Investigations into the role of the plastidial peptide methionine sulfoxide reductase in response to oxidative stress in Arabidopsis.

Peptidyl Met residues are readily oxidized by reactive oxygen species to form Met sulfoxide. The enzyme peptide Met sulfoxide reductase (PMSR) catalyzes the reduction of Met sulfoxides back to Met. In doing so, PMSR is proposed to act as a last-chance antioxidant, repairing proteins damaged from oxidative stress. To assess the role of this enzyme in plants, we generated multiple transgenic lines with altered expression levels of the plastid form of PMSR (PMSR4). In transgenic plants, PMSR4 expression ranged from 95% to 40% (antisense) and more than 600% (overexpressing lines) of wild-type plants. Under optimal growing conditions, there is no effect of the transgene on the phenotype of the plants. When exposed to different oxidative stress conditions-methyl viologen, ozone, and high light-differences were observed in the rate of photosynthesis, the maximum quantum yield (Fv/Fm ratio), and the Met sulfoxide content of the isolated chloroplast. Plants that overexpressed PMSR4 were more resistant to oxidative damage localized in the chloroplast, and plants that underexpressed PMSR4 were more susceptible. The Met sulfoxide levels in proteins of the soluble fraction of chloroplasts were increased by methyl viologen and ozone, but not by high-light treatment. Under stress conditions, the overexpression of PMSR4 lowered the sulfoxide content and underexpression resulted in an overall increase in content.

Ozone-induced ethylene and foliar injury responses are altered in 1-aminocyclopropane-1-carboxylate synthase antisense potato plants.

• In potato (Solanum tuberosum), two 1-aminocyclopropane-1-carboxylate synthase (ACS) genes are induced by ozone (O3 ). Antisense inhibition of these ethylene (C2 H4 ) biosynthetic genes allowed us to examine the relationship between O3 -induced C2 H4 emission and foliar injury. • Thirty-two lines (antisense for ST-ACS4 or ST-ACS5) were screened in the glasshouse for acute O3 -induced C2 H4 and lesions. Stomatal conductance and ACS transcripts were quantified for selected C2 H4 -altered lines. Six lines were field-tested for chronic O3 effects. • Ten lines produced less, and four lines produced more, acute-O3 -induced C2 H4 than nontransformed (NT) plants. Ethylene levels did not appear to be correlated with stomatal conductance. ST-ACS4 and -5 transcript were reduced in transgenic plants, except in two C2 H4 over-producing lines. In the field, these C2 H4 over-producing lines displayed stunting and leaf rolling in charcoal-filtered (CF) air and chronic O3 , and they sustained the most severe O3 injury. • When C2 H4 production was strongly suppressed or enhanced, corresponding reductions or increases in lesion severity were observed, suggesting a critical role for C2 H4 in the lesion formation process during O3 stress.

The impact of ozone on a salt marsh cordgrass (Spartina alterniflora).

Spartina alterniflora plants were collected from salt marshes within New Jersey, South Carolina, and Georgia USA and shipped to The Pennsylvania State University. New plants were grown from rhizomes in six open-top field chambers. Three chambers received charcoal-filtered air, and three received charcoal-filtered air plus 80 ppb ozone, 8 h/day for 65 days. Flower, leaf, and shoot number per plant were recorded weekly. Photosynthetic rates were measured in week 5, and foliar injury was assessed during week 9. Final dry weight of roots, shoots, and rhizomes were determined. While ozone-treated plants from all states expressed symptoms of ozone injury, plants from South Carolina exhibited no effect of ozone on any other measured variable. Plants from the Georgia site showed ozone-induced reductions in all measured variables except leaf dry weight. Ozone-treated plants from New Jersey showed reductions in photosynthetic rate, leaf and shoot number, and root dry weights. Only plants from New Jersey produced flowers, with ozone treatment causing delay in flowering and reduction in the number of flower spikes produced.

Ozone-induced changes in biosynthesis of Rubisco and associated compensation to stress in foliage of hybrid poplar.

Experiments were conducted during the growing seasons of 1993-1995 to determine whether exposure to ozone (O(3)) affected the synthesis of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) in hybrid poplar, Populus maximowizii A. Henry x trichocarpa Torr. & A. Gray, Clone 245. As the canopy aged, the concentration of Rubisco decreased at a more rapid rate in lower leaves of hyrid poplar ramets subjected to chronic O(3) exposure in open-top chambers than in comparable foliage of plants grown in charcoal-filtered air. There was no difference in rate of synthesis of Rubisco between treatments, suggesting that loss of this protein in O(3)-treated leaves was caused by an accelerated rate of proteolysis. In foliage higher in the canopy, both concentration and rate of synthesis of Rubisco were stimulated by O(3) for a brief period when the leaves were young. Quantification of mRNA for the small (rbcS) and large (rbcL) transcripts of Rubisco did not reveal changes that were likely to reflect altered synthesis of Rubisco as a prime response to O(3). Analyses of Rubisco concentration and rate of Rubisco synthesis in foliage connected by vascular traces within the canopy indicated that loss of Rubisco in older leaves was associated with an increase in this protein in younger leaves higher in the canopy. These data support the notion that accelerated senescence may provide some compensatory benefit to the plant. In 1995, the rate of synthesis of Rubisco was almost always higher in O(3)-treated foliage than in nontreated foliage, even when the concentration of Rubisco was adversely affected by the O(3) treatment. Because accelerated foliar abscission in response to O(3) was minimal in 1995 compared to other years, we speculate that, when abscission is delayed, Rubisco synthesis and concentration become uncoupled.

Activity of ribulose bisphosphate carboxylase/oxygenase from potato cultivars with differential response to ozone stress.

The effect of ozone (O3 ) on the carboxylation activity of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) from the fifth youngest leaf of four cultivars of Solarium tuberosum L. (potato) was investigated. Ozone-tolerant cultivars 'Superior' (SP) and 'Norgold Russet' (NR), and O3 -sensitive cultivars 'Cherokee' (CK) and 'Norland' (NL) were exposed to O3 for five consecutive days in controlled-environment chambers. Leaf samples were obtained during the O3 -exposure period and at 1, 5, and 10 d post-exposure. For both charcoal-filtered air (control) and O3 -treated foliage, total activity of Rubisco declined with increasing leaf age. Ozone induced an accelerated decline in the carboxylation activity of Rubisco in leaves obtained from CK and NL by the third day of O3 exposure. In contrast, O3 had no impact on the Rubisco activity from leaves of SP and NR as compared to non-treated foliage. Differential foliar conductance could not account for the response of the sensitive and tolerant cultivars. Rubisco was purified from cultivars SP and CK to determine whether protein sulphydryl content correlated with foliar O3 sensitivity. The number of SH groups did not differ between the two cultivars, either in chemically denatured or non-denatured Rubisco. Consequently, neither number nor availability of SH groups appear to be factors controlling sensitivity of Rubisco to oxidative damage by O3 .