Protoplast cultures are remarkable examples of plant cell dedifferentiation. The state of dedifferentiation is evidenced by changes in cell morphology, genome organization, as well as by the capability of protoplasts to differentiate into multiple types of cells (depending on the type of the stimulus applied). The first change in the genome structure is connected with large-scale chromatin decondensation, affecting chromocentres involving various types of these repetitive sequences. This paper describes not only the de- and recondensation of satellite DNA type I and 5S rDNA repetitive sequences, but it also compares the recondensation level of chromatin with the levels of oxidative stress which were decreased by using an antioxidant, as well as the capabilities of the antioxidative systems within protoplasts, during the first 72 h of their culture. It is demonstrated that the treatment of protoplasts with ascorbic acid not only decreased the level of oxidative stress but also positively stimulated the expression of the ascorbate peroxidase and catalase. It also led to a greater recondensation of the chromatin (when compared to the untreated protoplasts); in addition, it supported cell proliferation. It is concluded that large-scale genome relaxation is more directly connected with oxidative stress than with large changes in the expression of genes; and further, that its recondensation is related to the start of (as well as the level of) protection by the antioxidative systems.

Department of Botany Faculty of Science Palacký University Slechtitelů 11 Olomouc 783 71 Czech Republic

Zobrazit více v PubMed

Avramova ZJ. Heterochromatin in animals and plants. Similarities and differencies. Plant Physiology. 2002;129:40–49. PubMed PMC

Bat'ková P, Pospíšilová J, Synkvá H. Production of reactive oxygen species and development of antioxidative systems during in vitro growth and ex vitro transfer. Biologia Plantarum. 2008;52:413–422.

Debeaujon I, Branchard M. Induction of somatic embryogenesis and callogenesis from cotyledons and leaf protoplasts-derived colonies of melon (Cucumis melo L.) Plant Cell Reports. 1992;12:37–40. PubMed

de Marco A, Roubelakis-Angelakis KA. Specific features of the ascorbate/glutathione cycle in cultured protoplasts. Plant Cell Reporter. 1999;18:406–411.

Exner V, Hennig L. Chromatin rearrangements in development. Current Opinion in Plant Biology. 2008;11:64–69. PubMed

Gajdová J, Navrátilová B, Smolná J, Lebeda A. Factors affecting protoplast isolation and cultivation of Cucumis spp. Journal of Applied Botany and Food Quality. 2007;81:1–6.

Ganal M, Riede I, Hemleben V. Organization and sequence analysis of two related satellite DNAs in cucumber (Cucumis satiuvus L.) Journal of Molecular Evolution. 1986;23:23–30.

Gomes A, Femandes E, Lima JFC. Use of fluorescence probes for detection of reactive nitrogen species: a review. Journal of Fluorescence. 2006;16:119–139. PubMed

Grafi G. How cells dedifferentiate: a lesson from plants. Developmental Biology. 2004;268:1–6. PubMed

Grafi G. The complexity of cellular dedifferentiation: implications for regenerative medicine. Trends in Biotechnology. 2009;27:329–332. PubMed

Kim JH, Shin SH, Cho SH, Lee WS. DNA endonuclease- and active oxygen-associated degradation of chloroplast DNA in response to paraquat-induced oxidative stress. Journal of Plant Biology. 2000;43:226–231.

Maccarrone M, Van Zadelhoff G, Veldink GA, Vliegenthart JFG, Finazzi-Agró A. Early activation of lipoxygenase in lentil (Lens culinaris) root protoplasts by oxidative stress induces programmed cell death. European Journal of Biochemistry. 2000;267:5078–5084. PubMed

Mancini A, Buschini A, Restivo FM, Rossi C, Poli P. Oxidative stress as DNA damage in different transgenic tobacco plants. Plant Science. 2006;170:845–852.

Navrátilová B, Greplová M, Vyvadilová M, Klíma M, Gajdová J, Skálová D. Electrofusion of protoplasts in selected vegetables of Brassica, Cucumis, and Solanum genera. Acta Horticulturae. 2006;725:801–805.

Ondřej V, Lukášová E, Krejčí J, Matula P, Kozubek S. Lamin A/C and polymeric actin in genome organization. Molecules and Cells. 2008;26:356–361. PubMed

Ondřej V, Kitner M, Doležalová I, Nádvorník P, Navrátilová B, Lebeda A. Chromatin structural rearrangement during dedifferentiation of protoplasts of Cucumis sativus L. Molecules and Cells. 2009;27:443–447. PubMed

Papadakis AK, Roubelakis-Angelakis KA. Oxidative stress could be responsible for the recalcitrance of plant protoplasts. Plant Physiology and Biochemistry. 2002;40:549–559.

Potters G, Horemans N, Caubergs RJ, Asard H. Ascorbate and dehydroascorbate influence cell cycle progression in a tobacco cell suspension. Plant Physiology and Biochemistry. 2000;38:531–540. PubMed PMC

Rakoczy-Trojanowska M. Alternative methods of plant transformation: a short review. Cellular and Molecular Biology Letters. 2002;7:849–858. PubMed

Sedlářová M, Luhová L, Petřivalský M, Lebeda A. Localization and metabolism of reactive oxygen species during Bremia lactucae pathogenesis in Lactuca sativa and wild Lactuca spp. Plant Physiology and Biochemistry. 2007;45:607–616. PubMed

Tessadori F, Chupeau MCh, Chupeau Y, Knip M, Germann S, van Driel R, Fransz P, Gaudin V. Large-scale dissociation and sequential reassembly of pericentric heterochromatin in dedifferentiated Arabidopsis cells. Journal of Cell Science. 2007;120:1200–1208. PubMed

van Driel R, Fransz P. Nuclear architecture and genome functioning in plants and animals: what can we learn from both? Experimental Cell Research. 2004;296:86–90. PubMed

Zhao J, Morozova N, Williams L, Libs L, Avivi Y, Grafi G. Two phases of chromatin decondensation of plant cells. Journal of Biological Chemistry. 2001;276:22772–22778. PubMed