A novel mode of action for a microbial-derived immunotoxin: the cytolethal distending toxin subunit B exhibits phosphatidylinositol 3,4,5-triphosphate phosphatase activity

The Actinobacillus actinomycetemcomitans cytolethal distending toxin (Cdt) is a potent immunotoxin that induces G(2) arrest in human lymphocytes. We now show that the CdtB subunit exhibits phosphatidylinositol (PI)-3,4,5-triphosphate phosphatase activity. Breakdown product analysis indicates that CdtB hydrolyzes PI-3,4,5-P(3) to PI-3,4-P(2) and therefore functions in a manner similar to phosphatidylinositol 5-phosphatases. Conserved amino acids critical to catalysis in this family of enzymes were mutated in the cdtB gene. The mutant proteins exhibit reduced phosphatase activity along with decreased ability to induce G(2) arrest. Consistent with this activity, Cdt induces time-dependent reduction of PI-3,4,5-P(3) in Jurkat cells. Lymphoid cells with defects in SHIP1 and/or ptase and tensin homolog deleted on chromosome 10 (PTEN) (such as Jurkat, CEM, Molt) and, concomitantly, elevated PI-3,4,5-P(3) levels were more sensitive to the toxin than HUT78 cells which contain functional levels of both enzymes and low levels of PI-3,4,5-P(3). Finally, reduction of Jurkat cell PI-3,4,5-P(3) synthesis using the PI3K inhibitors, wortmannin and LY290004, protects cells from toxin-induced cell cycle arrest. Collectively, these studies show that the CdtB not only exhibits PI-3,4,5-P(3) phosphatase activity, but also that toxicity in lymphocytes is related to this activity.     

Effect of progestins on serum hormones,semen production,and agonistic behavior in the gerenuk (Litocranius walleri walleri)

The use of progestins to suppress endogenous testosterone production to reduce agonistic behavior and prevent semen production was studied in gerenuk. Five male gerenuk (20 months to 3 years of age), housed as a bachelor group, were treated with 3 monthly injections of medroxyprogesterone acetate (MPA; 2.5–20 mg/kg), followed by a melengestrol acetate implant (MGA; 0.3 g/kg) for 2 months. Blood samples collected monthly were assayed for serum testosterone and cortisol using enzyme‐linked immunoassays. Quantitative behavioral data were collected for 30 min 3/week starting 1 month before treatment. Body weight, testes volume, and semen traits were measured before treatment, after MPA treatment, and after MGA treatment. Results showed lower (P<0.05) mean serum testosterone concentrations after MPA (4.34 pg/ml) and MGA (5.02 pg/ml) treatment compared to pre‐treatment values (65.9 pg/ml) in four of five gerenuk. The remaining sub‐adult gerenuk had low testosterone initially (4.9 pg/ml) that did not decrease further with treatment (1.4 and 7.8 pg/ml for MPA and MGA, respectively). Mean serum cortisol concentrations decreased markedly after treatment with MPA (6.0±3.7 ng/ml) and MGA (0.8±0.3 ng/ml). Cortisol concentrations were regained rapidly post‐treatment (42.8±4.8 ng/ml) and were not significantly different from the pre‐treatment value (60.6±12.6 ng/ml; P>0.05). The mean incidence of combined aggressive/dominant behaviors (horning, sparring, supplanting, threat) was not different before and after treatment. Body weight, total numbers of spermatozoa produced per ejaculate, percent motility, and percent normal spermatozoa declined maximally 8 months after treatment. Mean testes volume decreased (P<0.05) after MGA treatment (10.53 cm³vs. 11.96 cm³ pre‐treatment). Elevated hepatic enzymes and bile acids were seen in three of five animals after progestin treatment and anorexia was noted in two males after MGA implant removal, however two of three males had elevated liver enzymes before progestin treatment began. Results show that reducing serum testosterone concentration does not seem to modify agonistic behavior in bachelor gerenuk groups. Zoo Biol 26:245–257, 2007. © 2007 Wiley‐Liss, Inc.     

Fish oil induced increase in walking distance, but not ankle brachial pressure index, in peripheral arterial disease is dependent on both body mass index and inflammatory genotype

Peripheral arterial disease (PAD) is an atherosclerotic disease. Evidence suggests that atherosclerosis is an inflammatory condition and long chain n-3 fatty acids, found in oily fish and fish oils, have been shown to reduce inflammation. Genetic and lifestyle factors such as body mass index (BMI) also influence inflammation. In this study we have examined the effect of fish oil in patients with claudication secondary to PAD. Fish oil supplementation, providing 1g EPA and 0.7 g DHA per day for 12 weeks, increased walking distance on a treadmill set at 3.2 km/h with a 7% incline. Walking distance to first pain increased from 76.2+/-8.5 m before fish oil to 140.6+/-25.5 m after fish oil (mean+/-SEM, p=0.004) and total distance walked increased from 160.0+/-21.5 m before fish oil to 242.1+/-34.5 m after fish oil (p=0.002). Fish oil supplementation also improved ankle brachial pressure index (ABPI) from 0.599+/-0.017 before fish oil to 0.776+/-0.030 after fish oil (p<0.001). The increase in walking distance was dependent on both BMI and genotype for single nucleotide polymorphisms in the genes encoding the pro-inflammatory cytokines tumour necrosis factor-alpha and interleukin (IL)-1beta and the anti-inflammatory cytokine IL-10 (detected using amplification refractory mutation system polymerase chain reaction). Neither BMI nor any of the genotypes examined affected the ability of fish oil to increase ABPI. The mechanisms by which fish oil affects walking distance and ABPI do not appear to be the same.     

Molecular basis for the functional interaction of dynein light chain with the nuclear-pore complex

Nucleocytoplasmic transport occurs through nuclear pore complexes (NPCs) embedded in the nuclear envelope. Here, we discovered an unexpected role for yeast dynein light chain (Dyn2) in the NPC. Dyn2 is a previously undescribed nucleoporin that functions as molecular glue to dimerize and stabilize the Nup82-Nsp1-Nup159 complex, a module of the cytoplasmic pore filaments. Biochemical analyses showed that Dyn2 binds to a linear motif (termed DID(Nup159)) inserted between the Phe-Gly repeat and coiled-coil domain of Nup159. Electron microscopy revealed that the reconstituted Dyn2-DID(Nup159) complex forms a rigid rod-like structure, in which five Dyn2 homodimers align like 'pearls on a string' between two extented DID(Nup159) strands. These findings imply that the rigid 20 nm long Dyn2-DID(Nup159) filament projects the Nup159 Phe-Gly repeats from the Nup82 module. Thus, it is possible that dynein light chain plays a role in organizing natively unfolded Phe-Gly repeats within the NPC scaffold to facilitate nucleocytoplasmic transport.     

Evaluation of Blast Furnace Slag as a Means of Reducing Metal Availability in a Contaminated Sediment for Beneficial Use Purposes

An attractive option for the management of dredged sediment involves the use of dredged sediment for beneficial use purposes, such as for fill material. Treatment (chemical amendment) of contaminated sediment may be necessary to limit the environmental and human availability (bioaccessibility, leachability, plant uptake) of heavy metals associated with the contaminated sediment before it is placed. A laboratory study was conducted to investigate the effect of admixing a specific chemical amendment (blast furnace slag) with slightly contaminated fresh-water sediment for reducing metal availability. Initial characterization tests of the un-amended sediment showed that the some of the metals analyzed were present in relatively available (non-residual) forms. Although sulfide was present in the un-amended sediment, the amount was not sufficient to bind all of the available metals. A series of metal availability testing methods indicated that the amendment of the sediment with blast furnace slag (4% on a dry weight ratio basis) had the potential to slightly reduce the availability of some, but not all of the available metals associated with the sediment. Results of the column and batch leaching tests showed that leachability of certain metals, such as barium, nickel and zinc, was reduced by the amendment, but the leachability of copper increased. The effect of the amendment for decreasing bioaccessibility for lead and arsenic was not demonstrated. The amended soil had a detrimental effect on most of the plant species that were evaluated. The metal availability results for the plant uptake tests were also mixed, with slightly lower uptake of certain metals by corn grown within the amended sediment.     

Murine Norovirus 1 (MNV1) Replication Induces Translational Control of the Host by Regulating eIF4E Activity during Infection

Protein synthesis is a tightly controlled process responding to several stimuli, including viral infection. As obligate intracellular parasites, viruses depend on the translation machinery of the host and can manipulate it by affecting the availability and function of specific eukaryotic initiation factors (eIFs). Human norovirus is a member of the Caliciviridae family and is responsible for gastroenteritis outbreaks. Previous studies on feline calicivirus and murine norovirus 1 (MNV1) demonstrated that the viral protein, genome-linked (VPg), acts to direct translation by hijacking the host protein synthesis machinery. Here we report that MNV1 infection modulates the MAPK pathway to activate eIF4E phosphorylation. Our results show that the activation of p38 and Mnk during MNV1 infection is important for MNV1 replication. Furthermore, phosphorylated eIF4E relocates to the polysomes, and this contributes to changes in the translational state of specific host mRNAs. We propose that global translational control of the host by eIF4E phosphorylation is a key component of the host-pathogen interaction.     

Submergence of black ash logs to control emerald ash borer and preserve wood for American Indian basketmaking

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Profiling for novel proteomics biomarkers in neurodevelopmental disorders

Protein biomarker discovery from biological fluids, such as serum, has been widely applied to disorders such as cancer and has more recently also been utilized in neuro-psychiatric disorders with relatively clear biological causes, such as Alzheimer's disease and schizophrenia. The application of the associated technologies for the identification of protein biomarker signatures in neurodevelopmental disorders, such as autism spectrum disorder and attention deficit hyperactivity disorder, is comparatively less well established. The aim of this article is to provide an overview of the various protocols available for such analysis, discuss reports in which these techniques have been previously applied in biomarker discovery/validation in neurodevelopmental disorders, and consider the future development of this area of research.     

BOLD temporal dynamics of rat superior colliculus and lateral geniculate nucleus following short duration visual stimulation

Background

The superior colliculus (SC) and lateral geniculate nucleus (LGN) are important subcortical structures for vision. Much of our understanding of vision was obtained using invasive and small field of view (FOV) techniques. In this study, we use non-invasive, large FOV blood oxygenation level-dependent (BOLD) fMRI to measure the SC and LGN''s response temporal dynamics following short duration (1 s) visual stimulation.

Methodology/Principal Findings

Experiments are performed at 7 tesla on Sprague Dawley rats stimulated in one eye with flashing light. Gradient-echo and spin-echo sequences are used to provide complementary information. An anatomical image is acquired from one rat after injection of monocrystalline iron oxide nanoparticles (MION), a blood vessel contrast agent. BOLD responses are concentrated in the contralateral SC and LGN. The SC BOLD signal measured with gradient-echo rises to 50% of maximum amplitude (PEAK) 0.2±0.2 s before the LGN signal (p<0.05). The LGN signal returns to 50% of PEAK 1.4±1.2 s before the SC signal (p<0.05). These results indicate the SC signal rises faster than the LGN signal but settles slower. Spin-echo results support these findings. The post-MION image shows the SC and LGN lie beneath large blood vessels. This subcortical vasculature is similar to that in the cortex, which also lies beneath large vessels. The LGN lies closer to the large vessels than much of the SC.

Conclusions/Significance

The differences in response timing between SC and LGN are very similar to those between deep and shallow cortical layers following electrical stimulation, which are related to depth-dependent blood vessel dilation rates. This combined with the similarities in vasculature between subcortex and cortex suggest the SC and LGN timing differences are also related to depth-dependent dilation rates. This study shows for the first time that BOLD responses in the rat SC and LGN following short duration visual stimulation are temporally different.