Type 1 human T-cell leukemia virus small envelope protein expressed in mouse cells by using a bovine papilloma virus-derived shuttle vector.

Abstract

In an attempt to express the small (transmembrane) envelope protein p2le of type 1 human T-cell leukemia (lymphotrophic) virus (HTLV-1) exclusive of other viral gene products, we have constructed a recombinant plasmid clone (pMBE-1) in a bovine papillomavirus-derived mammalian expression vector. Mouse C127 cells transfected with the pMBE-1 plasmid expressed the introduced HTLV-1 viral gene(s) as demonstrated by Northern blot and indirect immunofluorescence with natural human antisera. The transfected mouse cells were injected into BALB/c mice, and a monoclonal antibody was recovered which specifically recognizes a 21-kilodalton protein present in HTLV-1 virions, indicating that the pMBE-1 plasmid encodes the small envelope protein.Type 1 human T-cell leukemia (lymphotropic) virus (HTLV-1) is a retrovirus isolated from a variety of cell lines established from patients with adult T-cell leukemia or lymphoma (16,23). The HTLV-1 retroviral genome consists of four separate genes encoding distinct viral proteins (19), specifically, the gag, pol, and env genes and a region, pX, located between the 3' end of the envelope gene and the 3' long terminal repeat U3. Only recently have the viral envelope proteins been characterized (7,18,21). These include a glycosylated protein of 62 kilodaltons (kDa), derived from an amino acid backbone of 46 kDa, which serves as the precursor for the small envelope protein p21e and the large envelope protein gp46.The small envelope gene product (plSe) of murine and feline retroviruses has been associated with profound immunosuppressive effects (20). These effects include inhibition of lymphocyte blastogenesis (12) and depressed macrophage-mediated functions (2). The HTLV-1 small envelope protein p21e shares 73% amino acid sequence homology with the pl5e proteins of various murine and feline retroviral isolates (1). Infection of immunocompetent T cells in vitro with HTLV-1 results in the abrogation of various immune functions, including specific recognition of antigen, cytotoxic function, and the requirement for macrophages in antigen recognition (14). It is not clear whether the p2le of HTLV-1 shares the immune inhibitory properties of the plSe proteins. To begin to address this issue, we designed a mammalian expression vector (pMBE-1) to direct the synthesis of most of the HTLV-1 p21e exclusive of the rest of the envelope gene product. We show here results obtained by using a bovine papillomavirus (BPV)-derived vector in which an HTLV-1 DNA segment containing most of the coding region of the putative small envelope protein, the pX region, and the 3' long terminal repeat is located downstream from a mouse metallothionein promoter. The results demonstrate the utility of this vector system in expressing a subgenic region of the HTLV-1 genome in mammalian cells * Corresponding author. in the absence of other virally coded or induced cell proteins. In addition, it provides a rapid and simple means of producing specific monoclonal antibodies by using the transfected cells rather than cells i...

The Effect of Bovine Herpesvirus Type 1 Glycoproteins gI and gIII on Herpesvirus Infections

“…Although BPV vectors have been used to express the HSV-1 thymidine kinase gene (Lusky et al, 1983) and other surface proteins such as the envelope glycoprotein of human T cell leukaemia virus (Eiden et al, 1985), hepatitis B surface protein (Wang et al, 1983), and the influenza virus haemagglutinin (Sambrook et al, 1985), this represents the first expression of a herpesvirus glycoprotein gene using a BPV vector. The gI and glII proteins expressed by our transfected bovine cells were indistinguishable from native BHV-1 gI and glII by comparison of Mr and antigenic areas defined by all monoclonal antibodies used to date.…”

“…1 a). Plasmid pBPV-C # 1 was constructed by ligating the gIII gene (Zamb, 1987) into p341 (Eiden et al, 1985). The resultant plasmid, pMC10B, was digested, and the fragment containing the MMT promoter, gIII gene and SV40 polyadenylation site was isolated and ligated into the BamHI site of pCGBPV9AB5 (Matthias et al, 1983).…”

“…The plasmid pBPV-B# 1 was constructed by ligating the gI gene (Zamb, 1987) into p341 (Eiden et al, 1985) between the MMT promoter and the simian virus 40 (SV40) polyadenylation site. This plasmid was ligated to the BPV fragment of BamHI-digested pNeoBPV100 (Lusky & Botchan, 1984).…”

The bovine papillomavirus genome and its uses as a eukaryotic vector

“…A similar pattern of cell surface expression has been seen for the vesicular stomatitis virus glycoprotein [92,93]. Other cell surface markers include the human heavy chain of an HLA human histocompatibility antigen [64], the transmembrane envelope protein p2le of type 1 human T-cell leukaemia virus [89] and the interleukin 2 receptor [90]. While such studies indicate the utility of BPV expression vector in studying membrane events, the host cell range may limit the potential in this area.…”

“…They also include the viral glycoproteins hepatitis B surface antigen, HBSag [12,66,87,88], influenza virus haemagglutinin and cap-recognizing proteins [61,68] and type 1 human T-cell leukaemia virus small envelope protein [89]. The system has in addition been used to express normal cell surface markers such as the heavy chain of an HLA human histocompatability antigen [64] and the human interleukin 2 receptor [90].…”