Oxidative post translational modifications of proteins related to cell cycle are involved in cadmium toxicity in wheat seedlings.

Abiotic stress is greatly associated with plant growth inhibition and redox cell imbalance. In the present work, we have investigated in which way oxidative posttranslational modifications (PTM) of proteins related to cell cycle may be implicated in post-germinative root growth reduction caused by cadmium, by methyl viologen (MV) and by hydrogen peroxide (H2O2) in wheat seedlings. Although cadmium is considered a redox inactive metal, reactive oxygen species were detected in the apex root of metal-treated seedlings. Oxidative stress hastened cells displacement from the cell division zone to elongation/differentiation zone, resulting in a shortened meristem. The number of cells in the proliferation zone was lower after MV, H2O2 and 10 µM Cd²âº treatments compared to control. All treatments increased protein carbonylation. Although no modification in total Ub-conjugated proteins was detected, oxidative treatments reduced cyclin D and CDKA protein ubiquitination, concomitantly with a decrease in expression of cyclin D/CDKA/Rb/E2F-regulated genes. We postulate that ROS and oxidative PTM could be part of a general mechanism, specifically affecting G1/S transition and progression through S phase. This would rapidly block cell cycle progression and would allow the cellular defence system to be activated.

Sunflower cotyledons cope with copper stress by inducing catalase subunits less sensitive to oxidation.

Copper is an essential trace element for living organisms, in excess, can be toxic to the cell because of its capacity to generate reactive oxygen species (ROS). Catalase (CAT) catalyzes the dismutation of hydrogen peroxide into water and dioxygen and in plants it is located in peroxisomes and glyoxysomes. Different metals can induce changes in CAT activity, but the mechanism underlying its changes is unclear. After 4h of treatment with 5 and 10 µM CuCl(2) a decrease in the specific CAT activity was detected in sunflower cotyledons of post-germinative heterotrophic seedlings. At 8h of treatment, 5 µM Cu(2+) produced an induction of CAT activity while only a complete recovery to control values was observed for 10 µM Cu(2+) treated seedlings. These activity variations were not related to the level of CAT protein expression, but they did correlate with the oxidative state of the CAT protein. This indicates that the mechanism of CAT inactivation by Cu(2+) involves oxidation of the protein structure. The level of the mRNA of CATA3 and CATA4 increased with the presence of the metal after 4h of exposure. These CAT genes code for the synthesis of CAT subunits less sensitive to oxidation, which would prevent the copper-induced oxidative inactivation of CAT.

Modifications in catalase activity and expression in developing sunflower seedlings under cadmium stress.

Catalase (CAT) dismutates the reactive oxygen species H2O2 into water and dioxygen and in plants; it is located in peroxisomes and glyoxysomes. In the present study, we investigated the effect of cadmium (a well-known oxidative stress inducer) on catalase in roots and cotyledons of developing sunflower seedlings, at 10 microM and 100 microM. Although germination was unaltered after 48 h of exposure to 100 microM Cd2+, root length was significantly reduced. CAT activity was also significantly reduced, but this activity was completely restored (10 microM treatment) or even enhanced (100 microM treatment) 24 h later. Although CAT protein abundance remained similar to control in roots and cotyledons of Cd-treated seedlings, cadmium produced CAT protein oxidation, indicating that the mechanism of CAT inactivation by Cd2+ involves oxidation of the protein structure. The transcripts of the four genes described for sunflower (CATA1 to CATA4) increased after cadmium treatment; CATA1 and CATA2 were the most overexpressed in cotyledon and root, respectively. The differential expression of catalase genes in sunflower seedlings under Cd stress might be related to the synthesis of CAT isoforms less sensitive to oxidation, which would prevent enzyme inactivation and H2O2 accumulation.

Mechanism of CATA3 induction by cadmium in sunflower leaves.

One of the main antioxidant enzymes, catalase (CAT, EC 1.11.1.6), is capable of catalyzing the dismutation of H(2)O(2). This enzyme is involved in signal transduction pathway in plants, controlling the cellular level of this reactive oxygen species. Four different genes, CATA1-CATA4, were identified in Helianthus annuus L. cotyledons. Incubation of sunflower leaf discs with 300 and 500 microM CdCl(2) under light conditions increased CATA3 transcript level. However, it was not induced by Cd(2+) in etiolated plants. This Cd(2+)-induced increase was reverted by adding 10mM ascorbate. Treatments with 0.4 and 10 microM rose bengal (a generator of (1)O(2)) did not activate CATA3, but 10 microM methyl viologen (an enhancer of O(2)(-) production) and 10 mM H(2)O(2) increased its expression. In isolated chloroplasts, Cd(2+) and methyl viologen produced oxidation of the probe 2',7'-dichlorofluorescein diacetate indicating ROS formation. Besides, Cd(2+) treatment of leaf discs under light decreased CAT activity and increased carbonyl groups content, thus suggesting that enzyme inactivation could be due - in part - to a protein oxidation. These results indicate that light is involved in Cd(2+)-induced CATA3 enhancement, which leads to the synthesis of CAT isoforms less sensible to oxidation, and that chloroplast might be the main source of ROS responsible for this process.