In JoVE (1)

Other Publications (10)

Articles by Angélique Millon-Frémillon in JoVE

Other articles by Angélique Millon-Frémillon on PubMed

Unraveling ICAP-1 Function: Toward a New Direction?

European Journal of Cell Biology. Apr, 2006  |  Pubmed ID: 16546571

Cell adhesion to either the extracellular matrix (ECM) or to neighboring cells is of critical importance during both physiological and pathological situations. Integrins are a large family of cell adhesion receptors composed of two non-covalently linked alpha and beta subunits. They have a well-identified dual function of mediating both firm adhesion and signaling. The short cytoplasmic domain of integrin can interact with cytoplasmic proteins that are either shared by several different integrins or specific for one type of integrin. Integrin cytoplasmic domain-associated protein-1 (ICAP-1) is a small cytoplasmic protein that specifically interacts with the beta1 integrin subunit. In this review we will discuss recent findings on ICAP-1, not only at the structural and functional level, but also its possible interconnection in other signaling pathways such as those that control cell proliferation.

Cell Adaptive Response to Extracellular Matrix Density is Controlled by ICAP-1-dependent Beta1-integrin Affinity

The Journal of Cell Biology. Jan, 2008  |  Pubmed ID: 18227284

Cell migration is an integrated process requiring the continuous coordinated assembly and disassembly of adhesion structures. How cells orchestrate adhesion turnover is only partially understood. We provide evidence for a novel mechanistic insight into focal adhesion (FA) dynamics by demonstrating that integrin cytoplasmic domain-associated protein 1 (ICAP-1) slows down FA assembly. Live cell imaging, which was performed in both Icap-1-deficient mouse embryonic fibroblasts and cells expressing active beta(1) integrin, shows that the integrin high affinity state favored by talin is antagonistically controlled by ICAP-1. This affinity switch results in modulation in the speed of FA assembly and, consequently, of cell spreading and migration. Unexpectedly, the ICAP-1-dependent decrease in integrin affinity allows cell sensing of matrix surface density, suggesting that integrin conformational changes are important in mechanotransduction. Our results clarify the function of ICAP-1 in cell adhesion and highlight the central role it plays in the cell's integrated response to the extracellular microenvironment.

Podosome-type Adhesions and Focal Adhesions, So Alike Yet So Different

European Journal of Cell Biology. Sep, 2008  |  Pubmed ID: 18417250

Cell-matrix adhesions are essential for cell migration, tissue organization and differentiation, therefore playing central roles in embryonic development, remodeling and homeostasis of tissues and organs. Matrix adhesion-dependent signals cooperate with other pathways to regulate biological functions such as cell survival, cell proliferation, wound healing, and tumorigenesis. Cell migration and invasion are integrated processes requiring the continuous, coordinated assembly and disassembly of integrin-mediated adhesions. An understanding of how integrins regulate cell migration and invasiveness through the dynamic regulation of adhesions is fundamental to both physiological and pathological situations. A variety of cell-matrix adhesions has been identified, namely, focal complexes, focal adhesions, fibrillar adhesions, podosomes, and invadopodia (podosome-type adhesions). These adhesion sites contain integrin clusters able to develop specialized structures, which are different in their architecture and dynamics although they share almost the same proteins. Here we compare recent advances and developments in the elucidation of the organization and dynamics of focal adhesions and podosome-type adhesions, in order to understand how such subcellular sites - though closely related in their composition - can be structurally and functionally different. The underlying question is how their respective physiological or pathological roles are related to their distinct organization.

Specificities of β1 Integrin Signaling in the Control of Cell Adhesion and Adhesive Strength

European Journal of Cell Biology. Feb-Mar, 2011  |  Pubmed ID: 20971526

Cells exert actomyosin contractility and cytoskeleton-dependent force in response to matrix stiffness cues. Cells dynamically adapt to force by modifying their behavior and remodeling their microenvironment. This adaptation is favored by integrin activation switch and their ability to modulate their clustering and the assembly of an intracellular hub in response to force. Indeed integrins are mechanoreceptors and mediate mechanotransduction by transferring forces to specific adhesion proteins into focal adhesions which are sensitive to tension and activate intracellular signals. α(5)β(1) integrin is considered of major importance for the formation of an elaborate meshwork of fibronectin fibrils and for the extracellular matrix deposition and remodeling. Here we summarize recent progress in the study of mechanisms regulating the activation cycle of β(1) integrin and the specificity of α(5)β(1) integrin in mechanotransduction.

Osteoblast Mineralization Requires Beta1 Integrin/ICAP-1-dependent Fibronectin Deposition

The Journal of Cell Biology. Jul, 2011  |  Pubmed ID: 21768292

The morphogenetic and differentiation events required for bone formation are orchestrated by diffusible and insoluble factors that are localized within the extracellular matrix. In mice, the deletion of ICAP-1, a modulator of β1 integrin activation, leads to severe defects in osteoblast proliferation, differentiation, and mineralization and to a delay in bone formation. Deposition of fibronectin and maturation of fibrillar adhesions, adhesive structures that accompany fibronectin deposition, are impaired upon ICAP-1 loss, as are type I collagen deposition and mineralization. Expression of β1 integrin with a mutated binding site for ICAP-1 recapitulates the ICAP-1-null phenotype. Follow-up experiments demonstrated that ICAP-1 negatively regulates kindlin-2 recruitment onto the β1 integrin cytoplasmic domain, whereas an excess of kindlin-2 binding has a deleterious effect on fibrillar adhesion formation. These results suggest that ICAP-1 works in concert with kindlin-2 to control the dynamics of β1 integrin-containing fibrillar adhesions and, thereby, regulates fibronectin deposition and osteoblast mineralization.

Calcium and Calmodulin-dependent Serine/threonine Protein Kinase Type II (CaMKII)-mediated Intramolecular Opening of Integrin Cytoplasmic Domain-associated Protein-1 (ICAP-1α) Negatively Regulates β1 Integrins

The Journal of Biological Chemistry. Jul, 2013  |  Pubmed ID: 23720740

Focal adhesion turnover during cell migration is an integrated cyclic process requiring tight regulation of integrin function. Interaction of integrin with its ligand depends on its activation state, which is regulated by the direct recruitment of proteins onto the β integrin chain cytoplasmic domain. We previously reported that ICAP-1α, a specific cytoplasmic partner of β1A integrins, limits both talin and kindlin interaction with β1 integrin, thereby restraining focal adhesion assembly. Here we provide evidence that the calcium and calmodulin-dependent serine/threonine protein kinase type II (CaMKII) is an important regulator of ICAP-1α for controlling focal adhesion dynamics. CaMKII directly phosphorylates ICAP-1α and disrupts an intramolecular interaction between the N- and the C-terminal domains of ICAP-1α, unmasking the PTB domain, thereby permitting ICAP-1α binding onto the β1 integrin tail. ICAP-1α direct interaction with the β1 integrin tail and the modulation of β1 integrin affinity state are required for down-regulating focal adhesion assembly. Our results point to a molecular mechanism for the phosphorylation-dependent control of ICAP-1α function by CaMKII, allowing the dynamic control of β1 integrin activation and cell adhesion.

Microtubule-dependent Modulation of Adhesion Complex Composition

PloS One. 2014  |  Pubmed ID: 25526367

The microtubule network regulates the turnover of integrin-containing adhesion complexes to stimulate cell migration. Disruption of the microtubule network results in an enlargement of adhesion complex size due to increased RhoA-stimulated actomyosin contractility, and inhibition of adhesion complex turnover; however, the microtubule-dependent changes in adhesion complex composition have not been studied in a global, unbiased manner. Here we used label-free quantitative mass spectrometry-based proteomics to determine adhesion complex changes that occur upon microtubule disruption with nocodazole. Nocodazole-treated cells displayed an increased abundance of the majority of known adhesion complex components, but no change in the levels of the fibronectin-binding α5β1 integrin. Immunofluorescence analyses confirmed these findings, but revealed a change in localisation of adhesion complex components. Specifically, in untreated cells, α5-integrin co-localised with vinculin at peripherally located focal adhesions and with tensin at centrally located fibrillar adhesions. In nocodazole-treated cells, however, α5-integrin was found in both peripherally located and centrally located adhesion complexes that contained both vinculin and tensin, suggesting a switch in the maturation state of adhesion complexes to favour focal adhesions. Moreover, the switch to focal adhesions was confirmed to be force-dependent as inhibition of cell contractility with the Rho-associated protein kinase inhibitor, Y-27632, prevented the nocodazole-induced conversion. These results highlight a complex interplay between the microtubule cytoskeleton, adhesion complex maturation state and intracellular contractile force, and provide a resource for future adhesion signaling studies. The proteomics data have been deposited in the ProteomeXchange with identifier PXD001183.

Isolation of Integrin-based Adhesion Complexes

Current Protocols in Cell Biology. Mar, 2015  |  Pubmed ID: 25727331

The integration of cells with their extracellular environment is facilitated by cell surface adhesion receptors, such as integrins, which play important roles in both normal development and the onset of pathologies. Engagement of integrins with their ligands in the extracellular matrix, or counter-receptors on other cells, initiates the intracellular assembly of a wide variety of proteins into adhesion complexes such as focal contacts, focal adhesions, and fibrillar adhesions. The proteins recruited to these complexes mediate bidirectional signaling across the plasma membrane, and, as such, help to coordinate and/or modulate the multitude of physical and chemical signals to which the cell is subjected. The protocols in this unit describe two approaches for the isolation or enrichment of proteins contained within integrin-associated adhesion complexes, together with their local plasma membrane/cytosolic environments, from cells in culture. In the first protocol, integrin-associated adhesion structures are affinity isolated using microbeads coated with extracellular ligands or antibodies. The second protocol describes the isolation of ventral membrane preparations that are enriched for adhesion complex structures. The protocols permit the determination of adhesion complex components via subsequent downstream analysis by western blotting or mass spectrometry.

Definition of a Consensus Integrin Adhesome and Its Dynamics During Adhesion Complex Assembly and Disassembly

Nature Cell Biology. Dec, 2015  |  Pubmed ID: 26479319

Integrin receptor activation initiates the formation of integrin adhesion complexes (IACs) at the cell membrane that transduce adhesion-dependent signals to control a multitude of cellular functions. Proteomic analyses of isolated IACs have revealed an unanticipated molecular complexity; however, a global view of the consensus composition and dynamics of IACs is lacking. Here, we have integrated several IAC proteomes and generated a 2,412-protein integrin adhesome. Analysis of this data set reveals the functional diversity of proteins in IACs and establishes a consensus adhesome of 60 proteins. The consensus adhesome is likely to represent a core cell adhesion machinery, centred around four axes comprising ILK-PINCH-kindlin, FAK-paxillin, talin-vinculin and α-actinin-zyxin-VASP, and includes underappreciated IAC components such as Rsu-1 and caldesmon. Proteomic quantification of IAC assembly and disassembly detailed the compositional dynamics of the core cell adhesion machinery. The definition of this consensus view of integrin adhesome components provides a resource for the research community.

Proteomic Analysis of Integrin-associated Complexes from Mesenchymal Stem Cells

Proteomics. Clinical Applications. Jan, 2016  |  Pubmed ID: 26147903

Multipotent mesenchymal stem cells (MSCs) have the capability to differentiate down adipocyte, osteocyte and chondrocyte lineages and as such offer a range of potential therapeutic applications. The composition and stiffness of the extracellular matrix (ECM) environment that surrounds cells dictates their transcriptional programme, thereby affecting stem cell lineage decision-making. Cells sense force via linkages between themselves and their microenvironment, and this is transmitted by integrin receptors and associated adhesion signalling complexes. To identify regulators of MSC force sensing, we sought to catalogue MSC integrin-associated adhesion complex composition.

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