Raf kinase inhibitory protein (RKIP) negatively regulates the MAP kinase (MAPK), G protein‐coupled receptor kinase‐2, and NF‐κB signalling cascades. RKIP has been implicated as a metastasis suppressor for prostate cancer, but the mechanism is not known. Here, we show that RKIP inhibits invasion by metastatic breast cancer cells and represses breast tumour cell intravasation and bone metastasis in an orthotopic murine model. The mechanism involves inhibition of MAPK, leading to decreased transcription of LIN28 by Myc. Suppression of LIN28 enables enhanced let‐7 processing in breast cancer cells. Elevated let‐7 expression inhibits HMGA2, a chromatin remodelling protein that activates pro‐invasive and pro‐metastatic genes, including Snail. LIN28 depletion and let‐7 expression suppress bone metastasis, and LIN28 restores bone metastasis in mice bearing RKIP‐expressing breast tumour cells. These results indicate that RKIP suppresses invasion and metastasis in part through a signalling cascade involving MAPK, Myc, LIN28, let‐7 , and downstream let‐7 targets. RKIP regulation of two pluripotent stem cell genes, Myc and LIN28, highlights the importance of RKIP as a key metastasis suppressor and potential therapeutic agent.

Increased proliferation and elevated levels of protein synthesis are characteristics of transformed and tumor cells. Though components of the translation machinery are often misregulated in cancers, what role tRNA plays in cancer cells has not been explored. We compare genome-wide tRNA expression in cancer-derived versus non-cancer-derived breast cell lines, as well as tRNA expression in breast tumors versus normal breast tissues. In cancer-derived versus non-cancer-derived cell lines, nuclear-encoded tRNAs increase by up to 3-fold and mitochondrial-encoded tRNAs increase by up to 5-fold. In tumors versus normal breast tissues, both nuclear- and mitochondrial-encoded tRNAs increase up to 10-fold. This tRNA over-expression is selective and coordinates with the properties of cognate amino acids. Nuclear- and mitochondrial-encoded tRNAs exhibit distinct expression patterns, indicating that tRNAs can be used as biomarkers for breast cancer. We also performed association analysis for codon usage-tRNA expression for the cell lines. tRNA isoacceptor expression levels are not geared towards optimal translation of house-keeping or cell line specific genes. Instead, tRNA isoacceptor expression levels may favor the translation of cancer-related genes having regulatory roles. Our results suggest a functional consequence of tRNA over-expression in tumor cells. tRNA isoacceptor over-expression may increase the translational efficiency of genes relevant to cancer development and progression.

The ten–eleven translocation (TET) family of methylcytosine dioxygenases initiates demethylation of DNA and is associated with tumorigenesis in many cancers; however, the mechanism is mostly unknown. Here we identify upstream activators and downstream effectors of TET1 in breast cancer using human breast cancer cells and a genetically engineered mouse model. We show that depleting the architectural transcription factor high mobility group AT-hook 2 ( HMGA2 ) induces TET1 . TET1 binds and demethylates its own promoter and the promoter of homeobox A ( HOXA ) genes, enhancing its own expression and stimulating expression of HOXA genes including HOXA7 and HOXA9 . Both TET1 and HOXA9 suppress breast tumor growth and metastasis in mouse xenografts. The genes comprising the HMGA2–TET1–HOXA9 pathway are coordinately regulated in breast cancer and together encompass a prognostic signature for patient survival. These results implicate the HMGA2–TET1–HOX signaling pathway in the epigenetic regulation of human breast cancer and highlight the importance of targeting methylation in specific subpopulations as a potential therapeutic strategy.

Tumour metastasis suppressors are inhibitors of metastasis but their mechanisms of action are generally not understood. We previously showed that the suppressor Raf kinase inhibitory protein (RKIP) inhibits breast tumour metastasis in part via let‐7. Here, we demonstrate an integrated approach combining statistical analysis of breast tumour gene expression data and experimental validation to extend the signalling pathway for RKIP. We show that RKIP inhibits let‐7 targets (HMGA2, BACH1) that in turn upregulate bone metastasis genes (MMP1, OPN, CXCR4). Our results reveal BACH1 as a novel let‐7‐regulated transcription factor that induces matrix metalloproteinase1 (MMP1) expression and promotes metastasis. An RKIP pathway metastasis signature (designated RPMS) derived from the complete signalling cascade predicts high metastatic risk better than the individual genes. These results highlight a powerful approach for identifying signalling pathways downstream of a key metastasis suppressor and indicate that analysis of genes in the context of their signalling environment is critical for understanding their predictive and therapeutic potential.

Transfer RNAs (tRNAs) are typically considered housekeeping products with little regulatory function. However, several studies over the past 10 years have linked tRNA misregulation to cancer. We have previously reported that tRNA levels are significantly elevated in breast cancer and multiple myeloma cells. To further investigate the cellular and physiological effects of tRNA overexpression, we overexpressed tRNA_i^Met in two human breast epithelial cell lines. We then determined tRNA abundance changes and performed phenotypic characterization. Overexpression of tRNA_i^Met significantly altered the global tRNA expression profile and resulted in increased cell metabolic activity and cell proliferation. Our results extend the relevance of tRNA overexpression in human cells and underscore the complexity of cellular regulation of tRNA expression.