Disulfide Bond Formation in the Herpes Simplex Virus 1 UL6 Protein Is Required for Portal Ring Formation and Genome Encapsidation

Albright, Brandon S.; Nellissery, Jacob; Szczepaniak, Renata; Weller, Sandra K.

doi:10.1128/jvi.00123-11

Cited by 25 publications

(31 citation statements)

References 47 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We have previously reported that mature HSV capsids are extensively cross-linked by disulfide bonds (9) and that the UL6 portal ring contains intersubunit disulfide bonds (10). This raises the question of how disulfide bond formation in the portal is regulated, especially in a cellular environment that is generally thought to be reducing (10).…”

Section: Resultsmentioning

confidence: 99%

“…The formation of inter-and intramolecular disulfide bonds is generally recognized to regulate a wide range of cellular processes. We have previously shown that disulfide bonds are essential for capsid stability and for the formation and/or stability of the UL6 portal ring (9,10). It is possible that the HSV protease is regulated by the redox state.…”

Section: Discussionmentioning

confidence: 99%

“…Capsids were isolated and separated by sucrose gradient sedimentation as previously described (10). Fractions were collected and prepared for Western blot analysis under reducing conditions as described above.…”

Section: Methodsmentioning

confidence: 99%

“…Cells were harvested and proteins were separated by reducing SDS-PAGE as previously described and transferred to a polyvinylidene difluoride membrane (10). Membranes were blocked with 5% milk and incubated overnight with the following antibodies: mouse monoclonal anti-HA (1:1,000; catalog number Sc-7392; Santa Cruz), rabbit polyclonal anti-HA (1:1,000; catalog number 631207; Clontech), rabbit polyclonal anti-UL32 (1:1,000; antibody to synthetic antigenic peptide was generated by Open Biosystems), mouse monoclonal anti-ICP8 (1: 2,000; catalog number ab20194; Abcam), mouse monoclonal anti-VP5 (1:1,000; catalog number HA018; EastCoast Bio), mouse monoclonal anti-VP24 9-2 (1:1,000) (37), and mouse anti-actin (1:10,000; catalog number A5441; Sigma).…”

Section: Methodsmentioning

confidence: 99%

“…We have recently demonstrated disulfide linkages between capsid proteins VP5, VP23, and VP19C and other components of the mature virion, including UL17, UL25, and tegument proteins; furthermore, we showed that disulfide bond formation is required for capsid stability (9). Intersubunit disulfide bonds also exist between monomeric subunits of the portal ring and contribute to ring formation and/or stability (10). The disulfide bonds in the portal ring can be detected in the nucleus.…”

mentioning

confidence: 95%

See 4 more Smart Citations

The Putative Herpes Simplex Virus 1 Chaperone Protein UL32 Modulates Disulfide Bond Formation during Infection

Albright

Kosinski

Szczepaniak

et al. 2015

J Virol

Self Cite

View full text Add to dashboard Cite

During DNA encapsidation, herpes simplex virus 1 (HSV-1) procapsids are converted to DNA-containing capsids by a process involving activation of the viral protease, expulsion of the scaffold proteins, and the uptake of viral DNA. Encapsidation requires six minor capsid proteins (UL6, UL15, UL17, UL25, UL28, and UL33) and one viral protein, UL32, not found to be associated with capsids. Although functions have been assigned to each of the minor capsid proteins, the role of UL32 in encapsidation has remained a mystery. Using an HSV-1 variant containing a functional hemagglutinin-tagged UL32, we demonstrated that UL32 was synthesized with true late kinetics and that it exhibited a previously unrecognized localization pattern. At 6 to 9 h postinfection (hpi), UL32 accumulated in viral replication compartments in the nucleus of the host cell, while at 24 hpi, it was additionally found in the cytoplasm. A newly generated UL32-null mutant was used to confirm that although B capsids containing wild-type levels of capsid proteins were synthesized, these procapsids were unable to initiate the encapsidation process. Furthermore, we showed that UL32 is redox sensitive and identified two highly conserved oxidoreductase-like C-X-X-C motifs that are essential for protein function. In addition, the disulfide bond profiles of the viral proteins UL6, UL25, and VP19C and the viral protease, VP24, were altered in the absence of UL32, suggesting that UL32 may act to modulate disulfide bond formation during procapsid assembly and maturation. IMPORTANCEAlthough functions have been assigned to six of the seven required packaging proteins of HSV, the role of UL32 in encapsidation has remained a mystery. UL32 is a cysteine-rich viral protein that contains C-X-X-C motifs reminiscent of those in proteins that participate in the regulation of disulfide bond formation. We have previously demonstrated that disulfide bonds are required for the formation and stability of the viral capsids and are also important for the formation and stability of the UL6 portal ring. In this report, we demonstrate that the disulfide bond profiles of the viral proteins UL6, UL25, and VP19C and the viral protease, VP24, are altered in cells infected with a newly isolated UL32-null mutant virus, suggesting that UL32 acts as a chaperone capable of modulating disulfide bond formation. Furthermore, these results suggest that proper regulation of disulfide bonds is essential for initiating encapsidation.T he products of herpes simplex virus 1 (HSV-1) DNA replication are head-to-tail concatemers that are resolved into monomeric genomic units and packaged into a procapsid shell in the nucleus of the infected cell (reviewed in references 1 to 3). The procapsid is comprised of the major capsid protein (VP5), triplex proteins (VP19C and VP23), and a dodecameric UL6 portal ring. The precursor of the viral protease (UL26) and the scaffolding protein (UL26.5) form a scaffold around which the capsid shell assembles (3, 4). During DNA encapsidation, the viral protease (V...

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 95%

See 3 more Smart Citations

The Putative Herpes Simplex Virus 1 Chaperone Protein UL32 Modulates Disulfide Bond Formation during Infection

Albright

Kosinski

Szczepaniak

et al. 2015

J Virol

Self Cite

View full text Add to dashboard Cite

show abstract

Monosynaptic restriction of the anterograde herpes simplex virus strain H129 for neural circuit tracing

Fischer

Collins

Pang

et al. 2023

J of Comparative Neurology

View full text Add to dashboard Cite

Identification of synaptic partners is a fundamental task for systems neuroscience.To date, few reliable techniques exist for whole brain labeling of downstream synaptic partners in a cell-type-dependent and monosynaptic manner. Herein, we describe a novel monosynaptic anterograde tracing system based on the deletion of the gene UL6 from the genome of a cre-dependent version of the anterograde Herpes Simplex Virus 1 strain H129. Given that this knockout blocks viral genome packaging and thus viral spread, we reasoned that co-infection of a HSV H129 ΔUL6 virus with a recombinant adeno-associated virus expressing UL6 in a cre-dependent manner would result in monosynaptic spread from target cre-expressing neuronal populations. Application of this system to five nonreciprocal neural circuits resulted in labeling of neurons in expected projection areas. While some caveats may preclude certain applications, this system provides a reliable method to label postsynaptic partners in a brain-wide fashion.

show abstract

Human herpesvirus portal proteins: Structure, function, and antiviral prospects

Kornfeind

Visalli

2018

Reviews in Medical Virology

View full text Add to dashboard Cite

Herpesviruses (Herpesvirales) and tailed bacteriophages (Caudovirales) package their dsDNA genomes through an evolutionarily conserved mechanism. Much is known about the biochemistry and structural biology of phage portal proteins and the DNA encapsidation (viral genome cleavage and packaging) process. Although not at the same level of detail, studies on HSV-1, CMV, VZV, and HHV-8 have revealed important information on the function and structure of herpesvirus portal proteins. During dsDNA phage and herpesviral genome replication, concatamers of viral dsDNA are cleaved into single length units by a virus-encoded terminase and packaged into preformed procapsids through a channel located at a single capsid vertex (portal). Oligomeric portals are formed by the interaction of identical portal protein monomers. Comparing portal protein primary aa sequences between phage and herpesviruses reveals little to no sequence similarity. In contrast, the secondary and tertiary structures of known portals are remarkable. In all cases, function is highly conserved in that portals are essential for DNA packaging and also play a role in releasing viral genomic DNA during infection. Preclinical studies have described small molecules that target the HSV-1 and VZV portals and prevent viral replication by inhibiting encapsidation. This review summarizes what is known concerning the structure and function of herpesvirus portal proteins primarily based on their conserved bacteriophage counterparts and the potential to develop novel portal-specific DNA encapsidation inhibitors.

show abstract

Disulfide Bond Formation in the Herpes Simplex Virus 1 UL6 Protein Is Required for Portal Ring Formation and Genome Encapsidation

Cited by 25 publications

References 47 publications

The Putative Herpes Simplex Virus 1 Chaperone Protein UL32 Modulates Disulfide Bond Formation during Infection

The Putative Herpes Simplex Virus 1 Chaperone Protein UL32 Modulates Disulfide Bond Formation during Infection

Monosynaptic restriction of the anterograde herpes simplex virus strain H129 for neural circuit tracing

Human herpesvirus portal proteins: Structure, function, and antiviral prospects

Contact Info

Product

Resources

About