Deletion of the UL21 gene in Pseudorabies virus results in the formation of DNA-deprived capsids: 
an electron microscopy study

The attenuated pseudorabies virus (PRV) strain Bartha contains several characterized mutations that affect its virulence and ability to spread through neural circuits. This strain contains a small genomic deletion that abrogates anterograde spread and is widely used as a retrograde-restricted neural circuit tracer. Previous studies showed that the retrograde-directed spread of PRV Bartha is slower than that of wild-type PRV. We used compartmented neuronal cultures to characterize the retrograde defect and identify the genetic basis of the phenotype. PRV Bartha is not impaired in retrograde axonal transport, but transneuronal spread among neurons is diminished. Repair of the U L 21 locus with wild-type sequence restored efficient transneuronal spread both in vitro and in vivo. It is likely that mutations in the Bartha U L 21 gene confer defects that affect infectious particle production, causing a delay in spread to presynaptic neurons and amplification of infection. These events manifest as slower kinetics of retrograde viral spread in a neural circuit.

Section: Discussionmentioning

confidence: 99%

Repair of the U _L 21 Locus in Pseudorabies Virus Bartha Enhances the Kinetics of Retrograde, Transneuronal Infection In Vitro and In Vivo

Curanovic

Lyman

Bou-Abboud

et al. 2009

“…Previously, the PrV UL21 protein had been described as a capsid protein involved in DNA cleavage and packaging (15). In the absence of UL21, formation of DNAdeprived capsids was observed in a cell type-dependent manner (57). In contrast, a UL21 mutant virus we isolated earlier exhibited only a slight effect on replication in cell culture but was strongly attenuated in the natural host (26).…”

Section: Vol 79 2005mentioning

confidence: 92%

“…Electron microscopy showed that UL21-negative virions produced on porcine SK-6 cells or lung alveolar macrophages contained little or no DNA. In contrast, on nasal mucosa explants this defect was not observed, which could indicate that UL21 is involved in packaging of newly replicated DNA in a cell type-specific manner (57).…”

mentioning

confidence: 86%

“…It has been described as a 61-kDa capsid-associated protein which is involved in capsid maturation (15,57). A UL21 mutant generated by linker insertion mutagenesis with stop codons inserted after codon 291 of the UL21 open reading frame showed a 10-fold reduction in virus titer but produced wild-type-size plaques, while a mutant truncated after codon 4 replicated only poorly and formed small plaques in cultured cells.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Complex Formation between the UL16 and UL21 Tegument Proteins of Pseudorabies Virus

Klupp

Böttcher

Granzow

et al. 2005

The products of the UL16 and UL21 genes represent tegument proteins which are conserved throughout the mammalian herpesviruses. To identify and functionally characterize the respective proteins in the alphaherpesvirus pseudorabies virus, monospecific antisera against bacterially expressed fusion proteins were generated. In immunoblots the UL16 antiserum detected a ca. 40-kDa protein in infected cells and purified virion preparations, whereas the anti-UL21 serum recognized a protein of approximately 60 kDa. Interestingly, in immunoprecipitations using either antiserum, both proteins were coprecipitated, demonstrating the formation of a physical complex. To investigate protein function, viruses lacking either UL16, UL21, or both were constructed. Mutant viruses could be propagated on noncomplementing cells, indicating that these proteins, either alone or in combination, are not required for viral replication in cell culture. However, plaque sizes and viral titers were reduced. Electron microscopy showed only slight alterations in cytoplasmic virion morphogenesis, whereas intranuclear maturation stages were not affected. Similar results were obtained with a triple mutant simultaneously lacking the three conserved tegument proteins UL11, UL16, and UL21. In summary, our results uncover a novel interaction between conserved herpesvirus tegument proteins that increases the complexity of the intricate network of protein-protein interactions involved in herpesvirus morphogenesis.

“…The pUL21 protein of the alphaherpesvirus pseudorabies virus (PrV) has been described as a capsid-associated protein involved in capsid maturation during cleavage and encapsidation of the viral genome (3,33). However, a UL21 mutant of PrV is only slightly impaired in replication in cell culture (13,14) and in a murine intranasal infection model (12) but is strongly attenuated in PrV's natural host, the pig (13).…”

mentioning

confidence: 99%

Efficient Incorporation of Tegument Proteins pUL46, pUL49, and pUS3 into Pseudorabies Virus Particles Depends on the Presence of pUL21

Michael

Klupp

Karger

et al. 2007

The mature virion of the alphaherpesvirus pseudorabies virus (PrV) contains a minimum of 31 structural proteins which are recruited into the virus particle by a network of protein-protein interactions which is only incompletely understood. We show here that deletion of the tegument protein pUL21 resulted in a drastic decrease in the incorporation of the pUL46, pUL49, and pUS3 tegument components into mature virions. Moreover, the attenuated PrV strain Bartha (PrV-Ba), which, among other defects, carries mutations in pUL21, also fails to package pUL46, pUL49, and pUS3 efficiently. By the reconstitution of wild-type pUL21 expression to PrV-Ba and the transfer of mutated PrV-Ba pUL21 into wild-type PrV, we demonstrate that this phenotype is due to the mutated pUL21.