In situ Localization of the Major Capsid Protein during Lytic Infection by Herpes Simplex Virus

Bibor-Hardy, Viviane; Dagenais, André; Simard, René

doi:10.1099/0022-1317-66-4-897

Cited by 15 publications

(6 citation statements)

References 6 publications

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“…Yamauchi et al (1985) previously reported on the intracellular transport of the HCMV MCP into the nucleus using electron microscopy and separation of nuclear fractions in protein gels. In some immunofluorescence experiments using HSV-1-infected cells incubated with HCMV MCP antibodies, we observed a distinct nuclear staining as has been described for VP5 by Bibor-Hardy et al (1985) and for the MCP of EBV by Vroman et al (1985). In addition, a monoclonal antibody (IV.…”

Section: Hsv-i H C Msupporting

confidence: 84%

Prokaryotic expression of the major capsid protein of human cytomegalovirus and antigenic cross-reactions with herpes simplex virus type 1

Rudolph¹,

Kühn²,

Korn³

et al. 1990

Journal of General Virology

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The major capsid protein (MCP) of human cytomegalovirus (HCMV) was expressed in three portions as fl-galactosidase fusion proteins, covering about 75 % of the open reading frame (ORF). Fusion protein SH 1 contained nucleotides 101 to 1243 of the ORF, fusion protein FS 1 contained nucleotides 1944 to 3089 and fusion protein SS 1 covered nucleotides 2624 to 3793. The recombinant proteins were tested for their immunoreactivity with human sera. Fusion protein FS 1 was found to represent the immunodominant region. The recombinant proteins were used to generate polyvalent rabbit antisera to investigate cross-reactivities with the major capsid protein (VP5) of herpes simplex virus type 1 (HSV-1). A monospecific antiserum raised against the fusion protein close to the N terminus of the MCP, as well as a monoclonal antibody and a monospecific rabbit antiserum directed against the viral MCP, cross-reacted with the VP5 as shown by immunoblotting and immunofluorescence. In order to detect common epitopes of the major capsid proteins of HCMV and HSV-1, the recombinant proteins were conjugated to CNBr-activated Sepharose and taken for purification of MCP antibodies from HCMV and HSV-1 seropositive individuals. Using this affinity chromatography method, cross-reactivity could be observed with HCMV-and HSV-positive human antisera in immunoblot experiments.

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Section: Hsv-i H C Msupporting

confidence: 84%

Prokaryotic expression of the major capsid protein of human cytomegalovirus and antigenic cross-reactions with herpes simplex virus type 1

Rudolph¹,

Kühn²,

Korn³

et al. 1990

Journal of General Virology

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“…Activation of PKA substrates during We show here that phosphorylation of matrin 3 is common to all three herpesviruses and that for VZV it occurs through an indirect pathway other than PKA activation. Interaction with matrix proteins has been described for many viruses, including HIV, simian virus 40 (SV40), PRV, HCMV, human papillomavirus (7,27,51,63,66,74), and HSV-1 (9,10,14,49,83,84). At least two forms of matrin 3, which are suspected to reflect additional phosphorylation events, were detected in our studies.…”

Section: Discussionmentioning

confidence: 57%

“…Recently, a comparable role has been proposed for lamin A/C: serving as a scaffold for HSV genomes and nuclear factors that reduce heterochromatin assembly on viral promoters (70). Matrin 3 may also have a role in capsid assembly, since capsid proteins localize to nuclear matrix fractions (9,14). This would correlate with lines of evidence suggesting that ORF66 is involved in capsid assembly: in VZV ORF66-deficient infected T cells from SCID-hu T cell xenografts, there is greatly reduced intranuclear assembly of nucleocapsids (65).…”

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confidence: 99%

The Alphaherpesvirus US3/ORF66 Protein Kinases Direct Phosphorylation of the Nuclear Matrix Protein Matrin 3

Erazo

Yee

Banfield

et al. 2011

J Virol

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The protein kinase found in the short region of alphaherpesviruses, termed US3 in herpes simplex virus type 1 (HSV-1) and pseudorabies virus (PRV) and ORF66 in varicella-zoster virus (VZV), affects several viral and host cell processes, and its specific targets remain an area of active investigation. Reports suggesting that HSV-1 US3 substrates overlap with those of cellular protein kinase A (PKA) prompted the use of an antibody specific for phosphorylated PKA substrates to identify US3/ORF66 targets. HSV-1, VZV, and PRV induced very different substrate profiles that were US3/ORF66 kinase dependent. The predominant VZV-phosphorylated 125-kDa species was identified as matrin 3, one of the major nuclear matrix proteins. Matrin 3 was also phosphorylated by HSV-1 and PRV in a US3 kinase-dependent manner and by VZV ORF66 kinase at a novel residue (KRRRT150EE). Since VZV-directed T150 phosphorylation was not blocked by PKA inhibitors and was not induced by PKA activation, and since PKA predominantly targeted matrin 3 S188, it was concluded that phosphorylation by VZV was PKA independent. However, purified VZV ORF66 kinase did not phosphorylate matrin 3 in vitro, suggesting that additional cellular factors were required. In VZV-infected cells in the absence of the ORF66 kinase, matrin 3 displayed intranuclear changes, while matrin 3 showed a pronounced cytoplasmic distribution in late-stage cells infected with US3-negative HSV-1 or PRV. This work identifies phosphorylation of the nuclear matrix protein matrin 3 as a new conserved target of this kinase group.All herpesviruses studied to date encode proteins predicted to be protein kinases, based on the presence of conserved signature domains common to cellular kinases. Considerable interest in their roles and functions has developed, because they influence multiple cellular processes by controlling the phosphorylation of both cellular and viral proteins. Two serine/ threonine (S/T)-specific protein kinases are found in alphaherpesvirus genomes: the UL13 kinase of herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) (open reading frame 47 [ORF47] in varicella-zoster virus [VZV]) has recognizable homologues in all known herpesviruses, whereas the US3 kinase (ORF66 in VZV) is found in the short genomic region of members of the alphaherpesvirus subfamily. Both alphaherpesvirus kinases are genetically dispensable for viral growth in culture, but kinase-deficient viruses invariably show impaired replication in specific cell types and/or attenuation in animal models of disease (22,37,45,57,62,64,65,73,79), suggesting important roles that are dependent on the host cell environment.The 393-residue ORF66 protein kinase of VZV has such cell-specific importance. VZV causes chickenpox upon primary infection and herpes zoster upon reactivation from a prolonged neuronal latent state. VZVs expressing ORF66 that is translationally halted or kinase inactive show only modest growth impairment in cell cultures used for VZV propagation and in human skin in the SCID-hu mouse model but show s...

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“…The immune complexes were revealed with a 30-minute incubation at 21°C with streptavidin conjugated to 20-nm colloidal gold particles (1 : 20; Bethesda Research Laboratories, Gaithersburg, MD). Nuclei were fixed with 2.5% glutaraldehyde and processed for electron microscopy as described (13).…”

Section: Methodsmentioning

confidence: 99%

A novel autoantibody causing a peripheral fluorescent antinuclear antibody pattern is specific for nuclear pore complexes

Dagenais

Bibor-Hardy

Senécal

1988

Arthritis & Rheumatism

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We characterized serum from a patient with polymyositis, and found that it produced a peripheral (rim) fluorescent antinuclear antibody pattern on rat liver substrate. Indirect immunofluorescence analysis revealed a punctate pattern at the nuclear surface of PtK2, BHK-21, and HEp-2 cells. This pattern was still present after sequential extraction in situ with non-ionic detergent, DNase, RNase, and high ionic strength buffer (2M NaCI). Immunogold electron microscopic localization was specific for nuclear pore complexes. By immunoblot analysis, the antigens were polypeptides of 200 kd and 130 kd that were enriched in the nuclear fraction.Antinuclear antibodies (ANA) in human sera are routinely detected with indirect immunofluorescence microscopy on substrates such as rat liver sections (1). Among the patterns identified by this method, the peripheral (rim) pattern is of diagnostic importance because of its association with systemic lupus erythematosus (SLE) (1,2). Studies of the antigenic specificity of autoantibodies from sera causing a peripheral ANA pattern have shown a strong association with anti-DNA antibodies (2,3). In addition, autoantibodies to nuclear lamins, observed in sera from 1 patient with linear scleroderma (4) and 5 patients with SLE (5,6), were shown to cause a peripheral ANA pattern because of continuous labeling of the nuclear envelope.Nuclear pore complexes are structures localized at the points of fusion of the double nuclear membranes (7) and are thought to allow molecular exchanges across the nuclear envelope. We report the identification, in serum from a patient with polymyositis, of a novel autoantibody that causes a peripheral ANA pattern on rat liver sections and stains the nuclear envelope of various cell lines. This autoantibody reacts with nuclear pore complexes. PATIENTS AND METHODSPatient LL. The index serum was obtained from a 35-year-old white woman who, in December 1985, developed polymyositis, which was characterized by progressive proximal muscle weakness, elevated levels of serum creatine kinase (4,836 units/liter, normal 24-1 70), myopathic changes evident on electromyogram, and abnormal findings on quadriceps muscle biopsy. ANA were detected by indirect immunofluorescence on cryostat rat liver sections, at a titer of 1: 1,280 and with a peripheral pattern. Esophageal manometry revealed decreased peristaltic pressure in the distal two-thirds of the esophagus.Normal or negative results were noted on the following tests: hemoglobin, leukocyte and platelet

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In situ Localization of the Major Capsid Protein during Lytic Infection by Herpes Simplex Virus

Cited by 15 publications

References 6 publications

Prokaryotic expression of the major capsid protein of human cytomegalovirus and antigenic cross-reactions with herpes simplex virus type 1

Prokaryotic expression of the major capsid protein of human cytomegalovirus and antigenic cross-reactions with herpes simplex virus type 1

The Alphaherpesvirus US3/ORF66 Protein Kinases Direct Phosphorylation of the Nuclear Matrix Protein Matrin 3

A novel autoantibody causing a peripheral fluorescent antinuclear antibody pattern is specific for nuclear pore complexes

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