Morphogenesis of African swine fever virus in monkey kidney cells after reversible inhibition of replication by cycloheximide

Arzuza, Octavio; Urzainqui, Ana; Díaz-Ruíz, J. R.; Tabarés, E.

doi:10.1007/bf01309814

Cited by 15 publications

(15 citation statements)

References 31 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Such a model is divergent in some aspects from others previously reported. For example, Schloer (30) and Arzuza et al (2) include an additional membrane between the capsid and the external envelope. Such a membrane would be the outermost layer of the intracellular particles.…”

Section: Discussionmentioning

confidence: 99%

Assembly of African swine fever virus: role of polyprotein pp220

Andrés

Simón‐Mateo

Viñuela

1997

J Virol

100

View full text Add to dashboard Cite

Polyprotein processing is a common strategy of gene expression in many positive-strand RNA viruses and retroviruses but not in DNA viruses. African swine fever virus (ASFV) is an exception because it encodes a polyprotein, named pp220, to produce several major components of the virus particle, proteins p150, p37, p34, and p14. In this study, we analyzed the assembly pathway of ASFV and the contribution of the polyprotein products to the virus structure. Electron microscopic studies revealed that virions assemble from membranous structures present in the viral factories. Viral membranes became polyhedral immature virions after capsid formation on their convex surface. Beneath the lipid envelope, two distinct domains appeared to assemble consecutively: first a thick protein layer that we refer to as core shell and then an electron-dense nucleoid, which was identified as the DNA-containing domain. Immunofluorescence studies showed that polyprotein pp220 is localized in the viral factories. At the electron microscopic level, antibodies to pp220 labeled all identifiable forms of the virus from the precursor viral membranes onward, thus indicating an early role of the polyprotein pp220 in ASFV assembly. The subviral localization of the polyprotein products, examined on purified virions, was found to be the core shell. In addition, quantitative studies showed that the polyprotein products are present in equimolar amounts in the virus particle and account for about one-fourth of its total protein content. Taken together, these results suggest that polyprotein pp220 may function as an internal protein scaffold which would mediate the interaction between the nucleoid and the outer layers similarly to the matrix proteins of other viruses. MATERIALS AND METHODS Cells and viruses. Vero cells (ATCC CCL81) were cultured in Dulbecco's modified Eagle's medium supplemented with 10% newborn calf serum, which was reduced to 2% during viral infection. Swine alveolar macrophages, obtained by bronchoalveolar lavage, were maintained and infected in Dulbecco's modified Eagle's medium with 10% newborn calf serum. ASFV strain BA71V, adapted to grow in Vero cells, has been previously described (14). Highly purified ASFV was obtained by Percoll equilibrium centrifugation (7). Antibodies. Monoclonal antibody 18H.H7, against proteins pp220 and p150, as well as rabbit polyclonal anti-p150, anti-p37/p14, and anti-p34 sera, which also recognize the precursor form pp220, have been characterized previously (28, 33). Monoclonal antibody 19B.A2, against protein p72, has been described before (28). The anti-DNA immunoglobulin M (Ac 30-10

show abstract

Section: Discussionmentioning

confidence: 99%

Assembly of African swine fever virus: role of polyprotein pp220

Andrés

Simón‐Mateo

Viñuela

1997

J Virol

100

View full text Add to dashboard Cite

show abstract

“…Virion morphogenesis has been examined ultrastructurally, and the functions of specific viral proteins during this process have been identified either using immunoelectron microscopy and viral mutants lacking or inducibly expressing specific viral genes Arzuza et al 1992;Breese and DeBoer 1966;Brookes et al 1996;Garcia-Escudero et al 1998;Moura Nunes et al 1975;Rouiller et al 1998). ASFV replication primarily occurs in virus factories, elements of which are first observed by 6-8 h p.i.…”

Section: Virion Morphogenesismentioning

confidence: 99%

African Swine Fever Virus

Tulman

Delhon

et al. 2009

Current Topics in Microbiology and Immunology

134

117

View full text Add to dashboard Cite

African swine fever virus (ASFV) is a large, intracytoplasmicallyreplicating DNA arbovirus and the sole member of the family Asfarviridae . It is the etiologic agent of a highly lethal hemorrhagic disease of domestic swine and therefore extensively studied to elucidate the structures, genes, and mechanisms affecting viral replication in the host, virus-host interactions, and viral virulence. Increasingly apparent is the complexity with which ASFV replicates and interacts with the host cell during infection. ASFV encodes novel genes

show abstract

“…The progressive assembly of capsid and matrix proteins on the cytoplasmic face of these membranes leads to the production of fully assembled icosahedral particles that appear as 200-nm hexagons in cross-sections taken through virus assembly sites (49). Following assembly, mature ASFV particles leave the factory, move through the cytoplasm, and are eventually released from the cell by budding through the plasma membrane (6). Early studies have shown that ASFV particles associate with microtubules in vitro (16), and recent works suggest that retrograde dynein-dependent microtubule transport is required for the inward movement of the virus (3,26).…”

mentioning

confidence: 99%

Transport of African Swine Fever Virus from Assembly Sites to the Plasma Membrane Is Dependent on Microtubules and Conventional Kinesin

Jouvenet

Monaghan

Way

et al. 2004

J Virol

101

View full text Add to dashboard Cite

African swine fever virus (ASFV) is a large DNA virus that assembles in perinuclear viral factories located close to the microtubule organizing center. In this study, we have investigated the mechanism by which ASFV reaches the cell surface from the site of assembly. Immunofluorescence microscopy revealed that at 16 h postinfection, mature virions were aligned along microtubules. Furthermore, virus movement to the cell periphery was inhibited when microtubules were depolymerized by nocodazole. In addition, ASFV infection resulted in the increased acetylation of microtubules as well as their protection against depolymerization by nocodazole. Immunofluorescence microscopy showed that conventional kinesin was recruited to virus factories and to a large fraction of virus particles in the cytoplasm. Consistent with a role for conventional kinesin during ASFV egress to the cell periphery, overexpression of the cargo-binding domain of the kinesin light chain severely inhibited the movement of particles to the plasma membrane. Based on our observations, we propose that ASFV is recognized as cargo by conventional kinesin and uses this plus-end microtubule motor to move from perinuclear assembly sites to the plasma membrane.Viral replication and assembly often take place in specialized structures located in the nucleus or in the perinuclear cytoplasm. These specialized sites, called viral inclusions or virus factories, provide a focal point for the replication of virus genomes and the recruitment of structural proteins. Recent work has focused on understanding how viruses travel to, and from, these specialized sites during their infectious cycle. One common viral strategy uses microtubules for directed longrange transport of virus components inside cells (44,52,54). Directional movement along microtubules is mediated by motor proteins which hydrolyze ATP to induce the conformational changes necessary to move cargoes along microtubules (30,62).Movement of cellular materials towards the microtubule organizing center (MTOC) (retrograde transport) usually requires the minus-end directed microtubule motor dynein (62). The transport of cargoes by dynein requires association with dynactin, a heterooligomeric complex of at least nine different polypeptides. This has been demonstrated experimentally by the overexpression of the p50-dynamitin subunit of the dynactin complex, which dissociates the dynactin complex and severely affects vesicular trafficking and organelle distribution (10). Importantly, retrograde transport of several unrelated viruses, such as herpes simplex virus type 1 (HSV-1), adenovirus, vaccinia virus (VV), parvovirus, and human immunodeficiency virus type 1 is inhibited when the dynactin complex is dissociated, suggesting that dynein is required for inward transport of these viruses (18,38,43,57,59).The movement along microtubules, from the MTOC towards the plasma membrane (anterograde transport), involves the kinesin family of motor proteins (62). Conventional kinesin is a heterotetramer consisting of two heav...

show abstract

Morphogenesis of African swine fever virus in monkey kidney cells after reversible inhibition of replication by cycloheximide

Cited by 15 publications

References 31 publications

Assembly of African swine fever virus: role of polyprotein pp220

Assembly of African swine fever virus: role of polyprotein pp220

African Swine Fever Virus

Transport of African Swine Fever Virus from Assembly Sites to the Plasma Membrane Is Dependent on Microtubules and Conventional Kinesin

Contact Info

Product

Resources

About