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Acid invertase activities in roots and young seeds of a metalliferous population (MP) of Rumex dentatus were previously observed to be significantly higher than those of a non-metalliferous population (NMP) under Cu stress. To date, no acid invertase gene has been cloned from R. dentatus . Here, we isolated four full-length cDNAs from the two populations of R. dentatus , presumably encoding cell wall ( RdnCIN1 and RdmCIN1 from the NMP and MP, respectively) and vacuolar invertases ( RdnVIN1 and RdmVIN1 from the NMP and MP, respectively). Unexpectedly, RdnCIN1 and RdmCIN1 most likely encode special defective invertases with highly attenuated sucrose-hydrolyzing capacity. The transcript levels of RdmCIN1 were significantly higher than those of RdnCIN1 in roots and young seeds under Cu stress, whereas under control conditions, the former was initially lower than the latter. Unexpected high correlations were observed between the transcript levels of RdnCIN1 and RdmCIN1 and the activity of cell wall invertase, even though RdnCIN1 and RdmCIN1 do not encode catalytically active invertases. Similarly, the transcript levels of RdmVIN1 in roots and young seeds were increased under Cu stress, whereas those of RdnVIN1 were decreased. The high correlations between the transcript levels of RdnVIN1 and RdmVIN1 and the activity of vacuolar invertase indicate that RdnVIN1 and RdmVIN1 might control distinct vacuolar invertase activities in the two populations. Moreover, a possible indirect role for acid invertases in Cu tolerance, mediated by generating a range of sugars used as nutrients and signaling molecules, is discussed. [ABSTRACT FROM AUTHOR]

In the current study, we hypothesize that mine (metallicolous) populations of metallophytes form a trade-off between the roots and shoots when under copper (Cu) stress to adapt themselves to heavy metal contaminated habitats, and thus, differ from normal (non-metallicolous) populations in biomass allocation. To test the hypothesis, two populations of the metallophyte Kummerowia stipulacea, one from an ancient Cu mine (MP) and the other from a non-contaminated site (NMP), were treated with Cu2+ in hydroponic conditions. The results showed that MP plants had higher root/shoot biomass allocation and more complicated root system architecture compared to those of the NMP plants when under Cu stress. The net photosynthetic capacity was more inhibited in the NMP plants than in the MP plants when under Cu stress. The sugar (sucrose and hexose) contents and acid invertase activities of MP plants were elevated while those in NMP plants were inhibited after Cu treatment. The neutral/alkaline invertase activities and sucrose synthase level showed no significant differences between the two populations when under Cu stress. The results showed that acid invertase played an important role in biomass allocation and that the physiological responses were beneficial for the high root/shoot biomass allocation, which were advantageous during adaptive evolution to Cu-enriched mine soils. [Copyright &y& Elsevier]

The roots of metallophytes serve as the key interface between plants and heavy metal-contaminated underground environments. It is known that the roots of metallicolous plants show a higher activity of acid invertase enzymes than those of non-metallicolous plants when under copper stress. To test whether the higher activity of acid invertases is the result of increased expression of acid invertase genes or variations in the amino acid sequences between the two population types, we isolated full cDNAs for acid invertases from two populations of Kummerowia stipulacea (from metalliferous and non-metalliferous soils), determined their nucleotide sequences, expressed them in Pichia pastoris, and conducted real-time PCR to determine differences in transcript levels during Cu stress. Heterologous expression of acid invertase cDNAs in P. pastoris indicated that variations in the amino acid sequences of acid invertases between the two populations played no significant role in determining enzyme characteristics. Seedlings of K. stipulacea were exposed to 0.3µM Cu2+ (control) and 10µM Cu2+ for 7 days under hydroponics׳ conditions. The transcript levels of acid invertases in metallicolous plants were significantly higher than in non-metallicolous plants when under copper stress. The results suggest that the expression of acid invertase genes in metallicolous plants of K. stipulacea differed from those in non-metallicolous plants under such conditions. In addition, the sugars may play an important role in regulating the transcript level of acid invertase genes and acid invertase genes may also be involved in root/shoot biomass allocation. [Copyright &y& Elsevier]

Sucrose metabolism in roots of metallophytes is very important for root growth and maintenance of heavy metal tolerance. However, rare researches have been carried out on this topic so far. We tested here a hypothesis that roots of copper-tolerant plants should manifest higher activities of sucrose-cleaving enzymes than non-tolerant plants for maintaining root growth under Cu stress. Plants of two contrasting populations of metallophyte Rumex dentatus, one from an ancient Cu mine (MP) and the other from a non-mine site (NMP), were treated with Cu in controlled experiments. Cu treatment resulted in a higher root biomass and root/shoot biomass ratio in MP compared to NMP. More complicated root system architecture was showed in MP under Cu stress. Activities and transcript levels of acid invertase as well as contents of sucrose and reducing sugar in MP were elevated under Cu treatment, while activities of neutral/alkaline invertase and sucrose synthase showed no significant differences between two populations. The results indicate important roles of acid invertase in governing root growth under Cu stress. [Copyright &y& Elsevier]

The germination rate, mineral (Ca, Fe, K, Mn) and carbohydrate (starch, soluble sugars, sucrose, glucose, fructose) contents and hydrolase activities in cotyledons and embryonic axes of bean seeds subjected to cadmium stress were investigated. Compared to the control, Cd caused a reduction in germination percent, embryo growth and in distribution of biomass, mineral and sugars between cotyledon and embryonic axis and inhibited the activities of α-amylase and invertases: soluble acid (INV-AS), soluble neutral (INV-NS), cell wall bound acid (INV-AW). Moreover, the solute leakage into the germination medium was also used as bioindicator parameter to evaluate the toxicity of cadmium accumulation, which increased in different tissues of germinating seeds in the duration of treatment and provoked nutrient loss and, thereby, electrical conductivity enhancement in the imbibition medium. This was correlated with an impairment of membrane integrity, as evidenced by high malondialdehyde (MDA) content and lipoxygenase (LOX) activity in Cd-poisoned embryo. The contribution of solute loss at the expense of growing embryonic axis to failure in reserve mobilization after Cd exposure is emphasized in association with the delay in seed germination. [Copyright &y& Elsevier]

Ethylene signaling was addressed, for the first time, in plant responses to nitrogen dioxide (NO 2) by comparatively analyzing the performance of Arabidopsis ethylene insensitive 2 (ein2-1) with wild-type (WT) plants. Following NO 2 fumigation, severe leaf wilting and chlorosis occurred in WT plants, but much less symptoms were observed in ein2-1. The activities of superoxide dismutase (SOD), peroxidase (PRX) and catalase (CAT) were 39%, 92%, and 11% higher, respectively, in ein2-1 than in WT following NO 2 exposure. Although glutathione contents and the ratio of its reduced form (GSH) to oxidized form (GSSG) were decreased by NO 2 , an obviously alleviated degree was detected in ein2-1 relative to WT. Correspondingly, the contents of hydrogen peroxide (H 2 O 2) and malondialdehyde (MDA), and electrolyte leakage were 25%, 24%, and 29% lower, respectively, in ein2-1 than in WT. The difference of oxidative stress between two tested genotypes was also revealed by the leaf staining regarding the production and distribution of H 2 O 2 , superoxide anion (O 2 ˙ˉ), and cell death. The genes involved in antioxidation or oxidation-reduction processes mostly presented a stronger expression in ein2-1 than in WT under NO 2 stress. The photosynthesis-related parameters including chlorophyll and soluble sugar contents, net photosynthetic rate (P n), and ribulose bisphosphate carboxylase/oxygenase (Rubisco) activity and gene expression, and chlorophyll fluorescence parameters were affected, generally, to a lesser degree in ein2-1 than in WT following NO 2 fumigation. The enzymatic activities and gene expressions of invertases mostly displayed a higher level in ein2-1 relative to WT following NO 2 fumigation. For example, the activities of cytoplasmic, cell wall and vacuolar invertases were 76%, 26%, and 26% higher, respectively, in ein2-1 than in WT. Together, these data suggest that ethylene signal insensitivity efficiently improves plant tolerance to NO 2 exposure, and the possible mechanisms might be correlated with leaf antioxidative defense, photosynthesis-related processes, and sucrose metabolisms. • Ethylene insensitive mutation improves NO 2 tolerance. • The mechanism is correlated with antioxidative defense by regulating gene expression. • Differential cleavage of sucrose by invertases may play an important role. • The tolerance was positively correlated with photosynthesis efficiency. [ABSTRACT FROM AUTHOR]