The ability to non-invasively measure metabolic oxygen flux is a very important tool for physiologists interested in a variety of questions ranging from basic metabolism, growth/development, and stress adaptation. Technologies for measuring oxygen concentration near the surface of cells/tissues include electrochemical and optical techniques. A wealth of knowledge was gained using these tools for quantifying real-time physiology. Fiber-optic microprobes (optrodes) have recently been developed for measuring oxygen in a variety of biomedical and environmental applications. We have adopted the use of these optical microsensors for plant physiology applications, and used the microsensors in an advanced sensing modality known as self-referencing. Self-referencing is a non-invasive microsensor technique used for measuring realtime flux of analytes. This paper demonstrates the use of optical microsensors for non-invasively measuring rhizosphere oxygen flux associated with respiration in plant roots, as well as boundary layer oxygen flux in phytoplankton mats. Highly sensitive/selective optrodes had little to no hysteresis/calibration drift during experimentation, and an extremely high signal-to-noise ratio. We have used this new tool to compare various aspects of rhizosphere oxygen flux for roots of Glycine max, Zea mays, and Phaseolus vulgaris, and also mapped developmentally relevant profiles and distinct temporal patterns. We also characterized real-time respiratory patterns during inhibition of cytochrome and alternative oxidase pathways via pharmacology. Boundary layer oxygen flux was also measured for a phytoplankton mat during dark:light cycling and exposure to pharamacological inhibitors. This highly sensitive technology enables non-invasive study of oxygen transport in plant systems under physiologically relevant conditions.

Two year-old cloned plants of Dendrobium Second Love were submitted to 25 1C (light) and 10 1C (dark) under a 12 h photoperiod (60 mmol m À1 s À1 ) for 30 days. The endogenous levels of IAA, ABA, and the cytokinins Z, [9R]Z, iP, and [9R]iP were measured 15, 22, and 30 days after the start of the thermoperiodic treatment in lateral buds and leaves. The endogenous levels of IAA and cytokinins, especially the zeatin-derived forms, increased significantly in buds after 15 days of treatment. On the other hand, the amount of ABA decreased progressively and significantly throughout the treatment. The treatment conspicuously accelerated flower-bud development. The found correlation suggests that hormones are involved in the signal transduction pathway of thermoperiodic flowering control.

In Arabidopsis thaliana, the carotenoid cleavage dioxygenases (CCDs) MAX3 and MAX4 act together with MAX1 to produce a strigolactone signalling molecule required for the inhibition of axillary bud outgrowth. We show that both MAX3 and MAX4 transcripts are positively auxin regulated in a manner similar to the orthologous genes from pea (Pisum sativum) and rice (Oryza sativa), supporting evolutionary conservation of this regulation in plants. This regulation is important for branching control because large auxin-related reductions in these transcripts are associated with increased axillary branching. Both transcripts are up-regulated in max mutants, and consistent with max mutants having increased auxin in the polar auxin transport stream (PATS), this feedback regulation involves auxin signalling. We suggest that both auxin and strigolactone have the capacity to modulate each other's levels and distribution in a dynamic feedback loop required for the co-ordinated control of axillary branching.

Despite the considerable efforts directed over the past four decades to investigate, via correlative physiology and genetic engineering, the contribution of osmolytes to osmotolerance of higher plants, their physiological function(s) still remain a matter of debate. Here, we summarize the functions of proline and glycine betaine, recently documented in our group by using canola (Brassica napus L. var. oleifera), leaf discs treated in vitro with medium of low or high osmolarities. These discs accumulate proline when subjected to osmotic upshifts, whereas proline is rapidly consumed when the discs are further treated under non-stressing conditions. The period of time needed to osmoregulate the amount of proline is assumed to be too long to assign to the proline response a relevant function in turgor regulation which can be achieved with carbohydrates. In addition, proline accumulation is suppressed by supplying the osmotically stressing medium with polyamines, phyto-oxylipins and glycine betaine with no detrimental effect on the capacity to cope with osmotic stress. In opposition to proline, glycine betaine does not occur in canola. We take advantage of this situation to test the compatibility of exogenously supplied betaine in this plant material. Comparisons are done with spinach (Spinacia oleracea L.) leaf discs, which contain an endogenous amount of betaine and also exhibit the capacity to absorb a large amount of it. The betaine absorbed is not degraded and thus cannot be considered in this situation as a true osmoregulator. In turgid tissues of plants that do not produce betaine, this compound induces an elevated level of glutamine, a decreased level of protein, and photo-dependent glycine accumulation. This can rely on senescent-like effects and on a blockage in the glycolate cycle. Such damages to nitrogen metabolism are not observed in leaf explants treated simultaneously with betaine and the stressing agent. We also find that the treatments of turgid canola leaf discs with proline or betaine induce important changes in polyamine contents, mimicking, but with a higher order of magnitude, the effects of osmotic stress. Interestingly, this inducing activity is not observed when osmolytes are supplied under osmotic upshift conditions. Antagonistic effects between osmolytes and osmotic stress are described for the first time in higher plants. They suggest that proline and betaine can act as signal molecules in counteracting destabilizing effects to polyamine metabolism and lead us to decipher other counter-stress effects of nitrogenous osmolytes which, obviously, do not behave just as inert osmobalancers.

Proline (Pro) accumulation occurs in various plant organisms in response to environmental stresses. To identify the signaling components involved in the regulation of Pro metabolism upon water stress in Arabidopsis (Arabidopsis thaliana), a pharmacological approach was developed. The role of phosphoinositide-specific phospholipases C (PLCs) in Pro accumulation was assessed by the use of the aminosteroid U73122, a commonly employed specific inhibitor of receptor-mediated PLCs. We found that U73122 reduced pyrroline-5-carboxylate synthetase transcript and protein as well as Pro levels in salt-treated seedlings. Inhibition of PLC activity by U73122 was quantified by measuring the decrease of inositol 1,4,5-trisphosphate (InsP 3 ) levels. Moreover, the utilization of diacylglycerol kinase and InsP 3 -gated calcium release receptor inhibitors suggested that InsP 3 or its derivatives are essential for Pro accumulation upon salt stress, involving calcium as a second messenger in ionic stress signaling. This observation was further supported by a partial restoration of Pro accumulation in salt-and U73122-treated seedlings after addition of extracellular calcium, or when calcium homeostasis was perturbed by cyclopiazonic acid, a blocker of plant type IIA calcium pumps. Taken together, our data indicate that PLC-based signaling is a committed step in Pro biosynthesis upon salinity but not in the case of mannitol stress. Calcium acts as a molecular switch to trigger downstream signaling events. These results also demonstrated the specific involvement of lipid signaling pathway to discriminate between ionic and nonionic stresses.

Brefeldin A (BFA) inhibits exocytosis but allows endocytosis, making it a valuable agent to identify molecules that recycle at cell peripheries. In plants, formation of large intracellular compartments in response to BFA treatment is a unique feature of some, but not all, cells. Here, we have analyzed assembly and distribution of BFA compartments in development-and tissue-specific contexts of growing maize (Zea mays) root apices. Surprisingly, these unique compartments formed only in meristematic cells of the root body. On the other hand, BFA compartments were absent from secretory cells of root cap periphery, metaxylem cells, and most elongating cells, all of which are active in exocytosis. We report that cell wall pectin epitopes counting rhamnogalacturonan II dimers cross-linked by borate diol diester, partially esterified (up to 40%) homogalacturonan pectins, and (134)-␤-d-galactan side chains of rhamnogalacturonan I were internalized into BFA compartments. In contrast, Golgi-derived secretory (esterified up to 80%) homogalacturonan pectins localized to the cytoplasm in control cells and did not accumulate within characteristic BFA compartments. Latrunculin B-mediated depolymerization of F-actin inhibited internalization and accumulation of cell wall pectins within intracellular BFA compartments. Importantly, cold treatment and protoplasting prevented internalization of wall pectins into root cells upon BFA treatment. These observations suggest that cell wall pectins of meristematic maize root cells undergo rapid endocytosis in an F-actin-dependent manner. 228 -739004. Article, publication date, and citation information can be found at www.plantphysiol.org/cgi/

The legume-rhizobium symbiosis is initiated through activation of the Nod factor-signaling cascade, leading to a rapid reprogramming of host cell developmental pathways. In this work we combine transcriptome sequencing (RNA-seq) with molecular genetics and network analysis to quantify and categorize the transcriptional changes occurring in roots of barrel medic (Medicago truncatula) from minutes to days after inoculation with Sinorhizobium medicae. To identify the nature of the inductive and regulatory cues, we employed mutants with absent or decreased Nod factor sensitivities (nfp and lyk3, respectively) and an ethylene (ET)-insensitive, Nod factor hypersensitive mutant (skl). This unique dataset encompasses nine time points allowing observation of the symbiotic regulation of diverse biological processes with high temporal resolution. Among the many outputs of the study is the early Nod factor-induced, ET-regulated expression of ET signaling and biosynthesis genes. These results su...

Eleven samples of commercially available Greek wines were analyzed in order to determine the phenolic content. For the analysis, reversed phase-high performance liquid chromatography (RP-HPLC) coupled with diode array detection was used. The most abundant phenolic substance detected was (+)-catechin (11.8-40 mg l À1 ). Red wines contained more phenolic substances than white ones. The effect of aging in oak wood barrels on phenolics was also investigated. Specific types of Greek white wines, ''retsina'', were also analyzed. Pitch, which is added to such types of wines, contributes significantly to the increased amount of phenolic compounds present in the wine.