Structure of FcRY, an avian immunoglobulin receptor related to mammalian mannose receptors, and its complex with IgY

Abstract

Fc receptors transport maternal antibodies across epithelial cell barriers to passively immunize newborns. FcRY, the functional counterpart of mammalian FcRn (a major histocompatibility complex homolog), transfers IgY across the avian yolk sac, and represents a new class of Fc receptor related to the mammalian mannose receptor family. FcRY and FcRn bind immunoglobulins at pH ≤6.5, but not pH ≥7, allowing receptor-ligand association inside intracellular vesicles and release at the pH of blood. We obtained structures of monomeric and dimeric FcRY and an FcRY-IgY complex and explored FcRY's pH-dependent binding mechanism using electron cryomicroscopy (cryoEM) and small-angle X-ray scattering. The cryoEM structure of FcRY at pH 6 revealed a compact double-ring "head," in which the N-terminal cysteine-rich and fibronectin II domains were folded back to contact C-type lectin-like domains 1-6, and a "tail" comprising C-type lectin-like domains 7-8. Conformational changes at pH 8 created a more elongated structure that cannot bind IgY. CryoEM reconstruction of FcRY dimers at pH 6 and small-angle X-ray scattering analysis at both pH values confirmed both structures. The cryoEM structure of the FcRY-IgY revealed symmetric binding of two FcRY heads to the dimeric FcY, each head contacting the C(H)4 domain of one FcY chain. FcRY shares structural properties with mannose receptor family members, including a head and tail domain organization, multimerization that may regulate ligand binding, and pH-dependent conformational changes. Our results facilitate understanding of immune recognition by the structurally related mannose receptor family and comparison of diverse methods of Ig transport across evolution.

Fig. 1.

Composition and characterization of FcRY. (A) Schematic model of FcRY showing individual domains: CysR (red sphere labeled C), FNII (yellow oval labeled F), CTLDs 1–8 (cyan ovals labeled 1–8), the transmembrane region, and the cytoplasmic tail. (B) Ribbon diagrams of structures related to FcRY domains (CysR, PDB ID code 1DQO; FNII, PDB ID code 2FN2; CTLD, PDB ID code 2CL8). Electron density calculated to 25 Å from the coordinates is superimposed upon each ribbon diagram. (C) SDS/PAGE analysis of the purified FcRY ectodomain used for cryoEM and SAXS studies.

Fig. 2.

FcRY structure at pH 6. Electron density of the FcRY monomer from a reconstruction at 23 Å resolution is shown at low (Top) and high (Middle) contour levels. The domain arrangement of the FcRY monomer (derived by fitting low-resolution filtered structures into the cryoEM density and applying knowledge of the results of biochemical experiments; Fig. S3) is shown with numbers and letters corresponding to the domains in the Middle row and schematically with low resolution calculated envelopes (gray) representing each of the domains in the Bottom row. The individual domains of FcRY are color coded as in Fig. 1.

Fig. 3.

FcRY dimer structure. (A) Electron density (Middle) and possible domain arrangements for the FcRY dimer (Left, a non–domain-swapped dimer; Right, a domain-swapped dimer) from a reconstruction at 28 Å resolution. The domain arrangement was derived as described in Fig. 2. (B) Three different views of the FcRY dimer. Electron density is shown above the corresponding schematic domain arrangement in each view. The CysR (red) and FnII (yellow) domains were not labeled to indicate that the same domain arrangement would apply to a domain-swapped vs. non–domain-swapped dimer.

Fig. 4.

FcRY–IgY structure. (A, Middle) Electron density of the FcRY–IgY complex from a reconstruction at 25 Å resolution. Low, medium, and high contour levels (cyan, yellow, and red, respectively) are shown with the approximate locations of different portions of the complex structure indicated by lines. (Left and Bottom Right) electron density for the pH 6 FcRY monomer structure shown on both sides of the complex in the orientation in which it would interact with the FcY portion of IgY. (Top Right) FcY coordinates (PDB code 2W59) enclosed in low-resolution (25 Å) filtered electron density. (B) Fitting of the FcRY–IgY complex density with density from the FcRY structure and the calculated FcY density from its crystal structure as shown in A. (C) FcRY–IgY electron density (medium contour level) fit with FcY coordinates and the domain model of the pH 6 FcRY structure (as shown in Fig. 2 with a low-resolution calculated envelope in gray).

Fig. 5.

Likely orientations of FcRY and FcRY–IgY on a membrane. Schematic structures for the FcRY monomer and dimer are shown as the full-length proteins would be oriented on a membrane bilayer. The two FcRY monomers on the Right are shown in an orientation that would allow formation of a 2:1 FcRY–IgY complex.