Copyright°c 2002 IUBMB 1521-6543/02 $12.00 + .00 DOI: 10.1080/10399710290039007
Review Article
The Focal Adhesion Kinase—A Regulator of Cell Migration and Invasion
Christof R. Hauck,
1Datsun A. Hsia,
2and David D. Schlaepfer
21Research Center for Infectious Diseases, R¨ontgenring 11, 97070 W¨urzburg, Germany
2Department of Immunology, The Scripps Research Institute, La Jolla, California 92037
Summary
Cell migration plays an important role in embryonic devel- opment, wound healing, immune responses, and in pathological phenomena such as tissue invasion and metastasis formation. In this review, we summarize recent reports that connect the focal adhesion kinase (FAK) to cell migration and invasion. FAK is a nonreceptor protein tyrosine kinase involved in signal transduc- tion from integrin-enriched focal adhesion sites that mediate cell contact with the extracellular matrix. Multiple protein-protein in- teraction sites allow FAK to associate with adapter and structural proteins allowing for the modulation of mitogen-activated protein (MAP) kinases, stress-activated protein (SAP) kinases, and small GTPase activity. FAK-enhanced signals have been shown to me- diate the survival of anchorage-dependent cells and are critical for efficient cell migration in response to growth factor receptor and integrin stimulation. Elevated expression of FAK in human tumors has been correlated with increased malignancy and inva- siveness. Because recent findings show that FAK contributes to the secretion of matrix-metalloproteinases, FAK may represent an im- portant checkpoint in coordinating the dynamic processes of cell motility and extracellular matrix remodeling during tumor cell invasion.
IUBMB
Life
, 53: 115–119, 2002 Keywords Cell migration; focal adhesion kinase; integrins; invasion;matrix metalloproteinases; Src-family kinases.
Structural Characteristics of FAK-like Protein Tyrosine Kinases
Focal adhesion kinase (FAK)1 together with Pyk2 (1) form a subfamily of FAK-like protein-tyrosine kinases (PTKs). FAK
Received 12 December 2001; accepted 4 January 2002.
Address correspondence to David D. Schlaepfer, The Scripps Research Institute, Department of Immunology, IMM26, 10550 N.
Torrey Pines Rd., La Jolla, CA 92037. Fax: (858) 784-8227. E-mail:
dschlaep@scripps.edu
1Abbreviations: ERK, extracellular signal regulated kinase; FAK, focal ad- hesion kinase; FERM, band 4.1, ezrin, radixin, moesin; GAP, GTPase-activating
has been detected in man, mouse, chicken, frog, and fruitfly with a strong conservation of domain structure (Fig. 1). The N-terminal region harbors a FERM (band 4.1, ezrin, radixin, and moesin) homology domain (2) allowing FAK association with other PTKs and with the actin cytoskeletal-associated pro- tein ezrin (3–5). Based on the moesin FERM domain crys- tal structure, the FERM domain is subdivided into three lobes containing binding modules that either in combination or indi- vidually may enable FAK to recognize other proteins (6). The C-terminal region of FAK encompasses two proline-rich regions (amino acids 712–723 and 867–882) that serve as Src-homology 3 (SH3) binding sites for the adaptor molecule p130Cas (7 ), the Rho GTPase-activating protein (GAP) GRAF, or the Arf GAP ASAP1 (8, 9). The C-terminal FAK region also contains the fo- cal adhesion targeting or F.A.T. domain that spans binding sites for talin and paxillin (reviewed in 10, 11).
Regulation of FAK Activation
FAK activity and tyrosine phosphorylation are upregulated in response to cell-matrix contact, a number of soluble cellular acti- vators, as well as mechanical stimuli (reviewed in 12). However, we know surprisingly little about the molecular mechanisms of FAK activation. Nevertheless, a number of studies have shown that FAK activation and autophosphorylation at tyrosine-397 (Y-397) results in the Src homology 2 (SH2)-dependent recruit- ment and binding of Src-family PTKs. The signaling complex formed between FAK and c-Src leads to Src-mediated phos- phorylation of FAK at multiple sites in the kinase and C-terminal domain (12). Src-mediated phosphorylation of FAK at Tyr-925 creates a SH2 binding site for the Grb2 adapter protein and has been connected to the activation of the extracellular signal reg- ulated (ERK)/mitogen-activated protein (MAP) kinase pathway
protein; GEF, guanine nucleotide exchange factor; JNK, c-jun N-terminal kinase;
MAP kinase, mitogen-activated protein kinase; MMP, matrix metalloproteinase;
PTK, protein tyrosine kinase; SAP kinase, stress-activated protein kinase; SH, Src homology.
115 Konstanzer Online-Publikations-System (KOPS)
URL: http://www.ub.uni-konstanz.de/kops/volltexte/2007/4135/
URN: http://nbn-resolving.de/urn:nbn:de:bsz:352-opus-41359
Figure 1. Model of FAK structural features. FAK contains a central kinase domain flanked by N- and C-terminal sequences harboring a FERM domain, proline-rich SH3 binding motifs, and the focal adhesion targeting (FAT) domain. Proteins interact- ing with the different domains are indicated. The major tyrosine phosphorylation sites (Y-397, Y-407, Y-861, and Y-925) and a lysine residue critical for kinase activity (K-454) are marked.
FRNK, the FAK-related non-kinase, is expressed as an inde- pendent transcript encompassing the FAK C-terminal domain.
A point mutation in the FAT domain (Leu-1034 to Ser) im- pairs paxillin binding and abolishes focal adhesion targeting of FRNK.
(12). FAK also is highly serine phosphorylated in the C-terminal domain during the cell cycle and serine phosphorylation near the FAK proline-rich motifs may promote the dissociation of SH3-bound proteins such as p130Cas (13, 14).
FAK activity is tightly controlled in response to external stim- uli. Accordingly, FAK is rapidly dephosphorylated in cells de- prived of integrin-mediated cell adhesion and these events may be mediated by protein-tyrosine phosphatases such as PTEN or PTP-PEST (11). However, cells have evolved an additional means to control FAK activity through the autonomous expres- sion of the FAK C-terminal domain termed FRNK (10, 15).
FRNK lacks kinase activity and the Y-397 autophosphorylation site, but comprises the proline-rich sequences and the FAT do- main of FAK (Fig. 1). Therefore, FRNK strongly localizes to focal adhesions, but does not participate in a c-Src-containing signaling complex. It is believed that FRNK localization to fo- cal contacts displaces FAK from the binding partners of its FAT domain, leading to a dissociation from integrin control and pre- venting FAK activation (16–18). Notably, a point-mutated ver- sion of FRNK, FRNK S-1034, which does not localize to focal adhesions, does not inhibit FAK activation (16, 18, 19).
Downstream Signaling Events Involving the FAK-Src Complex
Signals from the FAK-Src PTK complex enhance and prolong activation of ERK/MAP kinases in cells plated on fibronectin-
activated integrins (reviewed in 12). Recent studies show that FAK also contributes to growth factor-initiated MAP kinase ac- tivation (16). Though it is not clear how FAK enhances growth factor-dependent signals, it is hypothesized that the FERM domain-mediated association of FAK with activated growth fac- tor receptors may contribute to this signaling connection. Ad- ditionally, FAK is also involved in integrin- and growth factor- initiated activation of the c-jun N-terminal kinase (JNK) (18, 20, 21). A possible link between FAK and JNK/stress-activated protein (SAP) kinase cascade activation is the tripartite com- plex formed between FAK, c-Src, and p130Cas (reviewed in 12, 22). Tyrosine phosphorylation of p130Cas promotes Crk adapter protein SH2 binding and the coordination of signals leading to the small GTPase Rac and JNK/SAP kinase activation (23, 24).
Supporting this hypothesis, a FAK-p130Cas complex activates JNK in response to fibronectin stimulation of primary synovial fibroblasts (21) and disruption of the FAK-Src-p130Cas com- plex in human A549 epithelial cells impairs epidermal growth factor-stimulated JNK activation (18).
Recent findings suggest that the FAK-Src PTK complex can also modulate small GTPase activity. Potential indirect connec- tions may be through paxillin in association with the p95PKL/
GIT1 adaptor protein and guanine nucleotide exchange factors (GEFs) for Rac and Cdc42 of the Cool/PIX family (reviewed in 25). A direct interaction of FAK and GIT1 also has been reported that promoted Cool/PIX recruitment to focal adhesions and sub- sequent stimulation of Rac GTPase activity (26). Additional un- characterized connections between FAK and Rho GTPases also exist, as FAK-deficient cells exhibit elevated Rho activity that is repressed upon FAK re-expression (27). Because the FAK- associated Rho-GAP protein Graf could represent a negative regulator of Rho GTPases, it is possible that FAK may modulate Rho activity via the recruitment and activation of Graf (8). It is likely that activation of Rac and Cdc42 activity coupled with the inhibition of Rho activity by FAK is an important linkage promoting the dynamic regulation of the actin cytoskeleton. In addition to effects on signaling cascades, FAK can also directly phosphorylate focal contact- and actin-associated molecules like paxillin orα-actinin (11, 28). Therefore, it is exciting to spec- ulate that FAK may also exert a direct regulatory function on focal contacts and the actin cytoskeleton through the tyrosine phosphorylation of key actin-associated proteins.
Role of FAK in Cell Migration
A number of in vitro studies have demonstrated a positive role for FAK in cell migration. FAK-deficient fibroblasts dis- play reduced migratory capacity in response to growth factor (chemotaxis), extracellular matrix protein (haptotaxis), and sub- strate rigidity-dependent (durotaxis) stimulation that is rescued by FAK re-expression (3, 19, 29, 30). Because FAK-null cells exhibit an overabundance of focal contacts and FAK can pro- mote the remodeling of cell-matrix contacts, it is believed that this linkage is one of the primary functions of FAK. Interference with FAK function via antisense oligonucleotides or by FRNK
expression inhibits the motility of various cell types (16–18).
FAK overexpression promotes haptotaxis migration that depends on FAK-p130Cas interactions and the integrity of the FAK Y-397 site (31). In particular, Src PTK recruitment to phos- phorylated Y-397 is critical for haptotaxis and chemotaxis of primary fibroblasts (3, 19). This suggests that signals coordi- nated by a combined FAK-Src PTK complex are responsible for increased cell motility and ongoing studies are aimed at eluci- dating the key targets responsible for promoting these events.
In addition to the modulation of Rac and Rho activity that can directly effect focal contact regulation, FAK-Src PTK connec- tions to the ERK/MAP kinase cascade is another linkage to a migration-promoting signaling pathway. Elevated ERK activity has been connected with increased actin-myosin cell contractil- ity needed by cells to create traction forces and the inhibition of ERK blocks FAK-Src PTK motility (16, 18, 24). Interestingly, expression of the FAK-related Pyk2 PTK is elevated in FAK-null cells, however, Pyk2 overexpression fails to promote cell motil- ity compared to cells reconstituted with FAK (12). Pyk2 does not strongly localize to focal contact sites in FAK-null cells and targeting the Pyk2 N-terminal and kinase domains to focal adhe- sions by fusion to the FAK C-terminal domain restored efficient cell migration and ERK activation in FAK-deficient fibroblasts (32). These results reinforce the importance of correct local- ization and spatial regulation of FAK activation necessary for cell migration. Mechanistically, time-lapse video microscopy of migrating fibroblasts showed that FAK promotes increased cell migration speed and that FAK extends the directional persis- tence of cell movement (30, 33). The simultaneous modulation of two important parameters of cell migration by FAK could explain its profound effect on the regulation of cell motility.
Role of FAK in Cell Invasion
Immunohistochemical staining of burn wounds showed that FAK expression was elevated in actively migrating keratinocytes and enhanced levels of FAK have also been found in smooth muscle cells during intimal hyperplasia, a pathological condi- tion involving the migration of cells from the media to the intima of the blood vessel wall (34). Studies with human tumor tissues and tumor-derived cell lines show that FAK expression is ele- vated during malignancy (reviewed in 35). High levels of FAK are found in prostate, breast, ovarian, and thyroid carcinomas as well as in metastases derived from colon carcinomas. In most cases, increased FAK expression levels correlate with enhanced tumor invasion (36). Parsons and coworkers reported that human prostate carcinoma cell lines have increased FAK expression resulting in enhanced FAK activity and tyrosine phosphoryla- tion (37). Elevated FAK levels were correlated with increased prostate carcinoma cell motility in vitro and either FRNK expres- sion or inhibition of Src family kinases inhibited cell migration.
As tumor cell invasion through matrix and tissue barriers requires combined effects of increased cell motility and regu- lated proteolytic degradation, it is reasonable to speculate that FAK-mediated signaling may contribute to multiple aspects of
the invasive process. In studying the effects of FRNK expres- sion in v-Src-transformed cells, we observed that FRNK in- terfered with in vitro invasion through a basement membrane barrier and in vivo experimental metastasis formation in nude mice (Hauck et al., submitted). Significantly, FRNK inhibition of v-Src-stimulated cell invasion resulted from reduced matrix met- alloproteinase 2 expression and was separate from measurable inhibitory effects on cell motility or growth. FAK antisense treatment as well as adenovirus-mediated expression of FRNK in human adenocarcinoma cells reduced EGF-stimulated cell motility, blocked in vitro basement membrane invasion, and also inhibited MMP-9 secretion from these cells (18).
Additionally, FAK antisense treatment of human ovarian car- cinoma cells inhibited integrin-stimulated secretion of matrix metalloproteinase-9 (MMP-9) in a Ras and ERK/MAP kinase- dependent manner (38). Although FAK may regulate MMP secretion independent of changes in MMP protein expression (39), the promoters of MMP genes are regulated in part by the MAP and SAP kinase signaling pathways. Notably, FRNK ex- pression v-Src-transformed fibroblasts resulted in the inhibition ERK2/MAP and JNK/SAP kinase signaling and resulted in re- duced MMP-2 mRNA production (Hauck et al., submitted).
The role of FAK in promoting cell invasion also applies to normal development, as investigations on the mechanisms of placental implantation strongly support a role of FAK in these processes (40). During pregnancy, placental cells from the fetus must invade the maternal uterus to gain access to blood vessels.
These invading cells termed cytotrophoblasts undergo a stepwise differentiation program resulting in a phenotype capable of inva- sive motility and recruitment of blood vessels. FAK phosphory- lation at Y-397 is strongly elevated in cytotrophoblasts invading into the uterus, indicating that FAK activity in the invasive cells is upregulated. Moreover, adenoviral-based FAK antisense treat- ment of cytotrophoblasts severely impaired the invasive abilities of these cells (40) supporting the conclusion that FAK plays an active and important role in cytotrophoblast-mediated cell invasion.
CONCLUDING REMARKS
We have documented a number of recent studies demon- strating a positive role for FAK in promoting cell migration and invasion. FAK integrates various physical and biochemical stimuli centering around focal adhesion sites to control the local- ized and transient assembly of a multi-protein complex (Fig. 2).
Signals initiated by this FAK-dependent complex contribute to the recruitment and modulation of small GTPases of the Rho and Ras families as well as MAP and SAP kinase pathways.
Consequently, FAK not only acts directly on the plasticity of cytoskeletal structures at focal adhesions, but mediates effects on gene expression that indirectly alter the ability of a cell to migrate and invade. Therefore, FAK is likely to constitute a ma- jor regulatory checkpoint coordinating the intracellular (integra- tion of the actin cytoskeleton at focal adhesions) and extracellu- lar (expression and secretion of extracellular matrix-degrading
Figure 2. Model of FAK connections to cell migration and in- vasion. FAK localized at focal contact sites can communicate with active growth factor receptors and integrins and via the FERM and FAT domains, respectively. FAK activation results in FAK phosphorylation at Tyr-397 and the recruitment of Src- family kinases into a signaling complex consisting of FAK, Src, and p130Cas. Phosphorylation of focal adhesion- and/or actin- associated substrates allows the binding of adaptor molecules leading to the direct or indirect modulation of small GTPases as well as the ERK2/MAP and JNK/SAP kinase cascades. Coor- dinated and localized stimulation of these cascades influences focal contact turnover and actin cytoskeleton dynamics in addi- tion to expression of motility- and invasion-associated proteins such as matrix metalloproteinases (MMPs).
proteases) requirements for efficient cell migration and invasion.
The prominent role FAK plays in these processes provides an attractive target for strategies that focus on blocking the FAK- initiated signaling complex to counter aberrant cell movement in vivo.
ACKNOWLEDGMENTS
David Schlaepfer is supported by grants from the National Cancer Institute (CA75240 and CA87038). This is manuscript 14622-IMM from the Scripps Research Institute.
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