Seed germination is a complicated biological process that requires regulated enzymatic and non-enzymatic reactions. The action of polyamine oxidase (PAO) produces hydrogen peroxide (H2O2), which promotes dicot seed germination. However, whether and, if so, how PAOs regulate monocot seed germination via H2O2 production is unclear. Herein, we report that the coleorhiza is the main physical barrier to radicle protrusion during germination of rice seed (a monocot seed) and that it does so in a manner similar to that of dicot seed micropylar endosperm. We found that H2O2 specifically and steadily accumulated in the coleorhizae and radicles of germinating rice seeds and was accompanied by increased PAO activity as the germination percentage increased. These physiological indexes were strongly decreased in number by guazatine, a PAO inhibitor. We also identified 11 PAO homologs (OsPAO1–11) in the rice genome, which could be classified into four subfamilies (I, IIa, IIb, and III). The OsPAO genes in subfamilies I, IIa, and IIb (OsPAO1–7) encode PAOs, whereas those in subfamily III (OsPAO8–11) encode histone lysine-specific demethylases. In silico-characterized expression profiles of OsPAO1–7 and those determined by qPCR revealed that OsPAO5 is markedly upregulated in imbibed seeds compared with dry seeds and that its transcript accumulated to a higher level in embryos than in the endosperm. Moreover, its transcriptional abundance increased gradually during seed germination in water and was inhibited by 5 mM guazatine. Taken together, these results suggest that PAO-generated H2O2 is involved in coleorhiza-limited rice seed germination and that OsPAO5 expression accounts for most PAO expression and activity during rice seed germination. These findings should facilitate further study of PAOs and provide valuable information for functional validation of these proteins during seed germination of monocot cereals.

In the present study, we evaluated the protective effect of exogenous heme oxygenase-1 (HO-1 EC 1.14.99.3) inducer hematin against mercury-induced oxidative damage in the roots of Alfalfa (Medicago sativa L.). Plants exposed to mercury (HgCl 2 ) exhibited a significant increase of lipid peroxidation, as well as inhibition of root elongation. However, hematin (50 µM) supplementation to HgCl 2 (100 µmol/L) treated plants effectively reduced the lipid peroxidation and partially increased the root elongation. These responses were mimicked by the application of aqueous solution of carbon monoxide (CO) with 50% saturation. Also, treatment with hematin could result in the potent induction of HO-1 transcript in the root tissues, as detected 12 h following treatment. Moreover, the activation of anti-oxidant enzyme, including glutathione reductase, monodehydroascorbate reductase and superoxide dismutase activities, and the decrease of lipoxygenase activity, were induced by hematin at 12 h or 24 h, which was further confirmed by histochemical staining for the detection of lipid peroxidation and loss of membrane integrity. Whereas, ascorbate peroxidase and guaiacol peroxidase isozyme activities or their transcripts were reduced, respectively, indicating that hydrogen peroxide might act as a signal to mediate Hgtolerance at the beginning of treatment. The ameliorating effects of hematin were specific, since the CO scavenger hemoglobin differentially reversed the above actions. Taken together, our results suggested that hematin exhibits a vital role in protecting the plant against Hg-induced oxidative damage.

Recent studies suggest that carbon monoxide (CO), which is mainly produced by heme oxygenase (HO EC 1.14.99.3), may function as a physiological messenger or bioactive molecule by interacting with nitric oxide (NO) in animal cells. In this study, we report that application of the hematin and hemin, two heme molecules cleaved by HO to yield CO in animals, dose-dependently induced the significant increase in wheat root elongation as well as the actions of IAA and NO donor sodium nitroprusside (SNP). These responses were mimicked by the application of aqueous solution of CO with different saturation. Also, above heme molecule-induced effect is specific for CO since the potent inhibitor of HO-1, zinc protoporphyrin-IX (ZnPPIX) or CO scavenger hemoglobin (Hb) blocked the action of hematin and hemin, respectively. Further results proved that treatment with hematin or IAA could result in either the potent induction of HO-1 transcript or CO releasing in wheat root segments, both of which were reversed by the addition of ZnPPIX. ZnPPIX with lower concentration could prevent the elongation induced by IAA, while in the SNP-treatment the prevention of root growth occurred solely at higher concentrations. Also, wheat root segments elongation induced by IAA, SNP or hematin, was blocked by the specific NO scavenger, inhibitors of NO synthase (NOS) and guanylate cyclase (GC), respectively. Meanwhile, production of reactive oxygen species (ROS) could be demonstrated in the growing zone of wheat root segments treated by hematin or SNP using specific histochemical assay combined with the inhibitor investigation. Taken together, above results suggested that CO produced by HO might mediate the induction of growth elongation of wheat root segments by IAA, which might be also related to NO/cGMPand even ROS-dependent pathways.

The aim of this study was to investigate whether presoaking with hemin, an inducer of heme oxygenase-1 (HO-1), could alleviate salinity damage during wheat seed germination in comparison with the pretreatment of a well-known nitric oxide (NO) donor sodium nitroprusside (SNP). The results showed that, compared with the samples upon 150 mM NaCl salt stress alone, both 10 lM hemin and 200 lM SNP pretreatments could (1) significantly attenuate the inhibition of seed germination and thereafter seedling growth; (2) induce HO expression; (3) enhance amylase activity, thus accelerating the formation of reducing sugar and total soluble sugar; and (4) increase the potassium (K) to sodium (Na) ratio, particularly in the shoot parts. Hemin and SNP could also increase antioxidant enzyme activities, including superoxide dismutase (SOD), catalase (CAT), guaiacol peroxidase (POD), and ascorbate peroxidase (APX), thus resulting in the alleviation of oxidative damage, as indicated by the decrease of thiobarbituric acid reactive substances (TBARS) content.Moreover, semi-quantitative RT-PCR and isozymatic analysis illustrated that hemin and SNP pretreatment were able to up-regulate the expression of Mn-SOD (especially) and Cu/Zn-SOD gene, and activate SOD isozymatic activities. Since the addition of the NO scavenger methylene blue (MB) differentially reversed the above effects, the protective roles of hemin might be related to the induction of endogenous NO signal. Meanwhile, hemindriven NO production was confirmed. Together, these results indicated that hemin exerted an advantageous effect on enhancing salinity tolerance during wheat seed germination, which might interact with NO.