Division of Virology, Institute of Biomedical and Life Sciences, University of Glasgow, Church Street, Glasgow G11 5JR, UK1
School of Animal and Microbial Science, The University of Reading, Whiteknights, PO Box 228, Reading RG6 5AJ, UK2
Complement Biology Group, Dept of Medical Biochemistry, University of Wales College of Medicine, 3rd Floor, Tenovus Building, Heath Park, Cardiff CF14 4XN, UK3
Author for correspondence: David Evans. Fax +44 141 330 6249. e-mail David.Evans{at}vir.gla.ac.uk
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Abstract |
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Introduction |
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The ability of a virus to bind to a target cell does not necessarily result in infection; the latter process may involve secondary events distinct from receptor binding, possibly mediated by additional cellular factors (Alkhatib et al., 1996 ; Bai et al., 1994
; Deng et al., 1996
; Feng et al., 1996
). Evidence is accumulating for the involvement of secondary factors for cell infection by certain picornaviruses, including the echoviruses that bind DAF (Evans, 1997
; Powell et al., 1997
) and coxsackievirus A21 (Shafren et al., 1997a
). Soluble DAF (sDAF) blocks virus binding to the cell surface by steric inhibition, which contrasts with the irreversible conformational changes induced in the poliovirus particle by soluble PVR (Kaplan et al., 1990
), or the rhinovirus particle by soluble derivatives of its receptor, ICAM-1 (Greve et al., 1991
; Hooverlitty & Greve, 1993
). In contrast, echovirus type 7 (EV7) binding of DAF at the cell surface results in the formation of 135S particles (Powell et al., 1997
), the altered sedimentation coefficient of these A particles reflects conformational changes that include the loss of the internal capsid protein VP4, indicative of an uncoating event involved in the infection process (Yafal et al., 1993
). Taken together these results suggest that additional determinants at the cell surface are required for infection by DAF-binding echoviruses. One potential candidate for a secondary factor is
2-microglobulin, antibodies to which block echovirus infection of rhabdomyosarcoma (RD) cells in a cell-specific manner (Ward et al., 1998
). The observed block occurs post-attachment but prior to RNA translation and replication, though the precise mechanism remains unclear. Our studies suggest that EV7 enters a receptor complex at the cell surface that is resistant to proteinase K and sDAF (Powell et al., 1998
; Ward et al., 1998
). Whether this complex consists solely of virus bound to DAF, or also contains the secondary factor(s) required for infection remains to be determined.
An involvement for 2-microglobulin was identified by the cloning and characterization of the ligand for an antibody raised to Ohio HeLa cell membrane fractions that blocked echovirus infection (Ward et al., 1998
). An alternative approach, used in this communication, is to determine the ability of antisera to cell surface proteins, known to co-localize with DAF, to block virus infection. The identification of such proteins may help in the identification of other components of the sDAF-resistant receptor complex that forms during echovirus infection of permissive cells. We report here that polyclonal antiserum to the complement control protein CD59 blocks infection of RD cells by several echoviruses. This block is not mediated at the level of attachment, but during a post-binding event necessary for the uncoating of the virus and the formation of intracellular A particles.
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Methods |
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Antibodies.
Polyclonal rabbit (purified Ig) and murine monoclonal anti-human CD59 antibodies (MEM 43, 43/5, YTH53.1, HelC1, HelC2, A35, 2/24) were obtained from B. P. Morgan, University of Wales College of Medicine, Cardiff, UK and P. J. Sims, Blood Research Institute, Milwaukee, USA. Anti-DAF (Ward et al., 1994 ) monoclonal antibody (MAb) 854 was obtained from P. D. Minor, NIBSC, Potters Bar, UK. Anti-
2-microglobulin MAb 1350 was obtained from Chemicon International, as was MAb PID6 directed against
v integrins, and an anti-CD44 MAb. Anti-enterovirus MAb 5-D8/1 was obtained from Dako, and goat anti-mouse-immunoglobulin (Ig)
-galactosidase was obtained from Harlan Sera-lab. MAbs 308 and DF1513, directed against aminopeptidase N and the transferrin receptor, respectively, were obtained from NeoMarkers. MAbs to CD97 and CD66 were obtained from Pharmingen and D. Fox (University of Reading, UK), and the anti-CD46 MAb J4-48 was obtained from Serotec. Polyclonal antiserum to
5 was kindly provided by B. Cushley (IBLS, University of Glasgow, UK) and the polyclonal antiserum to IgE was purchased from Dako.
Inhibition of infection with anti-CD59 antibodies.
Purified polyclonal anti-CD59 antibody was serially diluted twofold in DMEM and used to treat human RD cells in a 96-well format (105 cells per well) for 1 h at 37 °C. 104 TCID50 of virus was added and infection allowed to proceed for 24 h prior to staining. Soluble recombinant CD59 (sCD59), obtained from B. P. Morgan, University of Wales College of Medicine, Cardiff, UK, was incubated with the antibody for 30 min at room temperature prior to addition to the cells to remove CD59-specific antibodies. MAbs directed against CD59 were tested for their ability to block infection using essentially the same assay. Antibodies were diluted in DMEM, incubated with 105 RD cells for 1 h at 37 °C and washed prior to the addition of 104 TCID50 of virus. MAbs were cross-linked, where appropriate, by the addition of a saturating amount of secondary goat anti-mouse antiserum after washing. Incubation was continued for a further 1 h at 37 °C, at which point 104 TCID50 of virus was added in the presence of a 1/100 or 1/1000 dilution of the original primary MAbs.
Temporal analysis of the anti-CD59 block on infection.
RD cells were infected with EV7 at an m.o.i. of 1. Infection was allowed to proceed for 6 h at 37 °C before the cells were fixed and permeabilized with acetonemethanol. Intracellular virus antigen was detected using an anti-enterovirus VP1-specific MAb (5-D8/1, Dako) at a 1:400 dilution and an anti-mouse -galactosidase conjugate (Harlan Sera-labs). X-Gal was added and the assay allowed to develop overnight at room temperature. The blue product was solubilized by the addition of SDSNaOH (1%, 0·2 M), debris removed by centrifugation, and the absorbance of the samples measured at 560 nm. Parallel samples were treated for various lengths of time with CD59 antiserum by the addition of a 1/100 dilution of antiserum at appropriate times in all media, washes and virus preparation.
Radio-labelled virus binding assay.
Approximately 104 c.p.m. of purified 35S-labelled EV7 was incubated with 5x106 RD cells that had been pre-treated for 1 h with anti-DAF MAb 854 (1:1000 dilution), anti-CD59 polyclonal antiserum (1:100) or the DMEM control. Virus was allowed to bind for 1 h on ice, the cells washed twice with DMEM and the bound radioactivity quantified by scintillation counting. The percentage of bound virus was calculated relative to the mock-treated sample.
Virus entry assay.
Virus entry to RD cells was performed essentially as described previously (Ward et al., 1998 ). RD cells were treated with polyclonal anti-CD59 antiserum (1:100) or DMEM control and the susceptibility of bound radiolabelled virus to competition by sDAF assessed at various times after incubating cells at 37 °C.
Single-step growth curve.
RD cells (5x105) were treated with polyclonal anti-CD59 antiserum (1:100 dilution) or DMEM alone for 1 h at 37 °C. EV7 (m.o.i. of 3) was adsorbed at room temperature for 30 min in the presence or absence of anti-CD59 antiserum, the virus was removed, the cells washed, and either antiserum (1:100) or DMEM added back to the cell monolayers. Samples were removed at various time-points, freezethawed three times and the virus quantified by TCID50.
Cold synchronized eclipse products.
1x107 cells were either mock-treated with DMEM or treated with anti-CD59 antiserum (1:100) for 1 h at 37 °C with constant rotation (6 revs/min). The cells were pelleted and radiolabelled virus (approx. 2x105 c.p.m.) bound for 1 h on ice. Unbound virus was removed by washing and infection was allowed to proceed in the presence or absence of the polyclonal anti-CD59 antiserum for 1 h at 37 °C with constant rotation. Eluted virus was removed and cell-associated virus was released using 0·2% NP-40. Samples were sedimented through a linear 1545% sucrose gradient which was harvested in 1·5 ml fractions and the virus particles quantified by scintillation counting.
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Results |
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Polyclonal rabbit anti-human CD59 antiserum blocked infection of RD cells by similar titres of a range of echoviruses (Fig. 1a). The end-point antiserum titre varied from 1/400 for EV4 to 1/1600 for EV29 and EV6', and was highest for echoviruses that use DAF alone, or DAF and an unidentified receptor for cell entry. However, echoviruses that are known not to use DAF, such as EV4 and EV9, were also blocked at broadly similar end-point titres. The effect of the anti-CD59 antiserum was not batch-dependent, as antiserum from an independent source was also shown to inhibit infection in a similar fashion (data not shown). The specificity of this block was demonstrated in several ways. Poliovirus type 3 and coxsackievirus B2 and B3, which respectively use PVR, CAR and CAR+DAF as receptors (Bergelson et al., 1997
; Mendelsohn et al., 1989
; Shafren et al., 1997b
), were not inhibited from infecting RD cells under similar conditions. To confirm that the block was specific for the anti-CD59 components of the rabbit antiserum, antiserum was pre-incubated with purified soluble CD59 (sCD59). A 1/400 dilution of rabbit antiserum was incubated with varying concentrations of sCD59 and the remaining virus blocking activity tested against EV7 (Fig. 1b
). As little as 20 ng/ml of sCD59 abrogated the ability of anti-CD59 to block infection. Soluble CD59 alone, at a concentration of 200 ng/ml, had no effect upon EV7 infection of RD cells (Fig. 1b
). We investigated the nature of the affinity-purified sCD59 used in these assays to confirm that the blocking activity could not be attributed to a contaminating protein (see also Bodian et al., 1997
). Varying amounts of sCD59 were electrophoresed under reducing conditions and visualized by silver-staining (Fig. 1c
). Three bands were visible, corresponding to a trace of the dimeric form of CD59, the 20 kDa monomeric CD59 protein, and the slightly smaller deglycosylated form. This preparation of CD59 has previously been analysed by Western blot. Other than a small amount of non-glycosylated material, no contaminating proteins are visible (Bodian et al., 1997
), supporting our conclusion that the block to echovirus infection we observe is mediated by antibodies in the polyclonal antiserum specific for CD59.
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Antibodies to CD59 do not affect DAF binding by echovirus
Cell surface proteins involved in virus entry may have a role in virus binding or in a post-binding event required for a later stage of the infection process. To determine whether CD59 was implicated in virus binding we investigated the ability of polyclonal anti-CD59 rabbit antiserum to inhibit the binding of radiolabelled EV6 or EV7 to the surface of RD cells. EV6 was not blocked by either the anti-DAF MAb 854 or the anti-CD59 antiserum (Fig. 2); previous studies have demonstrated that this isolate does interact with DAF, as cell binding can be blocked with sDAF (Powell et al., 1998
). In contrast, although at least 90% of the EV7 binding was blocked by the anti-DAF MAb 854, there was no reduction in binding in the presence of the anti-CD59 antiserum. These results imply that CD59 probably does not function as a secondary attachment molecule.
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Discussion |
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CD59 is an 1820 kDa protein, widely expressed on a range of haematopoietic and non-haematopoietic cells. Like DAF, CD59 is anchored to the cell surface by a GPI tail and functions to regulate the complement cascade, albeit at a later stage than DAF, by interacting with C5b-8 and C5b-9 to block the incorporation and subsequent polymerization of C9 into functional C5b-9 complexes (Morgan & Meri, 1994 ), thereby preventing the formation of the membrane attack complex. The GPI anchor and functional similarity between DAF and CD59 possibly account for the observed similarity in the distribution patterns of the two molecules at the cell surface (Cerny et al., 1996
; Lisanti & Rodriguez-Boulan, 1991
), and prompted us to investigate whether CD59 has a role in echovirus infection.
We demonstrate that polyclonal antiserum to CD59 blocks infection of RD cells by a range of echoviruses, including representatives that use DAF alone for cell binding (e.g. EV7), those that bind DAF and an as yet an unidentified molecule (e.g. EV3, EV6, EV6') and some that do not bind DAF (e.g. EV9, EV4). We have not completed an extensive screen of all echoviruses, but preliminary experiments have shown that EV2, -17, -18 and -20 (all of which bind DAF with the exception of EV2; I. Goodfellow, unpublished data; Bergelson et al., 1994 ) are also blocked by CD59 antiserum (data not shown). Our results indicate that the block does not act by inhibiting virus binding to the cell surface. Neither EV6 or EV7 were inhibited from binding to RD cells in the presence of anti-CD59 antiserum, suggesting that CD59 is probably not involved in virus attachment. Of the viruses tested, the block to RD infection by EV9 always produced a characteristic patchy cytopathic effect, suggesting that although the monolayer was protected from direct infection, virus could still spread by cell-to-cell contact. This phenotype was not observed with the other non-DAF-binding virus tested (EV4) and is currently being investigated.
The specificity of the block by anti-CD59 antiserum was demonstrated by the failure to inhibit polio or coxsackievirus infection of RD cells, and by the ability of competing sCD59 to release the block. Furthermore, independently raised antiserum to CD59 also blocked infection by a range of echoviruses, suggesting that the inhibition was not a consequence of a minor contaminant of the immunogen used to generate the antiserum. Rabbit polyclonal antisera to other proteins expressed on the surface of RD cells, such as CD46 (MCP, a cellular receptor for measles virus) and 5 integrin, did not inhibit echovirus infection. Although polyclonal antiserum to CD59 efficiently blocked infection, none of the panel of anti-CD59 MAbs alone, in combination, or cross-linked, mediated the same effect. However, the MAbs screened were selected as blocking CD59 complement regulation, and are therefore directed against a limited region of CD59, all but MEM43/5 map to the active site of CD59 (Bodian et al., 1997
). It is probable that the polyclonal antiserum recognizes epitopes on CD59 outwith the active site that account for the block in infection. MAbs directed against a limited range of other cell surface markers, including CD44, CD46, CD66, the
2 and
v integrins, MHC-I, the transferrin receptor and aminopeptidase N all failed to block infection of RD cells by EV6, EV7 and EV9 (data not shown).
2-microglobulin has recently been implicated in echovirus infection of RD cells (Ward et al., 1998
) at a post-binding, pre-entry stage, involving the formation of a multi-component complex. Like
2-microglobulin, anti-CD59 antiserum blocked infection in a cell-specific manner, being restricted to RD cells, and having no effect on EV7 infection of Ohio Hela or HT29 cells. Both the latter express comparable levels of CD59 to RD cells, and we would speculate that the failure to block is a consequence of the significantly higher levels of DAF that these two cell lines express (data not shown). This could result in the virus using a route for cell infection that bypasses the requirement for CD59 or
2-microglobulin, thereby masking the blocking effect clearly demonstrable in RD cells.
Although not formally tested, the failure to inhibit EV7 cell infection with sCD59 alone suggests that, in solution at least, the virus does not irreversibly interact with CD59. We have also been unable to demonstrate an interaction between sCD59 and EV7 by surface plasmon resonance, or the binding of radiolabelled EV7 or EV12 to transfected murine cells expressing high levels of human CD59 (I. Goodfellow & B. Spiller, unpublished results). It further suggests that, if EV7 does interact with CD59 within a cell surface receptor complex, either the virus is inaccessible to sCD59 within the complex or sCD59 cannot precisely mimic the function(s) of the GPI-anchored protein, as has been suggested for sDAF (Medof et al., 1984 ; Moran et al., 1992
).
Anti-CD59 blocks EV7 infection of RD cells at an early stage, but does not inhibit the binding of EV7 to DAF. The precise stage at which the inhibition is effective remains to be determined, but our results demonstrate that the bound virus does not undergo the conformational changes that are associated with particle uncoating (Fig. 6). The absence of significant amounts of A particles in the eluted virus fraction (Fig. 6b
) suggests that such particles are not formed in the presence of anti-CD59 antiserum, rather than forming but not being retained at or within the cell. The inability to form A particles in the presence of anti-CD59 antiserum is similar to the inhibitory effect of anti-
2-microglobulin MAbs (T. Ward, unpublished results), though there are qualitative differences between the inhibition observed. In particular, the formation of an sDAF-resistant virusreceptor complex was not inhibited by anti-CD59, whereas we show here and previously that anti-
2-microglobulin retards the formation of this complex which remains partially sensitive to competing sDAF (Fig. 5
; Ward et al., 1998
). Whether this reflects qualitative differences in the reagents used for these experiments, or a true difference in the state of the virusreceptor complex is currently under investigation.
The mechanism by which anti-CD59 antiserum blocks echovirus infection of RD cells remains unclear. Identification of the cellular location of the blocked virusreceptor complexes, which is also unknown, may help determine how antiserum to CD59, and possibly also anti-2-microglobulin, blocks infection. The physical characteristics of the blocked particles suggest that this location occurs at, or before, the site at which A particles form. The resistance of the virusreceptor complexes to high levels of sDAF or protease digestion suggests that they are possibly located in cell surface endocytic compartments, such as clathrin-coated pits or caveolae, or are otherwise not exposed at the cell surface. However, we have previously demonstrated that inhibitors of pit or caveolae function do not prevent EV7 infection of RD cells and suggested that such compartments may not allow virus entry (Ward et al., 1998
). An alternative cellular location, with which both DAF and CD59 associate through the possession of GPI anchors, are lipid rafts. The latter consist of sphingolipid and cholesterol-rich microdomains that can be purified by resistance to non-ionic detergents such as Triton X-100 (Simons & Ikonen, 1997
) and which, although distinct from caveolae, can co-associate under certain conditions (Brown & London, 1998
). In particular, GPI-anchored proteins and glycosphingolipids associate in or near caveolae when cross-linked or clustered (Brown & London, 1998
; Mayor et al., 1994
; Schnitzer et al., 1995
; Wu et al., 1997
), a situation that could arise upon multi-valent binding to the icosahedral virus particle. We are currently investigating the cellular location of DAF and CD59 in the presence or absence of EV7 to determine whether there are distinct modifications in localization following virus binding. We are also screening antibodies to other GPI-anchored proteins to investigate whether these also co-localize with DAF and block infection by both DAF-binding and other enteroviruses. These studies may help determine whether the observed block to virus infection reported here is due to the direct inhibition of a critical stage in the infection pathway, or to a non-specific steric event that occurs due to the similar location of DAF and CD59 at the cell surface.
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Acknowledgments |
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Footnotes |
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References |
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Bai, M., Campisi, L. & Freimuth, P. (1994). Vitronectin receptor antibodies inhibit infection of HeLa and A549 cells by adenovirus type 12 but not by adenovirus type 2.Journal of Virology 68, 5925-5932.[Abstract]
Bergelson, J. M., Chan, M., Solomon, K. R., St John, N. F., Lin, H. & Finberg, R. W. (1994). Decay-accelerating factor (CD55), a glycosylphosphatidylinositol-anchored complement regulatory protein, is a receptor for several echoviruses.Proceedings of the National Academy of Sciences, USA 91, 6245-6249.[Abstract]
Bergelson, J. M., Cunningham, J. A., Droguett, G., Kurt-Jones, E. A., Krithivas, A., Hong, J. S., Horwitz, M. S., Crowell, R. L. & Finberg, R. W. (1997). Isolation of a common receptor for coxsackie B viruses and adenoviruses 2 and 5.Science 275, 1320-1323.
Bodian, D. L., Davis, S. J., Morgan, B. P. & Rushmere, N. K. (1997). Mutational analysis of the active site and antibody epitopes of the complement-inhibitory glycoprotein, CD59.Journal of Experimental Medicine 185, 507-516.
Brown, D. & London, E. (1998). Function of lipid rafts in biological membranes.Annual Review of Cell and Developmental Biology 14, 111-136.[Medline]
Cerny, J., Stockinger, H. & Horejsi, V. (1996). Noncovalent associations of T lymphocyte surface proteins.European Journal of Immunology 26, 2335-2343.[Medline]
Davitz, M. A., Low, M. G. & Nussenzweig, V. (1986). Release of decay-accelerating factor (DAF) from the cell-membrane by phosphatidylinositol-specific phospholipase-c (PIPLC) selective modification of a complement regulatory protein.Journal of Experimental Medicine 163, 1150-1161.[Abstract]
Deng, H. K., Liu, R., Ellmeier, W., Choe, S., Unutmaz, D., Burkhart, M., Di Marzio, P., Marmon, S., Sutton, R. E., Hill, C. M., Davis, C. B., Peiper, S. C., Schall, T. J., Littman, D. R. & Landau, N. R. (1996). Identification of a major co-receptor for primary isolates of HIV-1.Nature 381, 661-666.[Medline]
Evans, D. J. (1997). Picornavirus receptors, tropism and pathogenesis. In Molecular Aspects of HostPathogen Interactions, pp. 23-43. Edited by M. A. McCrae, J. R. Saunders, C. J. Smyth & N. D. Stow. Cambridge: Cambridge University Press.
Evans, D. & Almond, J. (1998). Cell receptors for picornaviruses as determinants of cell tropism and pathogenesis.Trends in Microbiology 6, 198-202.[Medline]
Feng, Y., Broder, C. C., Kennedy, P. E. & Berger, E. A. (1996). HIV-1 entry cofactor functional cDNA cloning of a 7-transmembrane, G-protein-coupled receptor.Science 272, 872-877.[Abstract]
Fricks, C. E. & Hogle, J. M. (1990). Cell-induced conformational change in poliovirus: externalization of the amino terminus of VP1 is responsible for liposome binding.Journal of Virology 64, 1934-1945.[Medline]
Greve, J. M., Forte, C. P., Marlor, C. W., Meyer, A. M., Hoover-Litty, H., Wunderlich, D. & McClelland, A. (1991). Mechanisms of receptor-mediated rhinovirus neutralization defined by two soluble forms of ICAM-1.Journal of Virology 65, 6015-6023.[Medline]
Holland, J. J. (1962). Irreversible eclipse of poliovirus by HeLa cells.Virology 16, 163-176.[Medline]
Hoover-Litty, H. & Greve, J. M. (1993). Formation of rhinovirus-soluble ICAM-1 complexes and conformational changes in the virion.Journal of Virology 67, 390-397.[Abstract]
Kaplan, G., Freistadt, M. S. & Racaniello, V. R. (1990). Neutralization of poliovirus by cell receptors expressed in insect cells.Journal of Virology 64, 4697-4702.[Medline]
Karnauchow, T. M., Tolson, D. L., Harrison, B. A., Altman, E., Lublin, D. M. & Dimock, K. (1996). The HeLa cell receptor for enterovirus 70 is decay accelerating factor (CD55).Journal of Virology 70, 5143-5152.[Abstract]
Lisanti, M. P. & Rodriguez-Boulan, E. (1991). Polarized sorting of GPI-linked proteins in epithelia and membrane microdomains.Cell Biology International Reports 15, 1023-1049.[Medline]
Mayor, S., Rothberg, K. G. & Maxfield, F. R. (1994). Sequestration of GPI-anchored proteins in caveolae triggered by cross-linking.Science 264, 1948-1951.[Medline]
Medof, M. E., Kinoshita, T. & Nussenzweig, V. (1984). Inhibition of complement activation on the surface of cells after incorporation of decay-accelerating factor (DAF) into their membranes.Journal of Experimental Medicine 160, 1558-1578.[Abstract]
Mendelsohn, C. L., Wimmer, E. & Racaniello, V. R. (1989). Cellular receptor for poliovirus: molecular cloning, nucleotide sequence, and expression of a new member of the immunoglobulin superfamily.Cell 56, 855-865.[Medline]
Moran, P., Beasley, H., Gorrell, A., Martin, E., Gribling, P., Fuchs, H., Gillett, N., Burton, L. E. & Caras, I. W. (1992). Human recombinant soluble decay accelerating factor inhibits complement activation in vitro and in vivo.Journal of Immunology 149, 1736-1743.
Morgan, B. P. & Meri, S. (1994). Membrane proteins that protect against complement lysis.Springer Seminars in Immunopathology 15, 369-396.[Medline]
Powell, R. M., Ward, T., Evans, D. J. & Almond, J. W. (1997). Interaction between echovirus 7 and its receptor, decay-accelerating factor (CD55): evidence for a secondary cellular factor in A-particle formation.Journal of Virology 71, 9306-9312.[Abstract]
Powell, R. M., Schmitt, V., Ward, T., Goodfellow, I., Evans, D. J. & Almond, J. W. (1998). Characterization of echoviruses that bind decay accelerating factor (CD55): evidence that some haemagglutinating strains use more than one cellular receptor.Journal of General Virology 79, 1707-1713.[Abstract]
Schnitzer, J. E., McIntosh, D. P., Dvorak, A. M., Liu, J. & Oh, P. (1995). Separation of caveolae from associated microdomains of GPI-anchored proteins [see comments].Science 269, 1435-1439.[Medline]
Shafren, D. R., Bates, R. C., Agrez, M. V., Herd, R. L., Burns, G. F. & Barry, R. D. (1995). Coxsackieviruses B1, B3, and B5 use decay accelerating factor as a receptor for cell attachment.Journal of Virology 69, 3873-3877.[Abstract]
Shafren, D. R., Dorahy, D. J., Ingham, R. A., Burns, G. F. & Barry, R. D. (1997a). Coxsackievirus A21 binds to decay-accelerating factor but requires intercellular adhesion molecule 1 for cell entry.Journal of Virology 71, 4736-4743.[Abstract]
Shafren, D. R., Williams, D. T. & Barry, R. D. (1997b). A decay-accelerating factor-binding strain of coxsackievirus B3 requires the coxsackievirusadenovirus receptor protein to mediate lytic infection of rhabdomyosarcoma cells.Journal of Virology 71, 9844-9848.[Abstract]
Simons, K. & Ikonen, E. (1997). Functional rafts in cell membranes.Nature 387, 569-572.[Medline]
Stang, E., Kartenbeck, J. & Parton, R. G. (1997). Major histocompatibility complex class I molecules mediate association of SV40 with caveolae.Molecular Biology of the Cell 8, 47-57.[Abstract]
Ward, T., Pipkin, P. A., Clarkson, N. A., Stone, D. M., Minor, P. D. & Almond, J. W. (1994). Decay accelerating factor (CD55) identified as the receptor for echovirus 7 using CELICS, a rapid immuno-focal cloning method.EMBO Journal 13, 5070-5074.[Abstract]
Ward, T., Powell, R. M., Pipkin, P. A., Evans, D. J., Minor, P. D. & Almond, J. W. (1998). Role for 2-microglobulin in echovirus infection of rhabdomyosarcoma cells.Journal of Virology 72, 5360-5365.
Wu, M., Fan, J., Gunning, W. & Ratnam, M. (1997). Clustering of GPI-anchored folate receptor independent of both cross-linking and association with caveolin.Journal of Membrane Biology 159, 137-147.[Medline]
Yafal, A. G., Kaplan, G., Racaniello, V. R. & Hogle, J. M. (1993). Characterization of poliovirus conformational alteration mediated by soluble cell receptors.Virology 197, 501-505.[Medline]
Received 26 April 1999;
accepted 19 January 2000.