10 February 2022 Latest Research Published: CD4 and CD8 co-receptors modulate functional avidity of CD1b-restricted T cells

Professor Tom Scriba has co-authored a research paper titled “CD4 and CD8 receptors modulate functional avidity of CD1b-restricted T-cells” appearing in the Nature Communications journal. 

Abstract

T cells recognize mycobacterial glycolipid (mycolipid) antigens presented by CD1b molecules, but the role of CD4 and CD8 co-receptors in mycolipid recognition is unknown. Here we show CD1b-mycolipid tetramers reveal a hierarchy in which circulating T cells expressing CD4 or CD8 co-receptor stain with a higher tetramer mean fluorescence intensity than CD4-CD8- T cells. CD4+ primary T cells transduced with mycolipid-specific T cell receptors bind CD1b-mycolipid tetramer with a higher fluorescence intensity than CD8+ primary T cells. The presence of either CD4 or CD8 also decreases the threshold for interferon-γ secretion. Co-receptor expression increases surface expression of CD3ε, suggesting a mechanism for increased tetramer binding and activation. Targeted transcriptional profiling of mycolipid-specific T cells from individuals with active tuberculosis reveals canonical markers associated with cytotoxicity among CD8+ compared to CD4+ T cells. Thus, expression of co-receptors modulates T cell receptor avidity for mycobacterial lipids, leading to in vivo functional diversity during tuberculosis disease

Discussion: In summary, we have shown that SGL-specific T cells analyzed directly ex vivo exhibit a bias towards CD4 expression, and our data reveal that CD4 SGL-specific T cells have a higher functional avidity than CD8 SGL-specific T cells. Whether we examined ex vivo, in vitro-derived, or transduced SGL-specific T cells, we observed a consistent hierarchy in which CD4 T cells bind SGLCD1b tetramer with a higher fluorescence intensity and have the lowest activation thresholds, followed by CD8 T cells, and DN T cells. In fact, our data show that the surface expression of a TCR co-receptor is associated with increased levels of TCR expression, revealing a potential mechanism for the increase in activation and functional tetramer avidity we observed. Finally, we examined the ex vivo transcriptional profiles of SGL-specific T cells in patients with active tuberculosis and found canonical transcription factor lineages among CD4 and CD8 CD1brestricted T cells. These data significantly advance our understanding of how co-receptors modulate the recognition of lipid antigens and reveal diverse functional profiles of CD1b-restricted T cells in humans. A previous report found that the CD4 co-receptor is not involved in lipid antigen recognition by CD1b-restricted T cells38. This study used blocking antibodies targeting the interaction between CD4 and MHC-II, which may not have reliably predicted the effect on CD1b. Further, the use of co-receptor blocking antibodies can have pleiotropic effects, including paradoxical activation, of T cells39. To avoid this limitation, we adopted a molecular approach to this question, and our results uncover a role for TCR co-receptors in lipid-specific T cell activation. Previous reports have shown that the CD4 co-receptor enhances iNKT cell activation, which the authors posit was through direct binding of CD4 to CD1d17,40. Our data extend these studies by revealing a potential molecular mechanism in the form of stabilized TCR expression at the surface of CD1brestricted T cells, which would not have emerged from experiments using blocking antibodies. In addition to increasing TCR expression, there may be other potential mechanisms that contribute to increasing the functional avidity of CD1b-restricted T cells, and these mechanisms are not mutually exclusive. First, TCR co-receptors may physically bind to CD1b and augment TCR affinity41. However, due to the structural differences between MHC-II and CD1b, putative binding sites for this interaction are not obvious. Second, precise spatial control of the immunologic synapse is important for tuning TCR sensitivity to antigen, possibly by controlling the level of cholesterol in lipid rafts42–44. Third, the rate of recycling in endosomes and activation state of T cells can influence TCR stability at the cell surface45,46. Due to the multifaceted nature of TCR engagement in live cells, the mechanism for the increase in functional avidity we report here is representative of in vivo antigen recognition and may be unrelated to alterations in the affinity between TCR and CD1b that could be measured using standard biophysical approaches47. Our data have important implications for antigen recognition in vivo. Previous studies have highlighted that subtle differences within the lipid tail can significantly influence TCR affinity for antigen, which translates into differences in function25,48–50. Our data suggest that co-receptor expression must also be factored into this calculation as our single-cell analyses of SGL- and GMM-specific T cells demonstrate that glycolipid-specific T cells that express CD4 and CD8 co-receptors are functionally distinct. Indeed, our data suggest that these subsets may align closely with peptide-specific T cells, rather than iNKT or MAIT cells51. Our data reveal previously unappreciated functional diversity of CD1b-restricted T cells as we detected expression of TBET, GATA3, RORC, and FOXP3 among circulating CD1b-restricted T cells ex vivo. Mycobacterial glycolipid-specific T cells that express the CD8 co-receptor are broadly cytotoxic T cells. This is consistent with the original description of an SGL-specific T cell clone, which produced IFN-γ in the presence of M.tb infected cells and reduced the bacterial burden of M.tb in culture24. Conversely, T cells that express the CD4 co-receptor are more diverse. At present, the only transcription factor that has been evaluated in CD1b-restricted T cells is PLZF, the transcription factor that regulates innate-like phenotypes in iNKT and MAIT cells52–54. In a humanized transgenic mouse model, CD1brestricted T cells did not express PLZF, which indicates that CD1b-restricted T cells may have a transcriptional landscape that is similar to MHC-restricted T cells55. Our findings that CD4 and CD8 CD1b-restricted T cells share major transcriptional pathways with MHC-restricted T cells is consistent with this model51. However, we were unable to detect an enrichment of RUNX3 in CD8 CD1b-restricted T cells, highlighting that there may also exist transcriptional pathways that are unique to CD1b-restricted T cells56 (Supplementary Table 4). Together, our data highlight the importance of considering co-receptor expression by CD1b-restricted T cells when evaluating lipid-containing vaccines in clinical and pre-clinical models as CD4 T cells may be more likely to be activated in the context of infection, possibly as a result of enhanced functional avidity. Lipid-based vaccines have shown promise in preclinical animal models57. CD4 or CD8 CD1b-restricted T cells could be differentially targeted by the adjuvant or delivery platform when ‘tuning’ the chemical structure of the antigen to modify TCR avidity and immunogenicity of these vaccine strategies.