any of the MMP inhibitor treatments, although GM6001 and MMP-2, MMP-2/29 inhibitors showed a slight reduction. All data shown are the mean 6 standard error of at least duplicate measurements and are representative of at least two independent experiments. (TIF) Table S1 Gelsolin modulates the expression of genes important for various cellular processes. Genes from the microarray output were classified under various biological pro- cesses using Gene Ontology annotation from DAVID bioinforma- tion resources. Average fold change in gene expression is References 1. Jemal A, Siegel R, Xu J, Ward E (2010) Cancer statistics, 2010. CA: a cancer journal for clinicians 60: 277–300. 2. Fearon ER, Vogelstein B (1990) A genetic model for colorectal tumorigenesis. Cell 61: 759–767. 3. Ashtonrickardt PG, Dunlop MG, Nakamura Y, Morris RG, Purdie CA, et al. (1989) High frequency of APC loss in sporadic colorectal carcinoma due to breaks clustered in 5Q21–22. Oncogene 4: 1169–1174. 4. Thiery JP (2002) Epithelial-mesenchymal transitions in tumour progression. Nat Rev Cancer 2: 442–454. 5. Nu¨ rnberg A, Kitzing T, Grosse R (2011) Nucleating actin for invasion. Nat Rev Cancer 11: 177–187. 6. Renoult C, Blondin L, Fattoum A, Ternent D, Maciver SK, et al. (2001) Binding of gelsolin domain 2 to actin. An actin interface distinct from that of gelsolin domain 1 and from ADF/cofilin. Eur J Biochem 268: 6165–6175. 7. Winston JS, Asch HL, Zhang PJ, Edge SB, Hyland A, et al. (2001) Downregulation of gelsolin correlates with the progression to breast carcinoma. Breast Cancer Res Treat 65: 11–21. 8. Dosaka-Akita H, Hommura F, Fujita H, Kinoshita I, Nishi M, et al. (1998) Frequent loss of gelsolin expression in non-small cell lung cancers of heavy smokers. Cancer Res 58: 322–327. 9. Tanaka H, Shirkoohi R, Nakagawa K, Qiao H, Fujita H, et al. (2006) siRNA gelsolin knockdown induces epithelial-mesenchymal transition with a cadherin switch in human mammary epithelial cells. Int J Cancer 118: 1680–1691. 10. Shieh DB, Godleski J, Herndon JE, Azuma T, Mercer H, et al. (1999) Cell motility as a prognostic factor in stage I nonsmall cell lung carcinoma - The role of gelsolin expression. Cancer 85: 47–57. 11. Visapaa H, Bui M, Huang Y, Seligson D, Tsai H, et al. (2003) Correlation of Ki- 67 and gelsolin expression to clinical outcome in renal clear cell carcinoma. Urology 61: 845–850. 12. Rao JY, David S, Visapaa H, Horvath S, Gaber F, et al. (2002) Alterations of cytoskeletal proteins Gelsolin and E-Cadherin as markers for bladder cancer progression: Tissue microarray analysis (TMA) in association with Ki67 and p53. Mod Pathol 15: 737. 13. Shieh DB, Chen IW, Wei TY, Shao CY, Chang HJ, et al. (2006) Tissue expression of gelsolin in oral carcinogenesis progression and its clinicopatholog- ical implications. Oral Oncol 42: 599–606. 14. Thor AD, Edgerton SM, Liu SQ, Moore DH, Kwiatkowski DJ (2001) Gelsolin as a negative prognostic factor and effector of motility in erbB-2-positive epidermal growth factor receptor-positive breast cancers. Clin Cancer Res 7: 2415–2424. 15. Witke W, Sharpe AH, Hartwig JH, Azuma T, Stossel TP, et al. (1995) Hemostatic, inflammatory, and fibroblast responses are blunted in mice lacking gelsolin. Cell 81: 41–51. 16. Arora PD, McCulloch CA (1996) Dependence of fibroblast migration on actin severing activity of gelsolin. J Biol Chem 271: 20516–20523. 17. De Corte V, Bruyneel E, Boucherie C, Mareel M, Vandekerckhove J, et al. (2002) Gelsolin-induced epithelial cell invasion is dependent on Ras-Rac signaling. EMBO J 21: 6781–6790. 18. Crowley J, Smith T, Fang Z, Takizawa N, Luna E (2009) Supervillin Reorganizes the Actin Cytoskeleton and Increases Invadopodial Efficiency. Mol Biol Cell: 948–962. 19. Van den Abbeele A, De Corte V, Van Impe K, Bruyneel E, Boucherie C, et al. (2007) Downregulation of gelsolin family proteins counteracts cancer cell invasion in vitro. Cancer Lett 255: 57–70. 20. Azuma T, Witke W, Stossel TP, Hartwig JH, Kwiatkowski DJ (1998) Gelsolin is a downstream effector of rac for fibroblast motility. EMBO J 17: 1362–1370. 21. Chellaiah M, Kizer N, Silva M, Alvarez U, Kwiatkowski D, et al. (2000) Gelsolin deficiency blocks podosome assembly and produces increased bone mass and strength. J cell biol 148: 665–678. 22. Chellaiah MA, Biswas RS, Yuen D, Alvarez UM, Hruska KA (2001) Phosphatidylinositol 3,4,5-trisphosphate directs association of Src homology 2- containing signaling proteins with gelsolin. J Biol Chem 276: 47434–47444. 23. Weaver AM (2006) Invadopodia: specialized cell structures for cancer invasion. Clin Exp Metastasis 23: 97–105. 24. Stylli SS, Kaye AH, Lock P (2008) Invadopodia: at the cutting edge of tumour invasion. J Clin Neurosci 15: 725–737. Gelsolin Induces Cancer Invasion via uPA Cascade determined from comparison between 4 gelsolin-overexpressing HCT116 clones and 2 vector-control HCT116 clones. (PDF) Author Contributions Conceived and designed the experiments: JLZ EHT CTY SM SK MST. Performed the experiments: JLZ EHT HKT LT YCL. Analyzed the data: JLZ EHT BY MJ CTY MST. Contributed reagents/materials/analysis tools: SM SCH. Wrote the paper: JLZ CTY. Review and critique of manuscript: CTY YCL YCG APK. 25. Tapia T, Ottman R, Chakrabarti R (2011) LIM kinase1 modulates function of membrane type matrix metalloproteinase 1: implication in invasion of prostate cancer cells. Mol Cancer 10: 6. 26. Clark ES, Weaver AM (2008) A new role for cortactin in invadopodia: regulation of protease secretion. Eur J Cell Biol 87: 581–590. 27. Webber MM, Waghray A (1995) Urokinase-mediated extracellular matrix degradation by human prostatic carcinoma cells and its inhibition by retinoic acid. Clin Cancer Res 1: 755–761. 28. Gay F, Estornes Y, Saurin JC, Joly-Pharaboz MO, Friederich E, et al. (2008) In colon carcinogenesis, the cytoskeletal protein gelsolin is down-regulated during the transition from adenoma to carcinoma. Hum Pathol 39: 1420–1430. 29. Porter RM, Holme TC, Newman EL, Hopwood D, Wilkinson JM, et al. (1993) Monoclonal antibodies to cytoskeletal proteins: an immunohistochemical investigation of human colon cancer. J Pathol 170: 435–440. 30. Brabletz T, Jung A, Reu S, Porzner M, Hlubek F, et al. (2001) Variable beta- catenin expression in colorectal cancers indicates tumor progression driven by the tumor environment. Proc Natl Acad Sci U S A 98: 10356–10361. 31. Tay PN, Tan P, Lan Y, Leung CH, Laban M, et al. (2010) Palladin, an actin- associated protein, is required for adherens junction formation and intercellular adhesion in HCT116 colorectal cancer cells. Int J Oncol 37: 909–926. 32. Van den Abbeele A, De Corte V, Van Impe K, Bruyneel E, Boucherie C, et al. (2007) Downregulation of gelsolin family proteins counteracts cancer cell invasion in vitro. Cancer Lett 255: 57–70. 33. Chou J, Stolz DB, Burke NA, Watkins SC, Wells A (2002) Distribution of gelsolin and phosphoinositol 4,5-bisphosphate in lamellipodia during EGF- induced motility. Int J Biochem Cell Biol 34: 776–790. 34. Andreasen PA, Kjøller L, Christensen L, Duffy MJ (1997) The urokinase-type plasminogen activator system in cancer metastasis: a review. Int J Cancer 72: 1– 22. 35. Kim MJ, Kim DH, Na HK, Surh YJ (2010) TNF-a induces expression of urokinase-type plasminogen activator and b-catenin activation through generation of ROS in human breast epithelial cells. Biochem Pharmacol 80: 2092–2100. 36. Katerinaki E, Evans GS, Lorigan PC, MacNeil S (2003) TNF-alpha increases human melanoma cell invasion and migration in vitro: the role of proteolytic enzymes. Br J Cancer 89: 1123–1129. 37. Jankun J, Skrzypczak-Jankun E (2001) Binding site of amiloride to urokinase plasminogen activator depends on species. Int J Mol Med 8: 365–371. 38. Delektorskaya VV, Golovkov DA, Kushlinskii NE (2008) Clinical significance of levels of molecular biological markers in zones of invasive front-line of colorectal cancer. Bull Exp Biol Med 146: 616–619. 39. Gaggioli C, Hooper S, Hidalgo-Carcedo C, Grosse R, Marshall JF, et al. (2007) Fibroblast-led collective invasion of carcinoma cells with differing roles for RhoGTPases in leading and following cells. Nat Cell Biol 9: 1392–1400. 40. Honda K, Yamada T, Hayashida Y, Idogawa M, Sato S, et al. (2005) Actinin-4 increases cell motility and promotes lymph node metastasis of colorectal cancer. Gastroenterology 128: 51–62. 41. Ambrosio EP, Rosa FE, Domingues MA, Villacis RA, Coudry RD, et al. (2011) Cortactin is associated with perineural invasion in the deep invasive front area of laryngeal carcinomas. Hum Pathol. 42. Vignjevic D, Schoumacher M, Gavert N, Janssen KP, Jih G, et al. (2007) Fascin, a novel target of beta-catenin-TCF signaling, is expressed at the invasive front of human colon cancer. Cancer Res 67: 6844–6853. 43. Chou J, Stolz DB, Burke NA, Watkins SC, Wells A (2002) Distribution of gelsolin and phosphoinositol 4,5-bisphosphate in lamellipodia during EGF- induced motility. Int J Biochem Cell Biol 34: 776–790. 44. Harvey SR, Sait SNJ, Xu Y, Bailey JL, Penetrante RM, et al. (1999) Demonstration of urokinase expression in cancer cells of colon adenocarcinomas by immunohistochemistry and in situ hybridization. Am J Pathol 155: 1115– 1120. 45. Grondahlhansen J, Ralfkiaer E, Kirkeby LT, Kristensen P, Lund LR, et al. (1991) Localization of urokinase-type plasminogen activator in stromal cells in adenocarcinomas of the colon in humans. Am J Pathol 138: 111–117. 46. Ramos-DeSimone N, Hahn-Dantona E, Sipley J, Nagase H, French DL, et al. 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