Focal adhesion kinase-like


Table of contents

Fak, the assembly of focal adhesions: The extracellular matrix signal through Fak to promote cell survival

pp125FAK is a tyrosine kinase that appears to regulate the assembly of focal adhesions and thereby promotes cell spreading on the extracellular matrix. In some cells, the C terminus of pp125FAK is expressed as a separate protein, pp41/43FRNK. Overexpression of pp41/43FRNK inhibits tyrosine phosphorylation of pp125FAK and paxillin and, in addition, overexpression delays cell spreading and focal adhesion assembly. Thus, pp41/43FRNK functions as a negative inhibitor of adhesion signaling and provides a tool to dissect the mechanism by which pp125FAK promotes cell spreading. The inhibitory effects of pp41/43FRNK expression can be rescued by the co-overexpression of wild-type pp125FAK and partially rescued by catalytically inactive variants of pp125FAK. However, both coexpression of a pp125FAK mutant for the autophosphorylation site that fails to bind the SH2 domain of pp60c-Src, or of a mutant that fails to bind paxillin, fail to promotes cell spreading. In contrast, expression of pp41/43FRNK and pp60c-Src reconstitute cell spreading and tyrosine phosphorylation of paxillin but do so without inducing tyrosine phosphorylation of pp125FAK. These data provide additional support for a model whereby pp125FAK acts as a "switchable adaptor" that recruits pp60c-Src to phosphorylate paxillin, promoting cell spreading. In addition, these data point to tyrosine phosphorylation of paxillin as being a critical step in focal adhesion assembly (Richardson, 1997).

In many malignant cells, both the anchorage requirement for survival and the function of the p53 tumor suppressor gene are subverted. These effects are consistent with the hypothesis that survival signals from extracellular matrix (ECM) suppress a p53-regulated cell death pathway. Survival signals from fibronectin are transduced by the focal adhesion kinase (FAK). If FAK or the correct ECM is absent, cells enter apoptosis through a p53-dependent pathway activated by protein kinase C lambda/iota and cytosolic phospholipase A2. This pathway is suppressible by dominant-negative p53 and Bcl2 but not CrmA. Upon inactivation of p53, cells survive even if they lack matrix signals or FAK. This is the first report that p53 monitors survival signals from ECM/FAK in anchorage- dependent cells (Ilic, 1998).

Fibronectins are widely expressed extracellular matrix ligands that are essential for many biological processes. Fibronectin-induced signaling pathways are elicited in diverse cell types when specific integrin receptors bind to the ninth and tenth FIII domains, FIII9-10. Integrin-mediated signal transduction involves activation of signaling pathways of the growth factor-dependent Ras-related small GTP-binding proteins Rho and Rac, and phosphorylation of focal adhesion kinase. The requirement of FIII9 and FIII10 for Rho and Rac activity and phosphorylation of focal adhesion kinase in BHK fibroblasts and Swiss 3T3 cells have been examined. FIII10 supports cell attachment but does not induce phosphorylation of focal adhesion kinase. In Swiss 3T3 cells, growth factor-independent phosphorylation of focal adhesion kinase and downstream adhesion events are dependent upon the presence of FIII9 in the intact FIII9-10 pair, whereas FIII10-mediated focal adhesion kinase phosphorylation requires a synergistic signal from growth factors. Furthermore, FIII10 is able to elicit cellular responses mediated by Rho, but not Rac, whereas FIII9-10 can elicit both Rho- and Rac-mediated responses. It is proposed that activation of specific integrin subunits by the FIII10 and FIII9-10 ligands elicits distinct signaling events. This may represent a general molecular mechanism for activation of receptor-specific signaling pathways by a multi-domain ligand (Hotchin, 1999).

Cell adhesive mechanisms which determine tissue architecture during morphogenesis are tightly regulated and have an impact on apoptosis, cell migration, proliferation, and differentiation. Bcl-2 is a death repressor that protects cells from apoptosis initiated by a variety of stimuli including loss of cell adhesion. Utilizing the kidney as a model of an organ that undergoes three-dimensional development it has been demonstrated that bcl-2 directly associates with paxillin. Focal adhesion kinase (FAK)(p125) and paxillin(p68) were highly expressed and tyrosine phosphorylated during development but decline to low levels following renal maturation (postnatal day 20) in normal mice. The decline in the expression of p125 FAK and p68 paxillin occurs together with an increase in specific cleavage products of lower molecular weights. Mice deficient in bcl-2 are born with renal hypoplasia and succumb to renal failure as a result of renal multicystic disease. In kidneys from postnatal day 20 bcl-2 -/- mice, tyrosine phosphorylation of p125 FAK and p68 paxillin is not down-regulated. However, the level of expression was similar to that of normal mice. These results demonstrate that the developmentally regulated expression and phosphorylation of FAK and paxillin, in the presence of bcl-2, is necessary for normal morphogenesis. The interaction of paxillin with bcl-2 during nephrogenesis may provide an alternative to integrin(s) signaling through paxillin/FAK thus bypassing the need for adhesion-mediated survival during three dimensional morphogenesis (Sorenson, 1999).

Signals from the extracellular matrix can modulate cellular differentiation and gene expression. In contrast to other extracellular matrix molecules pepsin-solubilized collagen VI (CVI) can stimulate DNA synthesis of various mesenchymal cell types, apparently independent of integrin-mediated signal transduction. In order to further elucidate collagen VI-induced signaling events, mouse 3T3 fibroblasts and human HT1080 fibrosarcoma cells were exposed to soluble CVI. CVI induces tyrosine phosphorylation of proteins that associate with focal adhesions, such as paxillin, focal adhesion kinase (FAK), and p130CAS (see CAS/CSE1 segregation protein). Furthermore, it activates the mitogen-activated protein kinase, erk2. Kinetic analysis shows that these phosphorylations are transient, reaching a maximum after 5 min for transformed HT1080 cells and 30 min for 3T3 fibroblasts. These effects are partly inhibited by a beta1-integrin function blocking antibody and by single chains of CVI. These results indicate that soluble fragments of native collagen VI, a ubiquitous component of the interstitial extracellular matrix, can mediate stimulation of DNA synthesis via tyrosine phosphorylation of paxillin, FAK, p130CAS, and erk2 in the absence of classical growth factors. Thus, CVI may serve as a matrix-derived sensor that allows for rapid reconstitution of a tissue defect by activating mesenchymal cells (Ruhl, 1999).

Focal adhesion kinase (FAK) is a nonreceptor protein tyrosine kinase involved in integrin-mediated control of cell behavior. Following cell adhesion to components of the extracellular matrix, FAK becomes phosphorylated at multiple sites, including tyrosines 397, 576, and 577. Tyr-397 is an autophosphorylation site that promotes interaction with c-Src or Fyn. Tyr-576 and Tyr-577 lie in the putative activation loop of the kinase domain, and FAK catalytic activity may be elevated through phosphorylation of these residues by associated Src family kinase. Recent studies have implicated FAK as a positive regulator of cell spreading and migration. To further study the mechanism of adhesion-induced FAK activation and the possible role and signaling requirements for FAK in cell spreading and migration, the tetracycline repression system was utilized to achieve inducible expression of either wild-type FAK or phosphorylation site mutants in fibroblasts derived from FAK-null mouse embryos. Using these Tet-FAK cells, it has been demonstrated that both the FAK autophosphorylation and activation loop sites are critical for maximum adhesion-induced FAK activation and FAK-enhanced cell spreading and migration responses. Negative effects on cell spreading and migration, as well as decreased phosphorylation of the substrate p130(Cas), are observed upon induced expression of the FAK autophosphorylation site mutant. These negative effects appear to result from an inhibition of integrin-mediated signaling by the FAK-related kinase Pyk2/CAKbeta/RAFTK/CadTK (Owen, 1999).

Regulation of Fak expression

T cell activation initiates signals that control gene expression of molecules important for T cell function. The focal adhesion kinase Pyk2 has been implicated in T cell signaling. To further analyze the involvement of Pyk2 in T cell processes, the effect of T cell stimulation on the expression of Pyk2 was examined. TCR ligation or PMA increases Pyk2 expression in Jurkat T cells and in normal T cells. In contrast, TCR ligation and PMA fails to induce any detectable increase in the expression of the other member of the focal adhesion kinase family, Fak, in Jurkat T cells and induces only a weak increase in Fak expression in normal T cells. The serine/threonine kinases, protein kinase C and mitogen-activated protein/extracellular signal-related kinase kinase (MEK), regulates Pyk2 expression, as inhibitors of these kinases block stimulus-induced Pyk2 expression. Cyclosporin A, FK506, and KN-62 do not block Pyk2 expression; thus, calcineurin and Ca2+/calmodulin-activated kinases are not critical for augmenting Pyk2 expression. TCR ligation increases Pyk2 mRNA, and the transcriptional inhibitor actinomycin D blocks Pyk2 expression. Strikingly, Ca2+ ionophores, at concentrations that in combination with other stimuli induce IL-2 expression, block TCR- and PMA-induced up-regulation of Pyk2 expression. Thus, the increase in Ca2+ has opposing effects on IL-2 and Pyk2 expression. Cyclosporin A and FK506, but not KN-62, block Ca2+ ionophore-mediated inhibition of Pyk2 expression, implicating calcineurin in down-regulating Pyk2 expression. These results show that TCR-triggered intracellular signals increase Pyk2 expression and shed light on the molecular mechanisms that regulate Pyk2 expression in T cells (Tsuchida, 1999).

Focal adhesion kinase (FAK) has been implicated in cellular processes that control cell adhesion, migration, cell cycle progression, and apoptosis. FRNK (FAK-related nonkinase) is the autonomously expressed, noncatalytic C-terminal portion of FAK. When ectopically expressed in cells, FRNK has been shown to act as a negative regulator of FAK activity, inhibiting cell spreading, migration, and cell cycle progression. The mechanisms that regulate FRNK expression during embryonic development and the functional role of FRNK in normal cell homeostasis remain poorly understood. FRNK expression in chicken cells is directed by an alternative promoter residing within an intron of FAK, positioned 3' of the exon encoding sequences for the catalytic domain and 5' of the exon encoding sequences for the C-terminal domain of FAK (e.g., FRNK). Using probes specific for FRNK, FRNK expression is shown to occur early in chicken embryogenesis, being readily detected at day 3, 6, or 9. Late in embryogenesis, at day 18, FRNK is expressed in a tissue-specific manner, predominately in lung and intestine cells. Western blot analysis of mouse tissues with a FAK-specific antibody reveals the expression of FRNK in the mouse lung. Reverse transcriptase PCR analysis of mouse lung RNA reveals the presence of spliced FRNK mRNAs containing 5' untranslated sequences derived from a positionally conserved exon present in the mouse genome. FAK is the first example of a tyrosine kinase regulated by a domain under the control of an alternative intronic promoter. It is also the first example of a focal adhesion-associated protein regulated by such a mechanism and thus represents a novel means for the modulation of cell adhesion signaling (Nolan, 1999).

FAK acts as a scaffold to assemble a complex of ERK and Calpain stimulating focal adhesion turnover

Cell migration on extracellular matrix requires the turnover of integrin-dependent adhesions. The nonreceptor tyrosine kinases Src and FAK regulate focal-adhesion turnover by poorly understood mechanisms. ERK/MAP kinase-mediated activation of the protease Calpain 2 also promotes focal-adhesion turnover; however, it is not known if this is linked to the activities of Src and FAK. Calpain 2 has previously been demonstrated to colocalize with focal-adhesion structures and can cleave several focal-adhesion complex components, including FAK. Studies utilizing Calpain inhibitors or Calpain-deficient cells confirm that Calpain's role in regulating focal-adhesion turnover is necessary for cell migration. A novel and kinase-independent function has been identified for FAK as an adaptor molecule that mediates the assembly of a complex consisting of at least Calpain 2 and p42ERK. Mutation of proline residues (Pro2) in the amino-terminal region of FAK blocks direct binding with Calpain 2 and also prevents formation of the Calpain 2/p42ERK complex in cells. Both complex formation and MEK/ERK activity are associated with Calpain-mediated proteolysis of FAK and focal adhesion turnover during transformation and migration. Furthermore, FAK is necessary for recruiting both Calpain 2 and p42ERK/MAPK to peripheral adhesion sites facilitating maximal Calpain activity (Carragher, 2003).

These results indicate that FAK combines to spatially couple Calpain 2 to its upstream regulator ERK/MAP kinase. In live cells fluorescence intensity relating to Calpain activity is visibly reduced in FAK-/- cells relative to wild-type MEFs. In addition, Calpain activity localizes to the cell membrane in wild-type MEFs but not in FAK-/- cells. Total Calpain activity in extracted cell lysates is reduced in FAK-/- cells relative to wild-type MEFs. These results are consistent with the conclusion that FAK is required for assembly of a complex containing both Calpain 2 and phospho-ERK, the upstream activating kinase Calpain, and for recruitment of this complex to the plasma membrane, which is known to promote full Calpain activation (Carragher, 2003). In summary, this study describes a novel function for FAK as an adaptor molecule that permits the assembly of a Calpain 2/FAK/p42ERK complex. FAK-dependent complex assembly and localization to the cell periphery facilitates ERK/MAPK-induced activation of Calpain. This subsequently permits Calpain-mediated cleavage of FAK (and most likely other adhesion components), focal adhesion turnover, and cell migration. These findings provide a mechanistic explanation for the similar adhesion turnover and migratory defects of FAK null and Calpain-deficient cells (Carragher, 2003).

Fak and adhesion, cell motility, invasion and oncogenic transformation

It has been proposed that the focal adhesion kinase (FAK) mediates focal adhesion formation through tyrosine phosphorylation during cell adhesion. This study investigated the role of FAK in focal adhesion structure and function. Loading cells with a glutathione-S-transferase fusion protein (GST-Cterm) containing the FAK focal adhesion targeting sequence, but not the kinase domain, decreased the association of endogenous FAK with focal adhesions. This displacement of endogenous FAK in both BALB/c 3T3 cells and human umbilical vein endothelial cells loaded with GST-Cterm decreased focal adhesion phosphotyrosine content. Neither cell type, however, exhibited a reduction in focal adhesions after GST-Cterm loading. These results indicate that FAK mediates adhesion-associated tyrosine phosphorylation, but not the formation of focal adhesions. The effect of inhibiting FAK function on other adhesion-dependent cell behavior was examined. Cells microinjected with GST-Cterm exhibited decreased migration. In addition, cells injected with GST-Cterm had decreased DNA synthesis compared with control-injected or noninjected cells. These findings suggest that FAK functions in the regulation of cell migration and cell proliferation (Gilmore, 1996).

Cell migration and invasion are fundamental components of tumor cell metastasis. Increased focal adhesion kinase (FAK) expression and tyrosine phosphorylation are connected with elevated tumorigenesis. Null mutation of FAK results in embryonic lethality, and FAK-/- fibroblasts exhibit cell migration defects in culture. Viral Src (v-Src) transformation of FAK-/- cells promotes integrin-stimulated motility equal to stable FAK reexpression. However, FAK-/- v-Src cells are not invasive, and FAK reexpression, Tyr-397 phosphorylation, and FAK kinase activity are required for the generation of an invasive cell phenotype. Cell invasion is linked to transient FAK accumulation at lamellipodia, formation of a FAK-Src-p130Cas-Dock180 signaling complex, elevated Rac and c-Jun NH2-terminal kinase activation, and increased matrix metalloproteinase expression and activity. These studies support a dual role for FAK in promoting cell motility and invasion through the activation of distinct signaling pathways (Hsia, 2003).

The ability of the focal adhesion kinase (FAK) to integrate signals from extracellular matrix and growth factor receptors requires the integrity of Tyr397, a major autophosphorylation site that mediates the Src homology 2-dependent binding of Src family kinases. However, the precise roles played by FAK in specific Src-induced pathways, especially as they relate to oncogenic transformation, remain unclear. The role of FAK in v-Src-induced oncogenic transformation was investigated by transducing temperature-sensitive v-Src (ts72v-Src) into p53-null FAK+/+ or FAK-/- mouse embryo fibroblasts (MEF). At the permissive temperature (PT), ts72v-Src induces abundant tyrosine phosphorylation, morphological transformation and cytoskeletal rearrangement in FAK-/- MEF, including the restoration of cell polarity, typical focal adhesion complexes, and longitudinal F-actin stress fibers. v-Src rescues the haptotactic (referring to cell-motility toward substratum-bound insolubilized extracellular matrix components), linear directional, and invasive motility defects of FAK-/- cells to levels found in FAK+/+ or FAK+/+-[ts72v-Src] cells, and, in the case of monolayer wound healing motility, there is an enhancement. Src activation failed to increase the high basal tyrosine phosphorylation of the Crk-associated substrate, CAS, found in FAK-/- MEF, indicating that CAS phosphorylation alone is insufficient to induce motility in the absence of FAK- or v-Src-induced cytoskeletal remodeling. Compared with FAK+/+[ts72v-Src] controls, FAK-/-[ts72v-Src] clones exhibited 7-10-fold higher anchorage-independent proliferation that could not be attributed to variations in either v-Src protein level or stability. Re-expression of FAK diminished the colony-forming activities of FAK-/-[ts72v-Src] without altering ts72v-Src expression levels, suggesting that FAK attenuates Src-induced anchorage independence. These data also indicate that the enhanced Pyk2 level found in FAK-/- MEF plays no role in v-Src-induced anchorage independence. Overall, these data indicate that FAK, although dispensable, attenuates v-Src-induced oncogenic transformation by modulating distinct signaling and cytoskeletal pathways (Moissoglu, 2003).

Cell migration is a complex, highly regulated process that involves the continuous formation and disassembly of adhesions (adhesion turnover). Adhesion formation takes place at the leading edge of protrusions, whereas disassembly occurs both at the cell rear and at the base of protrusions. Despite the importance of these processes in migration, the mechanisms that regulate adhesion formation and disassembly remain largely unknown. Quantitative assays have been developed to measure the rate of incorporation of molecules into adhesions and the departure of these proteins from adhesions. Using these assays, it has been shown that kinases and adaptor molecules, including focal adhesion kinase (FAK), Src, p130CAS, paxillin, extracellular signal-regulated kinase (ERK) and myosin light-chain kinase (MLCK) are critical for adhesion turnover at the cell front, a process central to migration (Webb, 2004).

Mice were generated with a floxed fak allele under the control of keratin-14-driven Cre fused to a modified estrogen receptor (CreERT2). 4-Hydroxy-tamoxifen treatment induces fak deletion in the epidermis, and suppresses chemically induced skin tumor formation. Loss of fak induced after benign tumors had formed inhibits malignant progression. Although fak deletion is associated with reduced migration of keratinocytes in vitro, no effect on wound re-epithelialization was found in vivo. However, increased keratinocyte cell death is observed after fak deletion in vitro and in vivo. This work provides the first experimental proof implicating FAK in tumorigenesis, and this is associated with enhanced apoptosis (McLean, 2005).

Fak and development

There are contrasting roles for integrin alpha subunits and their cytoplasmic domains in controlling cell cycle withdrawal and the onset of terminal differentiation. Ectopic expression of the integrin alpha5 or alpha6A subunit in primary quail myoblasts either decreases or enhances the probability of cell cycle withdrawal, respectively. The mechanisms by which changes in integrin alpha subunit ratios regulate this decision are addressed. Ectopic expression of truncated alpha5 or alpha6A indicate that the alpha5 cytoplasmic domain is permissive for the proliferative pathway, whereas the COOH-terminal 11 amino acids of alpha6A cytoplasmic domain inhibit proliferation and promote differentiation. The alpha5 and alpha6A cytoplasmic domains do not appear to initiate these signals directly, but instead regulate beta1 signaling. Ectopically expressed IL2R-alpha5 or IL2R-alpha6A have no detectable effect on the myoblast phenotype. However, ectopic expression of the beta1A integrin subunit or IL2R-beta1A, autonomously inhibits differentiation and maintains a proliferative state. Perturbing alpha5 or alpha6A ratios also significantly affects activation of beta1 integrin signaling pathways. Ectopic alpha5 expression enhances expression and activation of paxillin as well as mitogen-activated protein (MAP) kinase with little effect on focal adhesion kinase (FAK). In contrast, ectopic alpha6A expression suppresses FAK and MAP kinase activation with a lesser effect on paxillin. Ectopic expression of wild-type and mutant forms of FAK, paxillin, and MAP/erk kinase (MEK) confirm these correlations. These data demonstrate that (1) proliferative signaling (i.e., inhibition of cell cycle withdrawal and the onset of terminal differentiation) occurs through the beta1A subunit and is modulated by the alpha subunit cytoplasmic domains; (2) perturbing alpha subunit ratios alters paxillin expression and phosphorylation and FAK and MAP kinase activation; (3) quantitative changes in the level of adhesive signaling through integrins and focal adhesion components regulate the decision of myoblasts to withdraw from the cell cycle, in part via MAP kinase (Sastry, 1999).

Zebrafish focal adhesion kinase (Fak) has been cloned and its subcellular localization has been analyzed. Fak protein is localized at the cortex of notochord cells and at the notochord-somite boundary. During somitogenesis, Fak protein becomes concentrated at the basal region of epithelial cells at intersomitic boundaries. Phosphorylated Fak protein is seen at both the notochord-somite boundary and intersomitic boundaries, consistent with a role for Fak in boundary formation and maintenance. The localization of Fak protein to the basal region of epithelial cells in knypek;trilobite double mutant embryos shows that polarization of Fak distribution in the somite border cells is independent of internal mesenchymal cells. In addition, neither Notch signaling through Suppressor of Hairless nor deltaD is necessary for the wild-type segmental pattern of fak mRNA expression in the anterior paraxial mesoderm. However, nonsegmental expression of fak mRNA occurs with ectopic activation of Notch signaling through SuH and also in fused somite and beamter mutant embryos, indicating that there are multiple regulators of fak mRNA expression. These results suggest that Fak plays a central role in notochord and somite morphogenesis (Henry, 2001).

Targeted deletion of focal adhesion kinase (fak) in the developing dorsal forebrain results in local disruptions of the cortical basement membrane located between the neuroepithelium and pia-meninges. At disruption sites, clusters of neurons invade the marginal zone. Retraction of radial glial endfeet, midline fusion of brain hemispheres, and gliosis also occurs, similar to type II cobblestone lissencephaly as seen in congenital muscular dystrophy. Interestingly, targeted deletion of fak in neurons alone does not result in cortical ectopias, indicating that fak deletion from glia is required for neuronal mislocalization. Unexpectedly, fak deletion specifically from meningeal fibroblasts elicits similar cortical ectopias in vivo and altered laminin organization in vitro. These observations provide compelling evidence that FAK plays a key signaling role in cortical basement membrane assembly and/or remodeling. In addition, FAK is required within neurons during development because neuron-specific fak deletion alters dendritic morphology in the absence of lamination defects (Beggs, 2003).

FAK exerts its biological effects not only through its tyrosine kinase activity but also by acting as a scaffolding protein that connects cell surface proteins to the actin cytoskeleton in a large macromolecular complex. Disruption of FAK signaling may prevent the cytoskeletal reorganization and signal transduction cascades that are necessary to convey the bidirectional signals required for laminin organization and overall basement membrane stability. The dramatic reduction in adaptor protein p130CAS phosphorylation throughout fak-/- forebrains suggests that perturbations in localization or activities of this adaptor protein may play a significant role in the FAK phenotype. Although FAK can directly phosphorylate p130CAS, this reduction in phosphorylation may alternatively be due to the loss of Src recruitment into FAK/p130CAS signaling complexes. FAK and p130CAS have been shown to cooperate in the promotion of cell migration in many cell types. However, the mechanistic details of how FAK and p130CAS modulate the actin cytoskeleton are not fully known. Given that FAK signaling has been shown to both suppress and activate rho activity, FAK may serve as a regulatable switch dynamically controlling rho-mediated effects on the actin cytoskeleton. An intriguing further possibility is that FAK can act as a 'mechanosensor,' altering its link to the cytoskeleton in response to environmental tension or rigidity of the extracellular matrix. FAK deletion may disrupt this mechanosensory communication enough to alter the dynamic force necessary to assemble or remodel the matrix. This will be an interesting source of future study (Beggs, 2003).

Focal adhesion kinase (FAK) is a critical mediator of signal transduction by integrins and growth factor receptors in a variety of cells including endothelial cells (ECs). EC-specific knockout of FAK is described using a Cre-loxP approach. In contrast to the total FAK knockout, deletion of FAK specifically in ECs does not affect early embryonic development including normal vasculogenesis. However, in late embryogenesis, FAK deletion in the ECs leads to defective angiogenesis in the embryos, yolk sac, and placenta, impaired vasculature and associated hemorrhage, edema, and developmental delay, and late embryonic lethal phenotype. Histologically, ECs and blood vessels in the mutant embryos present a disorganized, detached, and apoptotic appearance. Consistent with these phenotypes, deletion of FAK in ECs isolated from the floxed FAK mice leads to reduced tubulogenesis, cell survival, proliferation, and migration in vitro. Together, these results strongly suggest a role of FAK in angiogenesis and vascular development due to its essential function in the regulation of multiple EC activities (Shen, 2005).

The metameric organization of the vertebrate body plan is established during somitogenesis as somite pairs sequentially form along the anteroposterior axis. Coordinated regulation of cell shape, motility and adhesion are crucial for directing the morphological segmentation of somites. Members of the Ena/VASP family of actin regulatory proteins are required for somitogenesis in Xenopus. Xenopus Ena (Xena) localizes to the cell periphery in the presomitic mesoderm (PSM), and is enriched at intersomitic junctions and at myotendinous junctions in somites and the myotome, where it co-localizes with beta1-integrin, vinculin and FAK. Inhibition of Ena/VASP function with dominant-negative mutants results in abnormal somite formation that correlates with later defects in intermyotomal junctions. Neutralization of Ena/VASP activity disrupts cell rearrangements during somite rotation and leads to defects in the fibronectin (FN) matrix surrounding somites. Furthermore, inhibition of Ena/VASP function impairs FN matrix assembly, spreading of somitic cells on FN and autophosphorylation of FAK, suggesting a role for Ena/VASP proteins in the modulation of integrin-mediated processes. Inhibition of FAK results in defects in somite formation, blocks FN matrix deposition and alters Xena localization. Together, these results provide evidence that Ena/VASP proteins and FAK are required for somite formation in Xenopus and support the idea that Ena/VASP and FAK function in a common pathway to regulate integrin-dependent migration and adhesion during somitogenesis (Kragtorp, 2006; full text of article).

The intestinal epithelium has a remarkable capacity to regenerate after injury and DNA damage. This study shows that the integrin effector protein Focal Adhesion Kinase (FAK) is dispensable for normal intestinal homeostasis and DNA damage signaling, but is essential for intestinal regeneration following DNA damage. Given Wnt/c-Myc signaling is activated following intestinal regeneration, the functional importance of FAK was investigated following deletion of the Apc tumor suppressor protein within the intestinal epithelium. Following Apc loss, FAK expression increased in a c-Myc-dependent manner. Codeletion of Apc and Fak strongly reduced proliferation normally induced following Apc loss, and this was associated with reduced levels of phospho-Akt and suppression of intestinal tumorigenesis in Apc heterozygous mice. Thus, FAK is required downstream of Wnt Signaling, for Akt/mTOR activation, intestinal regeneration, and tumorigenesis. Importantly, this work suggests that FAK inhibitors may suppress tumorigenesis in patients at high risk of developing colorectal cancer (Ashton, 2010).

Fak and dendrite growth

The rate and direction of axon and dendrite growth depend on multiple guidance signals and growth factors. Semaphorin 3A (Sema3A) acts as a repellent for axons and attractant for dendrites. This study shows that the requirement for integrin engagement distinguishes the response of axons and dendrites to Sema3A in hippocampal neurons. Sema3A promotes the extension of hippocampal dendrites by a pathway that requires focal adhesion kinase (FAK). The stimulation of dendrite growth and FAK phosphorylation by Sema3A depend on integrin engagement. Unlike their function as a target of Sema3A during the collapse of axonal growth cones, integrins facilitate the stimulation of dendrite extension. Conditional inactivation of the genes encoding β1 integrin or FAK blocks the growth-promoting effect of Sema3A but not the collapse of axonal growth cones. These results demonstrate that different pathways mediate the stimulation of dendrite growth and the collapse of axonal growth cones by Sema3A (Schlomann, 2009).

These results reveal a bidirectional interaction of integrins and semaphorin signalling. β1 integrins facilitate the effect of Sema3A on dendrites, whereas integrins are inhibited by Sema3A during growth-cone collapse and in endothelial cells. Sema3A stimulates dendrite extension only when neurons are cultured on the integrin ligand fibronectin. Fibronectin could be substituted by Mn2+ ions, which stabilise the active conformation of integrins, allowing Sema3A to promote dendrite extension on the non-specific adhesive PO. A soluble RGD peptide that blocks the interaction with integrins and deletion of the Itgb1 gene abrogated the stimulatory effect of Sema3A. By contrast, the collapse of axonal growth cones was independent of integrin engagement. Unlike the effect of cyclic nucleotides on the response to axon-guidance signals, β1-integrin engagement does not modulate the response of dendrites because Sema3A had no effect on dendrite length after inactivation of Itgb1 or Fak. The results show that the effect of Sema3A on dendrite growth requires a signalling pathway that is distinct from that involved in collapse of the growth cone (Schlomann, 2009).

Fak and apoptosis

Apoptotic cells undergo characteristic morphological changes that include detachment of cell attachment from the substratum and loss of cell-cell interactions. Attachment of cells to the extracellular matrix and to other cells is mediated by integrins. The interactions of integrins with the extracellular matrix activate focal adhesion kinase (FAK) and suppress apoptosis in diverse cell types. Members of the tumor necrosis family such as Fas and Apo-2L, also known as tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), induce apoptosis in both suspension and adherent cells through the activation of caspases. These caspases, when activated, cleave substrates that are important for the maintenance of nuclear and membrane integrity. FAK is sequentially cleaved into two different fragments early in Apo-2L-induced apoptosis. FAK cleavage is mediated by caspases, and FAK shows unique sensitivity to different caspases. These results suggest that disruption of FAK may contribute to the morphological changes observed in apoptotic suspension and adherent cells (Wen, 1997).

Focal adhesion kinase (FAK) has been implicated to play a role in suppression of apoptosis. UV irradiation induces cleavage of FAK and two of its interacting proteins Src and p130(Cas) in Madin-Darby canine kidney cells, concomitant with an increase in cell death. The cleavage of these proteins upon UV irradiation is completely inhibited by ZVAD-FMK, a broad range inhibitor of caspases, and apparently delayed by Bcl2 overexpression. To examine if FAK plays a role in suppressing UV-induced apoptosis, stable Madin-Darby canine kidney cell lines overexpressing FAK were established. A marked (30-40%) increase in cell survival upon UV irradiation is achieved by this strategy. In efforts to determine the mechanism by which FAK transduces survival signals to the downstream, it was found that a FAK mutant deficient in binding to phosphatidylinositol 3-kinase fails to promote cell survival. Moreover, the expression of the Src homology 3 domain of p130(Cas), which competes with endogenous p130(Cas) for FAK binding, abrogates the FAK-promoted cell survival. Together, these results suggest that the integrity of FAK and its binding to phosphatidylinositol 3-kinase and p130(Cas) are required for FAK to exert its antiapoptotic function (Chan, 1999).

The relationship between focal adhesion protein (FAK) activity and loss of cell-matrix contact during apoptosis is not entirely clear nor has the role of FAK in chemically induced apoptosis been studied. The status of FAK phosphorylation and cleavage in renal epithelial cells during apoptosis caused by the nephrotoxicant dichlorovinylcysteine (DCVC) has been investigated. DCVC treatment causes a loss of cell-matrix contact which is preceded by a dissociation of FAK from the focal adhesions and tyrosine dephosphorylation of FAK. Paxillin is also dephosphorylated at tyrosine. DCVC treatment activates caspase-3 which is associated with cleavage of FAK. However, FAK cleavage occurs after cells have already lost focal adhesions indicating that cleavage of FAK by caspases is not responsible for loss of FAK from focal adhesions. Accordingly, although inhibition of caspase activity with zVAD-fmk blocks activation of caspase-3, FAK cleavage, and apoptosis, it neither affects dephosphorylation nor translocation of FAK or paxillin. However, zVAD-fmk completely blocks the cell detachment caused by DCVC treatment. Orthovanadate prevents DCVC-induced tyrosine dephosphorylation of both FAK and paxillin; however, it does not inhibit DCVC-induced apoptosis and actually potentiates focal adhesion disorganization and cell detachment. Thus, FAK dephosphorylation and loss of focal adhesions are not due to caspase activation; however, caspases are required for FAK proteolysis and cell detachment (van de Water,1999).

Table of contents

Focal adhesion kinase-like: Biological Overview | Regulation | Developmental Biology | Effects of Mutation | References

Home page: The Interactive Fly © 1995, 1996 Thomas B. Brody, Ph.D.

The Interactive Fly resides on the
Society for Developmental Biology's Web server.