(Mela)statin’ the not so obvious: tumor suppressor hidden in intron

The complex molecular mechanisms involved in the development and progression of cutaneous malignant melanoma remain incompletely understood. Small 22-nucleotide microRNAs (miRNAs), which are becoming increasingly known to be associated with tumorigenesis, may now offer insight into the aggressively invasive behavior of melanoma. This new class of small RNAs targets mRNA turnover and translation thereby affecting proteins participating in cellular processes such as proliferation, apoptosis and differentiation. Alteration of the dynamics of these regulatory networks potentially contributes to malignant transformation and tumor progression. Nearly half of known human miRNA genes map to locations frequently linked to cancer, and a number of miRNAs were shown to act as tumor suppressors or oncogenes. Oncogenic miRNAs target mRNAs encoding proteins related to tumor suppression. Conversely, tumor suppressor activity of miRNAs could be a result of targeting mRNAs that encode proteins involved in tumor initiation and progression (Rovira et al., 2010). Two recently published studies have highlighted the role of miR-211 as a tumor suppressor in invasive melanoma (Levy et al., 2010; Mazar et al., 2010). miRNAs are located throughout the human genome and can be found in between genes (intergenic) or found located either within introns or exons of known genes (intronic). Intergenic miRNAs are generally transcribed by independent transcriptional regulatory components, and approximately half of catalogued miRNAs map to known intronic locations within protein-coding genes. Previous studies demonstrate that intronic miRNAs are usually expressed in coordination with their host gene mRNA, indicating that cotranscription from the host gene promoter is the most common process utilized for miRNA expression. However, 35% of intronic miRNAs have the ability to be transcribed independently of the host gene promoter, suggesting a possible explanation for why host gene mRNA and miRNA expression are not always parallel (Monteys et al., 2010). It has been proposed that cotranscribed intronic miRNAs offer a feedback mechanism such that a miRNA may target its host mRNA or control transcription factors, considered as ‘first-‘ or ‘second-order’ negative feedback (Hinske et al., 2010). This dynamic allows for a fine-tuning of the regulation of host genes. The loss or gain of function in the intronic miRNA dynamic may be an additional alteration in diseases, including cancer, ultimately contributing to a loss of feedback control systems and uncontrolled cell growth. The function of intronic cotranscribed miRNAs may not always be associated with their host gene. miR-211, located within an intron of the gene TRPM1, is one such example. In melanomas, TRPM1 (Melastatin) expression is decreased and was previously thought to act as a melanoma tumor suppressor. However, two recent studies point to miR-211, but not its host gene TRPM1, as a melanoma tumor suppressor. Levy et al. (2010) and Mazar et al. (2010) identified miR-211 as an important regulatory component of melanoma. Levy et al. (2010) sought to understand how decreased miR-211 signaling may play an independent role in melanoma invasiveness and metastasis. Screening a miRNA library transfected into a highly invasive melanoma cell line, they demonstrated that expression of miR-211 significantly decreased the invasiveness of the melanoma cells. Quantitative RT-PCR experiments with melanocytes and melanoma short-term cultures revealed that almost all malignant melanomas displayed decreased expression of TRPM1 and miR-211. Using two different cell populations of melanoma cultures, one with high expression of miR-211 and low invasive potential and vice versa, the expression of miR-211 was assessed. miR-211 function was either blocked using antagomirs or increased with mimics and expression of TRPM1 was blocked using siRNAs. Suppression of miR-211 resulted in an increase in the invasiveness of melanoma by 10-fold, while increasing miR-211 expression decreased the invasive potential by 2to 3-fold. Melanoma cell migration and other properties as a function of miR-211 and TRPM1 expression were analyzed. Decreasing miR-211 expression resulted in an increase in cell motility, migration and invasiveness. However, altering TRPM1 expression had no effect in either experiment. Using bioinformatics, predicted networks of genes expressed in melanoma based upon their roles in metastasis were identified. The most significant network included three central node genes, IGFR2, TGFBR2 and NFAT5, that exhibited a high degree of connectivity with other genes implicated in melanoma metastasis. miR-211 was shown to negatively regulate these genes, with TGFBR2 and NFAT5 (but not IGFR2) identified as direct targets of miR-211. Lastly, knocking down the central node genes in melanoma cells with high central node gene expression demonstrated an effect on melanoma invasive activity similar to increasing miR-211, suggesting that some of the tumor suppressor functions of miR-211 may be mediated by its regulation of IGFR2, TGFBR2, and NFAT5 (Figure 1). The tumor suppressor functions of miR-211 were also described by Mazar et al. (2010), who showed that the levels of miR-211 were consistently decreased in a majority of human melanoma cell lines and clinical samples when compared to normal melanocytes. Additionally, in melanoma cell lines with ectopic expression of miR-211, levels of predicted target mRNAs of miR-211 Coverage on: Mazar, J., Deyoung, K., Khaitan, D., Meister, E., Almodovar, A., Goydos, J., Ray, A., and Perera, R.J. (2010). The regulation of miRNA-211 expression and its role in melanoma cell invasiveness. PLoS ONE 5, e13779. Levy, C., Khaled, M., Iliopoulos, D., Janas, M.M., Schubert, S., Pinner, S., Chen, P.H., Li, S., Fletcher, A.L., Yokoyama, S., et al. (2010). Intronic miR-211 Assumes the Tumor Suppressive Function of Its Host Gene in Melanoma. Mol Cell 40, 841–849.