1. Mestrado Integrado em Medicina Biologia Celular e Molecular II 2009-2010

2. What is a microRNA?

3. RNA Coding RNA mRNA Non-Coding RNA Transcription RNAs tRNA rRNA Small RNAssiRNA microRNA snoRNAsnRNA

4. siRNA stands for a short interference RNA (therefore acting under the RNAi pathway) which is similar to the miRNA, catalysing the cleaveage of target mRNA under a post- transcriptional gene silencing mechanism; snoRNA acts on the modification of rRNA; snRNA is involved in splicing procedures of mRNA.

5. microRNAs (miRNAs) are a family of small non-coding RNAs with 21-25 nucleotides; negative gene regulators; They act as negative gene regulators; They are encoded by genes that are transcribed from DNA but that are not translated into proteins (non-coding RNA); Mature miRNA molecules are partially complementary to one or more messenger RNA (mRNA) molecules;

6. From the gene to the functional miRNA

8.  miRNAs are transcribed as a long RNA molecule called Pri-miRNA;

9.  Pri-miRNAs are processed in the nucleus into Pre-miRNAs by: ◦ a double-stranded RNA specific ribonuclease called Drosha; ◦ a double-stranded RNA binding protein called Pasha;

10.  The Pre-miRNAs are transported to the cytoplasm by an exportin 5-dependent mechanism;

11.  In the cytoplasm, the Pre-miRNAs are digested by another double-stranded RNA specific ribonuclease called Dicer;  The process results in a sequence-specific, single stranded, mature miRNA molecule.

12. The different action mechanisms

13. microRNAs act on a post- transcriptional level, negatively regulating gene expression; They generally target the 3’-UTR domain of the mRNA that they specifically regulate.

14.  The resulting miRNA is bound by a complex that is similar to the RNA-Induced Silencing Complex (RISC) that participates in RNA interference (RNAi)

15.  The complex-bound, single-stranded miRNA binds specific mRNAs through sequences;  These sequences are significantly (but not completely) complementary to the mRNA;

16.  The bound mRNA remains untranslated, resulting in reduced expression of the corresponding gene;

17. Regulation of cell proliferation and death

18. TS – Tumor Suppression | OG - Oncogene Recent evidence indicated that miRNAs can act as tumor supressours and oncogenes, being called oncomirs Expression profiling of miRNAs has been proved to be an accurate method to classify cancer subtypes and develop therapeutic approaches

19. Low levels or defective biogenesis of miRNA responsible for inhibiting oncogenes may lead to tumor formation; Other influences underlying decrease of apoptosis level, angiogenesis or de-differentiation processes may also lead to tumors.

20. The overexpression of a miRNA whose function would be to inhibit a tumor suppressor related translation process could also lead to tumor formation The question marks represent stages of miRNA biogenesis where the cause for its increased levels may be located.

21. In the future, anti-miRNA oligonucleotides (AMOs) that encode sequences fully complementary to miRNA may modulate their function and act as tumor suppressants (example)

22. A few key points concerning microRNAs

23. negative gene regulators; microRNAs (miRNAs) are a family of small non-coding RNAs (21-25 nucleotides) that act as negative gene regulators; miRNA biogenesis comprehends the processing of a primary DNA transcript by many enzyme complexes (such as Drosha and Dicer); The relation between miRNA and Cancer is yet to be totally explained but may hold the key to major therapeutic breakthroughs.

25. References and Sources

26.  Cooper, G. et al. (2007): "The Cell", 3 rd Edition  Chen, C. (2006): "microRNAs and the Post-transcriptional generic programs controlled by small RNAs"  Esquela-Kerscher, A. and Slack, F. (2006): "Oncomirs – microRNAs with a role in cancer", Nature Publications  He, L. and Hannon, J. (2006): "Small RNAs with a big role in gene regulation", Nature Publications  Chen, K. and Rajewsky, N. (2006): "The evolution of gene regulation", Nature Publications  Kim, V. (2005): "MicroRNA biogenesis: coordinated cropping and dicing", Nature Publications  Internet (as of March 2008): ◦ http://microrna.sanger.ac.uk/sequences/ http://microrna.sanger.ac.uk/sequences/ ◦ http://en.wikipedia.org/wiki/MicroRNA http://en.wikipedia.org/wiki/MicroRNA ◦ http://www.rosettagenomics.com/ http://www.rosettagenomics.com/