Talin controls integrin activation

2004 ◽  
Vol 32 (3) ◽  
pp. 434-437 ◽  
Author(s):  
D.A. Calderwood
Keyword(s):  
Conformational Changes ◽  
Critical Role ◽  
Dynamic Control ◽  
Common Element ◽  
Actin Binding ◽  
Cytoplasmic Domains ◽  
Integrin Activation ◽  
Intracellular Signals ◽  
Integrin Binding Site ◽  
Activation Pathways

Tight, dynamic control of the affinity of integrin adhesion receptors for their extracellular ligands (integrin activation) is essential for the development and functioning of multicellular organisms. Integrin activation is controlled by intracellular signals that, through their action on integrin cytoplasmic domains, induce conformational changes in integrin extracellular domains, resulting in increased affinity for the ligand. Recent results indicate that the binding of talin, a major actin-binding protein, to integrin β tails represents a final common step in integrin activation pathways. The major integrin-binding site lies within the talin FERM (four-point-one, ezrin, radixin, moesin) domain, and binding occurs via a variant of the classical PTB domain (phosphotyrosine-binding domain)–NPxY interaction. Formation of this talin–integrin complex plays a critical role in integrin activation, since mutations, in either talin or integrin β tails, which disrupt complex formation, inhibit integrin activation. Furthermore, use of RNA interference to knockdown talin expression selectively reveals that talin is essential for integrin activation in response to physiological agonists. Thus the association of the cytoskeletal protein talin with integrin β cytoplasmic domains is a critical step during integrin activation, and regulation of this step may be a final common element in the signalling pathways that control integrin activation.

New insights into the structural basis of integrin activation

Blood ◽  
2003 ◽  
Vol 102 (4) ◽  
pp. 1155-1159 ◽  
Author(s):  
Jian-Ping Xiong ◽  
Thilo Stehle ◽  
Simon L. Goodman ◽  
M. Amin Arnaout
Keyword(s):  
Conformational Changes ◽  
Structural Changes ◽  
Structural Basis ◽  
Integrin Activation ◽  
Cellular Functions ◽  
Electron Microscopic ◽  
The Past ◽  
Intracellular Signals ◽  
Bidirectional Signaling ◽  
New Hypothesis

Abstract Integrins are cell adhesion receptors that communicate biochemical and mechanical signals in a bidirectional manner across the plasma membrane and thus influence most cellular functions. Intracellular signals switch integrins into a ligand-competent state as a result of elicited conformational changes in the integrin ectodomain. Binding of extracellular ligands induces, in turn, structural changes that convey distinct signals to the cell interior. The structural basis of this bidirectional signaling has been the focus of intensive study for the past 3 decades. In this perspective, we develop a new hypothesis for integrin activation based on recent crystallographic, electron microscopic, and biochemical studies.

Abstract 496: Competition Between Filamin and Kindlin-2 Regulates Integrin Activation

2012 ◽  
Vol 32 (suppl_1) ◽  
Author(s):  
Mitali Das ◽  
Sujay Ithychanda ◽  
Kamila Bledzka ◽  
Jun Qin ◽  
Edward F Plow
Keyword(s):  
Cell Migration ◽  
K562 Cells ◽  
Cell Types ◽  
Suppressive Effect ◽  
Integrin Activation ◽  
Intracellular Signals ◽  
Integrin Binding Site ◽  
Multiple Cell ◽  
Β3 Integrin ◽  
Inside Out

Cell migration and adhesion during hemostasis, angiogenesis and inflammation are dynamically regulated by integrin heterodimeric adhesion receptors. Their interactions with cytosolic proteins, filamin (FLN), talin (TLN) and Kindlin (Kn2) enable them to convey intracellular signals (inside-out-signaling) to the external environment by engaging extracellular matrix ligands. While TLN and Kn2 activate integrins, FLN inhibits cell migration. TLN and Kn2 bind to membrane-proximal and -distal NPxY motifs of β integrin cytoplasmic tails (CTs), respectively, and an integrin binding site for FLN resides in between these two sequences. Competition between TLN and FLN regulates integrin activation, but it is unknown if FLN and Kn2 compete and regulate integrin inside-out signaling. This competition was tested using αIIbβ3 (platelet-specific) and β7 (lymphocyte-specific; strong FLN binder) integrins in multiple cell types. siRNA depletion of FLNA in K562 cells stably expressing αIIbβ3 integrin (K562-αIIbβ3) significantly enhanced PAC-1 (specific for activated αIIbβ3) binding compared to control siRNA, demonstrating its effect on β3 activation. In pulldown assays using GST-β3 CT, Kn2 bound β3 in CHO lysates transfected with Kn2, either alone or with FLN repeat 21; however, FLN binding to β3 CT was observed only when FLN repeat 21 was expressed alone. Under similar conditions using GST-β7 CT, FLN-β7 interaction was not perturbed by Kn2. This was more pronounced in endothelial cell lysates where GST-β7 CT bound endogenous FLNA but not Kn2. Weak talin-β7 CT binding in this assay was noted. Moreover, in K562-αIIbβ3 cells, exogenous Kn2 overcame the suppressive effect of FLN on αIIbβ3 activation. Overall, our data shows that FLN inhibits β3 integrin function, and competition between FLN and Kn2 can indeed regulate integrin activation.

scholarly journals Biased localization of actin binding proteins by actin filament conformation

2020 ◽  
Author(s):  
Andrew R Harris ◽  
Pamela Jreij ◽  
Brian Belardi ◽  
Andreas Bausch ◽  
Daniel A Fletcher
Keyword(s):  
Actin Filament ◽  
Single Molecule ◽  
Conformational Changes ◽  
Actin Filaments ◽  
Binding Proteins ◽  
Critical Role ◽  
Actin Binding ◽  
Cell Physiology ◽  
Physical Constraints

ABSTRACTThe assembly of actin filaments into distinct cytoskeletal structures plays a critical role in cell physiology, but how proteins localize differentially to these structures within a shared cytoplasm remains unclear. Here, we show that the actin-binding domains of accessory proteins can be sensitive to filament conformational changes. Using a combination of live cell imaging and in vitro single molecule binding measurements, we show that tandem calponin homology domains (CH1-CH2) can be mutated to preferentially bind actin networks at the front or rear of motile cells, and we demonstrate that the affinity of CH1-CH2 domain mutants varies as actin filament conformation is altered by perturbations that include stabilizing drugs, physical constraints, and other binding proteins. These findings suggest that conformational heterogeneity of actin filaments in cells could help to direct accessory binding proteins to different actin cytoskeletal structures through a biophysical feedback loop.

scholarly journals Biased localization of actin binding proteins by actin filament conformation

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Andrew R. Harris ◽  
Pamela Jreij ◽  
Brian Belardi ◽  
Aaron M. Joffe ◽  
Andreas R. Bausch ◽  
...  
Keyword(s):  
Actin Filament ◽  
Single Molecule ◽  
Conformational Changes ◽  
Actin Filaments ◽  
Binding Proteins ◽  
Critical Role ◽  
Actin Binding ◽  
Cell Physiology ◽  
Binding Domains

AbstractThe assembly of actin filaments into distinct cytoskeletal structures plays a critical role in cell physiology, but how proteins localize differentially to these structures within a shared cytoplasm remains unclear. Here, we show that the actin-binding domains of accessory proteins can be sensitive to filament conformational changes. Using a combination of live cell imaging and in vitro single molecule binding measurements, we show that tandem calponin homology domains (CH1–CH2) can be mutated to preferentially bind actin networks at the front or rear of motile cells. We demonstrate that the binding kinetics of CH1–CH2 domain mutants varies as actin filament conformation is altered by perturbations that include stabilizing drugs and other binding proteins. These findings suggest that conformational changes of actin filaments in cells could help to direct accessory binding proteins to different actin cytoskeletal structures through a biophysical feedback loop.

Integrin connections to the cytoskeleton through talin and vinculin

2008 ◽  
Vol 36 (2) ◽  
pp. 235-239 ◽  
Author(s):  
Wolfgang H. Ziegler ◽  
Alex R. Gingras ◽  
David R. Critchley ◽  
Jonas Emsley
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Critical Role ◽  
Cytoskeletal Proteins ◽  
Actin Binding ◽  
Helical Bundles ◽  
Helix Bundle ◽  
Integrin Binding Site ◽  
Β Chain ◽  
Actin Binding Site

Integrins are αβ heterodimeric receptors that mediate attachment of cells to the extracellular matrix and therefore play important roles in cell adhesion, migration, proliferation and survival. Among the cytoskeletal proteins that interact directly with the β-chain cytoplasmic domain, talin has emerged as playing a critical role in integrin activation and linkage to the actin cytoskeleton. Talin (2541 amino acids) is an elongated (60 nm) flexible antiparallel dimer, with a small globular head connected to an extended rod. The talin head contains a FERM (4.1/ezrin/radixin/moesin) domain (residues 86–400) with binding sites for several β integrin cytodomains and the talin rod contains a second lower-affinity integrin-binding site, a highly conserved C-terminal actin-binding site and also several binding sites for vinculin. We have determined previously the crystal structures of two domains from the talin rod, spanning residues 482–789. Talin-(482–655), which contains a VBS (vinculin-binding site), folds into a five-helix bundle whereas talin-(656–789) is a four-helix bundle. We have also reported the crystal structure of the N-terminal vinculin head domain in complex with an activated form of talin. In the present paper, we consider how binding sites buried within the folded helical bundles of talin and α-actinin form interactions with vinculin.

scholarly journals Integrin activation

2008 ◽  
Vol 36 (2) ◽  
pp. 229-234 ◽  
Author(s):  
Asoka Banno ◽  
Mark H. Ginsberg
Keyword(s):  
Integrin Receptors ◽  
Integrin Activation ◽  
Matrix Assembly ◽  
Hairpin Rna ◽  
Protein Kinase Cα ◽  
Integrin Binding Site ◽  
Cell Adhesion And Migration ◽  
And Migration ◽  
Activation Pathways ◽  
Rap1 Activation

Agonist stimulation of integrin receptors, composed of transmembrane α and β subunits, leads cells to regulate integrin affinity (‘activation’), a process that controls cell adhesion and migration, and extracellular matrix assembly. A final step in integrin activation is the binding of talin to integrin β cytoplasmic domains. We used forward, reverse and synthetic genetics to engineer and order integrin activation pathways of a prototypic integrin, platelet αIIbβ3. PMA activated αIIbβ3 only after expression of both PKCα (protein kinase Cα) and talin at levels approximating those in platelets. Inhibition of Rap1 GTPase reduced αIIbβ3 activation, whereas expression of constitutively active Rap1A(G12V) bypassed the requirement for PKCα. Overexpression of a Rap effector, RIAM (Rap1-GTP-interacting adaptor molecule), activated αIIbβ3 and bypassed the requirement for PKCα and Rap1. In addition, shRNA (short hairpin RNA)-mediated knockdown of RIAM blocked talin interaction with and activation of integrin αIIbβ3. Rap1 activation caused the formation of an ‘activation complex’ containing talin and RIAM that redistributed to the plasma membrane and activated αIIbβ3. The central finding was that this Rap1-induced formation of an ‘integrin activation complex’ leads to the unmasking of the integrin-binding site on talin, resulting in integrin activation.

Crystal structures of Magnaporthe oryzae trehalose-6-phosphate synthase (MoTps1) suggest a model for catalytic process of Tps1

2019 ◽  
Vol 476 (21) ◽  
pp. 3227-3240 ◽  
Author(s):  
Shanshan Wang ◽  
Yanxiang Zhao ◽  
Long Yi ◽  
Minghe Shen ◽  
Chao Wang ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Conformational Changes ◽  
Magnaporthe Oryzae ◽  
Structural Information ◽  
Complex Structure ◽  
Critical Role ◽  
Catalytic Process ◽  
Structural Basis ◽  
Salt Bridges ◽  
Terminal Domain

Trehalose-6-phosphate (T6P) synthase (Tps1) catalyzes the formation of T6P from UDP-glucose (UDPG) (or GDPG, etc.) and glucose-6-phosphate (G6P), and structural basis of this process has not been well studied. MoTps1 (Magnaporthe oryzae Tps1) plays a critical role in carbon and nitrogen metabolism, but its structural information is unknown. Here we present the crystal structures of MoTps1 apo, binary (with UDPG) and ternary (with UDPG/G6P or UDP/T6P) complexes. MoTps1 consists of two modified Rossmann-fold domains and a catalytic center in-between. Unlike Escherichia coli OtsA (EcOtsA, the Tps1 of E. coli), MoTps1 exists as a mixture of monomer, dimer, and oligomer in solution. Inter-chain salt bridges, which are not fully conserved in EcOtsA, play primary roles in MoTps1 oligomerization. Binding of UDPG by MoTps1 C-terminal domain modifies the substrate pocket of MoTps1. In the MoTps1 ternary complex structure, UDP and T6P, the products of UDPG and G6P, are detected, and substantial conformational rearrangements of N-terminal domain, including structural reshuffling (β3–β4 loop to α0 helix) and movement of a ‘shift region' towards the catalytic centre, are observed. These conformational changes render MoTps1 to a ‘closed' state compared with its ‘open' state in apo or UDPG complex structures. By solving the EcOtsA apo structure, we confirmed that similar ligand binding induced conformational changes also exist in EcOtsA, although no structural reshuffling involved. Based on our research and previous studies, we present a model for the catalytic process of Tps1. Our research provides novel information on MoTps1, Tps1 family, and structure-based antifungal drug design.

scholarly journals HIV-1 Envelope Conformation, Allostery, and Dynamics

Viruses ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 852
Author(s):  
Ashley Lauren Bennett ◽  
Rory Henderson
Keyword(s):  
Host Cell ◽  
Conformational Changes ◽  
Neutralizing Antibodies ◽  
Critical Role ◽  
Cell Receptor ◽  
Broadly Neutralizing Antibodies ◽  
Structural Mapping ◽  
Host Cell Receptor ◽  
Immunogen Design ◽  
Hiv 1

The HIV-1 envelope glycoprotein (Env) mediates host cell fusion and is the primary target for HIV-1 vaccine design. The Env undergoes a series of functionally important conformational rearrangements upon engagement of its host cell receptor, CD4. As the sole target for broadly neutralizing antibodies, our understanding of these transitions plays a critical role in vaccine immunogen design. Here, we review available experimental data interrogating the HIV-1 Env conformation and detail computational efforts aimed at delineating the series of conformational changes connecting these rearrangements. These studies have provided a structural mapping of prefusion closed, open, and transition intermediate structures, the allosteric elements controlling rearrangements, and state-to-state transition dynamics. The combination of these investigations and innovations in molecular modeling set the stage for advanced studies examining rearrangements at greater spatial and temporal resolution.

scholarly journals Dynamic changes in the osteoclast cytoskeleton in response to growth factors and cell attachment are controlled by β3 integrin

2003 ◽  
Vol 162 (3) ◽  
pp. 499-509 ◽  
Author(s):  
Roberta Faccio ◽  
Deborah V. Novack ◽  
Alberta Zallone ◽  
F. Patrick Ross ◽  
Steven L. Teitelbaum
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Cell Attachment ◽  
Cytoplasmic Tail ◽  
Cytoplasmic Domain ◽  
Small Gtpases ◽  
Cytoskeletal Reorganization ◽  
Integrin Activation ◽  
Intracellular Signals ◽  
Β3 Integrin

The β3 integrin cytoplasmic domain, and specifically S752, is critical for integrin localization and osteoclast (OC) function. Because growth factors such as macrophage colony–stimulating factor and hepatocyte growth factor affect integrin activation and function via inside-out signaling, a process requiring the β integrin cytoplasmic tail, we examined the effect of these growth factors on OC precursors. To this end, we retrovirally expressed various β3 integrins with cytoplasmic tail mutations in β3-deficient OC precursors. We find that S752 in the β3 cytoplasmic tail is required for growth factor–induced integrin activation, cytoskeletal reorganization, and membrane protrusion, thereby affecting OC adhesion, migration, and bone resorption. The small GTPases Rho and Rac mediate cytoskeletal reorganization, and activation of each is defective in OC precursors lacking a functional β3 subunit. Activation of the upstream mediators c-Src and c-Cbl is also dependent on β3. Interestingly, although the FAK-related kinase Pyk2 interacts with c-Src and c-Cbl, its activation is not disrupted in the absence of functional β3. Instead, its activation is dependent upon intracellular calcium, and on the β2 integrin. Thus, the β3 cytoplasmic domain is responsible for activation of specific intracellular signals leading to cytoskeletal reorganization critical for OC function.

scholarly journals Inhibition of acanthamoeba actomyosin-II ATPase activity and mechanochemical function by specific monoclonal antibodies.

1984 ◽  
Vol 99 (3) ◽  
pp. 1024-1033 ◽  
Author(s):  
D P Kiehart ◽  
T D Pollard
Keyword(s):  
Monoclonal Antibodies ◽  
Atpase Activity ◽  
Conformational Changes ◽  
Binding Sites ◽  
Polyclonal Antibodies ◽  
Myosin Ii ◽  
Antigenic Site ◽  
Actin Binding ◽  
Filamentous Form ◽  
Antibody Effects

Monoclonal and polyclonal antibodies that bind to myosin-II were tested for their ability to inhibit myosin ATPase activity, actomyosin ATPase activity, and contraction of cytoplasmic extracts. Numerous antibodies specifically inhibit the actin activated Mg++-ATPase activity of myosin-II in a dose-dependent fashion, but none blocked the ATPase activity of myosin alone. Control antibodies that do not bind to myosin-II and several specific antibodies that do bind have no effect on the actomyosin-II ATPase activity. In most cases, the saturation of a single antigenic site on the myosin-II heavy chain is sufficient for maximal inhibition of function. Numerous monoclonal antibodies also block the contraction of gelled extracts of Acanthamoeba cytoplasm. No polyclonal antibodies tested inhibited ATPase activity or gel contraction. As expected, most antibodies that block actin-activated ATPase activity also block gel contraction. Exceptions were three antibodies M2.2, -15, and -17, that appear to uncouple the ATPase activity from gel contraction: they block gel contraction without influencing ATPase activity. The mechanisms of inhibition of myosin function depends on the location of the antibody-binding sites. Those inhibitory antibodies that bind to the myosin-II heads presumably block actin binding or essential conformational changes in the myosin heads. A subset of the antibodies that bind to the proximal end of the myosin-II tail inhibit actomyosin-II ATPase activity and gel contraction. Although this part of the molecule is presumably some distance from the ATP and actin-binding sites, these antibody effects suggest that structural domains in this region are directly involved with or coupled to catalysis and energy transduction. A subset of the antibodies that bind to the tip of the myosin-II tail appear to inhibit ATPase activity and contraction through their inhibition of filament formation. They provide strong evidence for a substantial enhancement of the ATPase activity of myosin molecules in filamentous form and suggest that the myosin filaments may be required for cell motility.

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