Abstract by Fabian Pfersdorf
Chemokines and their receptors comprise a complex network, which controls cell migration in developmental processes, tissue repair, and immunological responses. Due to its involvement in a plethora of biological processes, the chemokine network is a key component in a wide variety of diseases, including e.g. autoimmune diseases and cancer. The signaling axis around the chemokine CXCL12 – including its two cognate receptors CXC-chemokine receptor 4 (CXCR4) and atypical chemokine receptor 3 (ACKR3) – is of specific interest due to its involvement in a broad range of health and disease mechanisms and its indispensability for life. Yet, there is a lot we don’t fully understand about the mechanisms underlying the modulation of this system. The objective of this thesis and the research presented herein is to shed light on some of these regulatory mechanisms.
For Manuscript I, we investigated the effects of receptor activity-modifying proteins (RAMPs) on CXCR4 and ACKR3 function. We found that CXCR4 and ACKR3 are differently modulated by RAMPs, with marked effects on both receptors’ β-arrestin recruitment and on ACKR3’s endocytosis and scavenging properties. In the case of ACKR3, RAMPs exert their effects at the cell membrane with only minor ligand-specific effects and no detectable involvement of the receptor’s N-terminus. We therefore suggest that this modulation occurs by allosteric interactions through the transmembrane domains.
For Manuscript II, we investigated how an altered receptor conformation affects the binding of ligands. For this purpose, we identified ACKR3D244A and ACKR3K247E as more and less constitutively active receptor variants, respectively, due to their effects on the receptor’s conformational equilibrium. Utilizing these conformation-biasing mutants, we found that ACKR3 conformation affects different agonists differently and influences the binding kinetics of CXCL12, with a slower chemokine dissociation from the more active receptor and vice versa. With this, we propose a mechanism by which the cellular surroundings of ACKR3 contribute to receptor activity and ligand preference.
In summary, the signaling axis around CXCL12 and its two cognate receptors – CXCR4 and ACKR3 – is modulated at different levels. RAMP binding to either receptor likely modulates receptor conformation, which affects not only receptor downstream function but also the upstream kinetics of chemokine binding.