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Title: Calcium-mediated signal transduction in transgenic Nicotiana plumbaginifolia
Author: Wood, Nicola T.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1997
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This study describes two approaches for the study of Ca2+-mediated signal transduction in stomatal guard cells. A novel assay for monitoring stimulus-induced changes in guard cell cytosolic Ca2+ ([Ca2+]cyt) was developed, using epidermal strips from a transgenic line of Nicotiana plumbaginifolia harbouring apoqequorin, the precursor of the Ca2+-sensitive photoprotein, aequorin. Results indicated that mechanical, low temperature and abscisic acid (AGA) signals, directly affected stomatal behaviour, promoting rapid closure, and that elevations of guard cell [Ca2+]cyt play a key role in the transduction of these signals. Studies with Ca2+ channel blockers and the Ca2+ chelator EGTA, further suggested that mechanical ABA signals primarily mobilise Ca2+ from intracellular store(s), whereas influx of extracellular Ca2+ was a key component in the transduction of low temperature signals. The stomatal response to low temperature was found to be significantly influenced by previous plant growth temperature. The second approach involved the production of transgenic lines of N. plumbaginifolia expressing apoaequorin under the control of two guard cell 'specific' promoters, the ABA-responsivecDeT6-19 promoter from C.plantagineum (Michel et al., 1994; Taylor et al., 1995) and the constitutive lipid transfer protein (LTP1) promoter from A. thaliana (Thoma et al., 1994), enabling stimulus-induced changes in guard cell [Ca2+]cyt to be monitored in intact seedlings. ABA-induction of cDET6-19 promoter activity did not affect the ability of stomata to respond to further applications of ABA. Results showed that mechanical and cold shock signals induced similar Ca2+ responses to those observed in epidermal strips, with similar sensitivities to Ca2+ antagonists. Studies with inhibitors of other putative components of the plant Ca2+ signaling pathway also implicated phosphoinositide turnover, possibly stimulated by a coupling G-protein, in the low temperature signaling pathway. The hypothesis that Ca2+ plays a central role in the molecular clock mechanism in plants has also been investigated. Transgenic N. plumbaginifolia seedlings expressing apoaequorin under the control of the CaMV 35S promoter were found to exhibit circadian oscillations in [Ca2+]cyt in free-running conditions, confirming previous findings of Johson et al (1995).
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available