Overproduction of the Membrane-bound Receptor-like Protein Kinase 1, RPK1, Enhances Abiotic Stress Tolerance in Arabidopsis

RPK1 (receptor-like protein kinase 1) localizes to the plasma membrane and functions as a regulator of abscisic acid (ABA) signaling in Arabidopsis. In our current study, we investigated the effect of RPK1 disruption and overproduction upon plant responses to drought stress. Transgenic Arabidopsis overexpressing the RPK1 protein showed increased ABA sensitivity in their root growth and stomatal closure and also displayed less transpirational water loss. In contrast, a mutant lacking RPK1 function, rpk1-1, was found to be resistant to ABA during these processes and showed increased water loss. RPK1 overproduction in these transgenic plants thus increased their tolerance to drought stress. We performed microarray analysis of RPK1 transgenic plants and observed enhanced expression of several stress-responsive genes, such as Cor15a, Cor15b, and rd29A, in addition to H₂O₂-responsive genes. Consistently, the expression levels of ABA/stress-responsive genes in rpk1-1 had decreased compared with wild type. The results suggest that the overproduction of RPK1 enhances both the ABA and drought stress signaling pathways. Furthermore, the leaves of the rpk1-1 plants exhibit higher sensitivity to oxidative stress upon ABA-pretreatment, whereas transgenic plants overproducing RPK1 manifest increased tolerance to this stress. Our current data suggest therefore that RPK1 overproduction controls reactive oxygen species homeostasis and enhances both water and oxidative stress tolerance in Arabidopsis.     

<Emphasis Type="Italic">CKB1</Emphasis> is involved in abscisic acid and gibberellic acid signaling to regulate stress responses in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Casein kinase II (CK2), an evolutionarily well-conserved Ser/Thr kinase, plays critical roles in all higher organisms including plants. CKB1 is a regulatory subunit beta of CK2. In this study, homozygous T-DNA mutants (ckb1-1 and ckb1-2) and over-expression plants (35S:CKB1-1, 35S:CKB1-2) of Arabidopsis thaliana were studied to understand the role of CKB1 in abiotic stress and gibberellic acid (GA) signaling. Histochemical staining showed that although CKB1 was expressed in all organs, it had a relatively higher expression in conducting tissues. The ckb1 mutants showed reduced sensitivity to abscisic acid (ABA) during seed germination and seedling growth. The increased stomatal aperture, leaf water loss and proline accumulation were observed in ckb1 mutants. In contrast, the ckb1 mutant had increased sensitivity to polyaluminum chloride during seed germination and hypocotyl elongation. We obtained opposite results in over-expression plants. The expression levels of a number of genes in the ABA and GA regulatory network had changed. This study demonstrates that CKB1 is an ABA signaling-related gene, which subsequently influences GA metabolism, and may play a positive role in ABA signaling.     

Carbon dioxide enrichment alleviates heat stress by improving cellular redox homeostasis through an ABA‐independent process in tomato plants

Plant responses to elevated CO₂ and high temperature are critically regulated through a complex network of phytohormones and redox homeostasis. However, the involvement of abscisic acid (ABA) in plant adaptation to heat stress under elevated CO₂ conditions has not been thoroughly studied. This study investigated the interactive effects of elevated CO₂ (800 μmol·mol^?1) and heat stress (42 °C for 24 h) on the endogenous level of ABA and the cellular redox state of two genotypes of tomato with different ABA biosynthesis capacities. Heat stress significantly decreased maximum photochemical efficiency of PSII (Fv/Fm) and leaf water potential, but also increased levels of malondialdehyde (MDA) and electrolyte leakage (EL) in both genotypes. Heat‐induced damage was more severe in the ABA‐deficient mutant notabilis (not) than in its parental cultivar Ailsa Craig (Ailsa), suggesting that a certain level of endogenous ABA is required to minimise the heat‐induced oxidative damage to the photosynthetic apparatus. Irrespective of genotype, the enrichment of CO₂ remarkably stimulated Fv/Fm, MDA and EL in heat‐stressed plants towards enhanced tolerance. In addition, elevated CO₂ significantly strengthened the antioxidant capacity of heat‐stressed tomato seedlings towards a reduced cellular redox state for a prolonged period, thereby mitigating oxidative stress. However, elevated CO₂ and heat stress did not alter the endogenous level of ABA or the expression of its biosynthetic gene NCED2 in either genotype, indicating that ABA is not involved in elevated CO₂‐induced heat stress alleviation. The results of this study suggest that elevated CO₂ alleviated heat stress through efficient regulation of the cellular redox poise in an ABA‐independent manner in tomato plants.     

OsGL1-3 is Involved in Cuticular Wax Biosynthesis and Tolerance to Water Deficit in Rice

Cuticular wax covers aerial organs of plants and functions as the outermost barrier against non-stomatal water loss. We reported here the functional characterization of the Glossy1(GL1)-homologous gene OsGL1-3 in rice using overexpression and RNAi transgenic rice plants. OsGL1-3 gene was ubiquitously expressed at different level in rice plants except root and its expression was up-regulated under ABA and PEG treatments. The transient expression of OsGL1-3–GFP fusion protein indicated that OsGL1-3 is mainly localized in the plasma membrane. Compared to the wild type, overexpression rice plants exhibited stunted growth, more wax crystallization on leaf surface, and significantly increased total cuticular wax load due to the prominent changes of C₃₀–C₃₂ aldehydes and C₃₀ primary alcohols. While the RNAi knockdown mutant of OsGL1-3 exhibited no significant difference in plant height, but less wax crystallization and decreased total cuticular wax accumulation on leaf surface. All these evidences, together with the effects of OsGL1-3 on the expression of some wax synthesis related genes, suggest that OsGL1-3 is involved in cuticular wax biosynthesis. Overexpression of OsGL1-3 decreased chlorophyll leaching and water loss rate whereas increased tolerance to water deficit at both seedling and late-tillering stages, suggesting an important role of OsGL1-3 in drought tolerance.     

Low concentration of exogenous abscisic acid increases lead tolerance in rice seedlings

The effects of exogenous abscisic acid (ABA) on lead tolerance in rice (Oryza sativa L.) seedlings were investigated. Pre-treatment with 0.1 g m³ ABA for 2 d restricted amount of Pb translocated from roots to shoots, decreased malondialdehyde and H₂O₂ contents in leaves, and alleviated Pb-induced decrease in plant growth and leaf chlorophyll content. Further, ABA pre-treatment adjusted leaf antioxidative enzyme activities (increased ascorbate peroxidase and catalase activities while decreased superoxide dismutase activity) and so alleviated oxidative stress.     

Water Relations and Growth of the flacca Tomato Mutant in Relation to Abscisic Acid

The flacca mutant in tomato (Lycopersicon esculentum Mill. cv Rheinlands Ruhm) was employed to examine the effects of a relatively constant diurnal water stress on leaf growth and water relations. As the mutant is deficient in abscisic acid (ABA) and can be phenotypically reverted to the wild type by applications of the growth substance, inferences can be made concerning the involvement of ABA in responses to water stress. Water potential and turgor were lower in leaves of flacca than of Rheinlands Ruhm, and were increased by ABA treatment. ABA decreased transpiration rates by causing stomatal closure and also increased the hydraulic conductance of the sprayed plants. Osmotic adjustment did not occur in flacca plants despite the daily leaf water deficits. Stem elongation was inhibited by ABA, but leaf growth was promoted. It is concluded that, in some cases, ABA may promote leaf growth via its effect on leaf water balance.     

Local coexpression domains in the genome of rice show no microsynteny with Arabidopsis domains

Plant responses to abiotic stress are determined both by the severity of the stress as well as the metabolic status of the plant. Abscisic acid (ABA) is a key component in integrating these various signals and controlling downstream stress responses. By screening for plants with decreased RD29A:LUC expression, we isolated two alleles, glutamate:glyoxylate transferase1-1 (ggt1-1) and ggt1-2, of a mutant with altered ABA sensitivity. In addition to reduced ABA induction of RD29A, ggt1-1 was altered in ABA and stress regulation of Δ ¹ -pyrroline-5-carboxylate synthase, proline dehydrogenase and 9-cis-epoxycarotenoid dioxygenase 3, which encode enzymes involved in Pro and ABA metabolsim, respectively. ggt1-1 also had altered ABA and Pro contents after stress or ABA treatments while root growth and leaf water loss were relatively unaffected. A light-dependent increase in H₂O₂ accumulation was observed in ggt1-1 consistent with the previously characterized role of GGT1 in photorespiration. Treatment with exogenous H₂O₂, as well as analysis of a mutant in nucleoside diphosphate kinase 2 which also had increased H₂O₂ content but is not involved in photorespiration or amino acid metabolism, demonstrated that the greater ABA stimulation of Pro accumulation in these mutants was caused by altered H₂O₂ content as opposed to other metabolic changes. The results suggest that metabolic changes that alter H₂O₂ levels can affect both ABA accumulation and ABA sensitivity. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

OsDMI3‐mediated activation of OsMPK1 regulates the activities of antioxidant enzymes in abscisic acid signalling in rice

In rice, the Ca²⁺/calmodulin (CaM)‐dependent protein kinase (CCaMK) OsDMI3 has been shown to be required for abscisic acid (ABA)‐induced antioxidant defence. However, it is not clear how OsDMI3 participates in this process in rice. In this study, the cross‐talk between OsDMI3 and the major ABA‐activated MAPK OsMPK1 in ABA‐induced antioxidant defence was investigated. ABA treatment induced the expression of OsDMI3 and OsMPK1 and the activities of OsDMI3 and OsMPK1 in rice leaves. In the mutant of OsDMI3, the ABA‐induced increases in the expression and the activity of OsMPK1 were substantially reduced. But in the mutant of OsMPK1, the ABA‐induced increases in the expression and the activity of OsDMI3 were not affected. Pretreatments with MAPKK inhibitors also did not affect the ABA‐induced activation of OsDMI3. Further, a transient expression analysis in combination with mutant analysis in rice protoplasts showed that OsMPK1 is required for OsDMI3‐induced increases in the activities of antioxidant enzymes and the production of H₂O₂. Our data indicate that there exists a cross‐talk between OsDMI3 and OsMPK1 in ABA signalling, in which OsDMI3 functions upstream of OsMPK1 to regulate the activities of antioxidant enzymes and the production of H₂O₂ in rice.     

Increased expression of Fe-chelatase leads to increased metabolic flux into heme and confers protection against photodynamically induced oxidative stress

Fe-chelatase (FeCh, EC 4.99.1.1) inserts Fe²⁺ into protoporphyrin IX (Proto IX) to form heme, which influences the flux through the tetrapyrrole biosynthetic pathway as well as fundamental cellular processes. In transgenic rice (Oryza sativa), the ectopic expression of Bradyrhizobium japonicum FeCh protein in cytosol results in a substantial increase of FeCh activity compared to wild-type (WT) rice and an increasing level of heme. Interestingly, the transgenic rice plants showed resistance to oxidative stress caused not only by the peroxidizing herbicide acifluorfen (AF) as indicated by a reduced formation of leaf necrosis, a lower conductivity, lower malondialdehyde and H₂O₂ contents as well as sustained F_v/F_m compared to WT plants, but also by norflurazon, paraquat, salt, and polyethylene glycol. Moreover, the transgenic plants responded to AF treatment with markedly increasing FeCh activity. The accompanying increases in heme content and heme oxygenase activity demonstrate that increased heme metabolism attenuates effects of oxidative stress caused by accumulating porphyrins. These findings suggest that increases in heme levels and porphyrin scavenging capacity support a detoxification mechanism serving against porphyrin-induced oxidative stress. This study also implicates heme as possibly being a positive signal in plant stress responses.     

Interplay Between Abscisic Acid and Jasmonic Acid and its Role in Water-oxidative Stress in Wild-type, ABA-deficient, JA-deficient, and Ascorbate-deficient Arabidopsis Plants

The interplay between jasmonic acid (JA) and abscisic acid (ABA) in plant responses to water stress and in water-stress-enhanced oxidative stress was investigated in Arabidopsis thaliana plants subjected to water stress by water deprivation. For this purpose a drought assay was conducted using Arabidopsis mutants impaired in ABA (aba2), JA (aos), and ascorbate (vtc1) biosynthesis. Our results show an interaction between ABA and JA during their biosynthesis. Moreover, the coordinated action of ABA and JA protected wild-type, aba2, and aos plants from the effects of stress. However, this effect was not observed in the vtc1 mutant, which showed a distinct decrease in the F _v/F _m ratio, concomitant with a marked fall in relative water content (RWC), despite high endogenous concentrations of JA and ABA. This finding indicates the relevance of ascorbate metabolism in plant acclimation to stress. Despite the interaction between the two phytohormones, drought-associated stomatal closure is regulated mainly by ABA and weakly by JA, whereas JA plays a role in the formation of antioxidants regulating ascorbate and glutathione metabolism. A time course analysis revealed the relevance of plant age and stress duration in the responses of the mutants compared to wild-type plants. Here we discuss the relationship between ABA, JA, ascorbate, and glutathione in plants under water stress.     

赤霉素调节植物对非生物逆境的耐性

赤霉素（GAs）是一类重要的植物激素,调控植物生长发育的诸多方面．最近的研究表明,GA也参与对生物与非生物胁迫的响应,然而GA参与非生物胁迫响应的遗传学证据及其机制有待于进一步研究．本实验室前期研究证明,水稻EullfELONGATEDUPPERMOSTINTERNODE）通过一个新的生化途径降解体内的活性赤霉素分子,并参与调控水稻对病原菌的基础抗病性．本研究发现,euil突变体对盐胁迫能力降低,而超表达EUll基因的水稻和拟南芥耐盐性显著提高．进一步研究发现,积累高含量赤霉素的水稻euil突变体对脱落酸（ABA）的敏感性下降,而赤霉素缺失的EUll超表达转基因水稻和拟南芥均改变了对于ABA的敏感性．EUll基因的转录受逆境诱导,其功能缺失与超表达调控了逆境标志基因的表达．综上推测,GA可能是通过影响ABA的信号途径从而改变了植物对非生物胁迫的响应．