Zoysiagrass (Zoysia spp.) is a common warm-season turfgrass well adapted in use of lawns and golf courses in many regions because of its excellent heat tolerance, density, low pesticide requirements, and minimal maintenance inputs (Patton et al., 2007a, 2007b; Zhang et al., 2009). Expanded use of zoysiagrass could play an important role in making golf courses and home lawns more environmentally friendly and sustainable (Patton and Reicher, 2007). However, the use of zoysiagrasses has been limited by its relatively weak winter hardiness compared with some other C4 turfgrass species, such as bafflograss, and freezing injury in many regions. Freezing temperature may cause damage by forming ice crystals, which result in rupture of cell membranes and cell dehydration (Fry and Huang, 2004; Zhang et al., 2008). Injury during exposure to chilling temperature also results from reduced or defective metabolic defense functions (Karpinski et al., 2002; Thomashow, 1999).
Zoysiagrasses undergo cold acclimation in late fall as temperature drop and photoperiod gets shorter. It has been well documented that increased cold acclimation could improve freeze tolerance of plants, including turfgrasses (Anderson et al., 2003). Plants possess various adaptive mechanisms for surviving freezing temperature such as increases in certain sugars or amino acids, increases in ABA content, and antioxidant capacity (Patton et al., 2007a, 2007b; Rogers et al., 1977; Zhang and Ervin, 2008; Zhang et al., 2009).
Various metabolites (such as proline) may accumulate during cold acclimation (Zhang and Ervin, 2008; Zhang et al., 2006, 2008). Proline, an amino acid, functions as osmoprotectant and antioxidant to protect cell membrane during dehydration. Patton et al. (2007a, 2007b) reported that proline content increased in response to cold acclimation in zoysiagrasses. Bermudagrass (Cynodon L.C. Rich) cultivars with higher stolon proline content exhibited greater freezing tolerance than those with lower proline during the winter (Munshaw et al., 2006).
Decreases in temperature and photoperiod in fall may create an imbalance, so that the energy absorbed through the light harvesting complex exceeds what can be dissipated or transduced by photosystem II (PSII; Karpinski et al., 2002; Zhang and Ervin, 2008). Excess energy may be directed to O2 and result in accumulation of toxic reactive oxygen species (ROS). To protect from oxidative stress, plants have developed efficient antioxidant defense systems to scavenge ROS such as superoxide radicals (O2−), hydrogen peroxide (H2O2), and hydroxyl radicals (HO−.) (McKersie and Bowley, 1997). The SOD (EC 1.15.11), a group of metalloenzymes, can convert O2− to H2O2, and considered as the “primary defense” against ROS (Perl-Treves and Perl, 2002; Zhang et al., 2008). The H2O2 is further reduced to water by the antioxidant enzymes—CAT (EC 18.104.22.168) and APX (EC 22.214.171.124). The CAT, localized in peroxisomes, scavenges H2O2 produced by glycolate oxidase in the C2 photorespiratory cycle (Perl-Treves and Perl, 2002). POD (EC 1.11.17) is also an important antioxidant enzyme for scavenging ROS. Antioxidant enzymes and metabolites have been shown to be associated with freezing tolerance in plants, including bermudagrass (Karpinski et al., 2002; Zhang and Ervin, 2008). Overexpression of a chloroplast Cu/Zn SOD gene increased resistance to chilling stress in tobacco (Gupta et al., 1993).
ABA and H2O2 have been considered as signaling molecules for inducing plant antioxidant defense systems against abiotic stresses (Xiong et al., 2002). Cytokinins exhibit antisenescence and antioxidant function. Heino et al. (1990) reported that ABA deficiency prevented development of freezing tolerance in Arabidopsis thaliana. Stolon ABA accumulation during cold acclimation is associated with freezing tolerance in bermudagrass (Zhang et al., 2008) and zoysiagrass (Zhang et al., 2009), and exogenous ABA increased freezing tolerance of bermudagrass (Zhang et al., 2008). Hu et al. (2006) found that ABA is a key inducer of H2O2 production in maize exposed to drought stress. Hsu and Kao (2010) indicated that ABA-induced leaf senescence of rice seedlings is due to H2O2 accumulation. Prasad et al. (1994) reported that accumulation of ABA and H2O2 protect mitochondria against CI in maize seedlings.
Cytokinins are adenine derivatives characterized by an ability to induce cell division in tissue culture (in the presence of auxin). They also promote shoot initiation, lateral bud growth, leaf expansion, nutrient mobilization, chloroplast differentiation, and activation of shoot meristems and delay senescence (Davies, 2010). Zeatin and zeatin riboside are some of the most important forms of cytokinins. Cytokinins are synthesized in root tips and developing seeds and transported from roots to shoots via the xylem (Davies, 2010). Taylor et al. (1990) measured cytokinin and ABA levels in field- and growth chamber-grown winter wheat plants. They found that ABA level increased, whereas cytokinin level declined during cold acclimation.
Research on changes of ABA, cytokinin, and H2O2 associated with antioxidant metabolism in zoysiagrass is lacking. Very few studies have been reported on antioxidant metabolism associated with cold acclimation and freezing tolerance in zoysiagrass. Investigations concerning the physiological responses of zoysiagrass to cold acclimation treatment would provide valuable selection information for turfgrass breeders and practitioners. The objectives of this study were to examine effect of cold acclimation treatment on the levels of ABA, cytokinin, and antioxidant enzyme activity and to investigate if cold treatment–induced changes of the hormones and antioxidants are associated with freezing tolerance in zoysiagrass.
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