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Topic: A natural metabolite-responsive ribozyme
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Matthew J. Brauer
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Member # 819
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posted 19. March 2004 13:41
From the March 18 2004 Nature
"Control of gene expression by a natural metabolite-responsive ribozyme."
Winkler, W.C., Nahvi, A., Collins, J.A., Breaker, R. R.
Abstract:
Most biological catalysts are made of protein; however, eight classes of natural ribozymes have been discovered that catalyse fundamental biochemical reactions. The central functions of ribozymes in modern organisms support the hypothesis that life passed through an 'RNA world' before the emergence of proteins and DNA. We have identified a new class of ribozymes that cleaves the messenger RNA of the glmS gene in Gram-positive bacteria. The ribozyme is activated by glucosamine-6-phosphate (GlcN6P), which is the metabolic product of the GlmS enzyme. Additional data indicate that the ribozyme serves as a metabolite-responsive genetic switch that represses the glmS gene in response to rising GlcN6P concentrations. These findings demonstrate that ribozyme switches may have functioned as metabolite sensors in primitive organisms, and further suggest that modern cells retain some of these ancient genetic control systems.
Tom Cech's News & Views article:
RNA finds a simpler way
Is there no end to the versatility of RNA? The latest feat to be revealed is RNA's ability to switch off genes through a neatly straightforward mechanism. So it isn't only proteins that can repress gene activity.
Organisms profit from synthesizing only those enzymes whose products are in demand. After all, if there's plenty of an amino acid around, why should a cell waste energy making the enzymes that are needed to generate it from precursors? A standard textbook mechanism by which this is achieved is repression. Repressor proteins sense when there is a build-up of a certain product, bind to the gene that encodes the enzyme that generates this product, and thereby inhibit the transcription of more messenger RNA (mRNA; Fig. 1a). On page 281 of this issue, Winkler et al.1 describe a bacterial gene-regulation system with a twist. The repressor is not a protein, but instead is a switchable (on–off) self-cleavage element within the mRNA itself.
The newly discovered molecular switch involves an RNA molecule with enzymatic activity. Self-cleavage by this ribozyme is accelerated 1,000-fold in the presence of glucosamine-6-phosphate (GlcN6P), a small sugar. GlcN6P is generated by the GlmS enzyme, which is encoded by a portion of the glmS mRNA downstream from the ribozyme sequence. So it is easy to envisage a gene-regulatory circuit in which the glmS mRNA is translated into GlmS protein until the GlcN6P product accumulates; at that point, GlcN6P binds to the special catalytic element in the mRNA, causing it to self-destruct (Fig. 1b). Although the cleavage event itself leaves the coding region of the message intact, the RNA is either destabilized and subject to degradation, or its ability to be translated into functional protein is compromised.
The minimal region of the RNA that can confer this regulatory activity is roughly 75 nucleotides long, the size of a transfer RNA. The chemical mechanism by which it is cleaved during repression has previously been seen in other ribozymes, such as the hammerhead, hairpin and hepatitis delta virus ribozymes2, but the structure of its catalytic fold appears to be distinct from these. When placed upstream of an un-related 'reporter' gene, the glmS ribozyme element also repressed its expression, and this required the same sequences that are required for glmS self-cleavage in vitro1. Thus, the active RNA element is modular and transplantable.
Although switching gene expression in this manner is new, there are precedents for the individual components of this regulatory circuit. First, consider GlcN6P binding. Even though it once seemed unlikely that RNA, with its limited diversity of chemical groups and its high negative charge, could specifically bind small molecules, we now know that some naturally occurring RNAs do just that. For example, a particular group of ribozymes forms a pocket that binds guanosine monophosphate, one of the four monomer building-blocks of RNA3. And a specific region of RNA from the human immunodeficiency virus binds a derivative of the amino acid arginine4. More recently, short (<100 nucleotides) RNA 'aptamers' have been identified that specifically bind everything from hydrophobic amino acids to small organic molecules to metal ions5, 6. In terms of specificity, an RNA aptamer can even distinguish the plant alkaloid theophylline from the closely related molecule caffeine7.
As a second precedent, aptamers found within some natural mRNAs bind small molecules as part of gene-regulatory feedback circuits. In the bacterium Escherichia coli, coenzyme B12 binds directly to, and thereby represses translation of, the mRNA coding for the protein that transports its precursor, cobalamin8. In Bacillus species, the synthesis of thiamin and riboflavin involves discrete genetic units. These operons are controlled by direct binding of thiamin pyrophosphate and flavin mononucleotide to leader sequences of the corresponding mRNAs, resulting in transcription coming to an early finish9. Although structured RNAs had previously been found to regulate gene expression at multiple levels — during gene transcription, splicing of the RNA and the translation of mRNA into protein — these new systems are different in that they consist entirely of RNA. No protein seems to be required for either sensing the concentration of the metabolic end-product or switching off gene expression.
Finally, several groups had previously engineered artificial riboswitches — RNA aptamers containing ribozyme sequences that, on binding small molecules, induce ribozyme-mediated cleavage of the RNA10. These findings might have prompted some to wonder why nature failed to use, or perhaps to retain, such an elegant mechanism. But it's clear now that this perhaps-ancient talent of RNA is alive and well, and is currently used by at least some bacteria to control their glmS genes. An exciting question is how widespread these control elements are in biology.
In their eloquent and prescient article on genetic regulatory mechanisms, François Jacob and Jacques Monod11 thought that genetic repressors might be RNA. They nonetheless much preferred the idea of a protein repressor, as, in their own words "the capacity to form stereospecific complexes with small molecules appears to be a privilege of proteins". When many repressors were subsequently identified as proteins, the idea that RNA repressors existed faded away. But more recently, the concept that RNA can in fact bind small molecules with high specificity and affinity has been established. In a sense, the recent discoveries of naturally occurring 'RNA biosensors' have gone full-circle back to Jacob and Monod — RNA can be the active element that switches off repressible genes in response to the concentration of a cellular metabolite.
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Pim van Meurs
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Member # 541
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posted 20. March 2004 15:03
I repeat the relevant sentence
These findings demonstrate that ribozyme switches may have functioned as metabolite sensors in primitive organisms, and further suggest that modern cells retain some of these ancient genetic control systems.
Suggesting that function switching or gain of function may be far more common that sometimes speculated. This also indicates that the definition of IC in which function is maintained, may not be relevant to evolutionary pathways.
quote:
In the postulated “RNA world” of 3.6 billion years ago, RNA was able to both store information and carry out chemical reactions, and life forms would not have used DNA or proteins, said Lilley. “You can get a very simple but crude genetic system that can evolve, and so you can build more complexity,” he said.
It was possible to imagine that these RNAs started recruiting extra molecules, rather as enzymes use cofactors in the modern world, he added. “The advantages of amino acids were such that they kind of took over, and probably the finest creation of this RNA world was, in fact, proteins.” Once there, it was possible very rapidly to get up to a modern, protein-based world, said Lilley.
Riboswitch ribozyme
[ 20. March 2004, 15:04: Message edited by: Pim van Meurs ]
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