Neutralization of feline immunodeficiency virus by antibodies targeting the V5 loop of Env

Abstract

Neutralizing antibodies (NAbs) play a vital role in vaccine-induced protection against infection with feline immunodeficiency virus (FIV). However, little is known about the appropriate presentation of neutralization epitopes in order to induce NAbs effectively; the majority of the antibodies that are induced are directed against non-neutralizing epitopes. Here, we demonstrate that a subtype B strain of FIV, designated NG4, escapes autologous NAbs, but may be rendered neutralization-sensitive following the insertion of two amino acids, KT, at positions 556-557 in the fifth hypervariable (V5) loop of the envelope glycoprotein. Consistent with the contribution of this motif to virus neutralization, an additional three subtype B strains retaining both residues at the same position were also neutralized by the NG4 serum, and serum from an unrelated cat (TOT1) targeted the same sequence in V5. Moreover, when the V5 loop of subtype B isolate KNG2, an isolate that was moderately resistant to neutralization by NG4 serum, was mutated to incorporate the KT motif, the virus was rendered sensitive to neutralization. These data suggest that, even in a polyclonal serum derived from FIV-infected cats following natural infection, the primary determinant of virus-neutralizing activity may be represented by a single, dominant epitope in V5. INTRODUCTIONFeline immunodeficiency virus (FIV) was first isolated in 1986 in Petaluma, CA, USA, from a cat with an immunodeficiency-like syndrome (Pedersen et al., 1987) and is an important pathogen of domestic cats worldwide. FIV is the feline homologue of human immunodeficiency virus (HIV) and these viruses share many biological and pathogenic features (Bendinelli et al., 1995;English et al., 1993;Johnson et al., 1994), typified by a selective targeting of CD4 + T lymphocytes and the induction of a progressive immunosuppression (Ackley et al., 1990).The induction of an effective humoral response against lentivirus infection is a key element in the immunological control of the disease, and the primary target for neutralizing antibodies (NAbs) is the virus envelope glycoprotein (Env) (Binley et al., 2004;Frost et al., 2005;Wei et al., 2003). Env varies by up to 30 % amongst FIV subtypes (Hosie & Beatty, 2007) and, thus, the preparation of an immunogen capable of inducing broadly neutralizing antibody responses against such diverse isolates of FIV would be of great value to the development of an FIV vaccine. Further, the development of a vaccine against FIV would have implications extending beyond veterinary medicine; FIV is the only non-primate lentivirus that induces AIDS-like symptoms in its natural host and, as such, is a valuable animal model for both prophylactic and therapeutic studies for HIV (Bendinelli et al., 1995;Elder et al., 1998;Okada et al., 1994). Moreover, cats have the advantage of being easier to breed and have shorter life cycles than other animal models currently used for HIV research (Miller et al., 2000).Previous studies showed that vaccination with whole inactivated va...

The Molecular Biology of Feline Immunodeficiency Virus (FIV)

“…Env is shrouded in carbohydrate, linked to the protein via glycosylation sites, and this appears to be very important in masking potential neutralising epitopes; de novo mutations appear altering the glycosylation pattern in response to host antibodies [20]. A single region in the FIV envelope V5 loop plays a dominant role in whether the virus is neutralised or escapes neutralisation: a long-term study of viral variants during infection of individual cats found the appearance of many new N -glycosylation sites within this loop [21] and an insertion of just two residues at position 556–7 re-sensitises escape mutants to neutralisation [22]. Mutation of the other variable loops is common with the V1,V2 and V4 regions also showing variations in glycosylation pattern [21].…”

Feline Immunodeficiency Virus (FIV) Neutralization: A Review

“…Infection with the residual, non-neutralized pseudotype was then quantified by measuring luciferase activity and the percent neutralization was calculated with reference to control wells containing no plasma. Plasmas were classified as strongly neutralizing (≥80%), moderately neutralizing (60–79%), and weakly neutralizing (40–59%) [35]. …”

“…The mechanism of escape from neutralization differed between variants, involving either shortening, lengthening or mutating of V5 (Figure 3). The importance of V5 in FIV neutralization is not restricted to subtype A viruses; in a separate study a neutralization-resistant Japanese subtype B strain of FIV (NG4) escaped neutralization by homologous and heterologous sera following the elimination of a potential site for N-linked glycosylation in V5 [35]. The V5 loop of FIV Env is highly variable among isolates, with variation occurring not only within the amino acid sequence but also in the length of the central region (up to 14 residues) towards the end of the loop structure formed by disulfide bond linkage.…”

“…One possibility is that the prototypic vaccines failed to induce neutralizing antibody against conformational epitopes that would be required to confer heterologous immunity; evidence to date would suggest that the majority of the neutralizing responses that develop in vivo are strain-specific and target V4 and V5 [35,52,65–69,101] while broadly neutralizing antibodies are rare. The breadth of the immunity induced by FL4-based vaccines has been extended by the introduction of heterologous strains of virus into the FL4 cell line [102–104], and such vaccines have afforded limited success.…”

“…Sequential plasma samples collected from three cats ( A , B —611, C , D —612, E , F —613) infected experimentally with a molecular clone of FIV GL8 were diluted 1:10 and examined for ability to neutralize HIV (FIV) pseudotypes bearing the GL8 Env as described [35]. Luciferase activity ( A , C , E ) was measured at three days post-infection and the percentage neutralization ( B , D , F ) calculated relative to a “no plasma” control.…”