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A role for ezrin in a neglected metastatic tumor function
Update
TRENDS in Molecular Medicine
249
Vol.10 No.6 June 2004
| Research Focus Response
A role for ezrin in a neglected metastatic tumor function Stefano Fais Department of Drug Research and Evaluation, Pharmacogenetic, Drug Resistance and Experimental Therapeutic Section, Istituto Superiore di Sanita, 00161 Rome, Italy
In the previous issue of Trends in Molecular Medicine, Kent Hunter discussed the potential effects of ezrin in tumor metastasis [1]. In his article, two recent papers were described, which identified ezrin as a key component in the metastasis of pediatric tumors [2,3]. The speculation concerning the role of ezrin in tumor metastasis was based on both the best-known ezrin functions and the ability of ezrin to confer metastatic capabilities to lowmetastatic tumors in experimental models. However, the most recognized ezrin function is to connect membrane proteins to the actin cytoskeleton [4 – 6], whereas the tumor functions that are directly related to metastatic behaviour are the capacity of tumor cells to migrate within tissues, transmigrate through vessels and to adhere to the metastatic organs. Therefore, the linkage between ezrin and metastatic behaviour is through the adhesion molecules that are known to depend on the ezrin-mediated linkage to actin, such as CD44, and are directly related to the invasion and metastasis of tumors [7]. However, in a recent paper, another ezrin-mediated function that might influence metastatic behaviour, which is related to a neglected tumor activity, has been proposed [8]. The phagocyte-like behavior of tumor cells was first observed a century ago [9,10]. Numerous studies have since described the phagocytic activity of various murine and human tumor cells against dead cells and undefined particles [11,12]. However, oncologists have generally considered the finding of phagocytosed nuclei, cell debris and matrix components within tumor cells a curiosity and the result of an aberrant or an inefficient activity within the malignant cell. Using several quantitative measures of phagocytic activity in tumor cell lines that were derived from either metastatic lesions or primary tumors, it was reported that cell lines derived exclusively from metastatic tumors showed strong phagocytic activity. On the basis of previous evidence on the essential role of ezrin in the phagocytic process of macrophages [13], the role of ezrin on the phagocytic activity of human tumor cells was examined. This data showed that ezrin is localized within the phagocytic vacuoles (Figure 1), together with lysosomal antigens [8]. Furthermore, by using oligo-antisense inhibition to block the synthesis of ezrin, the phagocytic activity of metastatic tumor cells was abrogated. However, the ezrin levels were comparable in both metastatic and non-metastatic tumors, suggesting that the overexpression of ezrin was not the sole cause of this phenomenon. In Corresponding author: Stefano Fais ([email protected]). www.sciencedirect.com
fact, contrasting results on the level of ezrin in adult tumors have been reported [14,15]. Therefore, it appears that a mechanism that is involved in ezrin activation and phosphorylation becomes deregulated in tumor cells, particularly in adult tumors. Rho-GTPases, which are known to be involved in ezrin– radixin – moesin (ERM) activation, are good candidates for the aberrant isoforms that are overexpressed in tumor metastasis [16], and Rho activation might induce aberrant phagocytic activity in non-professional phagocytes [17]. The role of phagocytic activity of tumors in metastasis needs to be clarified. Two major hypotheses can be proposed: (i) the ability of tumors to phagocytose might contribute to tumor escape from immune surveillance through the subtraction of antigenic material that is required for the processing activity of professional antigen-presenting cells, or (ii) a more complex hypothesis, in which the phagocytic activity allows tumors to survive in adverse environments, such as those that occur during hypoxic conditions because of the low blood supply during tumor growth or in metastatic organs, by ‘feeding’ on apoptotic cells and/or extracellular matrix components.
TRENDS in Molecular Medicine
Figure 1. Immunocytochemistry of a melanoma cell phagocytosing a latex bead. Note the precise localization of the ezrin staining (red) around the phagocytosed bead (indicated by the arrow), although ezrin staining is also spread within the melanoma-cell cytoplasm.
250
Update
TRENDS in Molecular Medicine
Ezrin is also involved in other functions, the alteration of which might be important in tumor malignancy. It is known that tumors are resistant to apoptotic stimuli, including those triggering Fas-mediated apoptosis [18]. Recent evidence suggests that the Fas– ezrin linkage and, in turn, the Fas – actin connection and the Fas-mediated polarization of a cell, are crucial requirements for rendering a cell prone to Fas-mediated apoptosis [19 – 21]. This suggests that an altered ezrin– Fas connection might contribute to the Fas-related resistance of tumors. Furthermore, ezrin is connected to P-glycoprotein [22], which regulates the efflux pumps that are responsible for some of the multidrug resistance mechanisms of tumors [23]. This suggests that ezrin overexpression and/or deregulation contribute to the development of efflux-pump-mediated resistance of tumors to cytotoxic agents. Therefore, it is conceivable that ezrin overexpression, or at least abnormal ezrin regulation, might affect many cellular functions that, together, contribute to the metastatic behaviour of tumors. References 1 Hunter, K.W. et al. (2004) Ezrin, a key component in tumor metastasis. Trends Mol. Med. 10, 201– 204 2 Khanna, C. et al. (2004) The membrane-cytoskeleton linker ezrin is necessary for osteosarcoma metastasis. Nat. Med. 10, 182 – 186 3 Yu, Y. et al. (2004) Expression profiling identifies the cytoskeletal organizer ezrin and the developmental homeoprotein Six-1 as key metastatic regulators. Nat. Med. 10, 175 – 181 4 Bretscher, A. (1999) Regulation of cortical structure by the ezrin – radixin – moesin protein family. Curr. Opin. Cell Biol. 11, 109 – 116 5 Gautreau, A. et al. (2002) ERM proteins and NF2 tumor suppressor: the Yin and Yang of cortical actin organization and cell growth signaling. Curr. Opin. Cell Biol. 1, 104– 109 6 Fais, S. and Malorni, W. (2003) Leukocyte uropod formation and membrane/cytoskeleton linkage in immune interactions. J. Leukoc. Biol. 73, 556– 563 7 Martin, T.A. et al. (2003) The role of the CD44/ezrin complex in cancer metastasis. Crit. Rev. Oncol. Hematol. 46, 165– 186 8 Lugini, L. et al. (2003) Potent phagocytic activity discriminate
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metastatic and primary human malignant melanoma: a key role of ezrin. Lab. Invest. 83, 1555 – 1567 Steinhaus, J. (1891) Ueber carcinom-einschlu¨sse. Virchows Arch. 126, 533 – 535 Stroebe, H. (1892) Zur kenntniss verschiedener cellula¨rer. Vorga¨nge und Erscheinungen in Geschwu¨lten. Beitr. Pathol. 11, 1 – 9 Marin-Padilla, M. (1977) Erythrophagocytosis by epithelial cells of brest carcinoma. Cancer 39, 1085– 1089 DeSimone, P.A. et al. (1980) Phagocytic tumor cell activity in oat cell carcinoma of the lung. Hum. Pathol. 11, 535– 539 Defacque, H. et al. (2000) Involvement of Ezrin/Moesin in de novo actin assembly on phagosomal membranes. EMBO J. 19, 199 – 212 Moilanen, J. et al. (2003) Ezrin immunoreactivity in relation to survival in serous ovarian carcinoma patients. Gynecol. Oncol. 90, 273– 281 Makitie, T. et al. (2001) Ezrin as a prognostic indicator and its relationship to tumor characteristics in uveal malignant melanoma. Invest. Ophthalmol. Vis. Sci. 42, 2442 – 2449 Clark, E.A. et al. (2000) Genomic analysis of metastasis reveals an essential role for RhoC. Nature 406, 532 – 535 Fiorentini, C. et al. (2001) Activation of Rho GTPases by cytotoxic necrotizing factor 1 induces macropinocytosis and scavenging activity in epithelial cells. Mol. Biol. Cell 12, 2061– 2073 Townson, J.L. et al. (2003) The role of apoptosis in tumor progression and metastasis. Curr. Mol. Med. 3, 631 – 642 Parlato, S. et al. (2000) CD95 (APO-1/Fas) linkage to the actin cytoskeleton through ezrin in human T lymphocytes: a novel regulatory mechanism of the CD95 apoptotic pathway. EMBO J. 19, 5123– 5134 Lozupone, F. et al. (2004) Identification and relevance of the CD95binding domain in the N-terminal region of Ezrin. J. Biol. Chem. 279, 9199– 9207 Luciani, F. et al. (2004) CD95/phosphorylated ezrin association underlies HIV-1 GP120/IL-2-induced susceptibility to CD95(APO-1/ Fas)-mediated apoptosis of human resting CD4þT lymphocytes. Cell Death Differ. 11, 574– 582 Luciani, F. et al. (2002) P-glycoprotein/actin association through ERM family proteins: a role in P-glycoprotein function in human cells of lymphoid origin. Blood 99, 641– 648 Gottesman, M.M. and Pastan, I. (1993) Biochemistry of multidrug resistance mediated by the multidrug transporter. Annu. Rev. Biochem. 62, 385– 427
1471-4914/$ - see front matter q 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.molmed.2004.04.005
TRENDS in Molecular Medicine
249
Vol.10 No.6 June 2004
| Research Focus Response
A role for ezrin in a neglected metastatic tumor function Stefano Fais Department of Drug Research and Evaluation, Pharmacogenetic, Drug Resistance and Experimental Therapeutic Section, Istituto Superiore di Sanita, 00161 Rome, Italy
In the previous issue of Trends in Molecular Medicine, Kent Hunter discussed the potential effects of ezrin in tumor metastasis [1]. In his article, two recent papers were described, which identified ezrin as a key component in the metastasis of pediatric tumors [2,3]. The speculation concerning the role of ezrin in tumor metastasis was based on both the best-known ezrin functions and the ability of ezrin to confer metastatic capabilities to lowmetastatic tumors in experimental models. However, the most recognized ezrin function is to connect membrane proteins to the actin cytoskeleton [4 – 6], whereas the tumor functions that are directly related to metastatic behaviour are the capacity of tumor cells to migrate within tissues, transmigrate through vessels and to adhere to the metastatic organs. Therefore, the linkage between ezrin and metastatic behaviour is through the adhesion molecules that are known to depend on the ezrin-mediated linkage to actin, such as CD44, and are directly related to the invasion and metastasis of tumors [7]. However, in a recent paper, another ezrin-mediated function that might influence metastatic behaviour, which is related to a neglected tumor activity, has been proposed [8]. The phagocyte-like behavior of tumor cells was first observed a century ago [9,10]. Numerous studies have since described the phagocytic activity of various murine and human tumor cells against dead cells and undefined particles [11,12]. However, oncologists have generally considered the finding of phagocytosed nuclei, cell debris and matrix components within tumor cells a curiosity and the result of an aberrant or an inefficient activity within the malignant cell. Using several quantitative measures of phagocytic activity in tumor cell lines that were derived from either metastatic lesions or primary tumors, it was reported that cell lines derived exclusively from metastatic tumors showed strong phagocytic activity. On the basis of previous evidence on the essential role of ezrin in the phagocytic process of macrophages [13], the role of ezrin on the phagocytic activity of human tumor cells was examined. This data showed that ezrin is localized within the phagocytic vacuoles (Figure 1), together with lysosomal antigens [8]. Furthermore, by using oligo-antisense inhibition to block the synthesis of ezrin, the phagocytic activity of metastatic tumor cells was abrogated. However, the ezrin levels were comparable in both metastatic and non-metastatic tumors, suggesting that the overexpression of ezrin was not the sole cause of this phenomenon. In Corresponding author: Stefano Fais ([email protected]). www.sciencedirect.com
fact, contrasting results on the level of ezrin in adult tumors have been reported [14,15]. Therefore, it appears that a mechanism that is involved in ezrin activation and phosphorylation becomes deregulated in tumor cells, particularly in adult tumors. Rho-GTPases, which are known to be involved in ezrin– radixin – moesin (ERM) activation, are good candidates for the aberrant isoforms that are overexpressed in tumor metastasis [16], and Rho activation might induce aberrant phagocytic activity in non-professional phagocytes [17]. The role of phagocytic activity of tumors in metastasis needs to be clarified. Two major hypotheses can be proposed: (i) the ability of tumors to phagocytose might contribute to tumor escape from immune surveillance through the subtraction of antigenic material that is required for the processing activity of professional antigen-presenting cells, or (ii) a more complex hypothesis, in which the phagocytic activity allows tumors to survive in adverse environments, such as those that occur during hypoxic conditions because of the low blood supply during tumor growth or in metastatic organs, by ‘feeding’ on apoptotic cells and/or extracellular matrix components.
TRENDS in Molecular Medicine
Figure 1. Immunocytochemistry of a melanoma cell phagocytosing a latex bead. Note the precise localization of the ezrin staining (red) around the phagocytosed bead (indicated by the arrow), although ezrin staining is also spread within the melanoma-cell cytoplasm.
250
Update
TRENDS in Molecular Medicine
Ezrin is also involved in other functions, the alteration of which might be important in tumor malignancy. It is known that tumors are resistant to apoptotic stimuli, including those triggering Fas-mediated apoptosis [18]. Recent evidence suggests that the Fas– ezrin linkage and, in turn, the Fas – actin connection and the Fas-mediated polarization of a cell, are crucial requirements for rendering a cell prone to Fas-mediated apoptosis [19 – 21]. This suggests that an altered ezrin– Fas connection might contribute to the Fas-related resistance of tumors. Furthermore, ezrin is connected to P-glycoprotein [22], which regulates the efflux pumps that are responsible for some of the multidrug resistance mechanisms of tumors [23]. This suggests that ezrin overexpression and/or deregulation contribute to the development of efflux-pump-mediated resistance of tumors to cytotoxic agents. Therefore, it is conceivable that ezrin overexpression, or at least abnormal ezrin regulation, might affect many cellular functions that, together, contribute to the metastatic behaviour of tumors. References 1 Hunter, K.W. et al. (2004) Ezrin, a key component in tumor metastasis. Trends Mol. Med. 10, 201– 204 2 Khanna, C. et al. (2004) The membrane-cytoskeleton linker ezrin is necessary for osteosarcoma metastasis. Nat. Med. 10, 182 – 186 3 Yu, Y. et al. (2004) Expression profiling identifies the cytoskeletal organizer ezrin and the developmental homeoprotein Six-1 as key metastatic regulators. Nat. Med. 10, 175 – 181 4 Bretscher, A. (1999) Regulation of cortical structure by the ezrin – radixin – moesin protein family. Curr. Opin. Cell Biol. 11, 109 – 116 5 Gautreau, A. et al. (2002) ERM proteins and NF2 tumor suppressor: the Yin and Yang of cortical actin organization and cell growth signaling. Curr. Opin. Cell Biol. 1, 104– 109 6 Fais, S. and Malorni, W. (2003) Leukocyte uropod formation and membrane/cytoskeleton linkage in immune interactions. J. Leukoc. Biol. 73, 556– 563 7 Martin, T.A. et al. (2003) The role of the CD44/ezrin complex in cancer metastasis. Crit. Rev. Oncol. Hematol. 46, 165– 186 8 Lugini, L. et al. (2003) Potent phagocytic activity discriminate
www.sciencedirect.com
Vol.10 No.6 June 2004
9 10 11 12 13
14
15
16 17
18 19
20
21
22
23
metastatic and primary human malignant melanoma: a key role of ezrin. Lab. Invest. 83, 1555 – 1567 Steinhaus, J. (1891) Ueber carcinom-einschlu¨sse. Virchows Arch. 126, 533 – 535 Stroebe, H. (1892) Zur kenntniss verschiedener cellula¨rer. Vorga¨nge und Erscheinungen in Geschwu¨lten. Beitr. Pathol. 11, 1 – 9 Marin-Padilla, M. (1977) Erythrophagocytosis by epithelial cells of brest carcinoma. Cancer 39, 1085– 1089 DeSimone, P.A. et al. (1980) Phagocytic tumor cell activity in oat cell carcinoma of the lung. Hum. Pathol. 11, 535– 539 Defacque, H. et al. (2000) Involvement of Ezrin/Moesin in de novo actin assembly on phagosomal membranes. EMBO J. 19, 199 – 212 Moilanen, J. et al. (2003) Ezrin immunoreactivity in relation to survival in serous ovarian carcinoma patients. Gynecol. Oncol. 90, 273– 281 Makitie, T. et al. (2001) Ezrin as a prognostic indicator and its relationship to tumor characteristics in uveal malignant melanoma. Invest. Ophthalmol. Vis. Sci. 42, 2442 – 2449 Clark, E.A. et al. (2000) Genomic analysis of metastasis reveals an essential role for RhoC. Nature 406, 532 – 535 Fiorentini, C. et al. (2001) Activation of Rho GTPases by cytotoxic necrotizing factor 1 induces macropinocytosis and scavenging activity in epithelial cells. Mol. Biol. Cell 12, 2061– 2073 Townson, J.L. et al. (2003) The role of apoptosis in tumor progression and metastasis. Curr. Mol. Med. 3, 631 – 642 Parlato, S. et al. (2000) CD95 (APO-1/Fas) linkage to the actin cytoskeleton through ezrin in human T lymphocytes: a novel regulatory mechanism of the CD95 apoptotic pathway. EMBO J. 19, 5123– 5134 Lozupone, F. et al. (2004) Identification and relevance of the CD95binding domain in the N-terminal region of Ezrin. J. Biol. Chem. 279, 9199– 9207 Luciani, F. et al. (2004) CD95/phosphorylated ezrin association underlies HIV-1 GP120/IL-2-induced susceptibility to CD95(APO-1/ Fas)-mediated apoptosis of human resting CD4þT lymphocytes. Cell Death Differ. 11, 574– 582 Luciani, F. et al. (2002) P-glycoprotein/actin association through ERM family proteins: a role in P-glycoprotein function in human cells of lymphoid origin. Blood 99, 641– 648 Gottesman, M.M. and Pastan, I. (1993) Biochemistry of multidrug resistance mediated by the multidrug transporter. Annu. Rev. Biochem. 62, 385– 427
1471-4914/$ - see front matter q 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.molmed.2004.04.005