the cell adhesion molecule e - cadherin is among the key molecules in the formation of the epithelial junctional complex [ 13 ] . e - cadherin forms homodimers in the extracellular domain between adjacent cells   and interacts intracellularly with a group of proteins collectively named catenins [ 5 , 6 ] . both the cadherin cytoplasmic domain and the associated catenins are required for full cell adhesion [ 7 , 8 ] , and -catenin is suggested to play a role in anchoring the cadherin - catenin complex to actin - based cytoskeletal components that include -actinin and vinculin . loss or significant reduction of e - cadherin expression has been observed in many epithelial cancers [ 1014 ] , and the -catenin gene is occasionally lost [ 14 , 15 ] or mutated in human cancer cells lines . unlike e - cadherin and -catenin , -catenin loses its function upon tyrosine phosphorylation in response to growth factors [ 1720 ] or v - src [ 21 , 22 ] . tyrosine phosphorylation of -catenin results in loss of anchoring of the e - cadherin - catenin complex to the network of actin filaments ( f - actin ) [ 23 , 24 ] . in normal epithelial cells in culture , tyrosine - phosphorylated -catenin needs to be dephosphorylated in order to link the e - cadherin - catenin complex to f - actin . in fact , several protein tyrosine phosphatases ( ptps ) , including dep-1 , ptp1b , and ptp , are upregulated with increasing cell density [ 25 , 26 ] , and they associate with the e - cadherin - catenin complex [ 2731 ] . therefore , constitutive tyrosine phosphorylation of -catenin and concomitant loss of ptp might cause loss of e - cadherin - mediated cell - cell adhesion in cancer cells . another mode of dysfunction of cell - cell adhesion is induced by perturbation of f - actin assembly to which the e - cadherin - catenin complex anchors . iqgap1 , the iq moti - containing guanine nucleotide - activating protein 1 ( iqgap1 ) , is an actin cross - linking protein [ 33 , 34 ] or scaffold protein . iqgap1 is recruited by protein phosphatase ( pp ) 2a to the e - cadherin - catenin complex that is constitutively associated with the small gtp - binding protein rac1 [ 36 , 37 ] , thereby leading to rearrangement of f - actin by iqgap1 in corporation with rac1 to which the e - cadherin - catenin complex anchors . because inhibition of pp2a activity or loss of pp2a expression results in e - cadherin endocytosis [ 37 , 38 ] , pp2a is considered to be involved in the establishment and maintenance of e - cadherin - mediated cell - cell adhesion as well as in a wide variety of biological processes such as tumor suppression [ 3941 ] , formation of tight junctions , and integrin - mediated cell - substratum adhesion [ 4348 ] . loss of cell - cell adhesion due to the internalization or endocytosis of e - cadherin without significant alterations in e - cadherin expression has been observed in many cells [ 38 , 4952 ] . despite this this implies that the loss of cell - cell adhesion is necessary but insufficient alone for cell migration and invasion .   in this paper , we focus on recent advances in the understanding of the mechanisms underlying regulation of cell migration and invasion that have been possible through the use of biochemical and pharmacological approaches . cell migration and invasion generally require rearrangement of f - actin at the leading edge of cells   and are associated with the formation of specific cellular protrusions termed lamellipodia or filopodia . rearrangement of f - actin is directed by the arp2/3 complex through induction of nucleation and branching of f - actin [ 5456 ] and is regulated by small gtp - binding proteins such as rac1 and cdc42   through wiskott - aldrich syndrome protein ( wasp ) and wasp verprolin homology proteins ( wave ) [ 55 , 5862 ] . n - wasp is a member of the wasp family of proteins [ 58 , 60 ] and is thought to be necessary for lamellipodia formation . wave family proteins , including wave1 , -2 , and -3 [ 58 , 61 ] , induce both lamellipodia [ 6466 ] and filopodia . among the wasp / wave family , wave2 has been the most intensively investigated and has been identified as functioning downstream of rac [ 68 , 69 ] . although direct interaction between rac1 and wave2 is not detected in human breast cancer cells , rac1 forms a complex with clip-170 , a microtubule - binding protein [ 70 , 71 ] , an actin cross - linking protein iqgap1 [ 33 , 34 ] , and kinesin-1 , one of the major motor proteins [ 72 , 73 ] , under the growth - arrested conditions   ( figure 1(a ) ) . clip-170 and iqgap1 bind microtubules and f - actin , respectively , and kinesin-1 transports many cytoplasmic vesicles , proteins , and mrnas as  cargo  toward the growing plus - ends of microtubules . therefore , formation of the rac1 , clip-170 , iqgap1 , and kinesin-1 complex induces linking of rac1 to both f - actin and microtubules by iqgap1 and clip-170 , respectively [ 74 , 75 ] . depletion of clip-170 expression causes growth factor - independent dissociation of iqgap1 and kinesin-1 from rac1 and the promotion of random lamellipodia formation and invasion . this suggests that clip-170 may play a role in preventing cells from growth factor - independent lamellipodia formation and invasion , by tethering iqgap1 and kinesin-1 to rac1 until stimulation by growth factors . following the stimulation of cells with growth factor , iqgap1 and kinesin-1 dissociate from the rac1-clip-170 complex   ( figure 1(b ) ) . as lamellipodia formation and dissociation of iqgap1 and kinesin-1 are inhibited by a phosphoinositide 3-kinase ( pi3k ) inhibitor , the dissociation of iqgap1 and kinesin-1 from the rac1-clip-170 complex is a prerequisite for lamellipodia formation in response to growth factor stimulation ; this depends on pi3k that is activated by the activated growth factor receptor ( figure 1(a ) ) . in order to form lamellipodia , wave2 needs to be transported to the leading edge of cells prior to lamellipodia formation [ 76 , 77 ] . wave2 is known to form multiprotein complexes that include abi-1 , sra-1 , nap-1 , and hspc300 [ 7880 ] . wave2 also forms a complex with iqgap1 and kinesin-1 in growth - arrested breast cancer cells ( figure 1(a ) ) , and the amounts of these proteins relative to wave2 increase after stimulation of the cells with growth factor   ( figure 1(b ) ) . as iqgap1 and kinesin-1 dissociate from the rac1-clip-170 complex upon growth factor stimulation , without significant alterations in the total amounts of iqgap1 and kinesin-1 , iqgap1 and kinesin-1 that are bound to wave2 might be proteins that previously dissociated from the complex ( figure 1(b ) ) . in spite of this , additional binding of iqgap1 and kinesin-1 to wave2 appears to be insufficient for wave2 translocation to the cell cortex . many investigations have revealed that a p21-activated protein kinase pak1 [ 81 , 82 ] , which is one of the downstream effectors of rac1 [ 81 , 83 , 84 ] , is constitutively associated with wave2   ( figure 2(a ) ) . pak1 is thought to regulate not only actin reorganization through several reported substrates , including lim kinase , p41-arc , and filamin , but also microtubule dynamics through stathmin [ 8890 ] , a microtubule destabilizing protein [ 91 , 92 ] . as direct interaction between rac1 and pak1 or wave2 is not detected in breast cancer cells , it is reasonable to assume that pix , the pak - interacting nucleotide exchange factor of rac / cdc42 , plays an important role in the signal transduction between rac1 and pak1 . pix is constitutively coupled to git1 and recruited to wave2-bound pak1 in response to growth factor stimulation . upon growth factor stimulation , the wave2-bound pak1 is activated and in turn phosphorylates stathmin   ( figures 2(a ) and 2(b ) ) . the putative phosphorylation sites within stathmin are ser16 , ser25 , ser38 , and ser63 . among these , growth factor stimulation causes phosphorylation at ser25 and ser38 , although phosphorylation at ser38 alone is dependent on pak1 in breast cancer cells . this implies that serine / threonine protein kinases other than pak1 phosphorylate stathmin at ser25 . in either case , the microtubule destabilizing activity of stathmin is inactivated by phosphorylation [ 8890 ] , and the phosphorylated stathmin is recruited to kinesin-1 ; this complexes with wave2 and pak1 and leads to the promotion of microtubule growth   ( figure 2(b ) ) . stathmin binding to kinesin-1 was thought to be transient and to play a role in the transportation of tubulin heterodimers to the microtubule ends ; however , stathmin binding to kinesin-1 and tubulin heterodimers is maintained after growth factor stimulation . this suggests the presence of a partner protein that facilitates the sustained linking of stathmin to the microtubule ends . the dynamic properties of microtubules are regulated by  plus - end - binding proteins  [ 9698 ] that include clip , clip - associated protein ( clasps ) , dynein / dynactin , apc ( adenomatous polyposis coli ) , and eb - family proteins [ 99101 ] . eb1 recognizes specific sites at the ends of growing microtubules   and promotes persistent microtubule growth . investigation of the stathmin - binding partners revealed that stathmin is constitutively associated with eb1 in the cytoplasm of growth - arrested cells . after phosphorylation of stathmin by pak1 , the phosphorylated stathmin - eb1 complex becomes associated , in an eb1-dependent manner , with the ends of microtubules that bear the wave2 complex . depletion of stathmin does not inhibit microtubule growth but inhibits wave2 translocation and lamellipodia formation , while eb1 depletion does not inhibit microtubule growth but inhibits stathmin binding to kinesin-1 at the microtubule ends , wave2 translocation , and lamellipodia formation . taken together , growth factor - induced and pak1-dependent phosphorylation of stathmin triggers the eb1-mediated specific binding of the phosphorylated stathmin - eb1 complex to the wave2-bearing microtubule ends , thereby leading to wave2 translocation and lamellipodia formation ( figure 2(b ) ) . the process of cancer cell migration and invasion involves intravasation and extravasation of cells into and out of blood vessels . therefore , it is highly likely that cancer cell migration and invasion are directionally controlled by extracellular stimuli such as serum growth factors . if this theory is correct , promotion of persistent growth of the wave2-bearing microtubules by the binding of the stathmin - eb1 complex seems to be insufficient for the directional membrane anchoring of microtubules , that is , without the aid of partner proteins . for example , the end - binding proteins clasps link microtubule ends to the cell cortex by the phosphatidylinosito 3,4,5-triphosphat- ( pip3- ) binding protein ll5 as a partner protein . pip3 , a lipid component of the cytoplasmic membrane produced by pi3k , is known to be the membrane target of many proteins [ 107111 ] that contain the pleckstrin homology ( ph ) domain ( e.g. , pkb / akt [ 112 , 113 ] ) , the basic region ( e.g. , wasp and wave2 [ 76 , 114 ] ) , or the irsp53/missing - in - metastasis ( mim ) domain imd ( e.g. , irsp53 [ 115 , 116 ] ) . in lamellipodia - forming breast cancer cells , irsp53 is constitutively associated with the wave2 complex that involves eb1 and plays an indispensable role in anchoring the complex to pip3 . irsp53 is a linker protein [ 118120 ] that interacts with the proline - rich region of wave2 through its central src - homology 3 ( sh3 ) domain [ 118 , 121 ] . therefore , directional cell migration through lamellipodia formation may be regulated by the interaction between wave2-bound irsp53 as the pip3-binding partner protein and pip3 as the membrane target molecule ; this suggests that the direction of membrane anchoring of microtubules that bear the wave2 complex is determined by the sites where pip3 is produced . in cells cultured in a chemotaxis chamber , only growth factor receptor in the membrane region facing high concentrations of growth factor was activated . local activation of the receptor causes recruitment of pi3k that in turn produces pip3 in close proximity to the activated receptor   ( figure 3(a ) ) , thereby leading to local interaction between irsp53-bound wave2 and pip3 , and thus the directional formation of lamellipodia towards the growth factor source ( figure 3(b ) ) .    in the context of pip3-binding proteins for lamellipodia formation , nonmuscle myosin iia heavy chain myh9 [ 122124 ] has the ability to bind to both pip3 and wave2 . whereas myh9 binding to wave2 is constitutive and requires the motor activity of myosin ii , myh9 binding to pip3 is induced only after wave2 membrane targeting by growth factor activation . the binding of myh9 to wave2 is probably through the interaction between the sh3-like domain of myh9 and the proline - rich domain of wave2 ; however , myh9 lacks a ph domain , unlike myosin 1b and myosin x , which bind to pip3 through their ph domains [ 126 , 127 ] . nevertheless , both myh9 and the motor activity of nonmuscle myosin ii are crucial for lamellipodia formation , as they induce convex f - actin arcs at the leading edge of cells   ( figure 4 ) . nonmuscle cells express multiple myosin ii proteins , including myosin iia , myosin iib , and myosin iic [ 122124 ] ; these are implicated in regulating many cellular processes , including cell spreading , migration , and cytokinesis [ 128131 ] , through generating the intracellular contractile forces and tension as conventional motor proteins by associating with f - actin . thus , myosin iia might also play a crucial role in cell migration through lamellipodia formation by providing the contractile forces and tension for the f - actin network initially rearranged by wave2 , irsp53 , and pip3 through the variable turnover dynamics of f - actin [ 132 , 133 ] to form a convex arc at the leading edge of cells ( figure 4 ) . as irsp53 exhibits f - actin - bundling activity [ 134 , 135 ] , the formation of a lining of f - actin at the leading edge of cells might be mediated by nonmuscle myosin iia in cooperation with irsp53 .   among the proteins and molecules that are involved in regulation of lamellipodia formation , wave2 , n - wasp , pi3k , rac1 , stathmin , and microtubules are also necessary for cell invasion [ 69 , 136139 ] . because cell invasion is accomplished by intensive accumulation of the f - actin bundles at the tips of cell protrusions where cells invade through the basement membrane matrix and narrow gaps , the signaling and regulatory molecules leading to cell invasion may share many common molecules , leading to cell migration through lamellipodia formation . cell migration and invasion are believed to require the formation of lamellipodia at the leading edge of cells by rearrangement of f - actin . wave2 forms multiprotein complexes consisting of iqgap1 , kinesin-1 , pak1 , irsp53 , and nonmuscle myosin iia . membrane targeting of the wave2 complex is triggered by binding of iqgap1 and kinesin-1 , which are dissociated from the rac1-clip-170 complex upon pi3k activation by the activated growth factor receptor . concomitantly , wave2-bound pak1 after rac1-dependent activation phosphorylates stathmin , followed by binding of the stathmin - eb1 complex to the microtubule ends that bear the wave2 complex ; this results in the promotion of persistent microtubule growth towards the cell cortex . the wave2 complex that targets the cell membrane is anchored by wave2-bound irsp53 and nonmuscle myosin iia to pip3 ; pip3 is produced by pi3k near the growth factor receptor that is locally activated in the membrane region facing higher concentrations of growth factor . colocalization of wave2 , irsp53 , nonmuscle myosin iia , and pip3 induces the directional formation of a convex f - actin arc lamellipodium . the discovery of many essential signaling and regulatory molecules for cell migration might have important implications for the development of effective therapeutic agents against cancer cell migration and invasion .
e - cadherin is one of the key molecules in the formation of cell - cell adhesion and interacts intracellularly with a group of proteins collectively named catenins , through which the e - cadherin - catenin complex is anchored to actin - based cytoskeletal components . although cell - cell adhesion is often disrupted in cancer cells by either genetic or epigenetic alterations in cell adhesion molecules , disruption of cell - cell adhesion alone seems to be insufficient for the induction of cancer cell migration and invasion . 
 a small gtp - binding protein , rac1 , induces the specific cellular protrusions lamellipodia via wave2 , a member of wasp / wave family of the actin cytoskeletal regulatory proteins . 
 biochemical and pharmacological investigations have revealed that wave2 interacts with many proteins that regulate microtubule growth , actin assembly , and membrane targeting of proteins , all of which are necessary for directional cell migration through lamellipodia formation . 
 these findings might have important implications for the development of effective therapeutic agents against cancer cell migration and invasion .