Impact of dimerisation on interactions in the CTLA-4, CD28 and PD-1 pathways 3533

Abstract Description CD80 and CD86, the ligands for CD28/CTLA-4, appear functionally similar yet possess well characterised biophysical differences. For example, CD80 exists as a non-covalent homodimer that interacts with covalent CTLA-4 dimers with high affinity and avidity. In contrast, CD86 is a weak monomeric ligand. While CD28 is a covalent dimer, it is thought to bind monovalently to its ligands. CD80 homodimerisation is also impacted by PD-L1, which forms heterodimers with CD80. Thus a complex series of affinity, avidity and valency effects can be envisaged. We have performed experiments assessing the impact of manipulating dimer states in these pathways. We show that intact CD80 homodimers drive the ubiquitylation of CTLA-4 whereas monomeric CD80 and CD86 do not. Surprisingly, mCD80 enhances the efficiency transendocytosis, allowing CTLA-4 to dissociate and be recycled in a manner similar to interactions with CD86. These features were also observed using CD80-PD-L1 heterodimers, which generates a physiological monovalent CD80 target for CTLA-4. We also tested CTLA-4 molecules with disrupted dimerisation and observed impaired transendocytosis of CD86. Our data indicate that dimerisation and avidity influence the outcome of CTLA-4 transendocytosis in general and also suggest the presence of feedback loops where CD80-PD-L1 interactions (which prevent PD-1 function) are counterbalanced by enhanced CTLA-4 removal of CD80 which restores PD-L1 availability and thereby PD-1 function. Funding Sources Supported by Wellcome grant number: 227384/Z/23/Z Topic Categories Immune Response Regulation: Molecular Mechanisms (IRM)