Cell migration is considered necessary for the invasion that accompanies the directional formation of the cellular protrusions termed lamellipodia. In invasive breast cancer MDA-MB-231 cells, lamellipodia formation is preceded by translocation of the actin cytoskeletal regulatory protein WAVE2 to the leading edge. WAVE2 translocation and lamellipodia formation require many signaling molecules, including PI3K, Rac1, Pak1, IRSp53, stathmin, and EB1, but whether these molecules are necessary for invasion remains unclear. In noninvasive breast cancer MCF7 cells, no lamellipodia were induced by IGF-I, whereas in MDA-MB-231 cells, Rac1, stathmin, and EB1 were overexpressed. Depletion of Rac1 or stathmin by small interfering RNA abrogated the IGF-I-induced invasion of MDA-MB-231 cells; however, depletion of EB1 did not, indicating the necessity of Rac1 and stathmin but not EB1 for invasion. The signaling pathway leading to cell invasion may not be identical but shares some common molecules, leading to cell migration through lamellipodia formation. 1. Introduction The formation of cellular protrusions such as lamellipodia at the leading edge of migrating cells is regulated by WASP/WAVE family of the actin cytoskeletal regulatory protein WAVE2 [1–3]. Before lamellipodia formation, WAVE2 is translocated to the leading edge along microtubules [4–6], which is mediated by many signaling and regulatory molecules. WAVE2 forms a complex with IQGAP1, the motor protein kinesin1 [6, 7], Pak1 [8], and IRSp53 [9] in the cytoplasm of quiescent cells and gathers additional IQGAP1 and kinesin1 [6], which are released from the Rac1-CLIP-170 complex [7], after stimulation of cells with HGF or IGF-I. Concomitantly, WAVE2-bound Pak1 is Rac1-dependently activated, which in turn inactivates stathmin, a microtubule-destabilizing protein [10, 11], by phosphorylation [8]. Stathmin is constitutively associated with the microtubule-end-binding protein EB1 [12], and the phosphorylated stathmin-EB1 complex is recruited to the microtubule ends that bear the WAVE2 complex after IGF-I stimulation [8]. Following translocation to the leading edge, WAVE2 is captured by PtdInsP3 through WAVE2-bound IRSp53 [13]. PtdInsP3 is produced by PI3K near the IGF-I receptor IGF-IR that is locally activated in the membrane region facing IGF-I [13]. These results indicate that many signaling and regulatory molecules, including IGF-IR, PI3K, Rac1, Pak1, IRSp53, stathmin, and EB1, are involved in inducing the directional lamellipodia formation in migrating cells. However, whether these molecules, except for
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