The Crystal Structure of the Costimulatory OX40-OX40L Complex

Abstract

OX40 is a T cell costimulator activated by OX40L. Blockade of the OX40L-OX40 interaction has ameliorative effects in animal models of T cell pathologies. In order to better understand the interaction between OX40 and OX40L, we have determined the crystal structure of murine OX40L and of the human OX40-OX40L complex at 1.45 and 2.4 A, respectively. These structures show that OX40L is an unusually small member of the tumor necrosis factor superfamily (TNFSF). The arrangement of the OX40L protomers forming the functional trimer is atypical and differs from that of other members by a 15 degrees rotation of each protomer with respect to the trimer axis, resulting in an open assembly. Site-directed changes of the interfacial residues of OX40L suggest this interface lacks a single "hot spot" and that instead, binding energy is dispersed over at least two distinct areas. These structures demonstrate the structural plasticity of TNFSF members and their interactions with receptors.

Assembly and structural properties of glucocorticoid-induced TNF receptor ligand: Implications for function

“…4B) and are consistent with a homology model of the hGITRL-GITR complex (Fig. 4C) that was generated by using the recently reported hOX40L-OX40 complex structure (12) as the template. The model suggests that solvent-accessible residues from the AAЈ, CD, DE, and GH loops of hGITRL lie in close proximity to hGITR and thus are poised to contribute to the hGITR recognition surface.…”

“…The structure also reveals that the hGITRL trimer interfaces are remarkably small and lack the tightly packed aromatic/hydrophobic residues commonly observed in traditional TNF trimer interfaces. An approximately similar expanded organization has been recently reported for OX40L (12), and sequence analysis suggests that OX40L, GITRL, and CD30L may represent a distinct subfamily within the TNF family that is characterized by a short (Ϸ120-130 residues) TNF homology domain (THD) compared with the traditional Ϸ150-residue THDs. Consistent with the relatively small intersubunit interface observed in the crystalline state, hGITRL also displays considerably weaker tendency to trimerize in solution compared with other TNF ligands.…”

“…6A) and contrasts with the more compact assembly of conventional TNF trimers. The only other TNF ligand that has been demonstrated to possess an approximately comparable organization is OX40L (12) for which the crystal structure has been solved recently (SI Fig. 6 C and D).…”

Structural basis for ligand-mediated mouse GITR activation

“…We have determined the crystal structure of mGITRL. We expected that mGITRL to form a trimer resembling the recently reported structure of OX40L (11). However, our results show that soluble mGITRL associates as a homodimer, which does not fit the paradigm.…”

Evolution of GITRL immune function: Murine GITRL exhibits unique structural and biochemical properties within the TNF superfamily

“…In the majority of cases, TNF ligands selfassemble as ''bell-shaped'' homotrimers in which monomeric subunits are noncovalently associated through interactions between highly conserved hydrophobic surfaces (2,18). Recently, the structures of OX40L (19) and human GITRL (15) revealed an expanded trimeric arrangement that more closely resemble an ''open flower-like'' assembly. Despite the somewhat altered organization, their gross overall quaternary features, in combination with the location of their receptor binding surfaces, clearly indicate that OX40L and human GITRL direct formation of a 3:3 receptor:ligand complex with overall similarity to other TNF:TNFR complexes.…”