Fenarimol, a Pyrimidine-Type Fungicide, Inhibits Brassinosteroid Biosynthesis

The plant steroid hormone brassinosteroids (BRs) are important signal mediators that regulate broad aspects of plant growth and development. With the discovery of brassinoazole (Brz), the first specific inhibitor of BR biosynthesis, several triazole-type BR biosynthesis inhibitors have been developed. In this article, we report that fenarimol (FM), a pyrimidine-type fungicide, exhibits potent inhibitory activity against BR biosynthesis. FM induces dwarfism and the open cotyledon phenotype of Arabidopsis seedlings in the dark. The IC50 value for FM to inhibit stem elongation of Arabidopsis seedlings grown in the dark was approximately 1.8 ± 0.2 μM. FM-induced dwarfism of Arabidopsis seedlings could be restored by brassinolide (BL) but not by gibberellin (GA). Assessment of the target site of FM in BR biosynthesis by feeding BR biosynthesis intermediates indicated that FM interferes with the side chain hydroxylation of BR biosynthesis from campestanol to teasterone. Determination of the binding affinity of FM to purified recombinant CYP90D1 indicated that FM induced a typical type II binding spectrum with a Kd value of approximately 0.79 μM. Quantitative real-time PCR analysis of the expression level of the BR responsive gene in Arabidopsis seedlings indicated that FM induces the BR deficiency in Arabidopsis

YCZ-18 is a new brassinosteroid biosynthesis inhibitor.

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

Clear cell adenocarcinoma of the colon: A case report and review of literature

A primary clear cell adenocarcinoma of the colon is a rare oncologic entity. We herein report a case of such a tumor of the sigmoid colon in a 71-year-old woman who was successfully treated by an endoscopic polypectomy in our hospital. We also reviewed the published reports regarding cases of primary clear cell tumors in the colon

YCZ-18-treated plants display the BR-deficient phenotype.

<p><b>YCZ-18</b>-treated plants (0.3, 1, 3 μM), Brz220-treated plants (3 μM) and brassinosteroid-deficient mutant (<i>det2</i>) plants were grown for 6 days in the dark (A) and for 10 days in the light (B-G) on medium containing the chemical indicated. The control plants (Cont) were untreated. Scale bar = 5 mm.</p

YCZ18-treated Arabidopsis in response to cathasterone (CT) and teasterone (TE).

<p>Five-day-old <i>Arabidopsis</i> seedlings (8A, 8E, white bar), treated with 0.5 μM <b>YCZ-18</b> (8B, 8E, yellow bar), treated with 0.5 μM <b>YCZ-18</b> together with 30 μM cathasterone (CT) (8C, 8E, red bar), or treated with 0.5 μM <b>YCZ-18</b> together with 10 μM teasterone (TE) (8D, 8E, blue bar). Data are the means ± s.e. obtained from 30 seedlings. Scale bar = 3 mm.</p

Effect of YCZ-18 on the growth of Arabidopsis in soil.

<p>The application of <b>YCZ-18</b> on wild-type <i>Arabidopsis</i> was performed by spraying an aqueous solution of <b>YCZ-18</b> (5 μM) onto ten-day-old wild-type <i>Arabidopsis</i> plants (approximately 0.2 pmol/plant), as indicated in the methods section. Four-week-old <i>Arabidopsis</i> seedlings (A), four-week-old <i>Arabidopsis</i> treated with <b>YCZ-18</b> (B), six-week-old <i>Arabidopsis</i> (C), six-week-old <i>Arabidopsis</i> seedlings treated with <b>YCZ-18</b> (D), rosette leaf number of six-week-old <i>Arabidopsis</i> at bolting from three plants (E). Data are the means ± s.e. obtained from 3 plants. Scale bar = 1 cm.</p

Binding of YCZ-18 to CYP90D1.

<p>Absorption spectra of oxidized CYP90D1 (blue line) and its <b>YCZ-18</b> complex (pink line). Recombinant CYP90D1 (3.5 μM) was dissolved in 50 mM NaH₂PO₄ (pH 7.0) with 0.1% Tween 20 containing 20% glycerol, and <b>YCZ-18</b> was added to CYP90D1 at a final concentration of 16 μM (A). Spectrophotometric titration of CYP90D1 with <b>YCZ-18</b> induced spectral changes in CYP90D1. <b>YCZ-18</b> was added to CYP90D1 (3.5 μM) at various final concentrations (a, 0.7; b, 1; c, 2; d, 4; e, 8; f, 12; g, 16 μM) (B). The spectral dissociation constant was calculated from a double reciprocal plot of absorbance differences, ΔA (436–415 nm) versus the <b>YCZ-18</b> concentrations given 0.79 μm (C). The experiment was duplicated to establish reproducibility.</p

YCZ-18 regulates the expression of BR-responsive genes.

<p>Quantitative RT-PCR experiment measuring the relative expression levels of a BR-upregulated gene (<i>TCH4</i>) (A), a BR biosynthetic gene (<i>DWF4</i>) (B) and two photosynthesis genes (<i>LHCP</i> (C) and <i>rbcS</i> (D)) of the wild-type plant (Cont), <b>YCZ-18</b>-treated (0.3, 1, 3 μM), Brz-treated (3 μM) and brassinosteroid-deficient mutant <i>det2</i>. Plants were grown for 10 days in the light (A, B) and for 6 days in the dark (C, D) on a medium containing the chemical indicated. All results are means ± s.e.</p