The RNA World Sidney Altman 1989 Nobel Laureate in Chemistry

6. The phrase "The RNA World" was coined by Walter Gilbert in 1986 in a commentary on the then recent observations of the catalytic properties of various RNAs. The RNA World referred to an hypothetical stage in the origin of life on Earth. During this stage, proteins were not yet engaged in biochemical reactions and RNA carried out both the information storage task of genetic information and the full range of catalytic roles necessary in a very primitive self-replicating system. Gilbert pointed out that neither DNA nor protein were required in such a primitive system if RNA could perform as a catalyst. At that time, it had only been demonstrated that RNA could cleave or ligate phosphodiester bonds. Nevertheless, as is a frequent occurrence in science, a general hypothesis was constructed from a few specific instances of a phenomenon. The RNA World Sidney Altman1989 Nobel Laureate in Chemistry

10. Cell. 1986 Sep 12;46(6):819-26 Major transcript of the frameshifted coxII gene from trypanosome mitochondria contains four nucleotides that are not encoded in the DNA Benne R, Van den Burg J, Brakenhoff JP, Sloof P, Van Boom JH, Tromp MC. The mitochondrial cytochrome oxidase (cox) subunit II gene from trypanosomes contains aframeshift at amino acid 170. This gene is highly conserved in different trypanosome species, suggesting that it is functional. Sequence determination of coxII transcripts of T. brucei and C. fasciculata reveals four extra, reading frame-restoring nucleotides at the frameshift position that are not encoded in the DNA. Southern blot analysis of DNA of both trypanosome species failed toshow the existence of a second version of the coxII gene. We conclude, therefore, that the extranucleotides are added during or after transcription of the frameshift gene by an RNA-editing process. Cell. 1989 May 5;57(3):355-66. RNA editing and the mitochondrial cryptogenes of kinetoplastid protozoa. Simpson L, Shaw J.

18. . 1989Jun;217(2-3):499-504Mol Gen Genet • Mol Gen Genet . 1979Jul 2;174(1):25-32 • Characterization of 10S RNA: a new stable rna molecule from Escherichia coli.Ray BK, Apirion D A precursor for a small stable RNA (10Sa RNA) of Escherichia coli.Subbarao MN, Apirion D

19. Science1996, 271, 990-3

21. RF-1 = UAG,UAA RF-2 = UGA,UAA RF-3 = RE\1\2

25. RNA interference mechanism and applications Micro RNA, RNAi, and SiRNA

26. Cell. 1993 Dec 3;75(5):843-54 The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14.Lee RC, Feinbaum RL, Ambros V.Harvard University, Department of Cellular and Developmental Biology, Cambridge, Massachusetts 02138. lin-4 is essential for the normal temporal control of diverse postembryonic developmental events in C. elegans. lin-4 acts by negatively regulating the level of LIN-14 protein, creating a temporal decrease in LIN-14 protein starting in the first larval stage (L1). We have cloned the C. elegans lin-4 locus by chromosomal walking and transformation rescue. We used the C. elegans clone to isolate the gene from three other Caenorhabditis species; all four Caenorhabditis clones functionally rescue the lin-4 null allele of C. elegans. Comparison of the lin-4 genomic sequence from these four species and site-directed mutagenesis of potential open reading frames indicated that lin-4 does not encode a protein. Two small lin-4 transcripts of approximately 22 and 61 nt were identified in C. elegans and found to contain sequences complementary to a repeated sequence element in the 3' untranslated region (UTR) of lin-14 mRNA, suggesting that lin-4 regulates lin-14 translation via an antisense RNA-RNA interaction.

30. NATURE | VOL 391 | 19 FEBRUARY 1998

31. miRNA genes • Most miRNA genes come from regions of the genome quite distant from previously annotated genes, implying they derive from different transcription units (TUs); • The miRNAs within a genomic cluster are often related to each other (but not always); • Not all of the cloned miRNAs are conserved even in very closely related animals, such as human/mouse, C. elegans/C. briggsae;

32. miRNA transcription and maturation For Metazoan miRNA: Nuclear gene to pri-miRNA(1); cleavage to miRNA precursor by Drosha RNaseIII(2); actively (5’-p, ~2nt 3’overhang) transported to cytoplasm by Ran-GTP/Exportin5 (3); loop cut by dicer(RNaseIII)(4); *duplex is generally short-lived, by Helicase to single strand RNA, forming RNA-Induced Silencing Complex, RISC/maturation (5-6).

33. The actions of small silencing RNA A, mRNA cleavage specified by a miRNA/siRNA; B, translational repression specified by miRNAs/siRNAs; C, transcriptional silencing, thought to be specified by heterochromatic siRNAs

37. Computational program to identify miRNA genes • Significant progress has been made in miRNA research since the report of the lin-4 RNA(1993). About 300 miRNAs have been identified in different organisms to date. • However, experimental identification miRNAs is still slow since some miRNAs are difficult to isolate by cloning due to low abundance /stability/ expression pattern/cloning procedure. Thus, computational identification of miRNAs from genomic sequences provide a valuable complement to cloning.

38. MiRNA: today Number of miRNA found: Computational approach experimentally validated Human:200-255 175 Nematods:103-120 100 Flies:96-124 77 miRNA functions Flies:control of cell proliferation, cell death and fat metabolism Nematods: neuronal patterning and embryonic development. Mammals: hematopoietic lineage differentiation. Plants: flower development, leaf development and axial meristem initiation