The “small RNA revolution” 50 23 87 Over the following years, many new small functional RNAs have been found. RNA is usually thought of as messenger RNA that serves as the template for translation of genes into proteins. In contrast, functional or non-coding RNA molecules are transcribed from a DNA sequence, but not translated into protein. The encoding DNA sequence is often referred to as an RNA gene. Functional RNA genes in the human genome include transfer RNA (tRNA), ribosomal RNA (rRNA), and various other small non-coding RNAs. Several hundred genes in our genome encode small functional RNA molecules collectively called microRNAs (miRNAs). Precursors of these miRNA molecules form structures of double-stranded RNA that can activate the RNA interference machinery. MicroRNAs downregulate gene expression either by degradation of messenger RNA through the RNA interference pathway or by inhibiting protein translation. 42 Caenorhabditis elegans 42 lin-4 70 let-7 C. elegans lin-4 let-7 let-7 65 let-7 lin-4 let-7 Drosophila 38 40 41 38 40 41 38 40 41 48 71 66 75 46 68 1 1 mir mir-1 1 43 6 28 1 Fig. 1 a pri-miRNA pre-miRNA b c RISC  10 1 30 67 1 lin-4 9 33 42 7 17 59 60 8 11 MicroRNAs and cancer C. elegans Drosophila 9 42 14 12 24 52 MicroRNAs as causal cancer genes at genomic breakpoints 13 13 mir-15 mir-16 mir-17-92 27 63 let-7 32 The mir-17-92 cluster—small RNAs with oncogenic potential mir-17-92 64 27 mir-17-92 mir-17-92 27 mir-17-92 27 mir-17-92 mir-17-92 55 76 19 63 mir-17-92 mir-17-92 mir-17-92 MicroRNAs with tumor suppressor potential let-7 32 let-7 let-7 let-7 let-7 32 let-7 77 Global loss of miRNA expression in cancer 52 52 24 24 37 37 37 let-7 MicroRNAs in the p53 tumor suppressor network 16 26 26 16 Together, these data indicate that altered expression of miRNAs is not simply a secondary event that reflects the less differentiated state of cancer cells. In contrast, at least in some cases, miRNA expression is specifically driven by tumor suppressors and oncogenes. MicroRNAs with a role in tumor invasion and metastasis 54 54 Regulation of miRNAs in cancer—who regulates the regulators? 27 13 53 73 mir-124a mir-127 73 56 44 let-7 let-7 44 56 81 81 MicroRNA profiling—implications for cancer diagnosis 52 52 52 49 78 MicroRNAs—novel therapeutic targets? Regulatory RNAs may also have therapeutic applications by which disease-causing miRNAs could be antagonized or functional miRNAs restored. The most intuitive choice of molecules to correct altered miRNA–messenger RNA interactions are RNA oligonucleotides. These oligonucleotides need to be chemically modified to allow for stability in serum and cellular uptake. Modified antisense oligonucleotides are already being developed to utilize the intrinsic RNAi pathway for delivery of gene therapy. If the delivery problem can be overcome, then miRNA therapies may also be possible. O 29 57 36 36 21 21 83 83 One limitation of antisense RNA therapies is the restricted number of cells that can be targeted. Any approach to knock down a particular miRNA with antisense oligonucleotides will only result in partial knockdown. This may represent a limitation for cancer therapies. It remains to be seen whether indirectly mediated bystander effects on cancer cells that have not been directly targeted may partly overcome this limitation. In contrast, a partial effect on function may be of therapeutic value in neurodegenerative diseases, such as Parkinson’s or Alzheimer’s disease. A partial restoration of dopamine production by antisense therapy might result in a significant clinical improvement in Parkinson patients. Similarly, a partial reduction of the disease-causing proteins in Alzheimer’s disease may lead to a clinical improvement and might be achievable by RNA based or miRNA gene therapy. Techniques and approaches to study miRNAs 25 3 5 38 51 4 5 58 MicroRNA expression studies 42 84 15 60 62 82 52 52 79 31 35 61 Functional characterization of miRNAs 34 39 86 29 45 57 22 22 36 88 37 72 80 85 88 mir-155 72 80 72 80 85 88 MicroRNA target sites 2 20 3 10 69 74 10 47 49 10 18 55 56 76 Conclusions Over recent years, miRNAs have emerged as major players in the complex networks of gene regulation and have been implicated in various aspects of human disease. Only 5 years after the first study reported a direct involvement of miRNAs in cancer, these small RNAs have already significantly improved our understanding of carcinogenesis. In addition to protein-coding oncogenes and tumor suppressor genes, we will have to take into account miRNAs and their regulatory networks if we aim to understand the complex processes underlying malignant transformation.