Toca-1 is suppressed by p53 to limit breast cancer cell invasion and tumor metastasis

Abstract

IntroductionTransducer of Cdc42-dependent actin assembly-1 (Toca-1) recruits actin regulatory proteins to invadopodia, and promotes breast tumor metastasis. Since metastatic breast tumors frequently harbor mutations in the tumor suppressor p53, we tested whether p53 regulates Toca-1 expression.MethodsNormal mammary epithelial cells (HBL-100, MCF10A) and breast cancer cell lines expressing wild-type (WT) p53 (DU4475, MTLn3) were treated with camptothecin or Nutlin-3 to stabilize p53 to test effects on Toca-1 mRNA and protein levels. Chromatin immunoprecipitation (ChIP) assays were performed to identify p53 binding site in Toca-1 gene. Stable silencing of p53 and Toca-1 were performed in MTLn3 cells to test effects on invadopodia and cell invasion in vitro, and tumor metastasis in vivo.ResultsWe observed that breast cancer cell lines with mutant p53 have high levels of Toca-1 compared to those with WT p53. Stabilization of WT p53 led to further reduction in Toca-1 mRNA and protein levels in normal breast epithelial cells and breast cancer cells. ChIP assays revealed p53 binding within intron 2 of toca1, and reduced histone acetylation within its promoter region upon p53 upregulation or activation. Stable silencing of WT p53 in MTLn3 cells led to increased extracellular matrix degradation and cell invasion compared to control cells. Interestingly, the combined silencing of p53 and Toca-1 led to a partial rescue of these effects of p53 silencing in vitro and reduced lung metastases in mice. In human breast tumors, Toca-1 levels were high in subtypes with frequent p53 mutations, and high Toca-1 transcript levels correlated with increased risk of relapse.ConclusionsBased on these findings, we conclude that loss of p53 tumor suppressor function in breast cancers leads to upregulation of Toca-1, and results in enhanced risk of developing metastatic disease.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-014-0503-x) contains supplementary material, which is available to authorized users.

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“…Although there are published data suggesting a role for TOCA-1 in endocytosis ( Bu et al , 2009 ), there is much stronger evidence that this protein plays a role in other aspects of plasma membrane dynamics, such as neurite elongation and filopodia formation ( Kakimoto et al , 2006 ; Bu et al , 2009 ; Chander et al , 2014 ). Given that epithelial cells in a monolayer shuffle and slide with respect to each other ( Timpe et al , 1978 ), we tested the possibility that TOCA-1 might play a role in tight junction membrane contact dynamics associated with normal cell movement.…”

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