Efficiency of signalling through cytokine receptors depends critically on receptor orientation

Abstract

Human erythropoietin is a haematopoietic cytokine required for the differentiation and proliferation of precursor cells into red blood cells. It activates cells by binding and orientating two cell-surface erythropoietin receptors (EPORs) which trigger an intracellular phosphorylation cascade. The half-maximal response in a cellular proliferation assay is evoked at an erythropoietin concentration of 10 pM, 10(-2) of its Kd value for erythropoietin-EPOR binding site 1 (Kd approximately equal to nM), and 10(-5) of the Kd for erythropoietin-EPOR binding site 2 (Kd approximately equal to 1 microM). Overall half-maximal binding (IC50) of cell-surface receptors is produced with approximately 0.18 nM erythropoietin, indicating that only approximately 6% of the receptors would be bound in the presence of 10 pM erythropoietin. Other effective erythropoietin-mimetic ligands that dimerize receptors can evoke the same cellular responses but much less efficiently, requiring concentrations close to their Kd values (approximately 0.1 microM). The crystal structure of erythropoietin complexed to the extracellular ligand-binding domains of the erythropoietin receptor, determined at 1.9 A from two crystal forms, shows that erythropoietin imposes a unique 120 degrees angular relationship and orientation that is responsible for optimal signalling through intracellular kinase pathways.

Synthetic Multivalent Ligands as Probes of Signal Transduction

“…The relative orientations of the receptor-associated enzyme active site and the substrate may influence the amount of product generated; this enzymatic efficiency, in turn, will influence signal transduction. [232][230] [229][225] When a signaling complex assembles, it is often unclear whether its activity is due to a specific orientation of receptors within the assembled complex or simply the localization of signaling components. [233] Thus, questions remain about the roles of receptor orientation in signaling.…”

The molecular and cellular biology of thrombopoietin: the primary regulator of platelet production

“…Careful study of the ways in which such molecules trigger receptor activation have also yielded important new insights into the structural biology of hematopoietic cytokines (Syed et al, 1998;Remy et al, 1999). Continued research in this ®eld is certain to bring new therapeutic molecules and a greater understanding of hematopoiesis.…”

Discovery and basic pharmacology of erythropoiesis-stimulating agents (ESAs), including the hyperglycosylated ESA, darbepoetin alfa: an update of the rationale and clinical impact

“…EPOR (erythropoietin receptor), which is found on the surface of erythroid precursor cells, is a cell- Fig. 1 Amino acid sequence of human erythropoietin (EPO) with glycosylation at three specific asparagines residues (Asn24, Asn34, and Asn83) and one serine residue (Ser126) [3] membrane-bound protein that is activated when one EPO molecule binds to two receptors in a process called homodimerization [8,11,12]. When EPO receptors on erythroid precursor cells are activated by EPO, the cells can be stimulated to survive, proliferate, and differentiate into RBCs.…”