Gordon R. Gray

Cold Acclimation and Freezing Tolerance (A Complex Interaction of Light and Temperature)

By comparing growth under five different temperature and irradiance regimes (20[deg]C and 800, 250, and 50[mu]mol m-2 s-1 and 5[deg]C and 250 and 50 [mu]mul m-2 s-1), we have examined the effects of light, temperature, and the relative reduction state of photosystem II on plant morphology, freezing tolerance (lethal temperature at which freezing injury occurs [LT50]), transcript levels of Lhcb and two cold-stimulated genes (Wcs19 and Wcs120), and photosynthetic adjustment in winter rye (Secale cereale L. cv Musketeer). We show, for the first time to our knowledge, that in addition to adjustments in photosynthetic capacity, nonphotochemical quenching capacity and tolerance to photoinhibition, the accumulation of the cold-induced transcript Wcs19, and the compact plant morphology usually associated with cold-hardening are correlated with the relative reduction state of photosystem II rather than with growth temperature or growth irradiance per se. In contrast, the acquisition of maximal LT50, as well as Lhcb and Wcs120 mRNA accumulation, appears to be dependent on both growth temperature and growth irradiance but in an independent, additive manner. The results are discussed with respect to the possible role of the modulation of chloroplastic redox poise in photosynthetic acclimation to cold-hardening temperatures and the attainment of maximal LT50.

Transcriptional Profiling Implicates Novel Interactions between Abiotic Stress and Hormonal Responses in Thellungiella, a Close Relative of Arabidopsis

Thellungiella, an Arabidopsis (Arabidopsis thaliana)-related halophyte, is an emerging model species for studies designed to elucidate molecular mechanisms of abiotic stress tolerance. Using a cDNA microarray containing 3,628 unique sequences derived from previously described libraries of stress-induced cDNAs of the Yukon ecotype of Thellungiella salsuginea, we obtained transcript profiles of its response to cold, salinity, simulated drought, and rewatering after simulated drought. A total of 154 transcripts were differentially regulated under the conditions studied. Only six of these genes responded to all three stresses of drought, cold, and salinity, indicating a divergence among the end responses triggered by each of these stresses. Unlike in Arabidopsis, there were relatively few transcript changes in response to high salinity in this halophyte. Furthermore, the gene products represented among drought-responsive transcripts in Thellungiella associate a down-regulation of defense-related transcripts with exposure to water deficits. This antagonistic interaction between drought and biotic stress response may demonstrate Thellungiellas ability to respond precisely to environmental stresses, thereby conserving energy and resources and maximizing its survival potential. Intriguingly, changes of transcript abundance in response to cold implicate the involvement of jasmonic acid. While transcripts associated with photosynthetic processes were repressed by cold, physiological responses in plants developed at low temperature suggest a novel mechanism for photosynthetic acclimation. Taken together, our results provide useful starting points for more in-depth analyses of Thellungiellas extreme stress tolerance.

Photosystem II Excitation Pressure and Development of Resistance to Photoinhibition (II. Adjustment of Photosynthetic Capacity in Winter Wheat and Winter Rye)

Winter wheat (Triticum aestivum L. cv Monopol), spring wheat (Triticum aestivum L. cv Katepwa), and winter rye (Secale cereale L. cv Musketeer) grown at 5[deg]C and moderate irradiance (250 [mu]mol m-2 s-1) (5/250) exhibit an increased tolerance to photoinhibition at low temperature in comparison to plants grown at 20[deg]C and 250 [mu]mol m-2 s-1 (20/250). However, 5/250 plants exhibited a higher photosystem II (PSII) excitation pressure (0.32–0.63) than 20/250 plants (0.18–0.21), measured as 1 - qP, the coefficient of photochemical quenching. Plants grown at 20[deg]C and a high irradiance (800 [mu]mol m-2 s-1) (20/800) also exhibited a high PSII excitation pressure (0.32–0.48). Similarly, plants grown at 20/800 exhibited a comparable tolerance to photoinhibition relative to plants grown at 5/250. In contrast to a recent report for Chlorella vulgaris (D.P. Maxwell, S. Falk, N.P.A. Huner [1995] Plant Physiol 107: 687–694), this tolerance to photoinhibition occurs in winter rye with minimal adjustment to polypeptides of the PSII light-harvesting complex, chlorophyll a/b ratios, or xanthophyll cycle carotenoids. However, Monopol winter wheat exhibited a 2.5-fold stimulation of sucrosephosphate synthase activity upon growth at 5/250, in comparison to Katepwa spring wheat. We demonstrate that low-temperature-induced tolerance to photoinhibition is not a low-temperature-growth effect per se but, instead, reflects increased photosynthetic capacity in response to elevated PSII excitation pressure, which may be modulated by either temperature or irradiance.

Temperature-driven plasticity in growth cessation and dormancy development in deciduous woody plants: a working hypothesis suggesting how molecular and cellular function is affected by temperature during dormancy induction

The role of temperature during dormancy development is being reconsidered as more research emerges demonstrating that temperature can significantly influence growth cessation and dormancy development in woody plants. However, there are seemingly contradictory responses to warm and low temperature in the literature. This research/review paper aims to address this contradiction. The impact of temperature was examined in four poplar clones and two dogwood ecotypes with contrasting dormancy induction patterns. Under short day (SD) conditions, warm night temperature (WT) strongly accelerated timing of growth cessation leading to greater dormancy development and cold hardiness in poplar hybrids. In contrast, under long day (LD) conditions, low night temperature (LT) can completely bypass the short photoperiod requirement in northern but not southern dogwood ecotypes. These findings are in fact consistent with the literature in which both coniferous and deciduous woody plant species’ growth cessation, bud set or dormancy induction are accelerated by temperature. The contradictions are addressed when photoperiod and ecotypes are taken into account in which the combination of either SD/WT (northern and southern ecotypes) or LD/LT (northern ecotypes only) are separated. Photoperiod insensitive types are driven to growth cessation by LT. Also consistent is the importance of night temperature in regulating these warm and cool temperature responses. However, the physiological basis for these temperature effects remain unclear. Changes in water content, binding and mobility are factors known to be associated with dormancy induction in woody plants. These were measured using non-destructive magnetic resonance micro-imaging (MRMI) in specific regions within lateral buds of poplar under SD/WT dormancing inducing conditions. Under SD/WT, dormancy was associated with restrictions in inter- or intracellular water movement between plant cells that reduces water mobility during dormancy development. Northern ecotypes of dogwood may be more tolerant to photoinhibition under the dormancy inducing LD/LT conditions compared to southern ecotypes. In this paper, we propose the existence of two separate, but temporally connected processes that contribute to dormancy development in some deciduous woody plant: one driven by photoperiod and influenced by moderate temperatures; the other driven by abiotic stresses, such as low temperature in combination with long photoperiods. The molecular changes corresponding to these two related but distinct responses to temperature during dormancy development in woody plants remains an investigative challenge.

Acclimation to low temperature or high light mitigates sensitivity to photoinhibition: roles of the Calvin cycle and the Mehler reaction

Winter wheat (Triticum aestivum L cv. Monopol) plants grown under either control (20˚C, 250 PFD), low temperature (5˚C, 250 PFD) or high light conditions (20˚C, 800 PFD) were compared in order to assess the roles of the Calvin cycle and the Mehler reaction in the differential sensitivity to chronic photoinhibition. Despite similar photosynthetic responses to irradiance, the partial pressure of CO2 [p(CO2)] and photoinhibition, photosynthetic acclimation to cold temperature appears to be quite distinct from acclimation to high light. First, the lower ratio of Rubisco oxygenation/Rubisco carboxylation and the reduced effects of p(CO2) on number of electrons per mole of CO2 fixed in cold-acclimated compared to high light-grown wheat indicate that photorespiration is differentially suppressed in cold-acclimated Monopol. Second, inhibition of the Calvin cycle by glyceraldehyde during photo-inhibition indicated that the sensitivity of high light-acclimated Monopol to photoinhibition was more dependent on Rubisco activity than the sensitivity of cold-acclimated plants to photoinhibition. Third, cold-acclimated Monopol exhibited higher electron transport rates in the presence of either ambient CO2, 2 kPa O2 or N2, 2 kPa O2 (either 77% or 68%, respectively) relative to controls compared to high light-acclimated plants exposed to similar gaseous environments (either 57% or 38%, respectively). Last, the activation state of NADP–malate dehydrogenase indicated that the stroma is highly reduced during cold acclimation relative to either controls or high light-grown Monopol. Thus, in cold-acclimated wheat, the Mehler reaction appears to play an important role while photorespiration plays a minimal role in mitigating the sensitivity to photoinhibition. In contrast, both photorespiration and the Mehler reac-tion appear to mitigate the sensitivity to photoinhibition in high light-grown Monopol. This is consistent with the differential sensitivity to methylviologen and the differential SOD activity observed between cold-acclimated and high light-grown Monopol.

Acclimation to temperature and irradiance modulates PSII charge recombination.

Acclimation of wild type and the chlorina F2 mutant of barley to either high light or low temperature results in a 2‐ to 3‐fold increase in non‐photochemical quenching which occurred independently of either energy‐dependent quenching (qE), xanthophyll cycle‐mediated antenna quenching or state transitions. Results of in vivo thermoluminescence measurements used to address this conundrum indicated that excitation pressure regulates the temperature gap for S 2 Q B ‐ and S 2 Q A ‐ charge recombinations within photosystem II reaction centers. This is discussed in terms of photoprotection through non‐radiative charge recombination.

A role for the anaphase promoting complex in hormone regulation.

To increase our knowledge of anaphase promoting complex (APC/C) function during plant development, we characterized an Arabidopsis thaliana T-DNA-insertion line where the T-DNA fell within the 5′ regulatory region of the APC10 gene. The insert disrupted endogenous expression, resulting in overexpression of APC10 mRNA from the T-DNA- internal CaMV 35S promoter, and increased APC10 protein. Overexpression of APC10 produced phenotypes resembling those of known auxin and ethylene mutants, and increased expression of two tested auxin-regulated genes, small auxin up RNA (SAUR) 15 and SAUR24. Taken together, our data suggests that elevated APC10 likely mimics auxin and ethylene sensitive phenotypes, expanding our understanding of proteolytic processes in hormone regulation of plant development.

The PSBP2 protein of Chlamydomonas reinhardtii is required for singlet oxygen-dependent signaling

In the green alga Chlamydomonas reinhardtii, the cytosolic GLUTATHIONE PEROXIDASE 5 gene (GPX5) is known to be transcriptionally up-regulated in response to singlet oxygen (1O2). As demonstrated by previous studies, fusion of the promoter region of GPX5 to the ARYLSULFATASE 2 gene (ARS2) creates an effective reporter system that can be used to monitor 1O2-driven GPX5 expression. This system was also used in this study to generate a stably transformed C. reinhardtii strain which expresses ARS2 in a 1O2-dependent manner, resulting in the synthesis of a functional protein with detectable activity. Using the strain of C. reinhardtii harboring a 1O2-sensitive reporter construct, a secondary mutagenic screen was performed. This allowed identification of mutant cell lines that were unable to up-regulate expression of the GPX5–ARS2 fusion in response to 1O2. In one of these lines, the mutation was subsequently localized to the first exon of the PSBP-like gene (PSBP2). The PSBP2 gene is part of a small protein family in C. reinhardtii, also present in all angiosperms studied thus far. While each member of the PSBP protein family contains a similar domain to the PSBP1 protein, which is a member of the oxygen evolving complex of photosystem II (PSII), the PSBP2 protein does not appear to be involved in PSII function, but may function as a sensor and/or signal mediating molecule of the 1O2 generated in the chloroplast.

Accumulation of Phosphorus-Containing Compounds in Developing Seeds of Low-Phytate Pea (Pisum sativum L.) Mutants

Low phytic acid (lpa) crops are low in phytic acid and high in inorganic phosphorus (Pi). In this study, two lpa pea genotypes, 1-150-81, 1-2347-144, and their progenitor CDC Bronco were grown in field trials for two years. The lpa genotypes were lower in IP6 and higher in Pi when compared to CDC Bronco. The total P concentration was similar in lpa genotypes and CDC Bronco throughout the seed development. The action of myo-inositol phosphate synthase (MIPS) (EC 5.5.1.4) is the first and rate-limiting step in the phytic acid biosynthesis pathway. Aiming at understanding the genetic basis of the lpa mutation in the pea, a 1530 bp open reading frame of MIPS was amplified from CDC Bronco and the lpa genotypes. Sequencing results showed no difference in coding sequence in MIPS between CDC Bronco and lpa genotypes. Transcription levels of MIPS were relatively lower at 49 days after flowering (DAF) than at 14 DAF for CDC Bronco and lpa lines. This study elucidated the rate and accumulation of phosphorus compounds in lpa genotypes. The data also demonstrated that mutation in MIPS was not responsible for the lpa trait in these pea lines.

Acquisition of freezing tolerance in Arabidopsis and two contrasting ecotypes of the extremophile Eutrema salsugineum (Thellungiella salsuginea).

Eutrema salsugineum (Thellungiella salsuginea) is an extremophile, a close relative of Arabidopsis, but possessing much higher constitutive levels of tolerance to abiotic stress. This study aimed to characterize the freezing tolerance of Arabidopsis (Columbia ecotype) and two ecotypes of Eutrema (Yukon and Shandong) isolated from contrasting geographical locations. Under our growth conditions, maximal freezing tolerance was observed after two- and three-weeks of cold acclimation for Arabidopsis and Eutrema, respectively. The ecotypes of Eutrema and Arabidopsis do not differ in their constitutive level of freezing tolerance or short-term cold acclimation capacity. However Eutrema remarkably outperforms Arabidopsis in long-term acclimation capacity suggesting a wider phenotypic plasticity for the trait of freezing tolerance. The combination of drought treatment and one-week of cold acclimation was more effective than long-term cold acclimation in achieving maximum levels of freezing tolerance in Eutrema, but not Arabidopsis. Furthermore, it was demonstrated growth conditions, particularly irradiance, are determinates of the level of freezing tolerance attained during cold acclimation suggesting a role for photosynthetic processes in adaptive stress responses.