Nuclear localization of maspin is essential for its inhibition of tumor growth and metastasis

Abstract

Maspin (mammary serine protease inhibitor or SerpinB5) acts as a tumor suppressor when overexpressed in aggressive cancer cell lines. However, its role in human cancer is controversial. Maspin expression has been associated with a poor prognosis in some studies, whereas in others, with favorable outcome. The clinical data suggest, however, that nuclearlocalized maspin is associated with improved survival. We hypothesized that the tumor suppressor activity of maspin may require nuclear localization, and that the discordance between clinical and experimental reports is a consequence of the variable subcellular distribution of maspin. Furthermore, we surmized that nuclear maspin could function as a tumor suppressor through the regulation of genes involved in tumor growth and invasion. Maspin or maspin fused to a nuclear export signal were expressed in metastatic human breast and epidermoid carcinoma cell lines. We found that pan-cellular localized maspin inhibited in vivo tumor growth and metastasis when assessed in xenograft chicken embryo and murine mammary fat pad injection models. However, when maspin was excluded from the nucleus via a nuclear exclusion signal, it no longer functioned as a metastasis suppressor. Using chromatin immunoprecipitation, we show that nuclear maspin was enriched at the promoter of colony-stimulating factor-1 (CSF-1) and associated with diminished levels of CSF-1 mRNA. Our findings demonstrate that the nuclear localization of maspin is required for its tumor and metastasis suppressor functions in vivo, and suggest that its mechanism of action involves, in part, direct association of maspin with target genes. KEYWORDS: breast cancer; chromatin immunoprecipitation; maspin; metastasis; nuclear localization Maspin (mammary serine protease inhibitor or SerpinB5), a member of the serpin family of serine protease inhibitors, was identified in 1994 as a tumor and metastasis suppressor. 1Overexpression of maspin inhibits cell motility and invasion in vitro, and decreases tumor formation, metastasis and angiogenesis in vivo.2 Initially, secreted maspin was thought to inhibit migration and increase cell adhesion via interaction with b1-integrin and other extracellular matrix components. 2 However, the molecular mechanism of such activity has yet to be convincingly demonstrated, and remains the subject of debate. Indeed, the very existence of extracellular maspin has recently been questioned. 3 Maspin has also been detected in the nucleus. 2 Maspin expression has been characterized in different cancers, particularly in breast cancer. Several studies showed that maspin mRNA is reduced in primary tumors and undetectable in metastases. 4,5 Conversely, others have shown that increased maspin is associated with poor prognosis. 6,7 These conflicting observations might be explained by distinct subcellular localization of maspin in cancer cells. Indeed, recent studies indicate that nuclear maspin associates with well-differentiated phenotype and improved survival, whereas cytoplasmic maspin is associated...

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“…Maspin acts as a tumor suppressor when overexpressed in aggressive cancer cell lines and is thought to inhibit carcinoma invasion, metastasis and angiogenesis and induce apoptosis 10 , but its role in human cancer is controversial because in some studies maspin expression has been associated with a poor prognosis, whereas in others it is associated with a favorable outcome 22 . Some investigators found that intranuclear localization of maspin is associated with improved survival and emphasized that the tumor suppressor activity of maspin might require nuclear localization [22][23][24][25] .…”

“…Some investigators found that intranuclear localization of maspin is associated with improved survival and emphasized that the tumor suppressor activity of maspin might require nuclear localization [22][23][24][25] . In addition, Bettstetter and colleagues 26 found that elevated nuclear maspin expression was closely correlated with high microsatellite instability.…”

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“…Mammary serine protease inhibitor (maspin or SERPIN B5) is a multifaced protein able to induce apoptosis and suppress cell motility and metastasis [69,70,71,72,73,74]. The mechanisms by which maspin exert their functions are still under investigation but recent evidence suggest that the tumor suppressive and possibly the antimetastatic responses are associated with the nuclear localization of this protein [75]. It has been demonstrated that maspin physically associates with chromatin and functions as a histone-deacetylase (HDAC) inhibitor [75,76].…”

“…The mechanisms by which maspin exert their functions are still under investigation but recent evidence suggest that the tumor suppressive and possibly the antimetastatic responses are associated with the nuclear localization of this protein [75]. It has been demonstrated that maspin physically associates with chromatin and functions as a histone-deacetylase (HDAC) inhibitor [75,76]. The function of maspin as metastasis suppressor has been associated with the regulation of the urokinase-type plasminogen activator (uPA) and receptor (uPAR) protein system [77,78].…”