In an effort to understand the contribution of the primer-binding site (PBS) region to human immunodeficiency virus (HIV) replication, we have constructed a mutant HIV proviral DNA with an alteration in the 5' end of the PBS. The PBS mutant proviral DNA was characterized by transfection of the viral DNA into CD4' and non-CD4' target cells.The results indicate that mutation in the PBS reduced the level of viral particles released into the medium of transfected cells in comparison to wild-type proviral DNA. The viral particles were noninfectious upon transmission to established CD4' cell lines and phytohemagglutinin-stimulated peripheral blood lymphocytes. Electron microscopic analysis of the transfected cells revealed no abnormalities in the structure of the virion directed by the mutant proviral DNA. Also, the protein and RNA contents of the mutant virions were similar to the wild type. The quantitation of intracellular viral structural protein in the transfected cells, however, indicated that the PBS mutation may have an effect on the assembly of viral particles in addition to completely abolishing reverse transcription of viral RNA into DNA. These results provide evidence that the PBS region of the viral genome has multiple functions in HIV-I replication.

Herpes simplex virus type 1 (HSV-1) glycoprotein E (gE) functions as an immunoglobulin G (IgG) Fc binding protein and is involved in virus spread. Previously we studied a gE mutant virus that was impaired for IgG Fc binding but intact for spread and another that was normal for both activities. To further evaluate the role of gE in spread, two additional mutant viruses were constructed by introducing linker insertion mutations either outside the IgG Fc binding domain at gE position 210 or within the IgG Fc binding domain at position 380. Both mutant viruses were impaired for spread in epidermal cells in vitro; however, the 380 mutant virus was significantly more impaired and was as defective as gE null virus. gE mutant viruses were inoculated into the murine flank to measure epidermal disease at the inoculation site, travel of virus to dorsal root ganglia, and spread of virus from ganglia back to skin to produce zosteriform lesions. Disease at the inoculation and zosteriform sites was reduced for both mutant viruses, but more so for the 380 mutant virus. Moreover, the 380 mutant virus was highly impaired in its ability to reach the ganglia, as demonstrated by virus culture and realtime quantitative PCR. The results indicate that the domain surrounding amino acid 380 is important for both spread and IgG Fc binding and suggest that this domain is a potential target for antiviral therapy or vaccines.

Human submandibular saliva reduces human immunodeficiency virus type 1 (HIV-1) infection in vitro. To define the mechanism of inhibition, virus was incubated with saliva or medium, velocity sucrose gradient centrifugation was performed, and fractions were analyzed for p24 and gp120. The results show that after incubation with saliva, the envelope glycoprotein was displaced from both a laboratory-adapted and a low-passage clinical HIV-1 isolate. To identify the salivary protein(s) responsible, submandibular saliva was fractionated by anion- exchange chromatography. Protein fractions containing anti-HIV activity were assayed for their ability to strip gp120 from virus. The partially purified active fractions contained two high-molecular-weight sialyated glycoproteins identified as salivary agglutinin and mucin, as well as several lower-molecular-weight proteins. It thus appears that specific salivary proteins interact with HIV-1 to strip gp120 from the virus with a resultant decrease in infectivity.

Studies from a number of laboratories have shown the presence of factor(s) in whole, parotid, and submandibular human saliva capable of inhibiting HIV-1 infectivity in vitro. Data from our laboratory suggested that the level of anti-HIV-1 activity is higher in submandibular than parotid or whole saliva. Previous results obtained with pooled submandibular saliva from seronegative individuals included a filtration step following saliva-virus interaction. In this article, we present data on the HIV-1 inhibitory activity of individual submandibular saliva samples collected from 15 donors. We show that although anti-HIV activity is quantitatively similar in most individuals (9 of 15), some (4 of 15) are much less active than others and some (2 of 15) lack inhibitory activity. We also show that for most individuals the level of anti-HIV inhibitor is similar with or without a filtration step. However, 2 of the 15 samples demonstrated activity only after filtration. The quantitative and qualitative anti-HIV activity of individual saliva samples appeared to reflect differences in the individual donors. We further show that the anti-HIV activity of submandibular saliva is demonstrated not only against laboratory strains of HIV-1 but is similarly active against three clinical HIV-1 isolates. In contrast, submandibular saliva had little effect on the infectivity of HIV-2 or SIV.

Viruses have evolved strategies to evade immunity mediated by antibody and complement. Herpesviruses and coronaviruses encode IgG Fc binding proteins that inhibit IgG activity, enabling the virus or infected cell to escape antibody attack. Herpesviruses, vaccinia virus and HIV-1 have the capacity to interfere with complement, either by incorporation of cellular complement regulatory proteins into the virion envelope or cell membrane, or by expression of viral molecules that mimic functions of complement regulatory proteins. The structure and biological activities of herpes simplex virus type 1 (HSV-1) glycoproteins gE, gI and gC are described. These glycoproteins protect HSV from immune attack; HSV-1 gE/gI form a complex that binds the Fc domain of IgG while gC is a C3b binding complement regulatory protein, providing a survival advantage to the virus in vitro and in vivo by inhibiting immune functions.