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The large tegument protein pUL36 is essential for formation of the capsid vertex-specific component at the capsid-tegument interface of herpes simplex virus 1.

Fan WH, Roberts AP, McElwee M, Bhella D, Rixon FJ, Lauder R - J. Virol. (2014)

Bottom Line: In addition, the presence of full-length pUL36 results in weak density that extends the CVSC toward the penton, suggesting either that this extra density is formed directly by pUL36 or that pUL36 stabilizes other components of the vertex-tegument interface.Herpesviruses have complex particles that are formed as a result of a carefully controlled sequence of assembly steps.We show that the largest viral protein, pUL36, which occupies the layer of tegument closest to the capsid, is essential for formation of structurally normal connections to the capsid.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Glasgow, United Kingdom.

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Comparison of CVSC densities in HSV-1 WT and mutant virus capsids. A region around the vertex (indicated by the white square in Fig. S1B in the supplemental material) of icosahedrally reconstructed C-capsids purified from the cytoplasm (A and B) or nucleus (C) of cells infected with WT HSV-1 or the inner tegument deletion mutants ARΔUL36 and FRΔUL37 is shown. Panels A and C are shown radially colored. Panel B shows the cytoplasmic C-capsid maps in panel A superimposed on a B-capsid map (shown in gray). The locations of the penton (5), one of the five peripentonal hexons (P), and a Ta and Tc triplex are marked on the WT image in panel A. Each black arrowhead indicates the position of one CVSC density. Scale bar = 50 Å.
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Figure 1: Comparison of CVSC densities in HSV-1 WT and mutant virus capsids. A region around the vertex (indicated by the white square in Fig. S1B in the supplemental material) of icosahedrally reconstructed C-capsids purified from the cytoplasm (A and B) or nucleus (C) of cells infected with WT HSV-1 or the inner tegument deletion mutants ARΔUL36 and FRΔUL37 is shown. Panels A and C are shown radially colored. Panel B shows the cytoplasmic C-capsid maps in panel A superimposed on a B-capsid map (shown in gray). The locations of the penton (5), one of the five peripentonal hexons (P), and a Ta and Tc triplex are marked on the WT image in panel A. Each black arrowhead indicates the position of one CVSC density. Scale bar = 50 Å.

Mentions: After they have left the nucleus, cytoplasmic C-capsids become tegumented and enveloped to form mature virions. To analyze the sequence of events occurring during tegument addition, we determined the structures of cytoplasmic C-capsids prepared from BHK cells infected with wild-type (WT) HSV-1 and two inner tegument mutants with complete deletions of the UL36 (ARΔUL36) and UL37 (FRΔUL37) genes (20). Previously, we reported problems with isolating cytoplasmic C-capsids from cells infected for 24 h with WT and FRΔUL37 due to aggregation of the partially tegumented capsids (20). To overcome these problems, we prepared capsids at 16 h after infection, when the concentration of cytoplasmic capsids was lower. C-capsids were isolated on sucrose gradients, and electron cryomicroscopic images were collected. The majority of the capsids in these preparations were C-capsids, which could be readily distinguished from contaminating A- and B-capsids, which lack DNA (see Fig. S1A in the supplemental material). 3D reconstructions were generated to final resolutions of 20 Å for the WT and 18 Å for ARΔUL36 and FRΔUL37. As expected, the structures of all three showed typical capsid features with hexons, pentons, and triplexes arranged on a T=16 icosahedral lattice, and no differences were apparent between them other than at the capsid vertices, which are known to be sites of interaction between capsid and tegument (Fig. 1A; see also Fig. S1B). Comparison with the nuclear B-capsid revealed additional densities on the outer surface of the capsids occupying the space between the P-hexons that surround the penton (Fig. 1B). The WT and FRΔUL37 structures closely resembled each other, with relatively extensive additional densities that are arranged in a radial pattern around the vertices. In contrast, the additional densities in ARΔUL36, although occupying similar positions, were much smaller. The presence of additional “star-shaped” densities at the vertices of intact virions is well established (28), and they are thought to be composed of the minor capsid/packaging proteins pUL17 and pUL25 and the inner tegument protein, pUL36. pUL36 has been reported to account for the inner, penton-capping hub of the star (40), with pUL17 and pUL25 making up the CVSC, which corresponds to the outer arms of the star (36, 39). Although the vertex-associated densities of the WT and FRΔUL37 cytoplasmic C-capsids were similar to the archetypal CVSC previously described for WT nuclear C-capsids (11, 36–38), those present in ARΔUL36 were significantly smaller. For convenience, we refer here to all forms of vertex-associated additional density as CVSC.


The large tegument protein pUL36 is essential for formation of the capsid vertex-specific component at the capsid-tegument interface of herpes simplex virus 1.

Fan WH, Roberts AP, McElwee M, Bhella D, Rixon FJ, Lauder R - J. Virol. (2014)

Comparison of CVSC densities in HSV-1 WT and mutant virus capsids. A region around the vertex (indicated by the white square in Fig. S1B in the supplemental material) of icosahedrally reconstructed C-capsids purified from the cytoplasm (A and B) or nucleus (C) of cells infected with WT HSV-1 or the inner tegument deletion mutants ARΔUL36 and FRΔUL37 is shown. Panels A and C are shown radially colored. Panel B shows the cytoplasmic C-capsid maps in panel A superimposed on a B-capsid map (shown in gray). The locations of the penton (5), one of the five peripentonal hexons (P), and a Ta and Tc triplex are marked on the WT image in panel A. Each black arrowhead indicates the position of one CVSC density. Scale bar = 50 Å.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 1: Comparison of CVSC densities in HSV-1 WT and mutant virus capsids. A region around the vertex (indicated by the white square in Fig. S1B in the supplemental material) of icosahedrally reconstructed C-capsids purified from the cytoplasm (A and B) or nucleus (C) of cells infected with WT HSV-1 or the inner tegument deletion mutants ARΔUL36 and FRΔUL37 is shown. Panels A and C are shown radially colored. Panel B shows the cytoplasmic C-capsid maps in panel A superimposed on a B-capsid map (shown in gray). The locations of the penton (5), one of the five peripentonal hexons (P), and a Ta and Tc triplex are marked on the WT image in panel A. Each black arrowhead indicates the position of one CVSC density. Scale bar = 50 Å.
Mentions: After they have left the nucleus, cytoplasmic C-capsids become tegumented and enveloped to form mature virions. To analyze the sequence of events occurring during tegument addition, we determined the structures of cytoplasmic C-capsids prepared from BHK cells infected with wild-type (WT) HSV-1 and two inner tegument mutants with complete deletions of the UL36 (ARΔUL36) and UL37 (FRΔUL37) genes (20). Previously, we reported problems with isolating cytoplasmic C-capsids from cells infected for 24 h with WT and FRΔUL37 due to aggregation of the partially tegumented capsids (20). To overcome these problems, we prepared capsids at 16 h after infection, when the concentration of cytoplasmic capsids was lower. C-capsids were isolated on sucrose gradients, and electron cryomicroscopic images were collected. The majority of the capsids in these preparations were C-capsids, which could be readily distinguished from contaminating A- and B-capsids, which lack DNA (see Fig. S1A in the supplemental material). 3D reconstructions were generated to final resolutions of 20 Å for the WT and 18 Å for ARΔUL36 and FRΔUL37. As expected, the structures of all three showed typical capsid features with hexons, pentons, and triplexes arranged on a T=16 icosahedral lattice, and no differences were apparent between them other than at the capsid vertices, which are known to be sites of interaction between capsid and tegument (Fig. 1A; see also Fig. S1B). Comparison with the nuclear B-capsid revealed additional densities on the outer surface of the capsids occupying the space between the P-hexons that surround the penton (Fig. 1B). The WT and FRΔUL37 structures closely resembled each other, with relatively extensive additional densities that are arranged in a radial pattern around the vertices. In contrast, the additional densities in ARΔUL36, although occupying similar positions, were much smaller. The presence of additional “star-shaped” densities at the vertices of intact virions is well established (28), and they are thought to be composed of the minor capsid/packaging proteins pUL17 and pUL25 and the inner tegument protein, pUL36. pUL36 has been reported to account for the inner, penton-capping hub of the star (40), with pUL17 and pUL25 making up the CVSC, which corresponds to the outer arms of the star (36, 39). Although the vertex-associated densities of the WT and FRΔUL37 cytoplasmic C-capsids were similar to the archetypal CVSC previously described for WT nuclear C-capsids (11, 36–38), those present in ARΔUL36 were significantly smaller. For convenience, we refer here to all forms of vertex-associated additional density as CVSC.

Bottom Line: In addition, the presence of full-length pUL36 results in weak density that extends the CVSC toward the penton, suggesting either that this extra density is formed directly by pUL36 or that pUL36 stabilizes other components of the vertex-tegument interface.Herpesviruses have complex particles that are formed as a result of a carefully controlled sequence of assembly steps.We show that the largest viral protein, pUL36, which occupies the layer of tegument closest to the capsid, is essential for formation of structurally normal connections to the capsid.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Glasgow, United Kingdom.

Show MeSH
Related in: MedlinePlus