Reduced Activity of Antioxidant Machinery Is Correlated with Suppression of Totipotency in Plant Protoplasts

Abstract

We previously showed that during protoplast isolation, an oxidative burst occurred and the generation of active oxygen species was differentially mediated in tobacco (Nicotiana tabacum) and grapevine (Vitis vinifera), accompanied by significant quantitative differences (A.K. Papadakis, K.A. Roubelakis-Angelakis [1999] Plant Physiol 127: 197-205). We have now further tested if the expression of totipotency in protoplasts is related to the activity of cellular antioxidant machinery during protoplast culture. Totipotent (T) tobacco protoplasts had 2-fold lower contents of intracellular O 2 .Ϫ and H 2 O 2 and 7-fold lower levels of O 2 .Ϫ and H 2 O 2 in the culture medium, compared with non-totipotent (NT) tobacco protoplasts. Addition of alkaline dimethylsulfoxide, known to generate O 2 .Ϫ , resulted in isolation of tobacco protoplasts with reduced viability and cell division potential during subsequent culture. Active oxygen species levels decreased in tobacco and grapevine protoplasts during culturing, although higher contents of O 2 .Ϫ and H 2 O 2 were still found in NT-compared with T-tobacco protoplasts, after 8 d in culture. In T-tobacco protoplasts, the reduced forms of ascorbate and glutathione predominated, whereas in NT-tobacco and grapevine protoplasts, the oxidized forms predominated. In addition, T-tobacco protoplasts exhibited severalfold lower lipid peroxidation than NT-tobacco and grapevine protoplasts. Furthermore, several antioxidant enzyme activities were increased in T-tobacco protoplasts. Superoxide dismutase activity increased in tobacco, but not in grapevine protoplasts during culturing due to the increased expression of cytoplasmic Cu/Zn-superoxide dismutase. The increase was only sustained in T-tobacco protoplasts for d 8. Together, these results suggest that suppressed expression of totipotency in protoplasts is correlated with reduced activity of the cellular antioxidant machinery.

Plant regeneration from leaf-derived protoplasts within the Daucus genus: effect of different conditions in alginate embedding and phytosulfokine application

“…The oxidative stress induced during the isolation procedure may be long-term and individual species may respond to it differently, among others, producing and cumulating peroxides and other active oxygen species (Watanabe et al 2002;Davey et al 2005c). It was postulated that suppressed expression of totipotency in protoplasts was correlated with reduced activity of the cellular antioxidant machinery (Papadakis et al 2001). Protection from oxidative damage and, in consequence, overcoming protoplast recalcitrance may be possibly achieved by application of an exogenous antioxidant such as ascorbic acid, glutathione, tocopherol, polyamines, or n-propyl gallate.…”

Changes in H2O2 content and antioxidant enzyme gene expression during the somatic embryogenesis of Larix leptolepis

“…Recently, there have been many experiments carried out on several plant species with the objective of explaining the role of oxidative stress in plant morphogenesis (de Marco and Roubelakis-Angelakis 1996; Cui et al 1999;Papadakis et al 2001;Papadakis and Roubelakis-Angelakis 2002). Oxidative stress is inevitably present in somatic embryo development as a particular cell differentiation.…”

Influence of anti- and prooxidants on rhizogenesis from hypocotyls of Mesembryanthemum crystallinum L. cultured in vitro

“…They include the explants properties, the chemical composition of the medium as well as physical culture conditions. To improve the efficiency of regeneration, selected stress factors that cause overproduction of reactive oxygen species (ROS) have been applied with a positive effect (Papadakis et al 2001;Tian et al 2003;Mitrovic et al 2012). Among ROS, hydrogen peroxide (H 2 O 2 ) has been the most efficiently studied signaling molecule in plants (van Breusegem et al 2008;Schmidt and Schippers 2015).…”