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RIBOSWITCHES
Abrar AhmedPh. D student, SKIMS
13-03-2013
Riboswitches
Regulatory segment of an mRNA molecule that binds a small molecule (ligand), resulting in a change in production of the proteins encoded by the mRNA.
•conserved genetic control elements in the 5’
untranslated region (UTR) of mRNAs.
• are regulated by metabolites: coenzymes (B1, B2,
B12), amino acids, purine bases.
• consist of only RNA.
• estimation: > 2% of all genes are regulated by
riboswitches.
Organization and properties of riboswitches
Natural aptamers are the basis of riboswitches
Organization and properties of riboswitches
Binding domain (Aptamer):
Regulatory Domain:(Expression platform)
• structured binding pocket
• high affinity:
• high specificity reacts upon ligand binding by the aptamer
comformational change alters gene expression
• Typically provide a feedback mechanism for controlling the expression of metabolic genes
metabolic product of a pathway inhibits expression of proteins required to produce the metabolite.
• RNA domains that modulate gene expression in response to metabolite bindingRiboswitches
Pre-Quinosine-I
2’-deoxyguanosine
Winkler & Breaker (2005) Annu Rev Microbiol 59: 487-517Roth, et al. (2007) Nat Struct Mol Biol. 14: 308-317Kim et al. (2007) Proc. Natl. Acad. Sci. 104: 16092-16097
Known riboswitch aptamers
Known riboswitch aptamers
• RFN element in mRNAs of genes required for biosynthesis of riboflavin and FMN
• FMN required for repression of the ribD operon of B. subtilis
• Mutations in the RFN element eliminate FMN-mediated repression of the operon
Regulation of riboflavin biosynthesis in B. subtilis
•Ronald Breaker et al. used in-line probing technique to prove
that RFN element is an aptamer that binds directly to FMN.
•In-line probing: The efficient hydrolysis of a phosphodiester bond
in RNA needs a 180 degree (“in-line”) arrangement among the
attacking nucliophile (water), the phosphorus atom in the
phosphodiester bond, and the leaving hydroxyl group at the end
of one of the RNA fragments created by the hydrolysis.
RNA analysis by in-line probing
ribD mRNA
The RFN element is a riboswitchRonald R Breaker (2002), PNAS 99, 15908-13
How riboswitches regulate gene expression?
Gene regulation by riboswitches:
-Transcription termination (FMN riboswitch)
-Translation inhibition (thiamine riboswitch)
- RNA cleavage (glmS riboswitch)
Transcription termination
Repression
Activation
ON OFF
The FMN riboswitch terminates transcription
Translation inhibition
Repression
Activation
ON OFF
thi operon of B. subtilis
RNA cleavage
Tom Cech, Nature (2004) 428: 263-264.
Cleavage of RNA by a riboswitchWinkler and Dann (2006), Nat Struct Mol Biol 13, 569-71
Summary• regulatory elements which control a wide set of metabolic pathways in
bacteria (mostly).
• consist solely of RNA
• Aptamer domain: high affinity and specific binding of metabolites
• Expression platform: regulation of gene expression by control of
- transcription termination
- translation initiation
- RNA stability
SIGNIFICANCE OF RIBOSWITCHES
Barrick & Breaker (2006) Scientific American 296: 50-57
Thanks