Biochemistry; Cardiovascular Diseases; Cell Adhesion; Cell Biology; Cytoskeleton; Pharmacology; Integrins; Transcellular Cell Migration
Integrins, transmembrane adhesion receptors, mediate cell adhesion and permit bidirectional transmission of mechanical force and biochemical signals across the plasma membrane. Integrin-dependent cellular activities such as adhesion, migration, proliferation, and survival rely upon the dynamic interaction of integrin cytoplasmic tails with intracellular integrin tail-binding proteins.
We use cell-biological, biochemical, and structural techniques to identify and characterize the interactions of integrin cytoplasmic tails with intracellular ligands, and to decipher how these interactions are regulated. This has allowed us to establish talin as a key regulator of integrin activation; to show that integrin binding to the actin crosslinking protein filamin controls cell migration and modulates integrin-talin interactions; to reveal that kindlins modulate talin-mediated integrin activation in an integrin-specific fashion; and to characterize interactions of the integrin-linked kinase. Ongoing studies
aim to extend these observations, to characterize the molecular basis and functional significance of new interactions between integrin cytoplasmic tails and cytoskeletal and signaling proteins, and to identify novel mechanisms by which specific integrin-associated proteins are regulated.
Specialized Terms: Integrin; Cell adhesion; Cell migration; Cytoskeleton; Structural biology
Extensive Research Description
Integrins are essential heterodimeric adhesion receptors formed by the non-covalent association of a and ß subunits. Each subunit is a type I transmembrane glycoprotein that has relatively large extracellular domains and a generally short cytoplasmic tail. Humans contain 18 a and 8 ß subunits that combine to produce at least 24 different heterodimers, each of which can bind to a specific repertoire of cell surface, ECM or soluble protein ligands. Cell-cell and cell-substratum adhesion is mediated by the binding of integrin extracellular domains to diverse protein ligands, however, cellular control of these adhesive interactions and their translation into dynamic cellular responses, such as cell spreading or migration, requires the integrin cytoplasmic tails. These short tails bind to intracellular ligands that connect the receptors to signaling pathways and cytoskeletal networks. Hence, by binding both extracellular and intracellular ligands, integrins provide a transmembrane link for the bidirectional transmission of mechanical force and biochemical signals across the plasma membrane.
We have found that talin binds to integrin ß subunit cytoplasmic tails through the FERM domain within the talin head. Integrin binding occurs via a variant of the classical PTB domain-NPxY interaction and, in addition to linking integrins to actin stress fibers, this interaction induces conformational changes in the integrin ectodomains that regulate integrin ligand-binding affinity (integrin activation). Tight regulation of integrin activation is essential because it controls cell adhesion, migration, and assembly of an extracellular matrix. Hence integrin activation is a critical step in angiogenesis, tumor cell metastasis, embryonic development, cardiac function and the immune response, and cellular control of integrin activation plays important roles in health and disease throughout development and during the course of adult life.
More recently we have investigated the binding of another class of FERM domain
proteins, the kindlins. Kindlins, like talins, contain an atypical FERM domain and we predict them to be structurally closely related to the integrin-activating talin head domain. We find that kindlins bind integrin ß tails and regulate integrin activation and signaling. The molecular and structural basis for kindlins effects on integrins are the subject of ongoing work.
We have also found that actin cross-linking proteins of the filamin family (filamin A, B and C) bind integrin ß tails, that tight association of filamin with integrin ß tails inhibits cell migration and that filamins can inhibit integrin activation. The regulated binding of filamin to integrin ß tails may therefore provide a control point for regulation of cell migration. Our structural analyses of filamin-integrin interactions revealed the basis for integrin binding and identified regulatory mechanisms including, competition with talin (which provides a mechanism by which filamin binding can suppress integrin activation), auto-inhibition by adjacent filamin domains (which may be released by mechanical stretching) and competition with other filamin-binding proteins. Recently we have shown that filamins are important for initiation of cell migration and that during cell differentiation filamin levels can be controlled by poly-ubiquitinylation and proteasomal degradation. Investigations of the molecular bases for these effects are underway.
Finally we have initiated structural and functional studies on the integrin-linked kinase (ILK); a signaling protein implicated in both integrin activation, possibly through interactions with kindlins, and in signaling downstream of integrins. In collaboration with the Boggon Lab (Pharmacology, Yale) we have solved the structures of a complex between the ILK ankyrin-repeat domain and the 1st LIM domain of the ILK-binding protein PINCH1 or PINCH2. These structures revealed the molecular basis of ILK-PINCH interactions, which are essential for targeting of ILK-PINCH-parvin heterotrimeric complexes to adhesion sites where they act as key signaling nodes. Ongoing structural and functional studies focus on other ILK domains as well as larger multi-domain complexes.
Full List of PubMed Publications
- Huet-Calderwood C, Rivera-Molina F, Iwamoto DV, Kromann EB, Toomre D, Calderwood DA: Novel ecto-tagged integrins reveal their trafficking in live cells. Nat Commun. 2017 Sep 18; 2017 Sep 18. PMID: 28924207
- Draheim KM, Huet-Calderwood C, Simon B, Calderwood DA: Nuclear Localization of Integrin Cytoplasmic Domain-associated Protein-1 (ICAP1) Influences β1 Integrin Activation and Recruits Krev/Interaction Trapped-1 (KRIT1) to the Nucleus. J Biol Chem. 2017 Feb 3; 2016 Dec 21. PMID: 28003363
- Shi X, Mihaylova VT, Kuruvilla L, Chen F, Viviano S, Baldassarre M, Sperandio D, Martinez R, Yue P, Bates JG, Breckenridge DG, Schlessinger J, Turk BE, Calderwood DA: Loss of TRIM33 causes resistance to BET bromodomain inhibitors through MYC- and TGF-β-dependent mechanisms. Proc Natl Acad Sci U S A. 2016 Aug 2; 2016 Jul 18. PMID: 27432991
- Morse EM, Sun X, Olberding JR, Ha BH, Boggon TJ, Calderwood DA: PAK6 targets to cell-cell adhesions through its N-terminus in a Cdc42-dependent manner to drive epithelial colony escape. J Cell Sci. 2016 Jan 15; 2015 Nov 23. PMID: 26598554
- Calderwood DA: The Rap1-RIAM pathway prefers β2 integrins. Blood. 2015 Dec 17. PMID: 26679542
- Iwamoto DV, Calderwood DA: Regulation of integrin-mediated adhesions. Curr Opin Cell Biol. 2015 Oct; 2015 Jul 17. PMID: 26189062
- Draheim KM, Li X, Zhang R, Fisher OS, Villari G, Boggon TJ, Calderwood DA: CCM2-CCM3 interaction stabilizes their protein expression and permits endothelial network formation. J Cell Biol. 2015 Mar 30. PMID: 25825518
- Simpson MA, Bradley WD, Harburger D, Parsons M, Calderwood DA, Koleske AJ: Direct interactions with the integrin β1 cytoplasmic tail activate the Abl2/Arg kinase. J Biol Chem. 2015 Mar 27; 2015 Feb 18. PMID: 25694433
- Bancroft T, Bouaouina M, Roberts S, Lee M, Calderwood DA, Schwartz M, Simons M, Sessa WC, Kyriakides TR: Up-regulation of thrombospondin-2 in Akt1-null mice contributes to compromised tissue repair due to abnormalities in fibroblast function. J Biol Chem. 2015 Jan 2; 2014 Nov 11. PMID: 25389299
- Ellis SJ, Lostchuck E, Goult BT, Bouaouina M, Fairchild MJ, López-Ceballos P, Calderwood DA, Tanentzapf G: The talin head domain reinforces integrin-mediated adhesion by promoting adhesion complex stability and clustering. PLoS Genet. 2014 Nov; 2014 Nov 13. PMID: 25393120
- Huet-Calderwood C, Brahme NN, Kumar N, Stiegler AL, Raghavan S, Boggon TJ, Calderwood DA: Differences in binding to the ILK complex determines kindlin isoform adhesion localization and integrin activation. J Cell Sci. 2014 Oct 1; 2014 Aug 1. PMID: 25086068
- Uchil PD, Pawliczek T, Reynolds TD, Ding S, Hinz A, Munro JB, Huang F, Floyd RW, Yang H, Hamilton WL, Bewersdorf J, Xiong Y, Calderwood DA, Mothes W: TRIM15 is a focal adhesion protein that regulates focal adhesion disassembly. J Cell Sci. 2014 Sep 15; 2014 Jul 11. PMID: 25015296
- Lee MY, Skoura A, Park EJ, Landskroner-Eiger S, Jozsef L, Luciano AK, Murata T, Pasula S, Dong Y, Bouaouina M, Calderwood DA, Ferguson SM, De Camilli P, Sessa WC: Dynamin 2 regulation of integrin endocytosis, but not VEGF signaling, is crucial for developmental angiogenesis. Development. 2014 Apr; 2014 Mar 5. PMID: 24598168
- Draheim KM, Fisher OS, Boggon TJ, Calderwood DA: Cerebral cavernous malformation proteins at a glance. J Cell Sci. 2014 Feb 15; 2014 Jan 30. PMID: 24481819
- Morse EM, Brahme NN, Calderwood DA: Integrin cytoplasmic tail interactions. Biochemistry. 2014 Feb 11; 2014 Jan 27. PMID: 24467163
- Gao J, Ha BH, Lou HJ, Morse EM, Zhang R, Calderwood DA, Turk BE, Boggon TJ: Substrate and inhibitor specificity of the type II p21-activated kinase, PAK6. PLoS One. 2013; 2013 Oct 28. PMID: 24204982
- Calderwood DA, Campbell ID, Critchley DR: Talins and kindlins: partners in integrin-mediated adhesion. Nat Rev Mol Cell Biol. 2013 Aug; 2013 Jul 17. PMID: 23860236
- Liu W, Draheim KM, Zhang R, Calderwood DA, Boggon TJ: Mechanism for KRIT1 release of ICAP1-mediated suppression of integrin activation. Mol Cell. 2013 Feb 21; 2013 Jan 11. PMID: 23317506
- Stiegler AL, Grant TD, Luft JR, Calderwood DA, Snell EH, Boggon TJ: Purification and SAXS analysis of the integrin linked kinase, PINCH, parvin (IPP) heterotrimeric complex. PLoS One. 2013; 2013 Jan 31. PMID: 23383235
- Bouaouina M, Jani K, Long JY, Czerniecki S, Morse EM, Ellis SJ, Tanentzapf G, Schöck F, Calderwood DA: Zasp regulates integrin activation. J Cell Sci. 2012 Dec 1; 2012 Sep 19. PMID: 22992465
- Stiegler AL, Draheim KM, Li X, Chayen NE, Calderwood DA, Boggon TJ: Structural basis for paxillin binding and focal adhesion targeting of β-parvin. J Biol Chem. 2012 Sep 21; 2012 Aug 6. PMID: 22869380
- Brahme NN, Calderwood DA: Cell adhesion: a FERM grasp of the tail sorts out integrins. Curr Biol. 2012 Sep 11. PMID: 22974999
- Baldassarre M, Razinia Z, Brahme NN, Buccione R, Calderwood DA: Filamin A controls matrix metalloproteinase activity and regulates cell invasion in human fibrosarcoma cells. J Cell Sci. 2012 Aug 15; 2012 May 17. PMID: 22595522
- Li X, Zhang R, Draheim KM, Liu W, Calderwood DA, Boggon TJ: Structural basis for small G protein effector interaction of Ras-related protein 1 (Rap1) and adaptor protein Krev interaction trapped 1 (KRIT1). J Biol Chem. 2012 Jun 22; 2012 May 10. PMID: 22577140
- Bouaouina M, Goult BT, Huet-Calderwood C, Bate N, Brahme NN, Barsukov IL, Critchley DR, Calderwood DA: A conserved lipid-binding loop in the kindlin FERM F1 domain is required for kindlin-mediated αIIbβ3 integrin coactivation. J Biol Chem. 2012 Mar 2; 2012 Jan 10. PMID: 22235127
- Razinia Z, Mäkelä T, Ylänne J, Calderwood DA: Filamins in mechanosensing and signaling. Annu Rev Biophys. 2012; 2012 Feb 23. PMID: 22404683
- Bouaouina M, Harburger DS, Calderwood DA: Talin and signaling through integrins. Methods Mol Biol. 2012. PMID: 21909921
- Razinia Z, Baldassarre M, Bouaouina M, Lamsoul I, Lutz PG, Calderwood DA: The E3 ubiquitin ligase specificity subunit ASB2α targets filamins for proteasomal degradation by interacting with the filamin actin-binding domain. J Cell Sci. 2011 Aug 1; 2011 Jul 12. PMID: 21750192
- Bouaouina M, Calderwood DA: Kindlins. Curr Biol. 2011 Feb 8. PMID: 21300280
- Chiswell BP, Stiegler AL, Razinia Z, Nalibotski E, Boggon TJ, Calderwood DA: Structural basis of competition between PINCH1 and PINCH2 for binding to the ankyrin repeat domain of integrin-linked kinase. J Struct Biol. 2010 Apr; 2009 Dec 4. PMID: 19963065
- Baldassarre M, Razinia Z, Burande CF, Lamsoul I, Lutz PG, Calderwood DA: Filamins regulate cell spreading and initiation of cell migration. PLoS One. 2009 Nov 13; 2009 Nov 13. PMID: 19915675
- Harburger DS, Bouaouina M, Calderwood DA: Kindlin-1 and -2 directly bind the C-terminal region of beta integrin cytoplasmic tails and exert integrin-specific activation effects. J Biol Chem. 2009 Apr 24; 2009 Feb 23. PMID: 19240021
- Harburger DS, Calderwood DA: Integrin signalling at a glance. J Cell Sci. 2009 Jan 15. PMID: 19118207
- Chiswell BP, Zhang R, Murphy JW, Boggon TJ, Calderwood DA: The structural basis of integrin-linked kinase-PINCH interactions. Proc Natl Acad Sci U S A. 2008 Dec 30; 2008 Dec 12. PMID: 19074270
- Lad Y, Jiang P, Ruskamo S, Harburger DS, Ylänne J, Campbell ID, Calderwood DA: Structural basis of the migfilin-filamin interaction and competition with integrin beta tails. J Biol Chem. 2008 Dec 12; 2008 Sep 30. PMID: 18829455
- Bouaouina M, Lad Y, Calderwood DA: The N-terminal domains of talin cooperate with the phosphotyrosine binding-like domain to activate beta1 and beta3 integrins. J Biol Chem. 2008 Mar 7; 2007 Dec 28. PMID: 18165225
- Lad Y, Kiema T, Jiang P, Pentikäinen OT, Coles CH, Campbell ID, Calderwood DA, Ylänne J: Structure of three tandem filamin domains reveals auto-inhibition of ligand binding. EMBO J. 2007 Sep 5; 2007 Aug 9. PMID: 17690686
- Lad Y, Harburger DS, Calderwood DA: Integrin cytoskeletal interactions. Methods Enzymol. 2007. PMID: 17697880
- Calderwood DA, Tai V, Di Paolo G, De Camilli P, Ginsberg MH: Competition for talin results in trans-dominant inhibition of integrin activation. J Biol Chem. 2004 Jul 9; 2004 May 13. PMID: 15143061
- Calderwood DA: Integrin activation. J Cell Sci. 2004 Feb 15. PMID: 14754902