Improved procedure for detection of superoxide dismutase isoforms in potato, Solanum tuberosum L.

• Autorzy: Momcilovic I. , Pantelic D. , Hfidan M. , Savic J. , Vinterhalter D.
• Języki publikacji: EN

Abstrakty

EN

Superoxide dismutase (SOD) in-gel activity assay with selective inhibitors (KCN and H₂O₂) is one of the most commonly used methods for identification of SOD isoform types, i.e., FeSOD, MnSOD or Cu/ZnSOD, and evaluation of oxidative stress response in plants. However, there are potential pitfalls that surround this assay, such as problem to detect isoforms with low activity, comigration of SOD isoforms or application of inappropriate inhibitor concentration. We propose an improved method based on the combination of in-gel analysis of SOD activity and nativePAGE immunoblotting for identification of isoforms and determination of SOD isoenzyme activity pattern in potato. Depending on cultivar and growing conditions, one MnSOD, 3 FeSOD and 5–6 Cu/ZnSOD isoforms were identified in potato leaves. The most important qualitative difference between ex vitro- and in vitro-grown plants was the presence of additional FeSOD and Cu/ZnSOD isoforms in plantlets grown in vitro. Compared with results of in-gel activity assay with selective inhibitors, new method allowed accurate identification of comigrating FeSOD and Cu/ZnSOD isoforms and two protein bands of ambiguous identities. Potato SODs were also characterized by SDS-PAGE immunoblotting and single MnSOD (23.6 kDa), three Cu/ZnSOD polypeptides (17.9, 17 and 16.3 kDa) and single FeSOD (25.1 kDa) polypeptide were detected in leaves of four examined cultivars. The difference in the number of FeSOD and Cu/ZnSOD isoforms/polypeptides between native-PAGE and SDS-PAGE immunoblots suggests that SOD proteins may have undergone post-translational modifications affecting protein mobility or existence of isoforms that differ from each other in total protein charge, but not in molecular weight.

• Wydawca: -
• Czasopismo: Acta Physiologiae Plantarum
• Rocznik: 2014
• Tom: 36
• Numer: 08
• Opis fizyczny: p.2059-2066,fig.,ref.

Twórcy

autor

Momcilovic I.

• Institute for Biological Research "Sinisa Stankovic", University of Belgrade, Bulevar Despota Stefana 142, 11060 Belgrade, Serbia

autor

Pantelic D.

• Institute for Biological Research "Sinisa Stankovic", University of Belgrade, Bulevar Despota Stefana 142, 11060 Belgrade, Serbia

autor

Hfidan M.

• Faculty of Biology, Uiversity of Belgrade, Studentski Trg 3, 11000 Belgrade, Serbia

autor

Savic J.

• Institute for Biological Research "Sinisa Stankovic", University of Belgrade, Bulevar Despota Stefana 142, 11060 Belgrade, Serbia

autor

Vinterhalter D.

• Institute for Biological Research "Sinisa Stankovic", University of Belgrade, Bulevar Despota Stefana 142, 11060 Belgrade, Serbia

Bibliografia

• Abdel-Ghany SE, Müller-Moulé P, Niyogi KK, Pilon M, Shikanai T (2005) Two P-type ATPases are required for copper delivery in Arabidopsis thaliana chloroplasts. Plant Cell 17:1233–1251
• Alscher RG, Erturk N, Heath LS (2002) Role of superoxide dismutases (SODs) in controlling oxidative stress in plants. J Exp Bot 53:1331–1341
• Anderson MD, Prasad TK, Stewart CR (1995) Changes in isozyme profiles of catalase, peroxidase, and glutathione reductase during acclimation to chilling in mesocotyls of maize seedlings. Plant Physiol 109:1247–1257
• Beauchamp C, Fridovich I (1971) Superoxide dismutase: improved assays and an assay applicable to acrylamide gels. Anal Biochem 44:276–287
• Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254
• Drotar A, Phelps P, Fall R (1985) Evidence for glutathione peroxidase activities in cultured plant cells. Plant Sci 42:35–40
• Dugas DV, Bartel B (2008) Sucrose induction of Arabidopsis miR398 represses two Cu/Zn superoxide dismutases. Plant Mol Biol 67:403–417
• Fidalgo F, Santos A, Santos I, Salema R (2004) Effects of long-term salt stress on antioxidant defence systems, leaf water relations and chloroplast ultrastructure of potato plants. Ann Appl Biol 145:185–192
• Fridovich I (1986) Superoxide dismutases. Adv Enzymol Relat Areas Mol Biol 58(6):61–97
• Gill SS, Tuteja N (2010) Reactive oxygen species and antioxidant machinery in abiotic stress tolerance in crop plants. Plant Physiol Biochem 48:909–930
• Kukavica B, Mojović M, Vučinić Ž, Maksimović V, Takahama U, Veljović-Jovanović S (2009) Generation of hydroxyl radical in isolated pea root cell wall, and the role of cell wall-bound peroxidase, Mn-SOD and phenolics in their production. Plant Cell Physiol 50:304–317
• Martinez CA, Loureiro ME, Oliva MA, Maestri M (2001) Differential responses of superoxide dismutase in freezing resistant Solanum curtilobum and freezing sensitive Solanum tuberosum subjected to oxidative and water stress. Plant Sci 160:505–515
• Meloni DA, Oliva MA, Martinez CA, Cambraia J (2003) Photosynthesis and activity of superoxide dismutase, peroxidase and glutathione reductase in cotton under salt stress. Environ Exp Bot 49:69–76
• Miller A-F (2012) Superoxide dismutases: ancient enzymes and new insights. FEBS Lett 586:585–595
• Miszalski Z, Ślesak I, Niewiadomska E, Baczek-Kwinta R, Lüttge U, Ratajczak R (1998) Subcellular localization and stress responses of superoxide dismutase isoforms from leaves in the C3-CAM intermediate halophyte Mesembryanthemum crystallinum L. Plant, Cell Environ 21:169–179
• Momcilovic I, Ristic Z (2007) Expression of chloroplast protein synthesis elongation factor, EF-Tu, in two lines of maize with contrasting tolerance to heat stress during early stages of plant development. J Plant Physiol 164:90–99
• Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassay with tobacco tissue cultures. Physiol Plant 15:473–497
• Pan S-M, Yau Y–Y (1992) Characterization of superoxide dismutase in Arabidopsis. Taiwania 37:58–66
• Pilon M, Ravet K, Tapken W (2011) The biogenesis and physiological function of chloroplast superoxide dismutases. Biochim Biophys Acta 1807:989–998
• Queirós F, Rodrigues JA, Almeida JM, Almeida DPF, Fidalgo F (2011) Differential responses of the antioxidant defence system and ultrastructure in a salt-adapted potato cell line. Plant Physiol Biochem 49:1410–1419
• Queval G, Noctor G (2007) A plate reader method for the measurement of NAD, NADP, glutathione, and ascorbate in tissue extracts: application to redox profiling during Arabidopsis rosette development. Anal Biochem 363:58–69
• Rahnama H, Ebrahimzadeh H (2006) Antioxidant isozymes activities in potato plants (Solanum tuberosum L.) under salt stress. J Sci Islam Repub Iran 17:225–230
• Rojas-Beltran JA, Dejaeghere F, Abd Alla Kotb M, Du Jardin P (2000) Expression and activity of antioxidant enzymes during potato tuber dormancy. Potato Res 43:383–393
• Santos I, Almeida J, Salema R (1999) The influence of UV-B radiation on the superoxide dismutase of maize, potato, sorghum, and wheat leaves. Can J Bot 77:70–76
• Weisiger RA, Fridovich I (1973) Superoxide dismutase organelle specificity. J Biol Chem 248:3582–3592
• Yamakura F, Kawasaki H (2010) Post-translational modifications of superoxide dismutase. Biochim Biophys Acta 1804:318–325

Identyfikatory

DOI

10.1007/s11738-014-1583-z