INTEGRINS: A BEDSIDE TO BENCH TO BEDSIDE STORY

Abstract

I provide a narrative of the path I took to discover the membrane receptors that mediate leukocyte adhesion, now known as β2 integrins or CD11/CD18. We followed this discovery with the first determination of the 3-D structures of integrins. The latter advance provided the foundation for understanding the unique features of integrins as divalent cation-dependent signaling receptors and as mechanosensitive conduits between the extracellular matrix and the intracellular cytoskeleton. Our structural studies are now opening new paths for taming overactive integrins in disease while minimizing the collateral damage associated with the faulty pharmacodynamics of current integrin inhibitory drugs.

Fig. 1.

Mammalian integrins. This protein family now consists of 24 α/β heterodimeric receptors assembled from 18 α-subunits and eight β-subunits. Nine α-subunits (shaded) contain an extra von Willebrand factor (vWF) type A domain (αA).

Fig. 2.

Structural comparisons of liganded and unliganded αA domain from the integrin CD11b subunit. (a, b) The MIDAS motif in the liganded (a) and unliganded (b) states. The metal ion at MIDAS is coordinated by residues from three surface loops, with a carboxyl oxygen from ligand (E, ball, and stick) binding the metal ion monodentately, completing the metal octahedral coordinating sphere. αA-containing integrins only require a key glutamate (or aspartate) in ligand to bind. This endows αA-containing integrins with unique access to heavier ligands like bundled collagen fibers and immunoglobulin domains. In the unliganded state, an oxygen atom from a water molecule replaces the ligand oxygen, and D242 from the third surface loop moves in to coordinate the metal directly. Coordinating oxygen atoms are in red, and hydrogen bonds are shown by dashed red lines. Direct bonds to the metal ion are shown as blue sticks. Water molecules are labeled ω1–ω3. (c) Ribbon diagrams showing the superposed structures of unliganded (gray) and liganded (light blue) αA domain of CD11b. Ligand glutamate (E, ball, and stick) induces major conformational changes in strand A-loop-α1 helix and strand-loop-α7 helix (indicated by arrows). A, F, strands A and F. Spheres, represent the respective metal ions at MIDAS.

Fig. 3.

Structure of αvβ3 ectodomain and its ligand binding site. (a) Left panel, ribbon diagram of the crystal structure showing the bent αvβ3 ectodomain, shown here, and in subsequent figures, in blue (αv) and red (β3). Right panel, a model of the genuextended αvβ3 ectodomain. Calf1 and βTD would extend into the plasma membrane and the cytoplasm in the native integrin. The 12 subdomains are labeled. The metal ions are not shown. (b) Binding of arginine-glycine-aspartate (RGD)-containing ligand (shown in ball and stick) to αvβ3. The peptide aspartate (D) completes the metal ion coordination sphere at MIDAS (encircled) as in glutamate (E)-ligated αA domain (Figure 2a), and the ligand arginine inserts into a pocket in the propeller stabilized mainly by salt bridges. The three metal ions in liganded β3 A-domain at MIDAS, ADMIDAS, and LIMBS (or SyMBS) are shown in cyan, magenta, and gray, respectively. αv and β3 residues are labeled cornflower blue and pink, respectively. A portion of the respective α1 helix containing the activation sensitive Y¹²² (in sphere) and the F-strand-loop-α7 helix are shown. Oxygen and nitrogen atoms are in red and blue, respectively. Hydrogen bonds and salt bridges are represented with dotted lines.

Fig. 4.

Ligand-induced movements in the αA-lacking αvβ3. Left and right panels show ribbon diagrams of the unliganded and liganded states, respectively. The four metal ions at the base of the propeller are not shown. The binding of the ligand Arg-Gly-Asp (RGD) (carbon, oxygen, and nitrogen atoms in green, red, and blue spheres) induces inward movement of the strand A-loop-α1 helix toward MIDAS, reported by Y¹²². This movement is mechanically coupled to the downward displacement of the c-terminal α7 helix (in gold) and a swingout of the underlying hybrid domain (arrows). The latter movement triggers additional quaternary changes that travel down across the legs, TM, and cytoplasmic regions, allowing the establishment of new links of the integrin with the actin cytoskeleton that promote cell adhesion.

Fig. 5.

The ligand relay model. Left and right panels show ribbon diagrams of the respective unliganded and liganded states of the ligand-binding region of an αA-containing integrin. The metal ions at the base of the propeller are not shown. The downward movement (arrow) of the c-terminal α7 helix (in gold) of the αA domain triggered by the binding of glutamate (E)-containing ligand (atoms shown in spheres) allows the invariant glutamate (E) at the bottom of the α7 helix (in gold) to reach and ligate the βA MIDAS ion (cyan), thus relaying the ligand occupancy state of αA to βA. The activation-sensitive tyrosine (Y) of the α1-helix, and the metal ions at LIMBS (gray) and ADMIDAS (magenta) are shown. The conformational changes (arrows) triggered in ligated βA are the same as shown in Figure 4.

Fig. 6.

The ligand-induced conformational changes in a β3 integrin ectodomain. Ribbon diagrams of the bent (left) and genuextended (right) αvβ3 ectodomain modeled relative to a lipid bilayer are shown. The four metal ions at the base of the propeller and at the genu of αv are shown as pink spheres. Agonists acting via G-protein-coupled receptors or other receptors prime the bent integrin to engage ligand (e.g., cyclic RGD in green). This hair-triggers genuextension, hybrid swingout, and proadhesive outside-in signaling, the duration of which is limited by agonist engagement. With partial agonist drugs, the extended state is stabilized in circulating cells, allowing the binding of natural antibodies to neoepitopes causing immune thrombocytopenia, or leaving the integrin in the ligand-competent state after drug dissociation, allowing circulating fibrinogen to bind platelets, causing paradoxical thrombosis.