OsHK3 is a crucial regulator of abscisic acid signaling involved in antioxidant defense in rice.

In this study, the role of the rice (Oryza sativa L.) histidine kinase OsHK3 in abscisic acid (ABA)-induced antioxidant defense was investigated. Treatments with ABA, H2 O2 , and polyethylene glycol (PEG) induced the expression of OsHK3 in rice leaves, and H2 O2 is required for ABA-induced increase in the expression of OsHK3 under water stress. Subcellular localization analysis showed that OsHK3 is located in the cytoplasm and the plasma membrane. The transient expression analysis and the transient RNA interference test in rice protoplasts showed that OsHK3 is required for ABA-induced upregulation in the expression of antioxidant enzymes genes and the activities of antioxidant enzymes. Further analysis showed that OsHK3 functions upstream of the calcium/calmodulin-dependent protein kinase OsDMI3 and the mitogen-activated protein kinase OsMPK1 to regulate the activities of antioxidant enzymes in ABA signaling. Moreover, OsHK3 was also shown to regulate the expression of nicotinamide adenine dinucleotide phosphate oxidase genes, OsrbohB and OsrbohE, and the production of H2 O2 in ABA signaling. Our data indicate that OsHK3 play an important role in the regulation of ABA-induced antioxidant defense and in the feedback regulation of H2 O2 production in ABA signaling.

OsDMI3 is a novel component of abscisic acid signaling in the induction of antioxidant defense in leaves of rice.

Ca(2+) and calmodulin (CaM) have been shown to play an important role in abscisic acid (ABA)-induced antioxidant defense. However, it is unknown whether Ca(2+)/CaM-dependent protein kinase (CCaMK) is involved in the process. In the present study, the role of rice CCaMK, OsDMI3, in ABA-induced antioxidant defense was investigated in leaves of rice (Oryza sativa) plants. Treatments with ABA, H(2)O(2), and polyethylene glycol (PEG) induced the expression of OsDMI3 and the activity of OsDMI3, and H(2)O(2) is required for the ABA-induced increases in the expression and the activity of OsDMI3 under water stress. Subcellular localization analysis showed that OsDMI3 is located in the nucleus, the cytoplasm, and the plasma membrane. The analysis of the transient expression of OsDMI3 in rice protoplasts and the RNA interference (RNAi) silencing of OsDMI3 in rice protoplasts showed that OsDMI3 is required for ABA-induced increases in the expression and the activities of superoxide dismutase (SOD) and catalase (CAT). Further, the oxidative damage induced by higher concentrations of PEG and H(2)O(2) was aggravated in the mutant of OsDMI3. Moreover, the analysis of the RNAi silencing of OsDMI3 in protoplasts and the mutant of OsDMI3 showed that higher levels of H(2)O(2) accumulation require OsDMI3 activation in ABA signaling, but the initial H(2)O(2) production induced by ABA is not dependent on the activation of OsDMI3 in leaves of rice plants. Our data reveal that OsDMI3 is an important component in ABA-induced antioxidant defense in rice.

Analysis of DNA methylation of maize in response to osmotic and salt stress based on methylation-sensitive amplified polymorphism.

Water stress is known to alter cytosine methylation, which generally represses transcription. However, little is known about the role of methylation alteration in maize under osmotic stress. Here, methylation-sensitive amplified polymorphism (MSAP) was used to screen PEG- or NaCl-induced methylation alteration in maize seedlings. The sequences of 25 differentially amplified fragments relevant to stress were successfully obtained. Two stress-specific fragments from leaves, LP166 and LPS911, shown to be homologous to retrotransposon Gag-Pol protein genes, suggested that osmotic stress-induced methylation of retrotransposons. Three MSAP fragments, representing drought-induced or salt-induced methylation in leaves, were homologous to a maize aluminum-induced transporter. Besides these, heat shock protein HSP82, Poly [ADP-ribose] polymerase 2, Lipoxygenase, casein kinase (CK2), and dehydration-responsive element-binding (DREB) factor were also homologs of MSAP sequences from salt-treated roots. One MSAP fragment amplified from salt-treated roots, designated RS39, was homologous to the first intron of maize protein phosphatase 2C (zmPP2C), whereas - LS103, absent from salt-treated leaves, was homologous to maize glutathione S-transferases (zmGST). Expression analysis showed that salt-induced intron methylation of root zmPP2C significantly downregulated its expression, while salt-induced demethylation of leaf zmGST weakly upregulated its expression. The results suggested that salinity-induced methylation downregulated zmPP2C expression, a negative regulator of the stress response, while salinity-induced demethylation upregulated zmGST expression, a positive effecter of the stress response. Altered methylation, in response to stress, might also be involved in stress acclimation.