Glycoproteins M and N of Pseudorabies Virus Form a Disulfide-Linked Complex

Abstract

Genes homologous to the herpes simplex virus UL49.5 open reading frame are conserved throughout the Herpesviridae. In the alphaherpesvirus pseudorabies virus (PrV), the UL49.5 product is an O-glycosylated structural protein of the viral envelope, glycoprotein N (gN) (A. Jöns, H. Granzow, R. Kuchling, and T. C. Mettenleiter, J. Virol. 70:1237–1241, 1996). For functional characterization of gN, a gN-negative PrV mutant, PrV-gNβ, and the corresponding rescuant, PrV-gNβR, were constructed, gN-negative PrV was able to productively replicate on noncomplementing cells, and one-step growth in cell culture was only slightly reduced compared to that of wild-type PrV. However, penetration was significantly delayed. In indirect immunofluorescence assays with rabbit serum directed against baculovirus-expressed gN, specific staining of wild-type PrV-infected cells occurred only after permeabilization of cells, whereas live cells failed to react with the antiserum. This indicates the lack of surface accessibility of gN in the plasma membrane of a PrV-infected cell. Western blot analyses and radioimmunoprecipitation experiments under reducing and nonreducing conditions led to the discovery of a heteromeric complex composed of gM and gN. The complex was stable in the absence of 2-mercaptoethanol but dissociated after the addition of the reducing agent, indicating that the partners are linked by disulfide bonds. Finally, gN was absent from gM-negative PrV virions, whereas gM was readily detected in virions in the absence of gN. Thus, gM appears to be required for virion localization of gN.

Intracellular Trafficking and Localization of the Pseudorabies Virus Us9 Type II Envelope Protein to Host and Viral Membranes

“…The lipid envelope of PRV virions contains at least 11 virally encoded membrane glycoproteins (26). Recently, we have characterized a novel PRV membrane protein designated Us9 (6).…”

Viral Inhibition of the Transporter Associated with Antigen Processing (TAP): A Striking Example of Functional Convergent Evolution

“…Based on current knowledge, the UL49.5 molecules are the only herpesvirus-encoded TAP inhibitors that fulfill a dual role in viral infection. Within the infected cell, UL49.5 forms a heterodimeric complex with glycoprotein M (gM) and guides proper glycosylation and maturation of this protein [ 86 – 88 ]. Cells expressing both BoHV-1 UL49.5 and gM show reduced TAP inhibition when compared to cells expressing UL49.5 only, suggesting that the interaction between UL49.5 and gM interferes with the capacity of UL49.5 to block TAP [ 89 ].…”

Structural Proteomics of Herpesviruses

“…Surprisingly, we did not detect any peptide corresponding to gN either in MuHV-4 or in BoHV-4 virions. However, as previously mentioned, gN and gM form a complex in herpesviruses [ 43 , 44 ] and gN is needed for the proper processing of gM. Moreover, we readily detected gN by Western blotting on the same MuHV-4 virions preparations even after the proteinase K treatment [ 29 ].…”