Time course of endogenous nitric oxide (NO) generation induced by SNP, hematin and CO aqueous solution in wheat seedling leaves.

<p>Before starting the experiments, 14-day-old wheat seedlings cultured in the Hoagland solution were kept in the light (L, 300 μmmol m⁻²s⁻¹) or dark (D) for 5 days (25°C). Afterwards, seedlings were cultured in the Hoagland solution without or with 100 μΜ SNP (S), 10 μM HO-1 inducer hematin (H), and 1.0% CO aqueous solution (CO), in the light (L) or dark (D) for another 3 days. NO contents were detected by using Greiss reagent. Values were the mean ± SE for at least three independent experiments. Bars denoted by the same letter did not significantly differ at <i>P</i><0.05 according to Duncan's multiple range tests.</p

HO-1 inducer hematin, exogenous CO and light treatments induce <i>HO-1</i> gene expression in wheat seedling leaves after dark pretreatment.

<p>Before starting the experiments, 14-day-old wheat seedlings cultured in the Hoagland solution were kept in the light (L, 300 μmmol m⁻²s⁻¹) or dark (D) for 5 days. Afterwards, seedlings were cultured in the Hoagland solution without (D→D) or with HO-1 inducer hematin (H, 10 μM), and 1.0% CO-saturated aqueous solution (CO) for another 12 h. <i>HO-1</i> mRNA expression was analyzed by quantitative real-time RT-PCR as described in Materials and Methods. Three independent experiments were performed, bars denoted by the same letter did not significantly differ at <i>P<</i>0.05 according to Duncan's multiple range tests.</p

Comparative effects of HO-1 inducer hematin, CO-saturated aqueous solution, and light on the restoration of chlorophyll content in etiolated wheat seedling leaves.

<p>14-day-old wheat seedlings were kept at 25°C in continuous darkness for 5 days. After that, some were transferred into light, while others were still left in darkness without (D→D) or with different contentions of hematin (1.0, 10, and 100 μM, D→D+H1.0, 10, 100), and different saturations of CO aqueous solution (0.1, 1.0, 10, and 50%, D→D+CO0.1, 1.0, 10, 50%). L→L+ZnPPIX, D→D+ZnPPIX, and D→L+ZnPPIX D→H10+Hb stand for combination with HO-1 specific inhibitor ZnPPIX (100 μM), and CO/NO scavenger hemoglobin (Hb, 0.1 gL⁻¹), for additional 3 days. L→L stands for wheat seedlings were grown in normal light cycle. Bars denoted by the same letter did not significantly differ at <i>P</i><0.05 according to Duncan's multiple range tests.</p

Time course of chlorophyll accumulation following incubation in HO-1 inducer hematin in wheat seedling leaves.

<p>Before starting the experiments, 14-day-old wheat seedlings cultured in the Hoagland solution were kept in the light (L, 300 μmmol m⁻²s⁻¹) or dark for 5 days. Afterwards, seedlings were cultured in the Hoagland solution with or without 10 μM HO-1 inducer hematin (H) treatment, in the dark (D) for another 5 days. L→L stands for wheat seedlings were grown in normal light cycle. Chlorophyll was extracted and quantified at various times. Bars denoted by the same letter did not significantly differ at <i>P<</i>0.05 according to Duncan's multiple range tests.</p

Time course of <i>HO-1</i> gene expression in wheat seedling leaves during transition from dark to light for additional hours.

<p>A, <i>HO-1</i> mRNA expression was analyzed by quantitative real-time RT-PCR as described in Materials and Methods. B, CO release under different treatments at 12 h. Values were the mean ± SE of three independent experiments. Bars denoted by the different letter differed significantly at <i>P<</i>0.05 according to Duncan's multiple range tests.</p

Effects of NO donor SNP, hematin and CO-saturated aqueous solution on HO activities in wheat seedling leaves after dark pretreatment.

<p>Before starting the experiments, 14-day-old wheat seedlings cultured in the Hoagland solution were kept in the light (L, 300 μmmol m⁻²s⁻¹) or dark (D) for 5 days. Afterwards, seedlings were cultured in the Hoagland solution without (D→D) or with SNP (S, 100 μM), HO-1 inducer hematin (H, 10 μM), 1.0% CO-saturated aqueous solution (CO) in completely darkness for another 3 days. Values were the mean ± SE for at least three independent experiments. Bars denoted by the same letter did not significantly differ at <i>P<</i>0.05 according to Duncan's multiple range tests.</p

Effects of treatment with SNP, hematin, CO-saturated aqueous solution, ZnPPIX, and cPTIO on Pfr contents (3 d) and expression profiles of <i>PHYA</i> expression (2 d).

<p>14-day-old wheat seedlings were grown for 5 days at 25°C in continuous darkness (D) or light (L) at 300 μmmol m⁻²s⁻¹ before either being transferred into light or left in continuous darkness without or with 100 μM SNP, 10 μM hematin, 1.0% CO aqueous solution, 100 μM HO-1 specific inhibitor ZnPPIX, and 100 μM cPTIO, or their combination treatments, for another 3 days. <i>PHYA</i> mRNA expression was analyzed by quantitative real-time RT-PCR as described in Materials and Methods. Values were the mean ± SE for at least three independent experiments (n = 20). Bars denoted by the same letter did not significantly differ at <i>P<</i>0.05 according to Duncan's multiple tests.</p

NO accumulation in etiolated distilled water was taken as control.

<p>The distribution of nitric oxide (NO) in wheat seedling leaves induced by SNP, hematin, and CO aqueous solution in wheat seedling leaves. Before starting the experiments, 14-day-old wheat seedlings cultured in the Hoagland solution were kept in the light (L, 300 μmmol m⁻²s⁻¹) or dark (D) for 5 days. Afterwards, seedlings were cultured in the Hoagland solution without or with 100 μΜ SNP (S), 10 μM HO-1 inducer hematin (H), and 1.0% CO aqueous solution (CO), in the light (L) or dark (D) for another 2 days. A, The NO distribution was detected by fluorescence probe DAF-2 DA and negative probe AF 4-DA 2 days after different treatments under fluorescence microscopy (TCS-SP2 confocal laser scanning microscope; Leica Lasertechnik GmbH). B, Mean relative DAF-2 DA and AF 4-DA fluorescence densities corresponding to samples without cPTIO treatment was given. Values were the mean ± SE for at least three independent experiments. Bars denoted by the same letter did not significantly differ at <i>P</i><0.05 according to Duncan's multiple range tests. Bars = 20 μm.</p

SNP, hematin, CO-saturated aqueous solution, CO scavenger hemoglobin, mammalian NOS-like inhibitor L-NAME, and the NO specific scavenger cPTIO differentially influence the chlorophyll content in etiolated wheat seedling leaves after dark pretreatment.

<p>Before starting the experiments, 14-day-old wheat seedlings cultured in the Hoagland solution were kept in the light (L, 300 μmmol m⁻²s⁻¹) or dark (D) for 5 days. Afterwards, seedlings were cultured in the Hoagland solution without or with 100 μΜ SNP (S), 10 μM HO-1 inducer hematin (H), 1.0% CO aqueous solution (CO), 0.1 g L⁻¹ Hb, 200 μM L-NAME, 100 μM cPTIO, or the above combination treatments, in the light (L) or dark (D) for another 3 days. Values were the mean ± SE for at least three independent experiments. Bars denoted by the same letter did not significantly differ at <i>P</i><0.05 according to Duncan's multiple range tests.</p

Additional file 1: Table S1. of Hydrogen peroxide is involved in hydrogen sulfide-induced lateral root formation in tomato seedlings

The accession numbers and primer sequences of real-time RT-PCR (qPCR). (DOC 45 kb