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Cell News 1/2015
Research news
is not altered in ILK-deficient cells (Vouret-Craviari et al., 2004),
the defect in matrix assembly most likely results from the im-
pairment in force generation and the failure to form FBs.
Interestingly, it was recently shown that the recruitment of most
proteins to adhesion sites is dependent on myosin II activity
(Schiller and Fässler, 2013). However, ILK recruitment to FA sites
occurs independent of myosin II activity (Schiller and Fässler,
2013), suggesting that ILK might act upstream and be involved
in myosin II-dependent recruitment of other FA components.
Our finding that ILK is required for the generation of traction
forces provides further functional evidence for this notion (Fig.
2B). This inability to generate force is very likely to affect further
recruitment of FB-associated proteins, leading to the observed
failure in ECM remodeling in ILK-deficient fibroblasts (Fig. 2D).
Besides driving
α5β1
integrin segregation along the actin cyto-
skeleton, ILK could additionally be involved in mediating force-
dependent conformational changes in
α5β1
integrins, which
occur during fibrillogenesis (Clark et al., 2005).
Although
α5β1
integrin is the primary FN receptor (Huveneers
et al., 2008), integrin
αvβ3
(Wennerberg et al., 1996),
α4β1
(Sechler et al., 2000) and
αIIbβ3
(Olorundare et al., 2001) have
been shown to be involved in FN fibrillogenesis in vitro. Knock-
out studies in mice suggest overlapping as well as independent
functions for
α5
- and
αv
-class integrins in this process and only
the double knockout of
α5
- and
αv
- integrins in mice results
in loss of fibrillogenesis (Yang et al., 1999). It is tempting to
speculate that ILK, through its ability to bind both to
β1
- and
β3
-integrins, might be involved in their differential engagement
and thereby in fine-tuning the forces required for FN fibrillo-
genesis.
It has been proposed that the assembly state of FN fibers plays
an important role in regulating cell behavior by acting as a
checkpoint signal for subsequent ECM remodeling (Schwarz-
bauer and DeSimone, 2011). For instance, only the precise ratio
of FN fibril assembly ensures epithelial branching morphoge-
nesis during cleft formation (Sakai et al., 2003a). Furthermore,
FN fibrillogenesis regulates fibrillin-1 microfibril assembly (Kin-
sey et al., 2008) and could thereby impact the ability of these
Figure 2: Integrin-linked kinase is required for focal adhesion maturation, cellular force generation and extracellular matrix deposition
A. Immunofluorescence analysis of paxillin as a marker for focal adhesions and F-actin in fibroblasts. Wild type fibroblasts display small focal complexes
(arrow), focal adhesions (open arrowhead), and fibrillar adhesions (arrowhead), which are tightly connected to the actin cytoskeleton. Note lack of focal
complexes and fibrillar adhesions, reduced amount of actin stress fibers, and accumulation of large peripheral focal adhesions (open arrowhead) in ILK-
deficient cells. Scale bar 25 μm. B. Left panel shows a heat-scale map of traction stress magnitudes obtained using traction force microscopy. The color
code indicates local traction in kPa. Cell outlines are indicated by dotted lines. Right panel shows the quantification of total cellular traction forces (mean
± SEM, n>30, **p=0.0011). Deletion of ILK severely compromises the ability of fibroblasts to generate traction forces. C. Collagen gel contraction assay
with ILK f/f and -/- fibroblasts. Deletion of ILK impairs the ability of fibroblasts to contract collagen gels. D. Immunofluorescence staining of the fibronec-
tin matrix and the actin cytoskeleton (phalloidin) to visualize cell area. Note decreased matrix deposition in ILK -/- cells. Scale bar 100 μm (modified from
Radovanac et al., 2013).
