Plant virus infection development as affected by heavy metal stress

The work was focused on the investigation of possible dependencies between the development of viral infection in plants and the presence of high heavy metal concentrations in soil. Field experiments have been conducted in order to study the development of systemic tobacco mosaic virus (TMV) infection in Lycopersicon esculentum L. cv. Miliana plants under effect of separate salts of heavy metals Cu, Zn and Pb deposited in soil. As it is shown, simultaneous effect of viral infection and heavy metals in tenfold maximum permissible concentration leads to decrease of total chlorophyll content in experiment plants mainly due to the degradation of chlorophyll a. The reduction of chlorophyll concentration under the combined influence of both stress factors was more serious comparing to the separate effect of every single factor. Plants' treatment with toxic concentrations of lead and zinc leaded to slight delay in the development of systemic TMV infection together with more than twofold increase of virus content in plants that may be an evidence of synergism between these heavy metal's and virus' effects. Contrary, copper although decreased total chlorophyll content but showed protective properties and significantly reduced amount of virus in plants.     

NOX5 NAD(P)H oxidase regulates growth and apoptosis in DU 145 prostate cancer cells

Reactive oxygen species (ROS) appear to play an important role in regulating growth and survival of prostate cancer. However, the sources for ROS production in prostate cancer cells have not been determined. We report that ROS are generated by intact American Type Culture Collection DU 145 cells and by their membranes through a mechanism blocked by NAD(P)H oxidase inhibitors. ROS are critical for growth in these cells, because NAD(P)H oxidase inhibitors and antioxidants blocked proliferation. Components of the human phagocyte NAD(P)H oxidase, p22^phox and gp91^phox, as well as the Ca²⁺ concentration-responsive gp91^phox homolog NOX5 were demonstrated in DU 145 cells by RT-PCR and sequencing. Although the protein product for p22^phox was not detectable, both gp91^phox and NOX5 were identified throughout the cell by immunostaining and confocal microscopy and NOX5 immunostaining was enhanced in a perinuclear location, corresponding to enhanced ROS production adjacent to the nuclear membrane imaged by 2',7'-dichlorofluorescin diacetate oxidation. The calcium ionophore ionomycin dramatically stimulated ferricytochrome c reduction in cell media, further supporting the importance of NOX5 for ROS production. Antisense oligonucleotides for NOX5 inhibited ROS production and cell proliferation in DU 145 cells. In contrast, antisense oligonucleotides to p22^phox or gp91^phox did not impair cell growth. Inhibition of ROS generation with antioxidants or NAD(P)H oxidase inhibitors increased apoptosis in cells. These results indicate that ROS generated by the newly described NOX5 oxidase are essential for prostate cancer growth, possibly by providing trophic intracellular oxidant tone that retards programmed cell death. superoxide anion; diphenylene iodonium; p22^phox; gp91^phox; adenosine 3',5'-cyclic monophosphate response element; caspases     

A highly conserved nuclear gene for low-level phylogenetics: elongation factor-1 alpha recovers morphology-based tree for heliothine moths

Molecular systematists need increased access to nuclear genes. Highly conserved, low copy number protein-encoding nuclear genes have attractive features for phylogenetic inference but have heretofore been applied mostly to very ancient divergences. By virtue of their synonymous substitutions, such genes should contain a wealth of information about lower-level taxonomic relationships as well, with the advantage that amino acid conservatism makes both alignment and primer definition straightforward. We tested this postulate for the elongation factor-1 alpha (EF-1 alpha) gene in the noctuid moth subfamily Heliothinae, which has probably diversified since the middle Tertiary. We sequenced 1,240 bp in 18 taxa representing heliothine groupings strongly supported by previous morphological and allozyme studies. The single most parsimonious gene tree and the neighbor-joining tree for all nucleotides show almost complete concordance with the morphological tree. Homoplasy and pairwise divergence levels are low, transition/transversion ratios are high, and phylogenetic information is spread evenly across gene regions. The EF-1 alpha gene and presumably other highly conserved genes hold much promise for phylogenetics of Tertiary age eukaryote groups.      

Evidence for contractile protein translocation in macrophage spreading, phagocytosis, and phagolysosome formation

Macrophage pseudopodia that surround objects during phagocytosis contain a meshwork of actin filaments and exclude organelles. Between these pseudopodia at the base of developing phagosomes, the organelle exclusion ceases, and lysosomes enter the cell periphery to fuse with the phagosomes. Macrophages also extend hyaline pseudopodia on the surface of nylon wool fibers and secrete lysosomal enzymes into the extracellular medium instead of into phagosomes. To analyze biochemically these concurrent alterations in cytoplasmic architecture, we allowed rabbit lung macrophages to spread on nylon wool fibers and then subjected the adherent cells to shear. This procedure caused the selective release of β-glucoronidase into the extracellular medium and yielded two fractions, cell bodies and isolated pseudopod blebs resembling podosomes, which are plasma-lemma-bounded sacs of cortical cytoplasm. Cytoplasmic extracts of the cell bodies eluted from nylon fibers contained two-thirds less actin-binding protein and myosin, and approximately 20 percent less actin and two-thirds of the other two proteins were accounted for in podosomes. The alterations in protein composition correlated with assays of myosin-associated EDTA-activated adenosine triphosphatase activity, and with a diminution in the capacity of extracts of nylon wool fiber-treated cell bodies to gel, a property dependent on the interaction between actin-binding protein and F-actin. However, the capacity of the remaining actin in cell bodies to polymerize did not change. We propose that actin-binding protein and myosin are concentrated in the cell cortex and particularly in pseudopodia where prominent gelation and syneresis of actin occur. Actin in the regions from which actin-binding protein and myosin are displaced disaggregates without depolymerizing, permitting lysosomes to gain access to the plasmalemma. Translocation of contractile proteins could therefore account for the concomitant differences in organelle exclusion that characterize phagocytosis.     

Effects of PGE1 or PGE2 on luteal function in pseudopregnant rats

Effects of PGE₁ or PGE₂ on luteal function were studied in 163 pseudopregnant rats. PGE₁ (10, 100, or 300μg) given intrauterine every 6 hr did not shorten pseudopregnancy (P < 0.05), however, the same doses of PGE₂ given intrauterine every 6 hr advanced luteolysis (P < 0.05). PGE₁ (100 or 300μg) given every 4 hr intramuscular maintained levels of progesterone in peripheral blood above controls (P < 0.05) while 100 or 300μg of PGE₂ hastened the decline in progesterone (P < 0.05). The antiluteolytic effect of PGE₁ was not via an inhibition of PGF secretion (P < 0.05) by the uterus or by induction of ovulation in treated animals. Moreover, PGE₁ (100, 200, or 500μg) given intramuscular every 4 hr from day 4 of pseudopregnancy until the next proestrus delayed luteal regression around 3 days (P < 0.05). PGE₂ at doses of 100, 200, or 500μg every 4 hr given intramuscular consistently shortened pseudopregnancy (P < 0.05). Lower doses were without effect (P < 0.05). Based on the above data it is concluded that PGE₂ is consistently luteolytic whereas PGE₁ is not luteolytic in pseudopregnant rats and that PGE₁ may be an antiluteolysin.     

The eukaryotic replisome tolerates leading‐strand base damage by replicase switching

The high‐fidelity replicative DNA polymerases, Pol ε and Pol δ, are generally thought to be poorly equipped to replicate damaged DNA. Direct and complete replication of a damaged template therefore typically requires the activity of low‐fidelity translesion synthesis (TLS) polymerases. Here we show that a yeast replisome, reconstituted with purified proteins, is inherently tolerant of the common oxidative lesion thymine glycol (Tg). Surprisingly, leading‐strand Tg was bypassed efficiently in the presence and absence of the TLS machinery. Our data reveal that following helicase–polymerase uncoupling a switch from Pol ε, the canonical leading‐strand replicase, to the lagging‐strand replicase Pol δ, facilitates rapid, efficient and error‐free lesion bypass at physiological nucleotide levels. This replicase switch mechanism also promotes bypass of the unrelated oxidative lesion, 8‐oxoguanine. We propose that replicase switching may promote continued leading‐strand synthesis whenever the replisome encounters leading‐strand damage that is bypassed more efficiently by Pol δ than by Pol ε.     

Novel and functional regulatory SNPs in the promoter region of <Emphasis Type="Italic">FOXP3</Emphasis> gene in a Gabonese population

Parasites exert a selection pressure on their hosts and are accountable for driving diversity within gene families and immune gene polymorphisms in a host population. The overwhelming response of regulatory T cells during infectious challenges directs the host immune system to lose the ability to mount parasite specific T cell responses. The underlying idea of this study is that regulatory single nucleotide polymorphism (SNPs) can cause significant changes in gene expression in functional immune genes. We identified and investigated regulatory SNPs in the promoter region of the FOXP3 gene in a group of Gabonese individuals exposed to a variety of parasitic infections. We identified two novel and one promoter variants in 40 individual subjects. We further validated these promoter variants for their allelic gene expression using transient transfection assays. Two promoter variants, −794 (C/G) and the other variant −734/−540 (C/T) revealed a significant higher expression of the reporter gene at basal level in comparison to the major allele. The identification of SNPs that modify function in the promoter regions could provide a basis for studying parasite susceptibility in association studies.     

Association of CISH -292A/T genetic variant with hepatitis B virus infection

Cytokine-inducible SRC homology 2 domain protein (CISH) is a suppressor of cytokine signaling that controls interleukin-2 signaling pathway. We investigated the single nucleotide polymorphism (SNP) -292A>T in 473 Vietnamese hepatitis B virus (HBV) carriers and 416 healthy controls. CISH variants at -292A>T were associated to HBV infection (Allelic: OR, 1.22 95% CI, 1–1.49; P = 0.04; Recessive: OR, 1.69 95% CI 1.23–2.54; P = 0.007). A gene dose effect for the risk allele -292T was observed (P = 0.04). The level of interleukin 2 and liver enzymes such as alanine transaminase, aspartate transaminase, total bilirubin, and direct bilirubin were not associated to CISH polymorphism at position -292A>T This study associated the vital role of CISH SNP -292A>T variant to hepatitis B virus infection in a Vietnamese population.