Integrins and Ion Channels: Molecular Complexes and Signaling

  • Integrins are cell surface receptors mediating cell-cell and cell-matrix adhesion, composed of alpha and beta subunits.
  • Mammals have 18 alpha and 8 beta subunits, forming 24 heterodimers.
  • Integrins regulate cell development, tissue remodeling, and organogenesis by mediating cell adhesion to the extracellular matrix (ECM).
  • Rapid communication occurs between cell adhesion receptors and channel proteins, forming multiprotein membrane complexes.
  • This communication may regulate ion fluxes and intracellular signaling pathways, sometimes mediated by cellular messengers like G proteins.
  • Interactions are reciprocal: ion channel stimulation often controls integrin activation or expression.
  • Studying integrin-ion channel interplay clarifies how the ECM regulates muscle excitability, synaptic plasticity, and lymphocyte activation.
  • Derangement of these processes has implications for tumor invasiveness and cardiovascular/neurologic diseases.
  • Chapter 1 provides background on integrin structure and signal transduction between cell adhesion machinery and ion channels.
  • Historical introduction
    • Highlights transmembrane ion fluxes through channels/transporters and their control of cell potential and excitability
    • Mentions the asymmetrical arrangement of ion transport proteins in epithelial layers affecting transepithelial transport, axis establishment in embryos and organogenesis
    • Notes that ion channel malfunction can lead to serious diseases
  • Integrin Structure
    • Integrins are transmembrane proteins formed by noncovalently associated alpha and beta subunits, each containing a multidomain extracellular portion, a single-membrane spanning helix, and a short cytoplasmic tail.
    • The exception is β4 subunit, which has a large cytoplasmic domain.
    • Integrins bind to specific ECM ligands like fibronectin, collagen, or laminin, or cell surface counter-receptors like ICAM-1 or VCAM-1.
    • ECM binding is regulated by conformational changes in integrin extracellular domains.
    • Cell adhesion recruits cytoskeletal, scaffolding, and signaling proteins to integrin cytoplasmic face, forming Focal Adhesions (FAs).
    • A bidirectional transmission of mechanical force and biochemical signals occurs across the plasma membrane through integrins.
    • "Outside-in signaling" transduces ECM-dependent signals into cellular responses like cell spreading, migration, proliferation, and death.
    • "Inside-out signaling" modulates integrin affinity for ECM and surface expression, controlling cell adhesion.
  • Integrin-Mediated Cell Adhesion
    • Detailed studies on integrin-mediated adhesion-affected cell signaling lagged behind neurotransmission or growth factor responses.
    • Challenges in physiological/biochemical experiments on adherent cells contributed to the delay.
    • By 1990, cell physiology and biomolecular methods advanced; patch-clamp methods and intracellular probes improved study opportunities.
    • Studies from the early nineties showed integrin-mediated cell adhesion stimulates K+ channel activity and controls cell pH and cytosolic free Ca2+.
  • Integrin structure: Extracellular Domains and ECM Ligand Binding
    • X-ray crystallography and NMR studies offered insight into the three-dimensional structure of the extracellular portion of integrin receptors.
    • Two integrin classes exist depending on whether or not their α subunit contains an extracellular von Willebrand factor type A domain (αA or αI).
    • MIDAS (metal ion-dependent adhesion site) is present in αA and modulates divalent cation-dependence of ECM protein binding.
    • αA assumes low affinity ('closed') or high affinity ('open') conformation.
    • Transition to 'open' state rearranges MIDAS for acidic residue binding from ECM ligand.
    • AlphaV contains containing 'α-propeller', 'Thigh', Calf-1 and Calf-2 domains. β subunit has PSI, EGF-like domains and tail domain (βTD).
    • Association of propeller and βA domains forms a 'head' for formation of the αβ complex.
    • Structure of ligand-bound domains typically studied using Arg-Gly-Asp (RGD) sequence-containing peptides. RGD inserts between the propeller and βA domains.
    • A 'switchblade' activation model proposes extension of bent extrastruture upon integrin activation, while a 'deadbolt' model proposes quaternary conformational rearrangements.
  • Transmembrane and Intracellular Domains
    • TM helices from and subunits show a conserved pattern of hydrophobicity.
    • Single-pass TM segments may traverse membranes as α-helices.
    • The TM helices of the α subunits probably lack significant helix tilt.
    • Cysteine-scanning mutagenesis experiments suggests TM domains are bonded when inactive and separate when active.
    • FRET studies support TM domain separation upon activation.
    • Conformational changes in cytoplasmic tails affect the ectodomain.
    • Movement and separation of TM segments leads to integrin activation.
  • Inside-Out Activation by Talin
    • Inside-out activation requires interaction between cytoplasmic ligands and integrin tails.
    • Talin consists of a ~50 kDa head region and a ~200 kDa rod segment. Head contains FERM domain. Rod contains α-helical bundles.
    • Talin binds both to integrin cytoplasmic domains and to vinculin and actin filaments, connecting cytoskeleton and ECM.
    • Cleavage between head and rod domains by protease calpain results in a 16-fold increase in binding to β3 integrin tails.
    • Integrin activation is mediated by the head domain. FERM contains a phosphotyrosine binding domain (PTB) that binds the more N-terminal of the two NPxY motifs found in the cytoplasmic domains of integrins.
  • Binding to Other Membrane Proteins and Cytoskeleton
    • Integrins are linked to the actin cytoskeleton.Exceptions 5/4, coupled to intermediate filaments.
    • Talin connects most integrin types to F-actin filaments.
    • Both and cytoplasmic tails bind to many other proteins, with mechanical and signaling effects.
    • Formation of FAs is dependent on these tails.
    • There are more than 70 intracellular proteins reported to bind integrin tails, with variable specificities.
  • Physiological and Pathological Implications:
    • Cell adhesion machinery transduces mechanical strain into modification of ion fluxes, like increase of free [Ca2+]i, often involving tyrosine kinase cascades.
    • Human T-cells form a membrane complex with voltage-gated K+ channels to activate integrin function.
    • Channel-mediated regulation of integrin expression was first evidenced in FLG 29.1 cells.
    • hERG1 stimulation during integrin-mediated cell adhesion stimulates surface expression of integrins.
    • Leukemic cell lines form membrane complexes containing hERG1, integrin subunits, and other proteins, correlated with neoplastic transformation.