Migfilin, a Molecular Switch in Regulation of Integrin
Activation |
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Authors: | Sujay Subbayya Ithychanda Mitali Das Yan-Qing Ma Keyang Ding Xiaoxia Wang Sudhiranjan Gupta Chuanyue Wu Edward F Plow and Jun Qin |
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Institution: | ‡Department of Molecular Cardiology, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio 44195 and the §Department of Pathology, University of Pittsburgh, Pittsburgh, Pennsylvania 15261 |
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Abstract: | The linkage of heterodimeric (α/β) integrin receptors with their
extracellular matrix ligands and intracellular actin cytoskeleton is a
fundamental step for controlling cell adhesion and migration. Binding of the
actin-linking protein, talin, to integrin β cytoplasmic tails (CTs)
induces high affinity ligand binding (integrin activation), whereas binding of
another actin-linking protein, filamin, to the integrin β CTs negatively
regulates this process by blocking the talin-integrin interaction. Here we
show structurally that migfilin, a novel cytoskeletal adaptor highly enriched
in the integrin adhesion sites, strongly interacts with the same region in
filamin where integrin β CTs bind. We further demonstrate that the
migfilin interaction dissociates filamin from integrin and promotes the
talin/integrin binding and integrin activation. Migfilin thus acts as a
molecular switch to disconnect filamin from integrin for regulating integrin
activation and dynamics of extracellular matrix-actin linkage.Cells reside in a protein network, the extracellular matrix
(ECM).4 Cell-ECM
contact is crucial for many physiological and pathophysiological processes and
is primarily mediated by heterodimeric (α/β) transmembrane
receptors, the integrins (1).
Integrins engage a variety of ECM proteins via their extracellular domains
while connecting to the actin cytoskeleton via their small cytoplasmic tails
(CTs). The ability of integrins to bind to their ligands is uniquely
controlled by the integrin CTs via a process called “inside-out
signaling,” i.e. upon cellular stimulation, an integrin,
typically expressed in a latent state, can receive intracellular signal(s) at
its CT, which transmits through the transmembrane domain to the extracellular
domain thereby converting the receptor from a low to a high affinity state
(integrin activation). How such long range information transfer is initiated
and regulated has been the central topic of integrin/cell adhesion research
over the decades (for reviews see Refs.
2-5).
Structural/biochemical studies have indicated that the inside-out signaling
involves the unclasping of the integrin α/β CT complex
(6-9),
followed by extensive rearrangement of transmembrane domain and extracellular
domain
(10-13).
Talin, a large actin-linking protein, was found to play a key role in the
unclasping process by binding to the integrin β CTs
(7-8,
14). Talin activity appears to
be controlled by multiple factors or pathways
(15-20).Relevant to this study is the role of filamin, another major actin
cross-linking protein
(21-22),
in integrin activation. Filamin was found to share an overlapping binding site
on integrin β CTs with talin and thus suppress the talin-integrin
interaction (16). Gene
silencing of filamin in various cell lines to remove the filamin-integrin
connection enhances integrin activation
(16,
23), whereas increased
filamin-integrin interaction inhibits cell migration
(24), a process critically
dependent on integrin activation. Together these observations support the
notion that filamin binding to integrin serves as a cellular brake to control
the dynamics of the integrin activation by inhibiting talin function and
ECM-cytoskeleton communication. The mechanism as to how the filamin brake is
turned off to promote integrin activation and cell migration is not
understood.Filamin is known to contain an N-terminal actin binding domain (ABD) and a
long rod-like domain of 24 immunoglobulin-like repeats, of which repeat 21
(IgFLN21) was shown to play a key role in binding to integrin β CTs and
blocking the talin-integrin β CT interaction
(16). Interestingly, IgFLN21
also recognizes another intracellular protein called migfilin, which has been
shown to be an important regulator of integrin-mediated cytoskeletal
rearrangements, cell shape change
(25), and cell migration
(26).In an effort to dissect the complex intermolecular interactions between
migfilin, filamin, and integrin, we have undertaken a detailed
structural/functional analysis. Using NMR spectroscopy, we have mapped the
precise IgFLN21 binding region in migfilin, which is located at the extreme N
terminus (residues 1-24) of migfilin (migfilin-N), and we solved solution
structure of the IgFLN21-migfilin-N complex. To our surprise, despite little
sequence homology, migfilin binds to the same region in IgFLN21 where integrin
β CT binds. Detailed NMR and biochemical analyses demonstrate that the
migfilin-filamin interaction is an order of magnitude higher than the
integrin-filamin interaction and that the migfilin binding to filamin can
competitively dissociate filamin from integrin and thus promote the
talin-integrin interaction. Using multiple functional approaches, we further
show that migfilin, but not its filamin binding defective mutant,
significantly enhances integrin activation. These data suggest a novel
regulatory pathway in which the binding of filamin to its downstream target
migfilin switches off the integrin-filamin connection, thereby promoting talin
binding to and activation of integrins. |
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Keywords: | |
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