The activity of antioxidative enzymes, contents of H2O2 and of ascorbate in tomato leaves of cultivars more or less sensitive to infection with Botrytis cinerea

Abstract

Key words: an ti ox i dants, apoplast, hy dro gen perox ide, peroxidase, to mato, Bo try tis cinerea, re sistance Ab stractThe aim of the pres ent stud ies was to com pare H 2 O 2 and ascorbate con tents as well as peroxidase (PO) and catalase (CAT) ac tiv i ties in leaves of less sus cep ti ble cultivar Perkoz and more sus cep ti ble Corindo af ter B. cinerea in fec tion.In crease in H 2 O 2 con tents in both Perkoz and Corindo cytosol was ob served, how ever, it ap peared ear lier in the less sus cep tible cultivar. The in crease in PO ac tiv ity in the cytosol frac tion was ob served 48 hours af ter in fec tion in both cultivars but it was greater in the less sus cep ti ble Perkoz. No sig nif i cant differ ences be tween the tested cultivars were ob served in ascorbate peroxidase (APX) ac tiv ity and in re duced and oxidated ascorbate con tents. PO ac tiv ity was thor oughly an a lyzed in the apoplast frac tion. It was mea sured with syringaldazine (S), tetramethylbenzidine (TMB) and ferulic acid (FA) -sub strates char ac ter is tic of isoenzymes in volved in lig ni fi ca tion and stiffen ing of a cell wall. In crease in PO ac tiv ity with these substrates was ob served ear lier in cultivar Perkoz than in cultivar Corindo. Sim i larly, in crease in PO ac tiv ity with NADH appeared sig nif i cantly ear lier in cultivar Perkoz. Apoplastic PO was sep a rated with DEAE Sepharose and two frac tions binding and non-bind ing were ob tained. Bind ing PO frac tion was sig nif i cantly more ac tive es pe cially with S, TMB and NADH af ter B. cinerea in fec tion. The in crease in the en zyme ac tiv ity was mostly ob served in cultivar Perkoz. Bind ing PO was sep arated by elec tro pho re sis on acrylamide gel and re vealed six enzy matic forms from which three were much more ac tive af ter in fec tion in cultivar Perkoz. The ob tained re sults sug gest that cell wall strength en ing me di ated by apoplast PO is a key fac tor re spon si ble for dif fer ent re sis tance of to mato cultivars Perkoz and Corindo to B. cinerea in fec tion. In tro duc tionPatho gen in fec tion trig gers a se ries of de fen sive reac tions in plants. Ox y gen burst is the first symp tom of stress re sult ing from the ger mi na tion of pathogen fun gus spores (Baker et al. 1995). Ac tive ox ygen spe cies (O 2 . , OH . , H 2 O 2 ) which ap pear may be toxic to both patho gen and host cells (Peng and Kuć 1992) and they are in ten sively re moved by the plant de fen sive sys tem. Patho gen in va sion stim ulates the host en zyme sys tem in volved in their removal. The first lines of de fense against in fec tion are apopalstic spaces and a cell wall (Wojtaszek 1997). Im por tant roles in de fense are played by antioxi dative sys tems: ascorbate peroxidase (APX), cata lase (CAT), peroxidases (PO) and ascorbate (As) which are pres ent in a cell wall and apoplast as well as by H 2 O 2 .The rapid in crease in H 2 O 2 level re sult ing from the in fec tion may be coun ter acted by CAT as well as APX and sev eral PO isoenzymes for which H 2 O 2 is a sub st...

Biochemical changes in Quercus robur L. leaves after Erysiphe alphitoides infection

“…In order to visualize POX isoforms, gels were incubated in 50 mM acetate buffer, pH 5.6, with the addition of 30 mM H 2 O 2 and 0.1% 4-chloro-1-naphthol (dissolved in a minimal volume of 96% ethanol) for 15 min and subsequently washed with distilled water (Patykowski & Urbanek, 2005).…”