Structural biology of betaglycan and endoglin, membrane-bound co-receptors of the TGF-beta family

Abstract

Betaglycan and endoglin, membrane-bound co-receptors of the TGF-β family, are required to mediate the signaling of a select subset of TGF-β family ligands, TGF-β2 and InhA, and BMP-9 and BMP-10, respectively. Previous biochemical and biophysical methods suggested alternative modes of ligand binding might be responsible for these co-receptors to selectively recognize and potentiate the functions of their ligands, yet the molecular details were lacking. Recent progress determining structures of betaglycan and endoglin, both alone and as bound to their cognate ligands, is presented herein. The structures reveal relatively minor, but very significant structural differences that lead to entirely different modes of ligand binding. The different modes of binding nonetheless share certain commonalities, such as multivalency, which imparts the co-receptors with very high affinity for their cognate ligands, but at the same time provides a mechanism for release by stepwise binding of the signaling receptors, both of which are essential for their functions.

Impact statement: The TGF-β family is one of the most highly diversified signaling families, with essential roles in nearly all aspects of metazoan biology. Though functionally diverse, all 33 human TGF-β family ligands signal through a much more limited number of receptors. Thus the signaling repertoire is limited and cannot account for the functional diversity of signaling ligands in vivo. This mini review covers recent advances in our understanding of the structural basis by which two co-receptors of the family, betaglycan and endoglin, selectively recognize a limited subset of TGF-β family ligands and enable their functions in the cells and tissues in which they are expressed. The advances described also highlight gaps in current understanding of how the co-receptors are displaced upon engagement by the signaling receptors and how they function in a physiological environment, and thus suggest new avenues for investigation that will further illuminate how these essential co-receptors function in vivo.

Keywords: Structural biology; TGF-beta; betaglycan; cell signaling; co-receptor; endoglin; transforming growth factor-beta.

Figure 1.

TGF-β family proteins and their canonical Smad signaling mechanism. (a) Structure of a representative TGF-β family member, TGF-β1, with one of the monomers in red and the other in blue (PDB 1KLC). The terms used to describe the hand-like structure of one of the monomers are shown. (b) Structure of a representative type I:type II receptor heterotetratmeric complex; the extracellular component of the complex corresponds to TGF-β1 bound to the ectodomains of the type I and type II signaling receptors (TβRI and TβRII, shaded magenta and orange, respectively; PDB 3KFD). The type I and type II kinase domains shown correspond to those of the TGF-β type I and activin type IIB receptors (PDB 1IAS and 2QLU, respectively). The positioning of the kinase domains relative to one another are not experimentally determined. Two representative pro-complex structures, that of pro-TGF-β1 (c) and pro-activin A (d) (PDB 3RJR and 5HLZ). (A color version of this figure is available in the online journal.)

Figure 2.

Domain structure of betaglycan and endoglin. Domain structures of betaglycan (a) and endoglin (b). Domains responsible for directly contacting the ligands they bind are indicated. Vertical lines specify amino acid identity between the respective domains. (A color version of this figure is available in the online journal.)

Figure 3.

Mechanisms for betaglycan-potentiated type II receptor binding. (a) Proposed mechanism for betaglycan-potentiated antagonism of activin A by InhA. Proposed mechanisms for betaglycan-potentiated TGF-β2 receptor complex assembly, as initially proposed based on affinity labeling (b) or later proposed based on extensive direct and competition binding studies (c). (A color version of this figure is available in the online journal.)

Figure 4.

Mechanism for endoglin-potentiated type I receptor binding. Proposed mechanism for endoglin-potentiated BMP-9/-10 receptor complex assembly. (A color version of this figure is available in the online journal.)

Figure 5.

Structure of the endoglin and its complex with BMP-9. Structure (a) and schematic (b) of the endoglin orphan domain (PDB 5I04). The strand-bend-strand motif that exits each of the domains and extends into the other domain is shaded in lavender. (c) Structure of the 2:1 complex formed between the endoglin orphan domain and BMP-9 dimer. Pairing of the orphan domain 1 edge β-strand, β6, with the exposed finger 4 (F4) β-strand of the ligand is depicted in the inset (PDB 5HZW). (d) Structure of the covalent endoglin ZP domain homodimer. Structure shown was modeled based on the experimentally determined endoglin ZP monomer structure (PDB 5HZW) and by positioning the free cysteine residues responsible for covalent dimer formation (Cys⁵¹⁶ and Cys⁵⁸², approximate locations of which are shown in red and magenta spheres, respectively) within a distance compatible with disulfide bond formation. (e) Proposed model for the complex between full-length endoglin and BMP-9. Model was constructed from the experimentally determined structure of the 2:1 endoglin orphan domain:BMP-9 dimer complex (PDB 5HZW) and the model for the endoglin ZP dimer shown in panel D. (A color version of this figure is available in the online journal.)

Figure 6.

Binding site on TGF-β2 for the betaglycan ZP-C domain. (a) Structure of TGF-β2 with methyl-bearing residues in deuterated Ile, Leu, Val C-methyl protonated TGF-β2 identified by NMR that shifted either significantly (red) or not (blue) upon titration with unlabeled betaglycan ZP-C⁶⁵ (PDB 2TGI). (b) Structure of TGF-β2 with residues which led to a significant disruption (red) or not (blue) of betaglycan ZP-C binding upon substitution with alanine (as assessed by surface plasmon resonance with immobilized TGF-β2 single amino acid variants) (PDB 2TGI). (c) Structures of rat (left) or mouse (right) betaglycan ZP-C, with the proposed binding sites in the A-B or F-G loops highlighted in magenta and orange, respectively^, (PDB 3QW9 and 4AJV). (d) Alignment of residues from the finger region of all TGF-β family growth factors in humans; positions highlighted in color were either shown to shift upon titration of deuterated methyl-protonated TGF-β2 with unlabeled ZP-C or to be affected in their binding affinity for ZP-C upon substitution. Hydrophobic residues are colored green, acidic residues are colored red, basic residues are colored blue, and neutral residues are colored purple. Boxed residues highlight those that are either entirely (Lys⁹⁷) or mostly unique (Val⁹²/Ile⁹² and Glu⁹⁹) to TGF-βs and Inh α. Figure is adapted and reproduced with permission from Henen et al. (A color version of this figure is available in the online journal.)

Figure 7.

Structure of the betaglycan orphan domain and comparison with the endoglin orphan domain. (a) Side-by-side comparison of the overall structures of the zebrafish betaglyan (left) and human endoglin (right) orphan domains, which aside from a significantly different orientation of the two β-sandwich domains, have similar overall structures (PDB 6MZP and 5I04). (b) Superposition of domains 1 (top) and 2 (bottom) of zebrafish betaglycan (orange strands/blue helices) and human endoglin (green strands and light blue helices) orphan domains. Superposition highlights additional helix-strand motif present in the zebrafish betaglycan orphan OD-1, but not the human endoglin OD-1 (PDB 6MZP and 5I04, respectively). (c) Model of the interface between zebrafish betaglycan orphan domain and TGF-β2. Model was built assuming the same manner of super β-sheet formation as that for the human endoglin orphan domain bound to BMP-9. Model highlights steric clashes (red ovals) between the additional β-strand inserted in the betaglycan orphan domain and the β-strand that forms finger 4 of the ligand. (d) Model for the 1:2:1 TGF-β2:TβRII:betaglycan orphan domain complex constructed using the program pyDockSAXS, fitted into the electron density calculated from the SAXS scattering curve of the 1:2:1 TGF-β2:TβRII:betaglycan orphan domain complex used to guide the docking. In the model shown, the TGF-β2 monomers are depicted in orange and blue, TβRII in green, and the betaglycan orphan domain in magenta. Figure is adapted and reproduced with permission from Figures 4 and 7 of Kim et al. (A color version of this figure is available in the online journal.)