Transcript The Biography of Ribonuclease P

The Biography of Ribonuclease P
Marta Wegorzewska
Macromolecules
5.7.09
www.physorg.com/news144947904.html
Discovery: pre-tRNA
Sidney Altman
1971: Precursor t-RNA
www.oisb.ca/.../pic_members_Sidney_Altman.jpg
Altman et al., 1971
41 nt at 5’ end
Discovery:
3 nt at 3’ end
Isolation and purification of
pre-tRNA
Altman et al., 1971
Discovery:
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Evans et al., 2006
RNase P
Nuclear
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extracts
of E.coli
Altman et al., 1971
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Discovery: purification of
1972:
Purifying RNase P
Robertson et al., 1972
RNase P
Actions of purified RNase P
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
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
Conservation: RNA
Evans et al., 2006
Bacteria
Conservation: Protein
Hartmann et al., 2003
Summary:
Bacteria
RNA-
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Protein-
Hall et al., 2002
E.coli
M1 RNA; type A
(ancestral)
B. sub
type B (Bacillus)
E.coli
one protein:
C5
B. sub
one protein: P
protein
Archaea
M. the.
one RNA
subunit
Eukarya
H. sap.
H1 RNA
H. sap.
M. the.
4 proteins:
Pop4, Rpr2,
Rpp1, Pop5
10 proteins:
hPop1, Rpp29,
Rpp21, Rpp30,
hPop5, Rpp14,
Rpp20, Rpp25,
Rpp40, Rpp38
Function: ribonuclease
RNase P functions to remove extraneous
5' sequences from precursor tRNAs
to generate mature tRNA
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Rnase P, Mg2+
www.science.ca/images/altman_rnase.jpg
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Evans et al., 2006
Mechanism of Action:
1. Pre-tRNA binding to
S and C domains of
RNase P RNA
subunit
2. RNase P protein
subunit binding to 5’
end of pre-tRNA
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
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??
Guide sequences (GS):
External guide sequences
(EGS): exogenous GS recruits
endogenous RNase P
Requirements:
Cobaleda et al., 2001
1. Complementary to target
mRNA
2. 3’ sequence for recognition
by M1 RNA (EGS)
Internal guide sequences
(IGS): GS covalently linked to
M1RNA (M1GS)
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 sarcomaassociated herpesvirus
Anti-cancer potential: M1GS used
for destruction of chimeric mRNAs
created by chromosomal translocation
(BCR-ABL)
Cobaleda et al., 2001
Application:
Anti-cancer potential:
Ba/F3 cells
expressing
the human
BCRABLp190
+
M1GS against BCR-ABL p190
Cobaledo et al., 2000
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:
1. EGS uses endogenous RNase P (most abundant,
stable and efficient enzymes) resulting in
irreversible cleavage of target mRNA
2. Highly specific and does not mistarget (RNA i)
3. Little sign of cytotoxicity
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