Is GABA involved in regulating plant growth and development? /

Rapid and large accumulation of GABA (y-aminobutyric acid) in response to a number of plant stresses has been well documented. But the role(s) of GABA in plants is not well defined. In recent years, the possibility of GABA involvement in regulating plant growth and development has been raised. In the present study, this possibility was examined. First, to rapidly and accurately determine GABA levels in plant tissues, a spectrometric method for GABA determination was developed based on a commercially available enzyme Gabase. Seventy mM LaCb almost completely removed water-soluble pigments from plant tissues which greatly interfere with the absorbance reading at 340nm. Inactivation of GAD (glutamate decarboxylase) by immediately adding methanol to a frozen plant tissue powder was suggested to prevent GABA production during extraction. The recovery of GABA with this method was approximately 100%. Second, the relationship between GABA levels and hypocotyl elongation in soybean seedlings was analyzed using different approaches to regulate in vivo GABA levels and the elongation of hypocotyls. The following major observations were made. (1) Mechanical stimulation by stroking elevated GABA levels and concurrently induced a rapid and significant reduction in hypocotyl elongation. (2) External GABA was demonstrated to penetrate into the hypocotyls using '*C-GABA. Application of external GABA elevated in vivo GABA levels, but failed to inhibit hypocotyl elongation. (3) LaCla and blue light irradiation caused an inhibition in the elongation of dark-grown hypocotyls, whereas GABA levels were not significantly affected. (4) Ca^was suggested to be involved in the signal transduction pathway leading from mechanical stimulation to GABA production, as indicated by the ability of La'* to inhibit GABA production in stimulated hypocotyls. (5) Bicuculline, saclofen and baclofen (agonists and antagonists of GABA receptors in animals) had no effect on hypocotyl elongation. It might indicate that GABA-binding components which are structurally similar to animal GABA receptors and functionally capable of regulating plant growth may not exist in plants. Therefore, the conclusion was drawn that GABA alone is not sufficient to inhibit hypocotyl elongation. Third, chloride influx in isolated Asparagus cells was enhanced by lOmM GABA during a 3 hour incubation, but the effect was not specific for GABA. Chloride efflux was not influenced by GABA. Both influx and efflux of chloride were significantly inhibited by NPPB, a chloride channel blocker. These results suggest that GABA does not influence the activity of plant chloride channels.

The production of 4-adminobutyrate in response to treatments reducing cytosolic pH and the regulation of intracellular pH

GABA (4-aminobutyrate) is synthesized through the decarboxylation of LGlu- (L-Glu-+ H+ ---> GABA + C02), and compared to many free amino acids is present in high concentrations in plant cells. GABA levels rise rapidly and dramatically in response to varied stress conditions including anaerobiosis. Recent papers suggest that GABA production and associated H+ consumption are parts of a metabolic pH-stat mechanism which ameliorates the intracellular pH decline associated with anaerobiosis or other treatments. To test this hypothesis GABA production and efflux have been measured in isolated Asparagus sprengeri cells in response to three treatments which potentially cause intracellular acidification. Acid loads were imposed using 60 min of (i) anaerobiosis, (ii) H+/LGlu- cotransport, and (iii) treatment with permeant weak acids (butyric, acetic and propionic). Both intra- and extracellular GABA concentrations increased more than 100% after anaerobiosis, almost 1000% after H+/L-Glu- cotransport (light or dark) and almost 5000/0 after addition of 5 mM butyric acid at pH 5.0. HPLC analysis of amino acids indicates that as GABA concentrations increased in response to butyric acid addition, glutamate concentrations decreased. Time-course studies demonstrated that added butyric acid stimulates GABA production by 2800/0 within 15 seconds. A fluorescent determination of cytosolic pH indicates that addition of butyric or other weak acids resulted in a rapid reduction in cytosolic pH of 0.6 pH units. The half time for the response to butyric acid addition is 2.1 seconds, indicating that the decline in cytosolic pH is rapid enough to account for the rapid stimulation of GABA production. The acid load in response to butyric acid addition was assayed by measurements of 14C-butyric acid uptake. Calculations indicate that GABA production accounted for 45% of the imposed acid load. The biological significance of GABA efflux is not yet understood. The results support the original hypothesis suggesting a role for GABA production in cellular pH regulation.

Impact of different irrigation strategies on grapes and wine quality of four grapevine cultivars (Vitis sp.) in cool climate conditions. An investigation into the relationships among ABA, water status, grape cultivar and wine quality

Niagara Peninsula of Ontario is the largest viticultural area in Canada. Although it is considered to be a cool and wet region, in the last decade many water stress events occurred during the growing seasons with negative effects on grape and wine quality. This study was initiated to understand and develop the best strategies for water management in vineyards and those that might contribute to grape maturity advancement. The irrigation trials investigated the impact of time of initiation (fruit set, lag phase and veraison), water replacement level based on theoretical loss through crop evapotranspiration (ETc; 100,50 and 25%) and different irrigation strategies [partial root zone drying (PRD) versus regulated deficit irrigation (RD!)] on grape composition and wine sensory profiles. The irrigation experiments were conducted in a commercial vineyard (Lambert Vineyards Inc.) located in Niagara-on-the-Lake, Ontario, from 2005 through 2009. The two experiments that tested the combination of different water regimes and irrigation time initiation were set up in a randomized block design as follows: Baco noir - three replicates x 10 treatments [(25%, 50% and 100% of ETc) x (initiation at fruit set, lag phase and veraison) + control]; Chardonnay - three replicates x seven treatments [(25%, 50% and 100% of ETc) x (initiation at fruit set and veraison) + control]. The experiments that tested different irrigation strategies were set up on two cultivars as follows: Sauvignon blanc - four replicates x four treatments [control, fully irrigated (100% ETc), PRD (100% ETc) and RDI (25% ETc)]; Cabemet Sauvignon - four replicates x five treatments [control, fully irrigated (100% ETc), PRD (100% ETc), RDI (50% ETc) and RDI (25% ETc)]. The controls in each experiment were nonirrigated. The irrigation treatments were compared for many variables related to soil water status, vine physiology, berry composition, wine sensory profile, and hormone composition [(abscisic acid (ABA) and its catabolites]. Soil moisture profile was mostly affected by irrigation treatments between 20 and 60 em depth depending on the grapevine cultivar and the regime of water applied. Overall soil moisture was consistently higher throughout the season in 100 and 50% ETc compare to the control. Transpiration rates and leaf temperature as well as shoot growth rate were the most sensitive variables to soil water status. Drip irrigation associated with RDI treatments (50% ETc and 25% ETc) had the most beneficial effects on vine physiology, fruit composition and wine varietal typicity, mainly by maintaining a balance between vegetative and reproductive parts of the vine. Neither the control nor the 100 ETc had overall a positive effect on grape composition and wine sensory typicity. The time of irrigation initiation affected the vine physiology and grape quality, the most positive effect was found in treatments initiated at lag phase and veraison. RDI treatments were overall more consistent in their positive effect on grape composition and wine varietal typicity comparing to PRD treatment. The greatest difference between non-irrigated and irrigated vines in most of the variables studied was found in 2007, the driest and hottest season of the experimental period. Soil water status had a greater and more consistent effect on red grapevine cultivars rather than on white winegrape cultivars. To understand the relationships among soil and plant water status, plant physiology and the hormonal profiles associated with it, abscisic acid (ABA) and its catabolites [phaseic acid (PA), dihydrophaseic acid (DPA), 7-hydroxy-ABA (TOH-ABA), 8' -hydroxy-ABA, neophaseic acid and abscisic acid glucose ester (ABA-GE)] were analyzed in leaves and berries from the Baco noir and Chardonnay irrigation trials over two growing seasons. ABA and some of its catabolites accurately described the water status in the vines. Endogenous ABA and some of its catabolites were strongly affected in Baco noir and Chardonnay by both the water regime (i.e. ET level) and timing of irrigation initiation. Chardonnay grapevines produced less ABA in both leaves and berries compared to Baco noir, which indicated that ABA synthesis is also cultivar dependant. ABA-GE was the main catabolite in treatments with high water deficits, while PA and DPA were higher in treatments with high water status, suggesting that the vine produced more ABA-GE under water deficits to maintain rapid control of the stomata. These differences between irrigation treatments with respect to ABA and catabolites were particularly noticeable in the dry 2007 season. Two trials using exogenous ABA investigated the effect of different concentrations of ABA and organs targeted for spraying, on grape maturation and berry composition of Cabemet Sauvignon grapevines, in two cool and wet seasons (2008-2009). The fIrst experiment consisted of three replicates x three treatments [(150 and 300 mg/L, both applications only on clusters) + untreated control] while the second experiment consisted in three replicates x four treatments [(full canopy, only clusters, and only leaves sprayed with 300 ppm ABA) + untreated control]. Exogenous ABA was effective in hastening veraison, and improving the composition of Cabemet Sauvignon. Ability of ABA to control the timing of grape berry maturation was dependant on both solution concentration and the target organ. ABA affected not only fruit composition but also yield components. Berries treated with ABA had lower weight and higher skin dry mass, which constitutes qualitative aspects desired in the wine grapes. Temporal advancement of ripening through hormonal control can lead to earlier fruit maturation, which is a distinct advantage in cooler areas or areas with a high risk of early frost occurrence. Exogenous ABA could provide considerable benefits to wine industry in terms of grape composition, wine style and schedule activities in the winery, particularly in wet and cool years. These trials provide the ftrst comprehensive data in eastern North America on the response of important hybrid and Vitis vinifera winegrape cultivars to irrigation management. Results from this study additionally might be a forward step in understanding the ABA metabolism, and its relationship with water status. Future research should be focused on ftnding the ABA threshold required to trigger the ripening process, and how this process could be controlled in cool climates.

The neoliberalization of street vending policy in Lima, Peru: the politics of citizenship. property and public space in the production of a new urban marginality

Neoliberalism is having a significant and global impact on political, social and economic life across spaces. This work illustrates how neoliberalism is attempting to change the ways in which the urban poor - particularly those that participate in street vending - use urban spaces in Lima, Peru. Using municipal policies, newspaper articles and local academic texts I argue that there is a changing marginality in Lima that is being experienced by street vendors, and currently in los canas of Lima. In particular, I discuss formalization, a neoliberal strategy in street vending policy, which is used with eradication and social assistance strategies in attempts to re-regulate street vendors.

Trading nitrogen for carbon: Nitrogen and carbon translocation in a plant/fungal (Metarhizium spp.) symbiosis

While nitrogen is critical for all plants, they are unable to utilize organically bound nitrogen in soils. Therefore, the majority of plants obtain useable nitrogen through nitrogen fixing bacteria and the microbial decomposition of organic matter. In the majority of cases, symbiotic microorganisms directly furnish plant roots with inorganic forms of nitrogen. More than 80% of all land plants form intimate symbiotic relationships with root colonizing fungi. These common plant/fungal interactions have been defined largely through nutrient exchange, where the plant receives limiting soil nutrients, such as nitrogen, in exchange for plant derived carbon. Fungal endophytes are common plant colonizers. A number of these fungal species have a dual life cycle, meaning that they are not solely plant colonizers, but also saprophytes, insect pathogens, or plant pathogens. By using 15N labeled, Metarhizium infected, wax moth larvae (Galleria mellonella) in soil microcosms, I demonstrated that the common endophytic, insect pathogenic fungi Metarhizium spp. are able to infect living soil borne insects, and subsequently colonize plant roots and furnish ts plant host with useable, insect-derived nitrogen. In addition, I showed that another ecologically important, endophytic, insect pathogenic fungi, Beauveria bassiana, is able to transfer insect-derived nitrogen to its plant host. I demonstrated that these relationships between various plant species and endophytic, insect pathogenic fungi help to improve overall plant health. By using 13C-labeled CO2, added to airtight plant growth chambers, coupled with nuclear magnetic resosnance spectroscopy, I was able to track the movement of carbon from the atmosphere, into the plant, and finally into the root colonized fungal biomass. This indicates that Metarhizium exists in a symbiotic partnership with plants, where insect nitrogen is exchanged for plant carbon. Overall these studies provide the first evidence of nutrient exchange between an insect pathogenic fungus and plants, a relationship that has potentially useful implications on plant primary production, soil health, and overall ecosystem stability.