Human IL-34 and CSF-1 Establish Structurally Similar Extracellular Assemblies with Their Common Hematopoietic Receptor

Abstract

The discovery that hematopoietic human colony stimulating factor-1 receptor (CSF-1R) can be activated by two distinct cognate cytokines, colony stimulating factor-1 (CSF-1) and interleukin-34 (IL-34), created puzzling scenarios for the two possible signaling complexes. We here employ a hybrid structural approach based on small-angle X-ray scattering (SAXS) and negative-stain EM to reveal that bivalent binding of human IL-34 to CSF-1R leads to an extracellular assembly hallmarked by striking similarities to the CSF-1:CSF-1R complex, including homotypic receptor-receptor interactions. Thus, IL-34 and CSF-1 have evolved to exploit the geometric requirements of CSF-1R activation. Our models include N-linked oligomannose glycans derived from a systematic approach resulting in the accurate fitting of glycosylated models to the SAXS data. We further show that the C-terminal region of IL-34 is heavily glycosylated and that it can be proteolytically cleaved from the IL-34:hCSF-1R complex, providing insights into its role in the functional nonredundancy of IL-34 and CSF-1.

Solution Structures of Complement C2 and Its C4 Complexes Propose Pathway-specific Mechanisms for Control and Activation of the Complement Proconvertases

“…Recent work on two different extracellular receptors by Savvides and co-workers (28,29) has convincingly shown the importance of including Asn-linked glycans in SAXS rigid body modeling. The putative glycan content is 16% for C2 and 9 -10% for C4b2 and C4b2a, meaning that glycans contributed significantly to their scattering curves.…”

CSF-1 Receptor Signaling in Myeloid Cells

“…Binding of CSF-1 to the CSF-1R is exclusively via the D2 and D3 domains, the D4 domains mediating CSF-1R homotypic interactions, via a broad interaction interface, whereas the D1 and D5 domains point away from the complex (Fig. 2B) Elegheert et al 2011;Ma et al 2012;Felix et al 2013). D4 shares a dimerization domain sequence fingerprint that has been identified in other closely related RTK III receptors, Kit and PDGFR (Yuzawa et al 2007;Yang et al 2008), and the presence of domains D4 and D5 in the CSF-1:CSF-1R D1 -D5 significantly decreases the K d of interaction Elegheert et al 2011;Ma et al 2012) owing to CSF-1R homotypic interactions.…”

“…2B) Elegheert et al 2011;Liu et al 2012;Ma et al 2012;Felix et al 2013). Despite their similarities, the IL-34:CSF-1R complex differs from the CSF-1:CSF-1R complex because: (1) there is a 20˚rotation difference of their D3 domains when their D2 domains are superimposed, resulting in an elongated pose that differs from the kinked configuration of the CSF-1R:CSF-1 complex; (2) CSF-1 is clamped deeper by the CSF-1R D2-D3 junction than IL-34, so that overlapping, yet different CSF-1R segments are used by each ligand; (3) to compensate for loss of some D2 interactions, IL-34 adopts amino-terminal and carboxy-terminal extensions to contact D3 (Liu et al 2012;Ma et al 2012); and (4) the apparent rigidity of the IL-34 structure (Liu et al 2012;Ma et al 2012) differs from the more plastic CSF-1 structure, which undergoes local structural rearrangements for receptor binding .…”

“…Mutation of individual hydrophilic residues involved in IL-34 interactions with CSF-1R D2 failed to affect IL-34 biological activity, whereas mutations of residues interacting with D3 substantially reduced IL-34 activity suggesting that charge interactions with D2 may capture IL-34 for subsequent interaction with D3 (Liu et al 2012). Despite the differences between the IL-34:CSF-1R and CSF-1:CSF-1R complexes, the distance between the two D3-D4 junctions important for critical homotypic D4 interactions (Elegheert et al 2011;Felix et al 2013) is equivalent (62 Å and 60 Å , respectively) (Ma et al 2012). …”

IL-34 and M-CSF form a novel heteromeric cytokine and regulate the M-CSF receptor activation and localization

“…However, the main and original characteristic of IL-34 remains its structural plasticity, associated with its small and hydrophobic dimerization interface in its non-covalent dimeric form [15]. These two related ''twin'' cytokines use a similar bivalent mode for binding to the M-CSFR, leading to homotypic M-CSFR/M-CSFR interactions through their D4 domain [17]. The M-CSFR belongs to the class III receptor-type tyrosine kinases, exhibiting five Ig-like extracellular domains [18].…”