The present project is focused on analyzing the biological activity of singlet oxygen (1O2). 1O2 is generated mainly photochemically by photosensitization reactions that are the basis for photo- oxidative tissue damage in humans suffering from porphyria and that have been exploited for photodynamic therapies of various benign and malignant diseases. 1O2 has been largely considered to be detrimental to cells due to its high reactivity and potential toxicity. However, recent data suggest that 1O2 may also be perceived as a signal. So far two major obstacles have hampered the analysis of the biological activity of 1O2. First, in cells under stress several other chemically distinct ROS are generated simultaneously, thus making it very difficult to link a particular cellular response to 1O2. Second, it is difficult to define criteria that may be used to distinguish between the cytotoxicity and the signaling role of this ROS. These problems have been alleviated in the present research project by using the conditional flu mutant of Arabidopsis. In the dark the flu mutant accumulates protochlorophyllide, a potent photosensitizer that upon illumination generates 1O2. By varying the length of the dark period one can modulate noninvasively the level of the photosensitizer and define conditions that minimize the cytotoxicity of 1O2 and endorse its signaling. The genetic basis of 1O2- signaling is revealed by the executer1 mutation that is sufficient to abrogate 1O2-mediated stress responses. These responses largely depend on differentially regulated gene expression. Among the genes that are up-regulated right after the release of 1O2, those encoding transcription factors are clearly overrepresented, implicating an as yet largely unexplored transcriptional regulatory network with triggering 1O2-mediated responses. Regulatory modules will be described consisting of primary 1O2-responsive transcription factors and their target genes that play a key role in transforming 1O2- derived signals into 1O2-dependent physiological changes. These responses range from stress acclimation and modifying disease resistance to the initiation of programmed cell death and committing suicide. We will try to understand how a seemingly simple initial event, the release of singlet oxygen, gives rise to the genetically controlled activation of such diverse responses.
Increased levels of chemically distinct reactive oxygen species (ROS) in humans have been associated with various stress symptoms, cancer, cell death, ageing and age-related pathologies such as stroke, neurodegeneration and cardiovascular diseases. Understanding the different roles of each of these ROS during the emergence of such disorders would greatly impact the design of new remedies and treatments. In the present work one of these ROS, singlet oxygen, that formerly had been considered to affect cells primarily by causing oxidative damage, will be analyzed under conditions that minimize its cytotoxicity and reveal the genetic basis of its signaling role.
