Comparative studies of C3 and C4 Atriplex hybrids in the genomics era: physiological assessments.

We crossed the C3 species Atriplex prostrata with the C4 species Atriplex rosea to produce F1 and F2 hybrids. All hybrids exhibited C3-like δ(13)C values, and had reduced rates of net CO2 assimilation compared with A. prostrata. The activities of the major C4 cycle enzymes PEP carboxylase, NAD-malic enzyme, and pyruvate-Pi dikinase in the hybrids were at most 36% of the C4 values. These results demonstrate the C4 metabolic cycle was disrupted in the hybrids. Photosynthetic CO2 compensation points (Г) of the hybrids were generally midway between the C3 and C4 values, and in most hybrids were accompanied by low, C3-like activities in one or more of the major C4 cycle enzymes. This supports the possibility that most hybrids use a photorespiratory glycine shuttle to concentrate CO2 into the bundle sheath cells. One hybrid exhibited a C4-like Г of 4 µmol mol(-1), indicating engagement of a C4 metabolic cycle. Consistently, this hybrid had elevated activities of all measured C4 cycle enzymes relative to the C3 parent; however, C3-like carbon isotope ratios indicate the low Г is mainly due to a photorespiratory glycine shuttle. The anatomy of the hybrids resembled that of C3-C4 intermediate species using a glycine shuttle to concentrate CO2 in the bundle sheath, and is further evidence that this physiology is the predominant, default condition of the F2 hybrids. Progeny of these hybrids should further segregate C3 and C4 traits and in doing so assist in the discovery of C4 genes using high-throughput methods of the genomics era.

Influence of vesicular trichomes of Atriplex nummularia on photosynthesis, osmotic adjustment, cell wall elasticity and enzymatic activity.

Vesicular trichomes play a key role in excluding toxic ions from some halophyte species, preventing the essential processes and functions of plants from being altered. Thus, the present study aimed to evaluate the influence of these structures on Atriplex nummularia irrigated using waters with three levels of osmotic potential (-0.1, -1.4 and -2.7 MPa), formulated with NaCl in plants with vesicular trichomes and plants with partial removal of trichomes. The experiment was conducted in a protected environment and plants were evaluated for physiological parameters (water, osmotic and pressure potentials, relative water content, osmotic adjustment, pressure-volume curve, gas exchange), electrolyte leakage, lipid peroxidation and enzymatic activity (superoxide dismutase, ascorbate peroxidase, catalase). The results obtained made it possible to identify the strong contribution of vesicular trichomes to physiological and biochemical parameters, with indication of cell wall stiffening and maintenance of turgor. Furthermore, the evaluation of the osmotic potentials obtained in the study suggests that the contribution of vesicular trichomes to the salinity tolerance of the species is greater than that of osmotic adjustment. Furthermore, gas exchange results suggest that the presence of trichomes was able to regulate stomatal processes so that the plant maintains its photosynthetic performance. Evaluation of electrolyte leakage, together with the increase in malondialdehyde content, showed that the maintenance of trichomes reduces the probability of oxidative stress. The activity of antioxidant enzymes was efficient in eliminating reactive oxygen species, especially the activity of ascorbate peroxidase, which stood out in terms of hydrogen peroxide detoxification.

Combined effect of Cr-toxicity and temperature rise on physiological and biochemical responses of Atriplex halimus L.

An experiment was conducted to evaluate the combined effect of temperature (26 and 30 °C) and Cr toxicity (0, 100 and 1000 µM Cr) on growth, photosynthesis, water content, Cr and nutrients uptake and translocation. The role of antioxidative enzyme towards stresses tolerance was also investigated. Results showed that the maximum relative growth rate and leaf area per plant of Atriplex halimus L. were recorded at 100 µM Cr and 26 °C. However, presence of Cr reduced net photosynthetic and stomatal conductance rates. Overall, temperature rise enhanced the toxic effect of Cr by reducing growth and photosynthesis and inducing antioxidant enzymes activities. Furthermore, temperature rise increased nutrient uptake, as well as nutrient translocation to aboveground tissues; while it diminished Cr translocation. Finally, roots were the main sink for Cr accumulation in A. halimus. At 1000 µM Cr, root Cr concentrations reached 7.2 and 9.1 mg g-1 at 26 and 30 °C, respectively; while shoot Cr concentrations were 0.45 and 0.44 mg g-1 (26 and 30 °C, respectively). The high Cr-accumulation in roots suggests that A. halimus presents a great potential for phytoremediation, especially phytostabilisation of Cr contaminated soils.

Physiological and biochemical mechanisms preventing Cd-toxicity in the hyperaccumulator Atriplex halimus L.

The xero-halophyte Atriplex halimus L., recently described as Cd-hyperaccumulator, was examined to determine Cd toxicity threshold and the physiological mechanisms involved in Cd tolerance. An experiment was conducted to investigate the effect of cadmium from 0 to 1350 µM on chlorophyll fluorescence parameters, gas exchange, photosynthetic pigment concentrations and antioxidative enzyme activities of A. halimus. Cadmium, calcium, iron, manganese, magnesium, potassium, phosphorous, sodium and zinc concentrations were also analyzed. Plants of A. halimus were not able to survive at 1350 µM Cd and the upper tolerance limit was recorded at 650 µM Cd; although chlorosis was observed from 200 µM Cd. Cadmium accumulation increased with increase in Cd supply, reaching maxima of 0.77 and 4.65 mg g(-1) dry weight in shoots and roots, respectively, at 650 µM Cd. Dry mass, shoot length, specific leaf area, relative growth rate, net photosynthetic rate, stomatal conductance, pigments contents and chlorophyll fluorescence were significantly reduced by increasing Cd concentration. However, the activities of superoxide dismutase (SOD; EC1.15.1.1), catalase (CAT; EC1.11.1.6) and guaiacol peroxidase (GPx; EC1.11.1.7) were significantly induced by Cd. Exposures to Cd caused also a significant decrease in P contents in roots, Mg and Mn contents in shoots and Fe and K contents in roots and shoots and had no effect on Ca, Na and Zn contents. The tolerance of A. halimus to Cd stress might be related with its capacity to avoid the translocation of great amounts of Cd in its aboveground tissues and higher activities of enzymatic antioxidants in the leaf.

Similar regulation patterns of choline monooxygenase, phosphoethanolamine N-methyltransferase and S-adenosyl-L-methionine synthetase in leaves of the halophyte Atriplex nummularia L.

Glycinebetaine (betaine) highly accumulates as a compatible solute in certain plants and has been considered to play a role in the protection from salt stress. The betaine biosynthesis pathway of betaine-accumulating plants involves choline monooxygenase (CMO) as the key enzyme and phosphoethanolamine N-methyltransferase (PEAMT), which require S-adenosyl-L-methionine (SAM) as a methyl donor. SAM is synthesized by SAM synthetase (SAMS), and is needed not only for betaine synthesis but also for the synthesis of other compounds, especially lignin. We cloned CMO, PEAMT and SAMS isogenes from a halophyte Atriplex nummularia L. (Chenopodiaceous). The transcript and protein levels of CMO were much higher in leaves and stems than in roots, suggesting that betaine is synthesized mainly in the shoot. The regulation patterns of transcripts for SAMS and PEAMT highly resembled that of CMO in the leaves during and after relief from salt stress, and on a diurnal rhythm. In the leaves, the betaine content was increased but the lignin content was not changed by salt stress. These results suggest that the transcript levels of SAMS are co-regulated with those of PEAMT and CMO to supply SAM for betaine synthesis in the leaves.