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G McFadden

Publications and source records attributed to G McFadden.

At least 37 records · Page 2Linked to original sources
PubMed2000

Myxoma virus expresses a TNF receptor homolog with two distinct functions.

Myxoma virus, a member of the poxvirus family of DNA viruses, encodes many virulence factors to combat and evade the host immune responses. Among the virus-encoded immuno-modulators is M-T2, a tumor necrosis factor receptor (TNF-R) homologue. M-T2 is secreted as monomeric and dimeric species that bind and inhibit rabbit TNF in a species-specific manner. Deletion analysis indicates that the anti-TNF function is mediated by the first three of four cysteine rich domains (CRDs) of M-T2. In addition, the intracellular form of M-T2 has the ability to block virus-induced apoptosis in lymphocytes, and the first two CRDs appear to be sufficient for this function. Although the mechanisms for the anti-TNF and anti-apoptotic functions of M-T2 are not yet fully defined, we postulate that these dual activities of M-T2 are mediated through different functional motifs and abrogate distinct cellular responses to virus infection.

Amino Acid Sequence↗
PubMed1999

Use of chemokine receptors by poxviruses.

Chemokine receptors serve as portals of entry for certain intracellular pathogens, most notably human immunodeficiency virus (HIV). Myxoma virus is a member of the poxvirus family that induces a lethal systemic disease in rabbits, but no poxvirus receptor has ever been defined. Rodent fibroblasts (3T3) that cannot be infected with myxoma virus could be made fully permissive for myxoma virus infection by expression of any one of several human chemokine receptors, including CCR1, CCR5, and CXCR4. Conversely, infection of 3T3-CCR5 cells can be inhibited by RANTES, anti-CCR5 polyclonal antibody, or herbimycin A but not by monoclonal antibodies that block HIV-1 infection or by pertussis toxin. These findings suggest that poxviruses, like HIV, are able to use chemokine receptors to infect specific cell subtypes, notably migratory leukocytes, but that their mechanisms of receptor interactions are distinct.

3T3 Cells↗
PubMed1999

The complete DNA sequence of myxoma virus.

Myxomatosis in European rabbits is a severely debilitating disease characterized by profound systemic cellular immunosuppression and a high rate of mortality. The causative agent, myxoma virus, is a member of the poxvirus family and prototype of the Leporipoxvirus genus. As a major step toward defining the genetic strategies by which the virus circumvents host antiviral responses, the genomic DNA sequence of myxoma virus, strain Lausanne, was determined. A total of 171 open reading frames were assigned to cover the 161.8-kb genome, including two copies each of the 12 genes that map within the 11.5-kb terminal inverted repeats. Database searches revealed a central core of approximately 120 kb that encodes more than 100 genes that exhibit close relationships to the conserved genes of members of other poxvirus genera. Open reading frames with predicted signal sequences, localization motifs, or homology to known proteins with immunomodulatory or host-range functions were examined more extensively for predicted features such as hydrophobic regions, nucleic acid binding domains, ankyrin repeats, serpin signatures, lectin domains. and structural cysteine spacings. As a result, several novel, potentially immunomodulatory proteins have been identified, including a family with multiple ankyrin-repeat domains, an OX-2 like member of the neural cell adhesion molecule family, a third myxoma serpin, a putative chemokine receptor fragment, two natural killer receptor-like species, and a variety of species with domains closely related to diverse host immune regulatory proteins. Coupled with the genomic sequencing of the related leporipoxvirus Shope fibroma virus, this work affirms the existence of a conserved complement of poxvirus-specific core genes and expands the growing repertoire of virus genes that confer the unique capacity of each poxvirus family member to counter the immune responses of the infected host.

Amino Acid Sequence↗
PubMed1999

The complete genome sequence of shope (rabbit) fibroma virus.

We have determined the complete DNA sequence of the Leporipoxvirus Shope fibroma virus (SFV). The SFV genome spans 159.8 kb and encodes 165 putative genes of which 13 are duplicated in the 12.4-kb terminal inverted repeats. Although most SFV genes have homologs encoded by other Chordopoxvirinae, the SFV genome lacks a key gene required for the production of extracellular enveloped virus. SFV also encodes only the smaller ribonucleotide reductase subunit and has a limited nucleotide biosynthetic capacity. SFV preserves the Chordopoxvirinae gene order from S012L near the left end of the chromosome through to S142R (homologs of vaccinia F2L and B1R, respectively). The unique right end of SFV appears to be genetically unstable because when the sequence is compared with that of myxoma virus, five myxoma homologs have been deleted (C. Cameron, S. Hota-Mitchell, L. Chen, J. Barrett, J.-X. Cao, C. Macaulay, D. Willer, D. Evans, and G. McFadden, 1999, Virology 264, 298-318). Most other differences between these two Leporipoxviruses are located in the telomeres. Leporipoxviruses encode several genes not found in other poxviruses including four small hydrophobic proteins of unknown function (S023R, S119L, S125R, and S132L), an alpha 2, 3-sialyltransferase (S143R), a protein belonging to the Ig-like protein superfamily (S141R), and a protein resembling the DNA-binding domain of proteins belonging to the HIN-200 protein family S013L). SFV also encodes a type II DNA photolyase (S127L). Melanoplus sanguinipes entomopoxvirus encodes a similar protein, but SFV is the first mammalian virus potentially capable of photoreactivating ultraviolet DNA damage.

Amino Acid Sequence↗
PubMed1999

Role of the C-terminal RDEL motif of the myxoma virus M-T4 protein in terms of apoptosis regulation and viral pathogenesis.

The purpose of this study was to investigate the significance of the C-terminal RDEL motif of the myxoma virus M-T4 protein in terms of apoptosis regulation and role in viral virulence. To accomplish this, a recombinant myxoma virus was created in which the C-terminal RDEL motif of M-T4 was deleted and a selectable marker (Ecogpt) was inserted immediately downstream. We hypothesized that removal of the RDEL motif from M-T4 would alter the subcellular localization of the protein and provide insight into its antiapoptotic role. Surprisingly, removal of the RDEL motif from M-T4 did not affect localization of the protein within the endoplasmic reticulum (ER), but it did reduce the stability of the mutant protein. Pulse-chase immunoprecipitation and endoglycosidase H analysis coupled with confocal fluorescent light microscopy demonstrated that the M-T4 RDEL(-) mutant protein is retained in the ER like wildtype M-T4 and suggests that the C-terminal RDEL motif is not the sole determinant for M-T4 localization to the ER. Infection of cultured rabbit lymphocytes with the M-T4 RDEL(-) mutant virus results in an intermediate apoptosis phenotype compared with the wildtype and M-T4 knockout mutant viruses. A novel myxomatosis phenotype was observed in European rabbits when infected with the recombinant M-T4 RDEL(-) mutant virus. Rabbits infected with the M-T4 RDEL(-) virus on day 9 postinfection exhibited an exacerbated edematous and inflammatory response at secondary sites of infections, particularly the ears. Our results indicate that the C-terminal RDEL motif may not be solely responsible for retention of M-T4 to the ER and that M-T4 may have a dual function in protecting infected lymphocytes from apoptosis and in modulating the inflammatory response to virus infection.

Amino Acid Motifs↗
PubMed1999

Endosomal/lysosomal retention and degradation of major histocompatibility complex class I molecules is induced by myxoma virus.

The highly immunosuppressive leporipoxvirus myxoma, previously was shown to promote the loss of cell surface class I major histocompatibility complex (MHC I) molecules. Here, we show that myxoma virus induces the loss of both cell surface and intracellular post-Golgi, beta(2)-microglobulin-associated MHC I. Myxoma-induced loss of these MHC I molecules is abrogated by vacuolar ATPase inhibitors, NH(4)Cl, and leupeptin. Furthermore, immunofluorescence microscopic studies reveal that in myxoma-infected cells, beta(2)-microglobulin-associated MHC I accumulates in Lamp-1(+) vesicular structures, suggesting that myxoma virus targets MHC I for degradation in late endosomes and/or lysosomes. These events are regulated by early gene product or products because they occur unabated in cells infected with myxoma virus in the presence of cytosine arabinoside, an inhibitor of DNA synthesis. Studies with baby green monkey kidney cells transfected with wild-type and tail-less forms of a mouse MHC I molecule, H-2L(d), indicate that the MHC I cytoplasmic tail is required for myxoma-induced localization in Lamp-1(+) organelles. Myxoma-induced endocytosis and degradation of MHC I may provide the virus with a means of dispensing with cell surface MHC I molecules that were loaded with peptides derived from viral proteins synthesized early in infection.

Animals↗
PubMed1999

Role of the myxoma virus soluble CC-chemokine inhibitor glycoprotein, M-T1, during myxoma virus pathogenesis.

Myxoma virus is a poxvirus that causes a virulent systemic disease called myxomatosis in European rabbits. Like many poxviruses, myxoma virus encodes a variety of secreted proteins that subvert the antiviral activities of host cytokines. It was recently demonstrated that the myxoma virus M-T1 glycoprotein is a member of a large poxvirus family of secreted proteins that bind CC-chemokines and inhibit their chemoattractant activities in vitro. To determine the biological role of M-T1 in contributing to myxoma virus virulence, we constructed a recombinant M-T1-deletion mutant virus that was defective in M-T1 expression. Here, we demonstrate that M-T1 is expressed continuously during the course of myxoma virus infection as a highly stable 43-kDa glycoprotein and is dispensable for virus replication in vitro. Deletion of M-T1 had no significant effects on disease progression or in the overall mortality rate of infected European rabbits but heightened the localized cellular inflammation in primary tissue sites during the initial 2 to 3 days of infection. In the absence of M-T1 expression, deep dermal tissues surrounding the primary site of virus inoculation showed a dramatic increase in infiltrating leukocytes, particularly monocytes/macrophages, but these phagocytes remained relatively ineffective at clearing virus infection, likely due to the concerted properties of other secreted myxoma virus proteins. We conclude that M-T1 inhibits the chemotactic signals required for the influx of monocytes/macrophages during the acute-phase response of myxoma virus infection in vivo, as predicted by its ability to bind and inhibit CC-chemokines in vitro.

Animals↗
PubMed1999

Detection of transplant vasculopathy in a rat aortic allograft model by fluorescence spectroscopic optical analysis.

BACKGROUND AND OBJECTIVE: Transplant vasculopathy is a leading cause of late cardiac graft loss. We have examined laser-induced fluorescence (LIF) spectroscopy as an optical diagnostic tool for detection of intimal plaque development and inflammatory cellular invasion in a rat model of aortic allograft transplant. STUDY DESIGN/MATERIALS AND METHODS: Infrarenal aortic segments were transplanted from Lewis to Sprague Dawley rats. A range of vasculopathy development was produced by treatment with a viral anti-inflammatory protein. LIF spectra were recorded from the intima of aortic implants at 28 days. Fluorescence intensity was analyzed for correlation with vasculopathy development. RESULTS: Significant differences in LIF intensity at 400-450 nm (P < or = 0.05 by ANOVA) were detected. LIF emission was correlated with plaque growth (R2 = 0.980), vessel narrowing (R2 = 0.964), and cellular invasion (R2 = 0.971) by regression analysis. CONCLUSION: LIF optical analysis provides a nontraumatic diagnostic approach for detection of atherosclerosis prior to cardiac transplant or during development of vasculopathy after transplant.

Animals↗
PubMed1999

Apoptosis: an innate immune response to virus infection.

Viruses can induce apoptosis of infected cells either directly, to assist virus dissemination, or by inadvertently triggering cellular sensors that initiate cell death. Cellular checkpoints that can function as 'alarm bells' to transmit pro-apoptotic signals in response to virus infections include death receptors, protein kinase R, mitochondrial membrane potential, p53 and the endoplasmic reticulum.

Animals↗
PubMed1999

Immunomodulation by viruses: the myxoma virus story.

Myxoma virus is a poxvirus pathogen of rabbits that has evolved to replicate successfully in the presence of an active immune response by an infected host. To accomplish this, the virus has developed a variety of strategies to avoid detection by or obstruct specific aspects of the antiviral response whose consolidated action is antagonistic to virus survival. We describe two distinct viral strategies carried out by viral proteins with which myxoma virus subverts the host immune response. The first strategy is the production of virus-encoded proteins known as viroceptors or virokines that mimic host receptors or cytokines. These seek to actively block extracellular immune signals required for effective virus clearance and produce a local environment in the infected tissue that is "virus friendly". The second strategy, carried out by intracellular viral proteins, seeks to retard the innate antiviral responses such as apoptosis, and hinder attempts by the infected cell to communicate with the cellular arm of the immune system. By studying these viral strategies of immune evasion, the myxoma system can provide insights into virus-host interactions and also provide new insights into the complex immune system.

Amino Acid Sequence↗
PubMed1998

Functional comparisons among members of the poxvirus T1/35kDa family of soluble CC-chemokine inhibitor glycoproteins.

Many poxviruses express a 35-40-kDa secreted protein, termed "T1" (for leporipoxviruses) or "35kDa" (for orthopoxviruses), that binds CC-chemokines with high affinity but is unrelated to any known cellular proteins. Many previously identified poxvirus cytokine-binding proteins display strict species ligand-binding specificity. Because the T1 and 35kDa proteins share only 40% amino acid identity, we compared the abilities of purified myxoma virus-T1 (M-T1) and vaccinia virus (strain Lister)- and rabbitpox virus-35kDa proteins to inhibit human CC-chemokines in vitro. All three proteins were equally effective in preventing several human CC-chemokines from binding to target chemokine receptors and blocking subsequent intracellular calcium release. The inhibitory affinities were comparable (Ki = 0.07-1.02 nM). These proteins also displayed similar abilities to inhibit (IC50 = 6.3-10.5 nM) human macrophage inflammatory protein-1alpha-mediated chemotaxis of human monocytes. None of the viral proteins blocked interleukin-8-mediated calcium flux or chemotaxis of human neutrophils, confirming that the biological specificity of the T1/35kDa family is targeted inhibition of CC-chemokines. Despite the significant sequence divergence between the leporipoxvirus T1 and orthopoxvirus 35kDa proteins, our data suggest that their CC-chemokine binding and inhibitory properties appear to be species nonspecific and that the critical motifs most likely reside within the limited regions of conservation.

Calcium↗
PubMed1998

Inhibitory specificity of the anti-inflammatory myxoma virus serpin, SERP-1.

SERP-1 is a myxoma virus-encoded serpin, secreted from infected cells, that is required for virulence and has anti-inflammatory activity. We report that purified recombinant SERP-1 forms SDS-stable complexes with urokinase-type plasminogen activator (uPA), tissue-type plasminogen activator (tPA), plasmin, thrombin, and factor Xa. N-terminal sequencing confirmed Arg319-Asn320 as the site of reaction. Mutation of these residues to Ala-Ala abolished inhibitory activity but had no effect on the specific cleavage at Thr315-Leu316 seen with elastase and with cathepsin G. Kinetic analysis of the reactions with uPA, tPA, plasmin, thrombin, Xa, and C1s showed second-order rate constants to vary over 3 logs, from kinh = 3 x 10(5) M-1 s-1 with thrombin to approximately 600 M-1 s-1 with C1s, while steady-state inhibition constants ranged from KI = 10 pM with thrombin to approximately 100 nM with C1s. Stoichiometries of inhibition varied between SI = 1.4 +/- 0.1 for uPA to SI = 13 +/- 3 for thrombin. Analysis of the variations in inhibition kinetics shows that when serpins act at low concentrations, comparable with the target protease or with KI (as appears likely for SERP-1 in vivo), inhibitory specificity becomes less dominated by kinh and is increasingly dependent on partitioning within the branched reaction mechanism and on the lifetime of the inhibited complex.

Amino Acid Sequence↗
PubMed1998

Virus-encoded receptors for cytokines and chemokines.

A number of viruses, particularly members of the poxvirus, herpesvirus and retrovirus families, have adapted to the vertebrate immune responses by capturing and modifying cellular genes which regulate the host immune system. Included among these host-derived virus genes are modified versions of receptors for cytokines or chemokines. Most of these receptor homologs, also called viroceptors, are either secreted glycoproteins or are located at the infected cell surface. Although these viroceptors can act in different ways, collectively they function by modifying the cytokine network to the advantage of the virus rather than the host.

Animals↗
PubMed1998

Apoptosis regulators from DNA viruses.

The past year has seen an considerable expansion in knowledge about the field of apoptosis modulators expressed by DNA viruses. These diverse classes of virus-encoded regulators include caspase inhibitors, signal transduction effectors, Bcl-2 homologs, cell cycle control proteins, transcriptional regulators, reactive oxide scavengers, kinases, 'death factors' and novel host-range proteins.

Animals↗