Physiological responses of low- and high-cadmium accumulating Robinia pseudoacacia-rhizobium symbioses to cadmium stress.

The role of rhizobia in alleviating cadmium (Cd) stress in woody legumes is still unclear. Therefore, two types of black locust (Robinia pseudoacacia L.) with high and low Cd accumulation abilities were selected from 11 genotypes in China, and the effects of rhizobium (Mesorhizobium huakuii GP1T11) inoculation on the growth, CO2 and H2O gas exchange parameters, Cd accumulation, and the absorption of mineral elements of the high (SX) and low Cd-accumulator (HB) were compared. The results showed that rhizobium-inoculation significantly increased biomass, shoot Cd contents, Cd accumulation, root-to-shoot translocation factor (TF) and the absorption and accumulation of mineral elements in both SX and HB. Rhizobium-inoculation increased chlorophyll a and carotenoid contents, and the intercellular carbon dioxide concentrations in HB plants. Under Cd exposure, the high-accumulator SX exhibited a significant decrease in photosynthetic CO2 fixation (Pn) and an enhanced accumulation of Cd in leaves, but coped with Cd exposure by increasing chlorophyll synthesis, regulating stomatal aperture (Gs), controlling transpiration (Tr), and increasing the absorption and accumulation of mineral elements. In contrast, the low-accumulator HB was more sensitive to Cd exposure despite preferential accumulation of Cd in roots, with decreased chlorophyll and carotenoid contents, but significantly increased root biomass. Compared to the low-accumulator HB, non-inoculated Cd-exposed SX plants had higher chlorophyll contents, and rhizobium-inoculated Cd-exposed SX plants had higher Pn, Tr, and Gs as well as higher levels of P, K, Fe, Ca, Zn, and Cu. In conclusion, the high- and low-Cd-accumulator exhibited different physiological responses to Cd exposure. Overall, rhizobium-inoculation of black locust promoted the growth and heavy metal absorption, providing an effective strategy for the phytoremediation of heavy metal-contaminated soils by this woody legume.

The Rpf84 gene, encoding a ribosomal large subunit protein, RPL22, regulates symbiotic nodulation in Robinia pseudoacacia.

MAIN CONCLUSION: A homologue of the ribosomal protein L22e, Rpf84, regulates root nodule symbiosis by mediating the infection process of rhizobia and preventing bacteroids from degradation in Robinia pseudoacacia. Ribosomal proteins (RPs) are known to have extraribosomal functions, including developmental regulation and stress responses; however, the effects of RPs on symbiotic nodulation of legumes are still unclear. Ribosomal protein 22 of the large 60S subunit (RPL22), a non-typical RP that is only found in eukaryotes, has been shown to function as a tumour suppressor in animals. Here, a homologue of RPL22, Rpf84, was identified from the leguminous tree R. pseudoacacia. Subcellular localization assays showed that Rpf84 was expressed in the cytoplasm and nucleus. Knockdown of Rpf84 by RNA interference (RNAi) technology impaired the infection process and nodule development. Compared with the control, root and stem length, dry weight and nodule number per plant were drastically decreased in Rpf84-RNAi plants. The numbers of root hair curlings, infection threads and nodule primordia were also significantly reduced. Ultrastructure analyses showed that Rpf84-RNAi nodules contained fewer infected cells with fewer bacteria. In particular, remarkable deformation of bacteroids and fusion of multiple symbiosomes occurred in infected cells. By contrast, overexpression of Rpf84 promoted nodulation, and the overexpression nodules maintained a larger infection/differentiation region and had more infected cells filled with bacteroids than the control at 45 days post inoculation, suggesting a retarded ageing process in nodules. These results indicate for the first time that RP regulates the symbiotic nodulation of legumes and that RPL22 may function in initiating the invasion of rhizobia and preventing bacteroids from degradation in R. pseudoacacia.

Gas exchange by the mesic-origin, arid land plantation species Robinia pseudoacacia under annual summer reduction in plant hydraulic conductance.

The mesic-origin plantation species Robinia pseudoacacia L. has been successfully grown in many arid land plantations around the world but often exhibits dieback and reduced growth due to drought. Therefore, to explore the behavior of this species under changing environmental conditions, we examined the relationship between ecophysiological traits, gas exchange and plant hydraulics over a 3-year period in trees that experienced reduced plant hydraulic conductance (Gp) in summer. We found that the transpiration rate, stomatal conductance (Gs) and minimum leaf water potential (Ψlmin) decreased in early summer in response to a decrease in Gp, and that Gp did not recover until the expansion of new leaves in spring. However, we did not observe any changes in the leaf area index or other ecophysiological traits at the leaf level in response to this reduction in Gp. Furthermore, model simulations based on measured data revealed that the canopy-scale photosynthetic rate (Ac) was 15-25% higher than the simulated Ac when it was assumed that Ψlmin remained constant after spring but almost the same as the simulated Ac when it was assumed that Gp remained high even after spring. These findings indicate that R. pseudoacacia was frequently exposed to a reduced Gp at the study site but offset its effects on Ac by plastically lowering Ψlmin to avoid experiencing any further reduction in Gp or Gs.

Comparative mitochondrial proteomic, physiological, biochemical and ultrastructural profiling reveal factors underpinning salt tolerance in tetraploid black locust (Robinia pseudoacacia L.).

BACKGROUND: Polyploidy is an important phenomenon in plants because of its roles in agricultural and forestry production as well as in plant tolerance to environmental stresses. Tetraploid black locust (Robinia pseudoacacia L.) is a polyploid plant and a pioneer tree species due to its wide ranging adaptability to adverse environments. To evaluate the ploidy-dependent differences in leaf mitochondria between diploid and tetraploid black locust under salinity stress, we conducted comparative proteomic, physiological, biochemical and ultrastructural profiling of mitochondria from leaves. RESULTS: Mitochondrial proteomic analysis was performed with 2-DE and MALDI-TOF-MS, and the ultrastructure of leaf mitochondria was observed by transmission electron microscopy. According to 2-DE analysis, 66 proteins that responded to salinity stress significantly were identified from diploid and/or tetraploid plants and classified into 9 functional categories. Assays of physiological characters indicated that tetraploids were more tolerant to salinity stress than diploids. The mitochondrial ultrastructure of diploids was damaged more severely under salinity stress than that of tetraploids. CONCLUSIONS: Tetraploid black locust possessed more tolerance of, and ability to acclimate to, salinity stress than diploids, which may be attributable to the ability to maintain mitochondrial structure and to trigger different expression patterns of mitochondrial proteins during salinity stress.

Contrasting dynamics of leaf potential and gas exchange during progressive drought cycles and recovery in Amorpha fruticosa and Robinia pseudoacacia.

Leaf gas exchange is closely associated with water relations; however, less attention has been given to this relationship over successive drought events. Dynamic changes in gas exchange and water potential in the seedlings of two woody species, Amorpha fruticosa and Robinia pseudoacacia, were monitored during recurrent drought. The pre-dawn leaf water potential declined in parallel with gas exchange in both species, and sharp declines in gas exchange occurred with decreasing water potential. A significant correlation between pre-dawn water potential and gas exchange was observed in both species and showed a right shift in R. pseudoacacia in the second drought. The results suggested that stomatal closure in early drought was mediated mainly by elevated foliar abscisic acid (ABA) in R. pseudoacacia, while a shift from ABA-regulated to leaf-water-potential-driven stomatal closure was observed in A. fruticosa. After re-watering, the pre-dawn water potential recovered quickly, whereas stomatal conductance did not fully recover from drought in R. pseudoacacia, which affected the ability to tightly control transpiration post-drought. The dynamics of recovery from drought suggest that stomatal behavior post-drought may be restricted mainly by hydraulic factors, but non-hydraulic factors may also be involved in R. pseudoacacia.

Leaf defense system of Robinia pseudoacacia L. seedlings exposed to 3years of elevated atmospheric CO2 and Cd-contaminated soils.

Short-term exposure to elevated CO2 increases cadmium (Cd) uptake in some plant species (wheat, poplars, and willows), which triggers an increase in antioxidative system activity to deal with additional reactive oxygen species that are generated. Here, we examined leaf defenses in Robinia pseudoacacia L. seedlings exposed to elevated CO2+Cd for 3years. Three years of elevated CO2 decreased Cd uptake into leaves and the Cd content in soils and increased the pH of rhizosphere soil relative to ambient CO2. In plants exposed to Cd stress, leaf chlorophyll content was greater under elevated CO2 than under ambient CO2. Superoxide dismutase, peroxidase, and catalase activity increased, glutathione content increased, and malondialdehyde and phytochelatins contents decreased under elevated CO2+Cd relative to Cd alone. Proline, soluble sugars, flavonoids, saponins, and phenolic acids contents were greater under elevated CO2+Cd than under Cd alone, and condensed tannin content was lower. Overall, long-term elevation of CO2 enhanced the leaf defense system of R. pseudoacacia exposed to Cd by stimulating antioxidant enzyme activity, osmotic adjustment, and the production of glutathione, flavonoids and phenolic acids. Future research should focus on understanding the mechanisms involved in the decrease in Cd uptake into leaves and Cd content in soils and the increase in rhizosphere soil pH under long-term exposure to elevated CO2.

A translationally controlled tumor protein gene Rpf41 is required for the nodulation of Robinia pseudoacacia.

Translationally controlled tumor protein (TCTP) is fundamental for the regulation of development and general growth in eukaryotes. Its multiple functions have been deduced from its involvement in several cell pathways, but its potential involvement in symbiotic nodulation of legumes cannot be suggested a priori. In the present work, we identified and characterized from the woody leguminous tree Robinia pseudoacacia a homolog of TCTP, Rpf41, which was up-regulated in the infected roots at 15 days post-inoculation but decreased in the matured nodules. Subcellular location assay showed that Rpf41 protein was located in the plasma membrane, cytoplasm, nucleus, and also maybe in cytoskeleton. Knockdown of Rpf41 via RNA interference (RNAi) resulted in the impaired development of both nodule and root hair. Compared with wild plants, the root and stem length, fresh weight and nodule number per plant was decreased dramatically in Rpf41 RNAi plants. The number of ITs or nodule primordia was also significantly reduced in the Rpf41 RNAi roots. The analyses of nodule ultrastructure showed that the infected cell development in Rpf41 RNAi nodules remained in zone II, which had fewer infected cells. Furthermore, the symbiosomes displayed noticeable shrinkage of bacteroid and peribacteroid space enlargement in the infected cells of Rpf41 RNAi nodules. In the deeper cell layers, a more remarkable aberration of the infected cell ultrastructure was observed, and electron-transparent lesions in the bacteroid cytoplasm were detected. These results identify TCTP as an important regulator of symbiotic nodulation in legume for the first time, and it may be involved in symbiotic cell differentiation and preventing premature aging of the young nodules in R. pseudoacacia.