Anti-CD8 Antibodies Can Inhibit or Enhance Peptide-MHC Class I (pMHCI) Multimer Binding: This Is Paralleled by Their Effects on CTL Activation and Occurs in the Absence of an Interaction between pMHCI and CD8 on the Cell Surface

Abstract

Cytotoxic T lymphocytes recognize short peptides presented in association with MHC class I (MHCI) molecules on the surface of target cells. The Ag specificity of T lymphocytes is conferred by the TCR, but invariable regions of the peptide-MHCI (pMHCI) molecule also interact with the cell surface glycoprotein CD8. The distinct binding sites for CD8 and the TCR allow pMHCI to be bound simultaneously by both molecules. Even before it was established that the TCR recognized pMHCI, it was shown that CTL exhibit clonal heterogeneity in their ability to activate in the presence of anti-CD8 Abs. These Ab-based studies have since been interpreted in the context of the interaction between pMHCI and CD8 and have recently been extended to show that anti-CD8 Ab can affect the cell surface binding of multimerized pMHCI Ags. In this study, we examine the role of CD8 further using point-mutated pMHCI Ag and show that anti-CD8 Abs can either enhance or inhibit the activation of CTL and the stable cell surface binding of multimerized pMHCI, regardless of whether there is a pMHCI/CD8 interaction. We further demonstrate that multimerized pMHCI Ag can recruit CD8 in the absence of a pMHCI/CD8 interaction and that anti-CD8 Abs can generate an intracellular activation signal resulting in CTL effector function. These results question many previous assumptions as to how anti-CD8 Abs must function and indicate that CD8 has multiple roles in CTL activation that are not necessarily dependent on an interaction with pMHCI.

Availability of a Diversely Avid CD8+ T Cell Repertoire Specific for the Subdominant HLA-A2-Restricted HIV-1 Gag p2419–27 Epitope

“…4A). For each E:T cell combination, CD8 was more important for T cell activation under limiting Ag density, consistent with previous reports (51,65).…”

“…The degree to which individual CD8 ϩ T cells depend on the pMHCI/CD8 interaction for stable tetramer binding can be evaluated by using pMHCI tetramers carrying mutations in the ␣ 3 domain that abrogate CD8 binding without affecting TCR recognition (CD8 null ) (51,(63)(64)(65)(66). Fig.…”

Characterizing the Impact of CD8 Antibodies on Class I MHC Multimer Binding

“…However, these interpretations have been challenged by recent data from Wooldridge et al (12). These authors presented intriguing data that argued that CD8 Abs have dramatic effects on peptide/MHC multimer binding even when the multimer was not expected to engage CD8.…”

“…These authors suggested that, rather than directly influencing CD8-class I multimer binding, CD8 Ab binding was influencing the ability of the TCR to engage the multimer. They went on to propose that, although some of these effects could be mediated by steric hindrance, a more likely model was that anti-CD8 Abs were causing changes in TCR distribution on the cell surface, leading to altered multimer binding (12). These data raised serious doubts about the proposed role for direct CD8 participation in multimer binding and also the validity of using anti-CD8 Abs to explore such a role.…”

Selection of T Cell Clones Expressing High-Affinity Public TCRs within Human Cytomegalovirus-Specific CD8 T Cell Responses

“…Although there is a gross correlation between resistance to blocking anti-CD8 mAb and TCR affinity, anti-CD8 mAb may in some cases inhibit CD8-independent T cell responses, presumably through delivery of inhibitory signals or nonspecific steric hindrance (35). TCR affinity can be more accurately assessed by analyzing binding of "CD8-null" HLA-A2/peptide tetramers, in which the ␣3 domain of HLA-A2 has been replaced by the ␣3 domain derived from a murine H-2 allele (29).…”