Effects of Two Strains of Tobacco Mosaic Virus on Photosynthetic Characteristics and Nitrogen Partitioning in Leaves of Nicotiana tabacum cv Xanthi during Photoacclimation under Two Nitrogen Nutrition Regimes

Photoacclimation was studied in tobacco leaves (Nicotiana tabacum cv Xanthi) infected with two strains of tobacco mosaic virus (TMV) and grown under different light and nitrogen nutrition regimes. Photosynthetic acclimation measured by the quantum yield and the maximum rate in saturating light of CO2-saturated photosynthesis was impaired to a greater extent in tobacco leaves infected with TMV strain PV230 than in those infected with TMV strain PV42. Infection with TMV strain PV230 severely impaired photosynthetic acclimation at high light/low nitrogen and during transfer from low to high light. Expanding leaves showing chlorotic-mosaic symptoms had greatly reduced capacity to acclimate to high light compared with controls and with developed leaves without visible symptoms. We conclude that the failure of expanding leaves to acclimate was largely due to the destruction of chloroplasts in yellow areas of the tissue, accompanied by severe reduction in ribulose-1,5-bisphosphate carboxylase/oxygenase levels, and corresponding reduction in photosynthesis on a leaf-area basis. When corrected for areas of healthy green tissue, photoacclimation of infected leaves was the same as that of controls. Visible symptom development was greatest in high light/low nitrogen treatments. In developed leaves without visible symptoms, virus accumulation, which was as extensive as in expanding leaves, accelerated senescence and impaired photoacclimation during transfer from low light to high light. Generally, infection with TMV strain PV42 did not impair photosynthetic acclimation and even enhanced it in some treatments, even though virus accumulated to the same concentration as in PV230-infected leaves. These data show that TMV does not simply impair photoacclimation in tobacco by competing with chloroplasts for leaf nitrogen reserves. Rather, specific properties of severe strains, such as PV230, which lead to visible symptom development and patchy loss of photosynthetic activity in expanding leaves as well as general acceleration of chloroplast senescence in developed leaves, contribute to impaired photoacclimation, which is generally exacerbated by low nitrogen nutrition.     

Susceptibility of Tobacco Leaves to Photoinhibition following Infection with Two Strains of Tobacco Mosaic Virus under Different Light and Nitrogen Nutrition Regimes

Sensitivity to photoinhibition under high light stress (2000 [mu]mol photons m-2 s-1 for 2 h in air) and recovery from this stress were examined in leaves of control, uninfected tobacco (Nicotiana tabacum cv Xanthi) leaves and in leaves in tobacco plants infected with tobacco mosaic virus (TMV) when grown under low light (150-200 [mu]mol photons m-2 s-1) or high light (1200 [mu]mol photons m-2 s-1) with high (8.0 mM) or low (0.5 mM) nitrate supply. Photoinhibition was monitored using the dark-adapted fluorescence parameters variable fluorescence/maximum fluorescence, an indicator of photosynthetic efficiency that correlated well with the quantum yield of photosynthetic oxygen evolution, and initial fluorescence, potentially an indicator of photoinhibitory damage. Susceptibility to photoinhibition was greater in low light- and low nitrogen-grown control plants than in high light- or high nitrogen-treated plants. Compared with uninfected controls, infection with the masked strain PV42 increased susceptibility to photoinhibition only in plants grown under low light/low nitrogen conditions. In expanding leaves, infection with severe strain TMV PV230 markedly accelerated photoinhibition under these conditions and under high light/low nitrogen conditions, even before visible symptoms were evident. High nitrogen levels during growth protected against this accelerated photoinhibitory response to virus infection during light stress and generally promoted recovery, at least prior to symptom development. As symptoms developed, the yellow regions provided evidence for chronic photoinhibitory damage, prior to and during the stress treatment, irrespective of growth conditions. Green regions of leaves showing visible symptoms were generally indistinguishable from control, uninfected plants during photoinhibitory stress and recovery. In developed leaves that remained free of visible symptoms during the experiments, in spite of the accumulation of about the same amounts of virus protein (S. Balachandran, C.B. Osmond, A. Makino [1994] Plant Physiol 104: 1043-1050) infection led to an acceleration of photoinhibition during stress treatments, especially in low light/low nitrogen treatments, in which chronic photoinhibitory damage was evident. These studies suggest a role for photoinhibitory damage in the acceleration of visible symptom development following TMV PV230 infection of expanding leaves, as well as in acceleration of senescence in developed leaves without visible symptoms.     

Low codon bias and high rates of synonymous substitution in Drosophila hydei and D. melanogaster histone genes

We have evaluated codon usage bias in Drosophila histone genes and have obtained the nucleotide sequence of a 5,161-bp D. hydei histone gene repeat unit. This repeat contains genes for all five histone proteins (H1, H2a, H2b, H3, and H4) and differs from the previously reported one by a second EcoRI site. These D. hydei repeats have been aligned to each other and to the 5.0-kb (i.e., long) and 4.8-kb (i.e., short) histone repeat types from D. melanogaster. In each species, base composition at synonymous sites is similar to the average genomic composition and approaches that in the small intergenic spacers of the histone gene repeats. Accumulation of synonymous changes at synonymous sites after the species diverged is quite high. Both of these features are consistent with the relatively low codon usage bias observed in these genes when compared with other Drosophila genes. Thus, the generalization that abundantly expressed genes in Drosophila have high codon bias and low rates of silent substitution does not hold for the histone genes.      

Wax as a Mechanism for Protection against Photoinhibition — A Study of Cotyledon orbiculata

The leaves of the Crassulacean acid metabolism plant Cotyledon orbiculata have a waxy coating which is highly reflective but can be easily removed by brushing. This provided an ideal system in which to investigate the role of epidermal wax as a possible photoprotectant. Removal of the wax, prior to exposure to natural sunlight, resulted in substantial decreases in F_v/F_m and in severe cases evidence of photoinhibitory damage, as indicated by a rise in F_o. Leaves from which wax had been removed also showed higher conversion of violaxanthin to zeaxanthin than waxed leaves. Recovery of brushed leaves over a 12 day period was correlated with an increase in the total pool of xanthophyll cycle components. This study suggests that the presence of highly reflective wax on the epidermis may confer significant photoprotection to plants exposed to high solar radiation environments.     

Measurements of the Engagement of Cyanide-Resistant Respiration in the Crassulacean Acid Metabolism Plant Kalanchoë daigremontiana with the Use of On-Line Oxygen Isotope Discrimination

Discrimination against ¹⁸O during dark respiration in tissues of Kalanchoë daigremontiana, Medicago sativa, and Glycine max was measured using an on-line system that enabled direct measurements of the oxygen fractionation of samples in a gas-phase leaf disk electrode unit. Discrimination factors for cytochrome pathway respiration were 18.6 to 19.8%_o for all tissues. However, discrimination in cyanide-resistant respiration was significantly higher in green tissues (30.4-31.2%_o) compared with nongreen tissues (25.3-25.9%_o). Using these discrimination factors, the partitioning of electron transport to these pathways was calculated from measurements of discrimination in the absence of inhibitors. Changes in flux through the alternative pathway were measured during the light and dark phases of Crassulacean acid metabolism in leaf disks of K. daigremontiana. The flux of electrons through the alternative pathway was higher during deacidification than during the other phases of Crassulacean acid metabolism. The increase in alternative pathway electron flux accounted for all of the increased respiration in the light phase. Despite this increase, simultaneous measurements of malate concentration and respiratory flux confirm that only a small proportion of the total malate decarboxylation occurs in the mitochondria.     

Adaptive responses by mouse early embryos to maternal diet protect fetal growth but predispose to adult onset disease

Poor maternal nutrition during pregnancy can alter postnatal phenotype and increase susceptibility to adult cardiovascular and metabolic diseases. However, underlying mechanisms are largely unknown. Here, we show that maternal low protein diet (LPD), fed exclusively during mouse preimplantation development, leads to offspring with increased weight from birth, sustained hypertension, and abnormal anxiety-related behavior, especially in females. These adverse outcomes were interrelated with increased perinatal weight being predictive of later adult overweight and hypertension. Embryo transfer experiments revealed that the increase in perinatal weight was induced within blastocysts responding to preimplantation LPD, independent of subsequent maternal environment during later pregnancy. We further identified the embryo-derived visceral yolk sac endoderm (VYSE) as one mediator of this response. VYSE contributes to fetal growth through endocytosis of maternal proteins, mainly via the multiligand megalin (LRP2) receptor and supply of liberated amino acids. Thus, LPD maintained throughout gestation stimulated VYSE nutrient transport capacity and megalin expression in late pregnancy, with enhanced megalin expression evident even when LPD was limited to the preimplantation period. Our results demonstrate that in a nutrient-restricted environment, the preimplantation embryo activates physiological mechanisms of developmental plasticity to stablize conceptus growth and enhance postnatal fitness. However, activation of such responses may also lead to adult excess growth and cardiovascular and behavioral diseases.     

Conceival,a novel noncontraceptive vaginal vehicle for lipophilic microbicides

The objective of this study was to develop a nontoxic and noncontraceptive vaginal drug delivery vehicle for lipophilic anti-human immunodeficiency virus (HIV) microbicides. Three representative poorly water-soluble novel broad-spectrum anti-HIV microbicides, PHI-113, PHI-346, and PHI-443, were evaluated in 11 different solvent systems. Based on their solubility profiles, a novel non-spermicidal self-emulsifying gel (viz Conceival) composed of pharmaceutical excipients, sorbitol, polyethylene glycol 400, polysorbate 80, microcrystalline cellulose, xanthan gum, and water was optimized. Conceival enhanced the solubility of these poorly water-soluble (<0.001 mg/mL) anti-HIV drugs by at least 150- to 270-fold. Conceival was evaluated in vivo in the New Zealand white rabbit model for the preservation of sperm function based on pregnancy outcome and the potential for vaginal irritation following single and multiple intravaginal applications, respectively. Conceival administered intravaginally immediately prior to artificial insemination with semen had no adverse effects on subsequent reproductive performance, neonatal survival, or pup development when compared with untreated control group. Histologic evaluation of vaginal tissues of rabbits exposed intravaginally to Conceival for 14 consecutive days revealed lack of epithelial, submucosal, and vascular changes at the gel application site (total irritation score <3 out of a possible 16). These findings indicate that Conceival has potential to become a clinically useful, safe noncontraceptive vaginal vehicle for lipophilic microbicides. Published: September 20, 2005     

GPCR screening via ERK 1/2: a novel platform for screening G protein-coupled receptors

Discovery of novel agonists and antagonists for G protein-coupled receptors (GPCRs) relies heavily on cell-based assays because determination of functional consequences of receptor engagement is often desirable. Currently, there are several key parameters measured to achieve this, including mobilization of intracellular Ca2+ and formation of cyclic adenosine monophosphate or inositol triphosphate. However, no single assay platform is suitable for all situations, and all of the assays have limitations. The authors have developed a new high-throughput homogeneous assay platform for GPCR discovery as an alternative to current assays, which employs detection of phosphorylation of the key signaling molecule p42/44 MAP kinase (ERK 1/2). The authors show that ERK 1/2 is consistently activated in cells stimulated by Gq-coupled GPCRs and provides a new high-throughput platform for screening GPCR drug candidates. The activation of ERK 1/2 in Gq-coupled GPCR systems generates comparable pharmacological data for receptor agonist and antagonist data obtained by other GPCR activation measurement techniques.     

Slowly reversible de-epoxidation of lutein-epoxide in deep shade leaves of a tropical tree legume may 'lock-in' lutein-based photoprotection during acclimation to strong light

The kinetics of response to strong light have been examined in deeply shaded leaves of the tropical tree legume (Inga sp.) which have extraordinarily high levels of the alpha-xanthophyll lutein-epoxide that are co-located in pigment-protein complexes of the photosynthetic apparatus with the beta-xanthophyll violaxanthin. As in other species, rapidly reversible photoprotection (measured as non-photochemical chlorophyll fluorescence quenching) is initiated within the time frame of sun-flecks (minutes), before detectable conversion of violaxanthin to antheraxanthin or zeaxanthin. Photoprotection is stabilized within hours of exposure to strong light by simultaneously engaging the reversible violaxanthin cycle and a slowly reversible conversion of lutein-epoxide to lutein. It is proposed that this lutein 'locks in' a primary mechanism of photoprotection during photoacclimation in this species, converting efficient light-harvesting antennae of the shade plant into potential excitation dissipating centres. It is hypothesized that lutein occupies sites L2 and V1 in light-harvesting chlorophyll protein complexes of photosystem II, facilitating enhanced photoprotection through the superior singlet and/or triplet chlorophyll quenching capacity of lutein.