VPg of murine norovirus binds translation initiation factors in infected cells

BACKGROUND: Norovirus genomic and subgenomic RNAs are covalently linked at the 5' nucleotide to a 15 kD protein called VPg. VPg of two human norovirus strains binds translation initiation factor eIF3 and other eIFs in vitro, suggesting VPg functions in initiation of protein synthesis on viral RNA. Human norovirus strains are not cultivable, and thus experimental evidence of interactions between VPg and eIFs in infected cells has been lacking. We used the cultivable murine norovirus MNV-1 as a model to study interactions between VPg and eIFs in infected cells. RESULTS: As shown previously for human norovirus VPg, MNV-1 VPg bound eIF3, eIF4GI, eIF4E, and S6 ribosomal protein in cell extracts by GST pull-down assay. Importantly, MNV-1 VPg co-precipitated eIF4GI and eIF4E from infected macrophages, providing evidence that VPg interacts with components of the translation machinery in norovirus infected cells. CONCLUSION: The interactions between MNV-1 VPg and eIFs completely mimic those reported for the human norovirus VPg, illustrating the utility of MNV-1 as a relevant molecular model to study mechanisms of human norovirus replication

Natural products that reduce rotavirus infectivity identified by a cell-based moderate-throughput screening assay

BACKGROUND: There is widespread interest in the use of innate immune modulators as a defense strategy against infectious pathogens. Using rotavirus as a model system, we developed a cell-based, moderate-throughput screening (MTS) assay to identify compounds that reduce rotavirus infectivity in vitro, toward a long-term goal of discovering immunomodulatory agents that enhance innate responses to viral infection. RESULTS: A natural product library consisting of 280 compounds was screened in the assay and 15 compounds that significantly reduced infectivity without cytotoxicity were identified. Time course analysis of four compounds with previously characterized effects on inflammatory gene expression inhibited replication with pre-treatment times as minimal as 2 hours. Two of these four compounds, α-mangostin and 18-β-glycyrrhetinic acid, activated NFκB and induced IL-8 secretion. The assay is adaptable to other virus systems, and amenable to full automation and adaptation to a high-throughput format. CONCLUSION: Identification of several compounds with known effects on inflammatory and antiviral gene expression that confer resistance to rotavirus infection in vitro suggests the assay is an appropriate platform for discovery of compounds with potential to amplify innate antiviral responses

Rotavirus infection activates the UPR but modulates its activity

<p>Abstract</p> <p>Background</p> <p>Rotaviruses are known to modulate the innate antiviral defense response driven by IFN. The purpose of this study was to identify changes in the cellular proteome in response to rotavirus infection in the context of the IFN response. We also sought to identify proteins outside the IFN induction and signaling pathway that were modulated by rotavirus infection.</p> <p>Methods</p> <p>2D-DIGE and image analysis were used to identify cellular proteins that changed in levels of expression in response to rotavirus infection, IFN treatment, or IFN treatment prior to infection. Immunofluorescence microscopy was used to determine the subcellular localization of proteins associated with the unfolded protein response (UPR).</p> <p>Results</p> <p>The data show changes in the levels of multiple proteins associated with cellular stress in infected cells, including levels of ER chaperones GRP78 and GRP94. Further investigations showed that GRP78, GRP94 and other proteins with roles in the ER-initiated UPR including PERK, CHOP and GADD34, were localized to viroplasms in infected cells.</p> <p>Conclusions</p> <p>Together the results suggest rotavirus infection activates the UPR, but modulates its effects by sequestering sensor, transcription factor, and effector proteins in viroplasms. The data consequently also suggest that viroplasms may directly or indirectly play a fundamental role in regulating signaling pathways associated with cellular defense responses.</p

Morita Duality and Noncommutative Wilson Loops in Two Dimensions

We describe a combinatorial approach to the analysis of the shape and orientation dependence of Wilson loop observables on two-dimensional noncommutative tori. Morita equivalence is used to map the computation of loop correlators onto the combinatorics of non-planar graphs. Several nonperturbative examples of symmetry breaking under area-preserving diffeomorphisms are thereby presented. Analytic expressions for correlators of Wilson loops with infinite winding number are also derived and shown to agree with results from ordinary Yang-Mills theory.Comment: 32 pages, 9 figures; v2: clarifying comments added; Final version to be published in JHE

N-Acetylcysteine Increases the Frequency of Bone Marrow Pro-B/Pre-B Cells, but Does Not Reverse Cigarette Smoking-Induced Loss of This Subset

We previously showed that mice exposed to cigarette smoke for three weeks exhibit loss of bone marrow B cells at the Pro-B-to-pre-B cell transition, but the reason for this is unclear. The antioxidant N-acetylcysteine (NAC), a glutathione precursor, has been used as a chemopreventive agent to reduce adverse effects of cigarette smoke exposure on lung function. Here we determined whether smoke exposure impairs B cell development by inducing cell cycle arrest or apoptosis, and whether NAC treatment prevents smoking-induced loss of developing B cells.Groups of normal mice were either exposed to filtered room air or cigarette smoke with or without concomitant NAC treatment for 5 days/week for three weeks. Bone marrow B cell developmental subsets were enumerated, and sorted pro-B (B220(+)CD43(+)) and pre-B (B220(+)CD43(-)) cell fractions were analyzed for cell cycle status and the percentage of apoptotic cells. We find that, compared to sham controls, smoke-exposed mice have ∼60% fewer pro-B/pre-B cells, regardless of NAC treatment. Interestingly, NAC-treated mice show a 21-38% increase in total bone marrow cellularity and lymphocyte frequency and about a 2-fold increase in the pro-B/pre-B cell subset, compared to sham-treated controls. No significant smoking- or NAC-dependent differences were detected in frequency of apoptotic cells or the percentage cells in the G1, S, or G2 phases of the cycle.The failure of NAC treatment to prevent smoking-induced loss of bone marrow pre-B cells suggests that oxidative stress is not directly responsible for this loss. The unexpected expansion of the pro-B/pre-B cell subset in response to NAC treatment suggests oxidative stress normally contributes to cell loss at this developmental stage, and also reveals a potential side effect of therapeutic administration of NAC to prevent smoking-induced loss of lung function

Rotavirus NSP1 Inhibits NFκB Activation by Inducing Proteasome-Dependent Degradation of β-TrCP: A Novel Mechanism of IFN Antagonism

Mechanisms by which viruses counter innate host defense responses generally involve inhibition of one or more components of the interferon (IFN) system. Multiple steps in the induction and amplification of IFN signaling are targeted for inhibition by viral proteins, and many of the IFN antagonists have direct or indirect effects on activation of latent cytoplasmic transcription factors. Rotavirus nonstructural protein NSP1 blocks transcription of type I IFNα/β by inducing proteasome-dependent degradation of IFN-regulatory factors 3 (IRF3), IRF5, and IRF7. In this study, we show that rotavirus NSP1 also inhibits activation of NFκB and does so by a novel mechanism. Proteasome-mediated degradation of inhibitor of κB (IκBα) is required for NFκB activation. Phosphorylated IκBα is a substrate for polyubiquitination by a multisubunit E3 ubiquitin ligase complex, Skp1/Cul1/F-box, in which the F-box substrate recognition protein is β-transducin repeat containing protein (β-TrCP). The data presented show that phosphorylated IκBα is stable in rotavirus-infected cells because infection induces proteasome-dependent degradation of β-TrCP. NSP1 expressed in isolation in transiently transfected cells is sufficient to induce this effect. Targeted degradation of an F-box protein of an E3 ligase complex with a prominent role in modulation of innate immune signaling and cell proliferation pathways is a unique mechanism of IFN antagonism and defines a second strategy of immune evasion used by rotaviruses

Using global team science to identify genetic parkinson's disease worldwide.

No abstract available

Case Reports1. A Late Presentation of Loeys-Dietz Syndrome: Beware of TGFβ Receptor Mutations in Benign Joint Hypermobility

Background: Thoracic aortic aneurysms (TAA) and dissections are not uncommon causes of sudden death in young adults. Loeys-Dietz syndrome (LDS) is a rare, recently described, autosomal dominant, connective tissue disease characterized by aggressive arterial aneurysms, resulting from mutations in the transforming growth factor beta (TGFβ) receptor genes TGFBR1 and TGFBR2. Mean age at death is 26.1 years, most often due to aortic dissection. We report an unusually late presentation of LDS, diagnosed following elective surgery in a female with a long history of joint hypermobility. Methods: A 51-year-old Caucasian lady complained of chest pain and headache following a dural leak from spinal anaesthesia for an elective ankle arthroscopy. CT scan and echocardiography demonstrated a dilated aortic root and significant aortic regurgitation. MRA demonstrated aortic tortuosity, an infrarenal aortic aneurysm and aneurysms in the left renal and right internal mammary arteries. She underwent aortic root repair and aortic valve replacement. She had a background of long-standing joint pains secondary to hypermobility, easy bruising, unusual fracture susceptibility and mild bronchiectasis. She had one healthy child age 32, after which she suffered a uterine prolapse. Examination revealed mild Marfanoid features. Uvula, skin and ophthalmological examination was normal. Results: Fibrillin-1 testing for Marfan syndrome (MFS) was negative. Detection of a c.1270G > C (p.Gly424Arg) TGFBR2 mutation confirmed the diagnosis of LDS. Losartan was started for vascular protection. Conclusions: LDS is a severe inherited vasculopathy that usually presents in childhood. It is characterized by aortic root dilatation and ascending aneurysms. There is a higher risk of aortic dissection compared with MFS. Clinical features overlap with MFS and Ehlers Danlos syndrome Type IV, but differentiating dysmorphogenic features include ocular hypertelorism, bifid uvula and cleft palate. Echocardiography and MRA or CT scanning from head to pelvis is recommended to establish the extent of vascular involvement. Management involves early surgical intervention, including early valve-sparing aortic root replacement, genetic counselling and close monitoring in pregnancy. Despite being caused by loss of function mutations in either TGFβ receptor, paradoxical activation of TGFβ signalling is seen, suggesting that TGFβ antagonism may confer disease modifying effects similar to those observed in MFS. TGFβ antagonism can be achieved with angiotensin antagonists, such as Losartan, which is able to delay aortic aneurysm development in preclinical models and in patients with MFS. Our case emphasizes the importance of timely recognition of vasculopathy syndromes in patients with hypermobility and the need for early surgical intervention. It also highlights their heterogeneity and the potential for late presentation. Disclosures: The authors have declared no conflicts of interes