Supervillin modulation of focal adhesions involving TRIP6/ZRP-1

Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA 01605, USA.
The Journal of Cell Biology (Impact Factor: 9.83). 08/2006; 174(3):447-58. DOI: 10.1083/jcb.200512051
Source: PubMed


Cell-substrate contacts, called focal adhesions (FAs), are dynamic in rapidly moving cells. We show that supervillin (SV)--a peripheral membrane protein that binds myosin II and F-actin in such cells--negatively regulates stress fibers, FAs, and cell-substrate adhesion. The major FA regulatory sequence within SV (SV342-571) binds to the LIM domains of two proteins in the zyxin family, thyroid receptor-interacting protein 6 (TRIP6) and lipoma-preferred partner (LPP), but not to zyxin itself. SV and TRIP6 colocalize within large FAs, where TRIP6 may help recruit SV. RNAi-mediated decreases in either protein increase cell adhesion to fibronectin. TRIP6 partially rescues SV effects on stress fibers and FAs, apparently by mislocating SV away from FAs. Thus, SV interactions with TRIP6 at FAs promote loss of FA structure and function. SV and TRIP6 binding partners suggest several specific mechanisms through which the SV-TRIP6 interaction may regulate FA maturation and/or disassembly.

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Available from: Elizabeth J Luna, Oct 07, 2015
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    • "TRIP6, another member of the zyxin family LIM domain proteins, (Yi and Beckerle, 1998; Lin and Lin, 2011) can also be targeted to focal adhesions. TRIP6 binding partners include supervillin and lysophosphatidic acid (LPA) 2 receptor (Xu et al., 2004; Takizawa et al., 2006), and has been suggested to facilitate focal adhesion turnover in epithelial cells and to mediate LPA-induced cell migration (Xu et al., 2004; Takizawa et al., 2006). Similar to zyxin, RNAi mediated knock-down of TRIP6-induced increased cell adhesion to FN in a fibroblast-like cell line COS7 (Takizawa et al., 2006), implicating that TRIP6 may not contribute to the increased VSMC adhesion. "
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    ABSTRACT: Zyxin is a focal adhesion protein that has been implicated in the modulation of cell adhesion and motility, and is hypothesized to be a mechano-sensor in integrin-mediated responses to mechanical force. To test the functional role of zyxin in the mechanotransduction of microvascular smooth muscle cells (VSMC), we utilized atomic force microscopy (AFM) to apply localized pulling forces to VSMC through a fibronectin (FN) focal adhesion induced by a FN-coated bead on cell surface. Application of force with the AFM induced an increase of zyxin accumulation at the site of the FN-bead focal adhesion that accompanied the VSMC contractile response. Whereas, reduction of zyxin expression by using a zyxin-shRNA construct abolished the VSMC contractile response to AFM pulling forces, even though the zyxin-silenced VSMCs displayed increased adhesion to FN in both AFM adhesion assays and cell adhesion assays. The reduced zyxin expression significantly impaired cell spreading and reorganization of the actin cytoskeleton that could indicate a possible underlying reason for the loss of a contractile response to mechanical force. Consistent with these observations, in zyxin-silenced VSMC, we also observed a reduced expression of Rac1, which plays an important role in the actin reorganization in VSMC, but increased thyroid receptor-interacting proteins (TRIP6) and FAK expression, the latter being a major protein that promote cell adhesion. In conclusion, these data support an important enabling role for zyxin in VSMCs ability to mechanically respond to applied force.
    Frontiers in Physiology 12/2012; 3:472. DOI:10.3389/fphys.2012.00472 · 3.53 Impact Factor
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    • "Nevertheless, the current level of resolution is consistent with a model in which myosin II and actin may induce localized folding as they interact with the supervillin N-terminus at or near the ligand-binding sites predicted by ANCHOR. As noted above, the focal adhesion-targeting sequence of supervillin (Takizawa et al., 2006) is also associated with a region containing predicted disordered ligand-binding sites. "
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    ABSTRACT: Supervillin, the largest member of the villin/gelsolin family, is a cytoskeleton regulating, peripheral membrane protein. Supervillin increases cell motility and promotes invasive activity in tumors. Major cytoskeletal interactors, including filamentous actin and myosin II, bind within the unique supervillin amino terminus, amino acids 1-830. The structural features of this key region of the supervillin polypeptide are unknown. Here, we utilize circular dichroism and bioinformatics sequence analysis to demonstrate that the N-terminal part of supervillin forms an extended intrinsically disordered region (IDR). Our combined data indicate that the N-terminus of human and bovine supervillin sequences (positions 1-830) represents an IDR, which is the largest IDR known to date in the villin/gelsolin family. Moreover, this result suggests a potentially novel mechanism of regulation of myosin II and F-actin via the intrinsically disordered N-terminal region of hub protein supervillin.
    Journal of biomolecular Structure & Dynamics 10/2012; 31(10). DOI:10.1080/07391102.2012.726531 · 2.92 Impact Factor
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    • "However, multiple contractions apparently occur in supervillin-knockdown cells before cleavage fails (Fig. 8), implying that myosin II is appropriately activated. Assuming that supervillin can bind simultaneously to myosin II and EPLIN, which also is required for normal cytokinesis (Chircop et al. 2009), supervillin could bridge myosin II/L-MLCK with septins at the plasma membrane through EPLIN (Fig. 6) or other membrane interactors (Nebl et al. 2002;Pestonjamasp et al. 1997;Takizawa et al. 2006). Such a supervillin-based scaffold could work in parallel with scaffolds associated with anillin, another actin-, myosin II-, and membrane-binding protein required for early cytokinesis (Piekny and Glotzer 2008;von Dassow 2009). "
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    ABSTRACT: Supervillin, the largest member of the villin/gelsolin/flightless family, is a peripheral membrane protein that regulates each step of cell motility, including cell spreading. Most known interactors bind within its amino (N)-terminus. We show here that the supervillin carboxy (C)-terminus can be modeled as supervillin-specific loops extending from gelsolin-like repeats plus a villin-like headpiece. We have identified 27 new candidate interactors from yeast two-hybrid screens. The interacting sequences from 12 of these proteins (BUB1, EPLIN/LIMA1, FLNA, HAX1, KIF14, KIFC3, MIF4GD/SLIP1, ODF2/Cenexin, RHAMM, STARD9/KIF16A, Tks5/SH3PXD2A, TNFAIP1) co-localize with and mis-localize EGFP-supervillin in mammalian cells, suggesting associations in vivo. Supervillin-interacting sequences within BUB1, FLNA, HAX1, and MIF4GD also mimic supervillin over-expression by inhibiting cell spreading. Most new interactors have known roles in supervillin-associated processes, e.g. cell motility, membrane trafficking, ERK signaling, and matrix invasion; three (KIF14, KIFC3, STARD9/KIF16A) have kinesin motor domains; and five (EPLIN, KIF14, BUB1, ODF2/cenexin, RHAMM) are important for cell division. GST fusions of the supervillin G2-G3 or G4-G6 repeats co-sediment KIF14 and EPLIN, respectively, consistent with a direct association. Supervillin depletion leads to increased numbers of bi- and multi-nucleated cells. Cytokinesis failure occurs predominately during early cytokinesis. Supervillin localizes with endogenous myosin II and EPLIN in the cleavage furrow, and overlaps with the oncogenic kinesin, KIF14, at the midbody. We conclude that supervillin, like its interactors, is important for efficient cytokinesis. Our results also suggest that supervillin and its interaction partners coordinate actin and microtubule motor functions throughout the cell cycle.
    Cytoskeleton 06/2010; 67(6):346-64. DOI:10.1002/cm.20449 · 3.12 Impact Factor
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