N-WASP-mediated invadopodium formation is involved in intravasation and lung metastasis of mammary tumors

Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Journal of Cell Science (Impact Factor: 5.43). 02/2012; 125(Pt 3):724-34. DOI: 10.1242/jcs.092726
Source: PubMed


Invadopodia are proteolytic membrane protrusions formed by highly invasive cancer cells, commonly observed on substrate(s) mimicking extracellular matrix. Although invadopodia are proposed to have roles in cancer invasion and metastasis, direct evidence has not been available. We previously reported that neural Wiskott-Aldrich syndrome protein (N-WASP), a member of WASP family proteins that regulate reorganization of the actin cytoskeleton, is an essential component of invadopodia. Here, we report that N-WASP-mediated invadopodium formation is essential in breast cancer invasion, intravasation and lung metastasis. We established stable cell lines based on MTLn3 rat mammary adenocarcinoma cells that either overexpressed a dominant-negative (DN) N-WASP construct or in which N-WASP expression was silenced by a pSuper N-WASP shRNA. Both the N-WASP shRNA and DN N-WASP cells showed a markedly decreased ability to form invadopodia and degrade extracellular matrix. In addition, formation of invadopodia in primary tumors and collagen I degradation were reduced in the areas of invasion (collagen-rich areas in the invasive edge of the tumor) and in the areas of intravasation (blood-vessel-rich areas). Our results suggest that tumor cells in vivo that have a decreased activity of N-WASP also have a reduced ability to form invadopodia, migrate, invade, intravasate and disseminate to lung compared with tumor cells with parental N-WASP levels.

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Available from: Bojana Gligorijevic,
    • "Invadopodia presence was shown in vitro in 2D conditions, in 3D environment [18] and in vivo. Tumor cryosections from mice mammary carcinoma were used to show the presence of the invadopodial markers actin and cortactin accompanied by collagen degradation in the protrusions of cells that are adjacent to the tumor edge and to tumor blood vessels [2] [12] [19] [20]. Other recent work demonstrated that invadopodia Contents lists available at ScienceDirect journal homepage: www.elsevier.com/locate/yexcr "
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    ABSTRACT: Invadopodiaareactin-richprotrusionsformedbymesenchymallymigratingcancercells.Theyaremainly composed ofactin,actin-associatedproteins,integrinsandproteinsofsignalingmachineries.These protrusionsdisplayfocalizedproteolyticactivitytowardstheextracellularmatrix.Itiswellknownthat polymerized (F-)actinispresentinthesestructures,butthenatureoftheactinisoformhasnotbeen studied before.Wehereshowthatbothcytoplasmicactinisoforms, β- and γ-actin, arepresentinthe invadopodiaofMDA-MB-231breastcancercellsculturedona2D-surface,wheretheycolocalizewiththe invadopodialmarkercortactin.Invadopodialstructuresformedbythecellsina3D-collagenmatrixalso contain β- and γ-actin. Wedemonstratethisusingisoform-specific antibodiesandexpressionof fluor- escently-taggedactinisoforms.Additionally,usingsimultaneousexpressionofdifferentiallytagged β- and γ-actin incells,weshowthattheactinisoformsarepresenttogetherinasingleinvadopodium.Cells with anincreasedlevelof β- or γ-actin, displayasimilarincreaseinthenumberandsizeofinvadopodia in comparisontocontrolcells.Moreover,increasingthelevelofeitheractinisoformsalsoincreases invasionvelocity.
    Experimental Cell Research 11/2015; DOI:10.1016/j.yexcr.2015.11.003 · 3.25 Impact Factor
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    • "Some data also support the existence of podosome-like structures within tissue collected from mice and fixed without culturing (Quintavalle et al., 2010). To dissect the function of podosomes/invadopodia in vivo, several groups disrupted their formation by genetically manipulating the genes that are crucial for their assembly, such as Tks5 (Blouw et al., 2008; Murphy et al., 2011), N-WASP (Gligorijevic et al., 2012), and Arg kinase (Gil-Henn et al., 2013). Nevertheless, evidence of podosome function in vivo is still scarce. "

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    • "As in 2D, how the cellular localization of invadopodia is determined in 3D is unknown. However, invadopodia are composed of actin, cortactin, cofilin, Tks5, and MT1-MMP in both 2D and 3D cultures and in vivo settings (Blouw et al., 2008; Lizarraga et al., 2009; Magalhaes et al., 2011; Gligorijevic et al., 2012; Yu Machesky, 2012). "
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    ABSTRACT: The leading cause of death in cancer patients is metastasis. Invasion is an integral part of metastasis and is carried out by proteolytic structures called invadopodia at the cellular level. In this introductory review, we start by evaluating the definition of invadopodia. While presenting the upstream signaling events involved, we integrate current models on invadopodia. In addition, we discuss the significance of invadopodia in 2D and 3D and in vivo. We finally point out technical challenges and conclude with open questions in the field.
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