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JXB Advance Access originally published online on May 22, 2006
Journal of Experimental Botany 2006 57(8):1667-1676; doi:10.1093/jxb/erj194
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© The Author [2006]. Published by Oxford University Press [on behalf of the Society for Experimental Biology]. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

RESEARCH PAPER

Reduced expression of top1ß gene induces programmed cell death and alters ascorbate metabolism in Daucus carota cultured cells

Vittoria Locato1 *, Alma Balestrazzi2 *, Laura De Gara1,3,{dagger} and Daniela Carbonera2

1Department of Plant Biology and Pathology, University of Bari, Via Orabona 4, I-70125, Bari, Italy
2Department of Genetics and Microbiology, University of Pavia, Via Ferrata 1, I-27100, Pavia, Italy
3Interdisciplinary Center for Biomedical Research (CIR) Università Campus Biomedico Via Longoni 83, I-00155 Roma, Italy

{dagger}To whom correspondence should be addressed. E-mail: degara{at}botanica.uniba.it

Topoisomerase I (topo I) is a nuclear enzyme which plays a fundamental role in several pathways involving changes in DNA topology. The topo I-mediated reaction is accomplished by the transient covalent binding of the enzyme to DNA (topo I–DNA complex). Stabilization of the topo I–DNA complex, leading to irreversible double-strand breaks, has been reported to occur in animal cells under oxidative stress conditions and during apoptosis. In order to study the existence of a putative link between the topo I-mediated DNA damage and ascorbate (ASC) metabolism, also involved in the responses against oxidative stress and in the apoptotic process in plants, Daucus carota cells showing reduced expression of the top1ß gene encoding the topo Iß isoform were produced, using an antisense RNA strategy. Two independent transgenic lines (AT1-ß/22 and ß/36), characterized by a slow growth phenotype, resistance to camptothecin, a specific inhibitor of topo I, but sensitivity to etoposide, an inhibitor of topo II, were investigated in this study. In the absence of external stimuli, AT1-ß/22 and ß/36 cells underwent programmed cell death (PCD) in a precocious phase of the growth curve. ASC metabolism showed remarkable differences in AT1-ß/22 and ß/36 cells, compared with control, and the observed alterations were similar to those occurring in tobacco Yellow Bright-2 cells induced to enter PCD by exogenous stimuli. However, differently from other studied examples of PCD, overproduction of reactive oxygen species was not detected in AT1-ß/22 and ß/36 cells. The relevance of these findings in relation to the signalling pathways leading to PCD is discussed.

Key words: Antioxidants, ascorbate, Daucus carota, programmed cell death, topoisomerase


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 Proc. Natl. Acad. Sci. USAHome page
S. Sirikantaramas, M. Yamazaki, and K. Saito
From the Cover: Mutations in topoisomerase I as a self-resistance mechanism coevolved with the production of the anticancer alkaloid camptothecin in plants
PNAS, May 6, 2008; 105(18): 6782 - 6786.
[Abstract] [Full Text] [PDF]


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