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Discovering the Power of RNase P: A Multi-Faceted Tool in Molecular Biology

The biography of Ribonuclease P (RNase P) traces its discovery by Altman in 1971, through the isolation and purification phases, to identifying its RNA and protein subunits. This ribonuclease and ribozyme aid in cleaving extraneous sequences from pre-tRNAs, essential for generating mature tRNAs. RNase P, conserved in RNA and protein across domains of life, plays a crucial role in processing precursor tRNAs. Its mechanism of action involves pre-tRNA binding to RNase P's domains, with potential applications in antibacterial, antiviral, and anticancer therapeutics. By targeting specific mRNAs with guide sequences, such as External Guide Sequences (EGS) and Internal Guide Sequences (IGS) linked to M1RNA, RNase P can induce mRNA cleavage. This technique has shown promise in gene knockdown studies, RNA interference, and cancer research, highlighting RNase P's versatility as a powerful molecular tool.

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Discovering the Power of RNase P: A Multi-Faceted Tool in Molecular Biology

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  1. The Biography of Ribonuclease P Marta Wegorzewska Macromolecules 5.7.09 www.physorg.com/news144947904.html

  2. Discovery: pre-tRNA 1971: Precursor t-RNA Sidney Altman www.oisb.ca/.../pic_members_Sidney_Altman.jpg Altman et al., 1971

  3. 41 nt at 5’ end Discovery: 3 nt at 3’ end Isolation and purification of pre-tRNA Altman et al., 1971

  4. Discovery: -----------------------------------------> RNase P Evans et al., 2006 Nuclear extracts of E.coli --------> Altman et al., 1971

  5. Discovery: purification of RNase P 1972: Purifying RNase P Actions of purified RNase P Robertson et al., 1972

  6. Discovery: RNase P: RNA-protein mix 1978: Benjamin Stark (graduate student) identified the RNA and protein subunit of E.coli RNase P M2: methylene blue staining for nucleic acid stain C5: Coomassie brilliant blue staining for protein Stark et al., 1978

  7. Discovery: RNase P: a ribonuclease and ribozyme 1983: Cecilia Guerrier-Takada 6= E.coli RNA + protein 7= E.coli RNA (M1 RNA) 8= E.coli protein (C5) Pre-tRNA Mature tRNA Guerrier-Takada et al., 1983

  8. Conservation: RNA Evans et al., 2006

  9. Conservation: Protein Eukarya Archaea Bacteria Hartmann et al., 2003

  10. Summary: Bacteria Archaea Eukarya E.coli M1 RNA; type A (ancestral) B. sub type B (Bacillus) RNA- Protein- H. sap. H1 RNA M. the. one RNA subunit H. sap. 10 proteins: hPop1, Rpp29, Rpp21, Rpp30, hPop5, Rpp14, Rpp20, Rpp25, Rpp40, Rpp38 E.coli one protein: C5 B. sub one protein: P protein M. the. 4 proteins: Pop4, Rpr2, Rpp1, Pop5 Hall et al., 2002

  11. Function: ribonuclease RNase P functions to remove extraneous 5' sequences from precursor tRNAs to generate mature tRNA Rnase P, Mg2+ www.science.ca/images/altman_rnase.jpg Evans et al., 2006

  12. Mechanism of Action: Pre-tRNA binding to S and C domains of RNase P RNA subunit RNase P protein subunit binding to 5’ end of pre-tRNA

  13. Function: Human Rnase P: a transcription factor RNase P protein found at 5S rRNA, 7SL RNA and U6 snRNA genes (non-RNase P substrate genes) and tRNA genes (RNase P substrate genes) Jarrous et al., 2007

  14. Potential as a tool: potential as a antibacterial, antiviral and anticancer agent Cobaleda et al., 2001 Can we take advantage of the catalytic function of M1RNA to target bacterial, viral, oncogenic mRNAs??

  15. Guide sequences (GS): External guide sequences (EGS): exogenous GS recruits endogenous RNase P Internal guide sequences (IGS): GS covalently linked to M1RNA (M1GS) Cobaleda et al., 2001 Requirements: Complementary to target mRNA 3’ sequence for recognition by M1 RNA (EGS)

  16. Application: Anti-bacterial potential: Guerrier-Takada et al. 1995 showed specific targeting of EGS to B-galactosidase and alkaline phosphatase encoding genes (expressions were decreased by 50-60% in E.coli) Anti-viral potential: IGS and EGS used to target herpes simplex virus 1 (HSV-1), human immunodeficiency virus (HIV), human influenza virus, human cytomegalovirus and Kaposi's sarcoma-associated herpesvirus Anti-cancer potential: M1GS used for destruction of chimeric mRNAs created by chromosomal translocation (BCR-ABL) Cobaleda et al., 2001

  17. Application: Anti-cancer potential: Ba/F3 cells expressing the human BCR-ABLp190 + M1GS against BCR-ABL p190 Cobaledo et al., 2000

  18. Application: advantages for basic science research This sounds awesome! Why don’t hear about it as a tool for used in gene knockdown studies? RNA interference Advantages of EGS/M1GS: EGS uses endogenous RNase P (most abundant, stable and efficient enzymes) resulting in irreversible cleavage of target mRNA Highly specific and does not mistarget (RNA i) Little sign of cytotoxicity

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