Ren L, Hong SH, Cassavaugh J, Osborne T, Chou AJ, Kim SY, Gorlick R, Hewitt SM, Khanna CThe actin-cytoskeleton linker protein ezrin is regulated during osteosarcoma metastasis by PKC. Oncogene 28: 792-802

Tumor and Metastasis Biology Section, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
Oncogene (Impact Factor: 8.46). 02/2009; 28(6):792-802. DOI: 10.1038/onc.2008.437
Source: PubMed


Ezrin is a member of the ERM (ezrin, radixin, moesin) protein family and links F-actin to the cell membrane following phosphorylation. Ezrin has been associated with tumor progression and metastasis in several cancers including the pediatric solid tumors, osteosarcoma and rhabdomyosarcoma. In this study, we were surprised to find that ezrin was not constitutively phosphorylated but rather was dynamically regulated during metastatic progression in osteosarcoma. Metastatic osteosarcoma cells expressed phosphorylated ERM early after their arrival in the lung, and then late in progression, only at the invasive front of larger metastatic lesions. To pursue mechanisms for this regulation, we found that inhibitors of PKC (protein kinase C) blocked phosphorylation of ezrin, and that ezrin coimmunoprecipitated in cells with PKCalpha, PKCiota and PKCgamma. Furthermore, phosphorylated forms of ezrin and PKC had identical expression patterns at the invasive front of pulmonary metastatic lesions in murine and human patient samples. Finally, we showed that the promigratory effects of PKC were linked to ezrin phosphorylation. These data are the first to suggest a dynamic regulation of ezrin phosphorylation during metastasis and to connect the PKC family members with this regulation.

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Available from: Tanasa S Osborne, Sep 22, 2015
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    • "The phosphorylation step stabilizes ezrin binding to the actin cytoskeleton and restricts its binding to specialized membrane domains [15] where the active ezrin assembles and integrates signaling molecules to exert diverse downstream effects [58]. Importantly, while some of the functional studies observed that it was the phosphorylated form of ezrin to drive metastasis [42-45,47,48,51], the underlying mechanism is still unknown. "
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    ABSTRACT: Extracellular matrix (ECM) degradation is a critical process in tumor cell invasion and requires matrix degrading protrusions called invadopodia. The Na(+)/H(+) exchanger (NHE1) has recently been shown to be fundamental in the regulation of invadopodia actin cytoskeleton dynamics and activity. However, the structural link between the invadopodia cytoskeleton and NHE1 is still unknown. A candidate could be ezrin, a linker between the NHE1 and the actin cytoskeleton known to play a pivotal role in invasion and metastasis. However, the mechanistic basis for its role remains unknown. Here, we demonstrate that ezrin phosphorylated at T567 is highly overexpressed in the membrane of human breast tumors and positively associated with invasive growth and HER2 overexpression. Further, in the metastatic cell line, MDA-MB-231, p-ezrin was almost exclusively expressed in invadopodia lipid rafts where it co-localized in a functional complex with NHE1, EGFR, ß1-integrin and phosphorylated-NHERF1. Manipulation by mutation of ezrins T567 phosphorylation state and/or PIP2 binding capacity or of NHE1s binding to ezrin or PIP2 demonstrated that p-ezrin expression and binding to PIP2 are required for invadopodia-mediated ECM degradation and invasion and identified NHE1 as the membrane protein that p-ezrin regulates to induce invadopodia formation and activity.
    PLoS ONE 09/2013; 8(9):e75113. DOI:10.1371/journal.pone.0075113 · 3.23 Impact Factor
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    • "The presence of numerous actin-rich cellular protrusions, in which ezrin is localized in pCB6 colonies, is consistent with the previously reported localization of ezrin in invasive cancers [45,46]. In contrast, Y477F ezrin colonies showed no actin-rich protrusions, indicating a loss of invasive function; while strong actin staining in the cortical region of cells was evident. "
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    ABSTRACT: The membrane cytoskeletal crosslinker, ezrin, a member of the ERM family of proteins, is frequently over-expressed in human breast cancers, and is required for motility and invasion of epithelial cells. Our group previously showed that ezrin acts co-operatively with the non-receptor tyrosine kinase, Src, in deregulation of cell-cell contacts and scattering of epithelial cells. In particular, ezrin phosphorylation on Y477 by Src is specific to ezrin within the ERM family, and is required for HGF-induced scattering of epithelial cells. We therefore sought to examine the role of Y477 phosphorylation in ezrin on tumor progression. Using a highly metastatic mouse mammary carcinoma cell line (AC2M2), we tested the effect of over-expressing a non-phosphorylatable form of ezrin (Y477F) on invasive colony growth in 3-dimensional Matrigel cultures, and on local invasion and metastasis in an orthotopic engraftment model. AC2M2 cells over-expressing Y477F ezrin exhibited delayed migration in vitro, and cohesive round colonies in 3-dimensional Matrigel cultures, compared to control cells that formed invasive colonies with branching chains of cells and numerous actin-rich protrusions. Moreover, over-expression of Y477F ezrin inhibits local tumor invasion in vivo. Whereas orthotopically injected wild type AC2M2 tumor cells were found to infiltrate into the abdominal wall and visceral organs within three weeks, tumors expressing Y477F ezrin remained circumscribed, with little invasion into the surrounding stroma and abdominal wall. Additionally, Y477F ezrin reduces the number of lung metastatic lesions. Our study implicates a role of Y477 ezrin, which is phosphorylated by Src, in regulating local invasion and metastasis of breast carcinoma cells, and provides a clinically relevant model for assessing the Src/ezrin pathway as a potential prognostic/predictive marker or treatment target for invasive human breast cancer.
    BMC Cancer 03/2012; 12(1):82. DOI:10.1186/1471-2407-12-82 · 3.36 Impact Factor
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    • "Phosphorylated Ezrin is thought to play a role in early-stage metastasis by connecting with the target organ site and therefore is thought to be an ineffective treatment strategy as its function is completed once a metastasis is established [2]. However, Ren et al. discovered Ezrin phosphorylation is also present at the leading edge of large metastatic lesions [75]. Targeting Ezrin could be promising for managing lung metastases; however, we should shift the focus from targeting one protein to identifying the complexity of multiple protein interactions to improve the efficacy of treatment. "
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    ABSTRACT: Osteosarcoma is the most common primary bone cancer of children and is established during stages of rapid bone growth. The disease is a consequence of immature osteoblast differentiation, which gives way to a rapidly synthesized incompletely mineralized and disorganized bone matrix. The mechanism of osteosarcoma tumorogenesis is poorly understood, and few proteomic studies have been used to interrogate the disease thus far. Accordingly, these studies have identified proteins that have been known to be associated with other malignancies, rather than being osteosarcoma specific. In this paper, we focus on the growing list of available state-of-the-art proteomic technologies and their specific application to the discovery of novel osteosarcoma diagnostic and therapeutic targets. The current signaling markers/pathways associated with primary and metastatic osteosarcoma that have been identified by early-stage proteomic technologies thus far are also described.
    Sarcoma 03/2012; 2012:169416. DOI:10.1155/2012/169416
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