Identification of exposed epitopes on the envelope glycoproteins of human T-cell lymphotropic virus type I (HTLV-I)

Abstract

Several lines of evidence underscore the important role of the humoral response specific for HTLV‐I envelope protein in the protection against viral infection. One approach to producing efficient immunogens is to synthesize peptides corresponding to the primary amino‐acid sequence of neutralizing epitopes found in the external sub‐unit gp46. In this study, we have selected synthetic peptides overlapping the major linear neutralizing determinants described earlier and used them as immunogens in rabbits and mice. All rabbit polyclonal antisera raised against peptides recognized epitopes in a denaturated context as well as MAbs raised against the HB peptide (aa287‐311). By contrast, synthetic peptides O (aa89‐110), HH (aa190‐209), T (aa190‐212) and HB (aa287‐311) have generated antibodies efficiently binding their epitopes in a native context, suggesting that these domains are well exposed both at the heterodimer and at the oligomer surface. None of the antibodies induced by synthetic peptides show in vitro neutralizing properties, even those with a good capacity to bind the native form of HTLV‐I envelope proteins. Int. J. Cancer 75:804–813, 1998.© 1998 Wiley‐Liss, Inc.

HTLV-1 uses HSPG and neuropilin-1 for entry by molecular mimicry of VEGF165

“…Beads were washed 5 times in lysis buffer, and proteins were eluted by boiling for 5 minutes in Laemmli buffer. Immunoblot analysis were performed using either the anti-SU 4D4 monoclonal antibody (mAb) provided by Claude Desgranges (Institut Cochin, Paris, France) 34 or the anti-HA antibody (Roche Diagnostics). For quantification, briefly exposed films were scanned with an AGFA scanner, and signal densities of Env or HA-sNRP-1 bands were measured with ImageJ software using the same area.…”

The Humoral Immune Response to Human T-Cell Lymphotropic Virus Type 1 Envelope Glycoprotein gp46 Is Directed Primarily against Conformational Epitopes

“…One sequence located in the amino-terminal half of HTLV-1 gp46 has been shown to induce HTLV-1-specific neutralizing antibodies when it is inoculated into goats (amino acids 90 to 98 [28]). However, in a recent study mMAbs generated to a similar peptide (amino acids 89 to 110) did not immunoprecipitate native HTLV-I gp46 or recognize HTLV-I gp46 on the surfaces of infected cells (10). Consequently, this particular sequence may not be accessible in native gp46.…”

“…Most studies have focused on antibodies to linear epitopes located on the carboxy-terminal half of gp46 (amino acids 170 to 312 [3,4,5,7,8,10,15,17,19,27]). These antibodies are found in more than 95% of infected individuals (reviewed in references 11 and 18), but the majority of antibodies to these epitopes do not mediate virus neutralization (4,7,10). Linear epitopes located in the middle region of the envelope (amino acids 175 to 199), as defined by monoclonal antibodies, are more likely to have neutralizing activity (4).…”

Early Assembly Step of a Retroviral Envelope Glycoprotein: Analysis Using a Dominant Negative Assay

“…Immunoprecipitations of the HTLV-1 envelope glycoproteins were performed as described in our previous studies ( Pique et al, 1990 ), using protein A–Sepharose CL-4B beads ( Pharmacia Biotechnology ) coated with a pool of sera from HTLV-1–infected individuals (provided by J. Coste, CRTS, Montpellier, France). For coimmunoprecipitation experiments, protein A–Sepharose beads were coated with rabbit anti–mouse Ig (Dako SA) plus purified 4D4 mAb, which is a murine mAb raised against a synthetic peptide covering the COOH-terminal domain (amino acids 287– 311) of the HTLV-1 SU (kindly donated by C. Desgranges and M.-P. Grange, INSERM U271, Lyon, France) ( Grange et al, 1998 ). Immunoprecipitates were electrophoresed in SDS-13% polyacrylamide gels under reducing conditions (except where otherwise stated), and visualized by autoradiography.…”

“…Immunoprecipitates were electrophoresed in SDS-10% polyacrylamide gels under reducing conditions, and proteins were transferred onto membranes (Immobilon-P; Millipore Corp. ). After saturation in PBS containing 0.05% Tween-20 and 5% skim milk, the membranes were incubated with a 1:3,000 dilution of the mAb 4D4 ( Grange et al, 1998 ) for 90 min at room temperature. Excess antibody was removed with six washes, and a 1:1,000 dilution of the second-step antibody (peroxidase-conjugated goat anti–mouse IgG; Jackson ImmunoResearch Laboratories, Inc.) was allowed to bind for 60 min.…”