developed. BL binds to BRI1 and functions as its most potent agonist.…”