Chair: Keith Davis, Ohio State University
Keith Davis (The Ohio State University) described the response of Arabidopsis to ozone exposure and presented evidence that the pattern of gene expression observed in ozone-treated plants overlapped significantly with the pattern of gene expression observed during a hypersensitive response. Studies using virulent and avirulent bacterial strains demonstrated that ozone did induce an active defense response capable of restricting bacterial growth in infiltrated leaves. A rapid accumulation of salicylic acid was correlated with ozone-induced gene expression, suggesting that salicylic acid may be involved in the signal transduction pathway leading to gene activation. A similar requirement for salicylic acid production has been shown for pathogen induced gene expression. Further studies using transgenic plants expressing salicylic hydroxylase showed the inhibition of salicylic acid accumulation prevented the ozone-induced accumulation of both PR1 and GST mRNAs, whereas PAL mRNA induction was normal. A model was presented suggesting that there are at least 2 distinct, interacting signal transduction pathways are triggered by ozone exposure and that salicylic acid is a component of one of these pathways.
Anne Britt (University of California, Davis) provided an interesting overview of the changes in UV-irradiation that are expected to occur should the protective ozone layer continue to be degraded, and how these changes may affect plants with respect to DNA damage; there does not appear to be much to worry about. Recent results from the Britt laboratory indicate that there is an active, light dependent repair pathway for UV-induced photoproducts and that it possible to isolate mutants deficient for this repair pathway. Preliminary studies indicate that a second pathway for the repair of cyclobutyl dimers is present and may be most active on actively transcribed DNA sequences.
In a poster talk, Kathleen Smith (Massachusetts Institute of Technology) also discussed DNA repair and the possible function of a novel Arabidopsis gene, RAD51. RAD51 is a homolog of the yeast ScRAD51 gene and the bacterial RecA gene, and thus may be involved in the repair of double strand breaks in the genome. RAD51 transcripts were found at very low levels in most tissues, with the highest levels observed in flowers and young seedlings. RAD51 mRNA was rapidly and strongly induced by exposure of gamma-irradiation in a dose-dependent manner. Interestingly, 2 mutants which are hypersensitive to gamma-irradiation do not exhibit the induction of RAD51 in response to irradiation, thus supporting the suggestion that Arabidopsis RAD51 may be involved in DNA repair.
Mike Thomashow (Michigan State University) provided an overview of the mechanism of cold acclimation in plants, and how Arabidopsis has proven to be a useful model for studying this rather poorly understood phenomenon. One particularly controversial issue has been whether cold acclimation involves the action of genes that encode cryoprotective polypeptides. Thomashow presented results in support of this notion. In particular, he and his colleagues established that the cold-regulated COR15a gene of Arabidopsis encodes a polypeptide that is targeted to the stromal compartment of chloroplasts and found that constitutive expression of the COR15a gene results in the enhancement of chloroplast freezing tolerance in non-acclimated plants. Whether the COR15a polypeptide acts directly as a cryoprotectant remains to be established. However, results obtained in collaboration with Peter Steponkus (Cornell University) suggest the possibility that the COR15a polypeptide inhibits freeze-induced membrane fusion. Thomashow also presented data indicating that the COR15a gene has a cold-regulated promoter that functions in both Arabidopsis and tobacco, a chilling-sensitive plant that does not cold acclimate. Temperature regulation is imparted by a cis-acting element, termed a DRE or C-box, that also imparts drought-regulated gene expression. Nuclear extracts prepared from both cold-acclimated and non-acclimated Arabidopsis plants contain a protein(s) that binds to the C-box. Efforts to clone the protein(s) that binds to the C-box are in progress.
Janet Braam (Rice University) gave an animated and rapid summary of her groups efforts to characterize the regulation and function of the 'touch' (TCH) genes. Highlights included the determination that a subset of the touch genes are induced by cold shock and that changes in cytoplasmic calcium levels may be involved in this induction. Further studies of the previously isolated TCH3 and TCH4 genes revealed that TCH3, a putative calcium-binding protein, accumulates during development at sites that may be under mechanical stress or that are undergoing expansion. TCH4 appears to encode a xyloglucan endotransglycosylase (XET), and as such, may have an important role in modifying the cell wall during responses to environmental stimuli and during development. TCH4 appears to be a member of a large gene family, opening up the possibility for the differential regulation of XETs in response to a number of environmental or developmental cues. Further studies of this interesting gene family will no doubt provide insight into the mechanisms controlling cell wall modifications and how these modifications affect plant structures.
And last, but not least, a poster talk by John Sedbrook (University of Wisconsin) summarized work from Patrick Masson's laboratory on the plant response to anoxia. This group has utilized transgenic Arabidopsis plants expressing AEQUORIN, a sensitive monitor for changes in free calcium levels, to examine the potential role of calcium in regulating the changes in gene expression associated with anoxia. AEQUORIN-expressing Arabidopsis seedlings developed a biphasic luminescence response in cotyledons and leaves, but not in roots nor hypocotyls. This luminescence response was composed of a fast and transient first peak which occurred within minutes of anoxia, followed by a second prolonged luminescence response which lasts 1.5 to 4 hours. The calcium channel blockers Gd3+ and ruthenium red partially inhibit the first response, suggesting that the calcium originates from both intracellular and extracellular stores. In contrast, Gd3+ and ruthenium red promoted a larger and earlier second response, suggesting different mechanisms for these responses. Upon return to normoxia, the luminescence of cotyledons and leaves quickly drops, before the luminescence transiently increases in all plant organs. The Gd3+, La3+, EGTA, and ruthenium red-sensitivity of this normoxia-induced calcium response is consistent with it being associated with the formation of oxygen free radicals, thus adding anoxia as yet another stress that may utilize active oxygen species as signal molecules.