RESUMO
Raf-1 kinase inhibitor protein was initially discovered as a physiological kinase inhibitor of the MAPK signaling pathway and was later shown to suppress cancer cell invasion and metastasis. Yet, the molecular mechanism through which RKIP executes its effects is not completely defined. RhoA has both a pro- and anti-metastatic cell-context dependent functions. Given that Rho GTPases primarily function on actin cytoskeleton dynamics and cell movement regulation, it is possible that one way RKIP hinders cancer cell invasion/metastasis is by targeting these proteins. Here we show that RKIP inhibits cancer cell invasion and metastasis by stimulating RhoA anti-tumorigenic functions. Mechanistically, RKIP activates RhoA in an Erk2 and GEF-H1 dependent manner to enhance E-cadherin membrane localization and inhibit CCL5 expression.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/prevenção & controle , Regulação Neoplásica da Expressão Gênica , Proteína de Ligação a Fosfatidiletanolamina/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo , Animais , Apoptose , Biomarcadores Tumorais/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/secundário , Proliferação de Células , Feminino , Humanos , Camundongos , Proteína de Ligação a Fosfatidiletanolamina/genética , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto , Proteína rhoA de Ligação ao GTP/genéticaRESUMO
The role of RhoA GTPases in breast cancer tumorigenesis and metastasis is unclear. Early studies within which mutations in RhoA were designed based on cancer-associated mutations in Ras supported an oncogene role for RhoA. However, recent whole-genome sequencing studies of cancers raised the possibility that RhoA may have a tumor suppression function. Here, using a syngeneic triple negative breast cancer murine model we investigated the physiological effects of reduced RhoA expression on breast cancer tumorigenesis and metastasis. RhoA knockdown had no effect on primary tumor formation and tumor proliferation, concurring with our in vitro findings where reduced RhoA had no effect on breast cancer cell proliferation and clonogenic growth. In contrast, primary tumors with RhoA knockdown efficiently invaded sentinel lymph nodes and significantly metastasized to lungs compared to control tumors. Mechanistically, the current study demonstrated that this is achieved by promoting a pro-tumor microenvironment, with increased cancer-associated fibroblasts and macrophage infiltration, and by modulating the CCL5-CCR5 and CXCL12-CXCR4 chemokine axes in the primary tumor. To our knowledge, this is the first such mechanistic study in breast cancer showing the ability of RhoA to suppress chemokine receptor expression in breast tumor cells. Our work suggests a physiological lung and lymph node metastasis suppressor role for RhoA GTPase in breast cancer.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/patologia , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/secundário , Receptores CCR5/metabolismo , Receptores CXCR4/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo , Animais , Apoptose , Biomarcadores Tumorais/genética , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Proliferação de Células , Feminino , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Camundongos , Receptores CCR5/genética , Receptores CXCR4/genética , Transdução de Sinais , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto , Proteína rhoA de Ligação ao GTP/genéticaRESUMO
Recent high-throughput-sequencing of cancer genomes has identified oncogenic mutations in the B-Raf genetic locus as one of the critical events in melanomagenesis. B-Raf encodes a serine/threonine kinase that regulates the MAPK/ERK kinase (MEK) and extracellular signal-regulated kinase (ERK) protein kinase cascade. In normal cells, the activity of B-Raf is tightly regulated and is required for cell growth and survival. B-Raf gain-of-function mutations in melanoma frequently lead to unrestrained growth, enhanced cell invasion and increased viability of cancer cells. Although it is clear that the invasive phenotypes of B-Raf mutated melanoma cells are stringently dependent on B-Raf-MEK-ERK activation, the downstream effector targets that are required for oncogenic B-Raf-mediated melanomagenesis are not well defined. miRNAs have regulatory functions towards the expression of genes that are important in carcinogenesis. We observed that miR-10b expression correlates with the presence of the oncogenic B-Raf (B-RafV600E) mutation in melanoma cells. While expression of miR-10b enhances anchorage-independent growth of B-Raf wild-type melanoma cells, miR-10b silencing decreases B-RafV600E cancer cell invasion in vitro. Importantly, the expression of miR-10b is required for B-RafV600E-mediated anchorage independent growth and invasion of melanoma cells in vitro. Taken together our results suggest that miR-10b is an important mediator of oncogenic B-RafV600E activity in melanoma.