Transmembrane protein CD69 acts as an S1PR1 agonist

Abstract

The activation of Sphingosine-1-phosphate receptor 1 (S1PR1) by S1P promotes lymphocyte egress from lymphoid organs, a process critical for immune surveillance and T cell effector activity. Multiple drugs that inhibit S1PR1 function are in use clinically for the treatment of autoimmune diseases. Cluster of Differentiation 69 (CD69) is an endogenous negative regulator of lymphocyte egress that interacts with S1PR1 incisto facilitate internalization and degradation of the receptor. The mechanism by which CD69 causes S1PR1 internalization has been unclear. Moreover, although there are numerous class A GPCR structures determined with different small molecule agonists bound, it remains unknown whether a transmembrane protein per se can act as a class A GPCR agonist. Here, we present the cryo-EM structure of CD69-bound S1PR1 coupled to the heterotrimeric Gicomplex. The transmembrane helix (TM) of one protomer of CD69 homodimer contacts the S1PR1-TM4. This interaction allosterically induces the movement of S1PR1-TMs 5-6, directly activating the receptor to engage the heterotrimeric Gi. Mutations in key residues at the interface affect the interactions between CD69 and S1PR1, as well as reduce the receptor internalization. Thus, our structural findings along with functional analyses demonstrate that CD69 acts incisas a protein agonist of S1PR1, thereby promoting Gi-dependent S1PR1 internalization, loss of S1P gradient sensing, and inhibition of lymphocyte egress.

Nuclear corepressors NCOR1 and NCOR2 entrain thymocyte signaling, selection, and emigration

“…CD69 mediates internalization of S1PR1; co-expression of S1PR1 and CD69 on the cell surface should be mutually exclusive 54,57 . We were therefore surprised to observe co-expression of CD69 and S1PR1 on the surface of CD4 SP thymocytes from DKO mice.…”

“…Why S1PR1 remained elevated in the presence of CD69 is less clear. Because the internalization of S1PR1 is a Gi-dependent process 54,57 , we considered which genes upregulated following Gi inhibition may be de-repressed in the absence of NCORs. The coupling between S1PR1 and Gi proteins is disrupted by pertussis toxin, blocking its inhibition of adenylyl cyclase 91 .…”

Get me out of here: Sphingosine 1‐phosphate signaling and T cell exit from tissues during an immune response

“…S1PR1 and CD69 proteins interact, as measured by co-immunoprecipitation and by cryoEM, and the key protein domains have been defined. 18,19,21 The transmembrane domain of CD69 binds the fourth transmembrane domain of S1PR1, and remarkably this shifts S1PR1 into a conformation similar to the S1P-bound structure, induces G-protein activation, and results in loss of surface S1PR1 and S1PR1 degradation. 18,21 While the relationship between CD69 and S1PR1 is clear, it will be important to investigate how their interaction is regulated.…”

“…18,19,21 The transmembrane domain of CD69 binds the fourth transmembrane domain of S1PR1, and remarkably this shifts S1PR1 into a conformation similar to the S1P-bound structure, induces G-protein activation, and results in loss of surface S1PR1 and S1PR1 degradation. 18,21 While the relationship between CD69 and S1PR1 is clear, it will be important to investigate how their interaction is regulated. For example, CD69 and likely S1PR1 have other binding partners 22,23 -does their abundance affect the ability of CD69 to sequester S1PR1?…”

“…First, S1PR5 has not been found to interact with CD69, and this may enable cells expressing S1PR5 to exit a site of acute inflammation. 21,48 Second, S1PR5 seems far more resistant to ligand-induced downmodulation than S1PR1. 48 If tissue S1P rises enough to result in substantial S1PR1 internalization, cells expressing S1PR5 may still be able to follow the residual S1P gradients out of the tissue.…”