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Transcriptional homeostasis: a mechanism of protein quality control
Medical Hypotheses (2004) 63, 232–234
http://intl.elsevierhealth.com/journals/mehy
Transcriptional homeostasis: a mechanism of protein quality control Richard Sallie* Western Gastroenterology, Department of Hepatology, St. John of God Hospital, Suite 35, 95 Monash Avenue, Nedlands, Perth, Western Australia 6009, Australia Received 5 February 2004; accepted 19 February 2004
Summary Sickle cell anaemia confirms that minor mutations in protein sequence can have catastrophic effects. However, RNA transcription depends on polymerases that have low fidelity and introduce errors at a rate of 10 5 substitutions/base synthesised. For many large proteins translation of these errors could result in significant loss of function, while for others (for example, cell surface signalling proteins) variability of protein expression could be advantageous. This paper outlines a mechanism that enables proteins to modulate error incorporation (variability) into RNAs sysnthesised by RNA polymerase – transcriptional homeostasis – thereby modulating, and error correcting, their own sysnthesis. Transcriptional homeostasis is a fundamental regulatory mechanism relevant to control of gene expression. c 2004 Published by Elsevier Ltd.
Background Proteins synthesis occurs once RNAs are synthesised by RNA polymerases, enzymes that lack fidelity or proofreading function [1]. Non-faithful template copying by RNApol results in introduction of mutations at a rate of 1 10 5 mutations/base RNA synthesised [2], a rate that could impact significantly on the function of large proteins or proteins with high turnover rates. Additionally, many cellular processes, including cell to cell signalling and control of proliferation by contact inhibition, may require proteins with similar, but subtly different, protein sequences or conformations to permit differential signalling *
Tel.: +61893897101; fax: +61893897103. E-mail addresses: [email protected], [email protected].
0306-9877/$ - see front matter c 2004 Published by Elsevier Ltd. doi:10.1016/j.mehy.2004.02.024
to occur. Stable maintenance of these differences requires control over the fidelity rate of RNA polymerases. Most cellular enzymes are under some form of kinetic control, usually by product inhibition. While simple negative feedback is sufficient to control the velocity of enzyme activity, it is insufficient to ensure the functional quality of any complex molecules synthesised, especially proteins. The functionality of RNApol , output is determined by the proteins translated from any RNAs it synthesises. Modulation of its fidelity would result in control over protein sequence. However, regulation of this adaptability requires that viral RNApol , function be dynamically modifiable and controllable and that requires a close nexus between the functional output of polymerases (i.e. the functionality of any resulting protein) and the fidelity of that polymerase.
Transcriptional homeostasis
The mechanism The mechanism of regulation proposed – transcriptional homeostasis – results from minor differential interactions of wild type (wt) and mutant (mt) proteins on RNApol in a series of feedback epicycles that link and modulate RNApol function, RNA replication and protein synthesis and is outlined schematically (Fig. 1). In principle, homeostatic systems depend on: (i) an efferent arm that produces a desired change in response to perturbations of equilibria; (ii) an afferent arm to monitor the systems response to that change; (iii) mechanism(s) by which (i) and (ii) can communicate. The mechanism described here to regulate protein expression requires the following: i) that translated proteins interact with RNApol ; (ii) that these interactions alter both polymerase processivity and fidelity; (iii) that wild-type protein/ RNApol interaction are more avid and replicate cognate RNAs more rapidly than mutant protein/ RNApol interactions. RNA polymerase function is
233 known to be responsive to and influenced by accessory proteins that induce conformational changes and modulate both processivity and fidelity [3], an experimental finding that provides the basis for the mechanism proposed. Transcriptional homeostasis is an adaptation that facilitates stable protein expression in cells and permits depends on, protein (phenotype) expression to inducing, and subtly influencing, rates of genetic mutation at an RNA level. If protein expression (i.e. phenotype) modulates RNApol properties in a manner contingent on the proteins functionality, and modulates the sequence of RNA templates RNApol synthesises, a subtle form of “quality control” is exerted over protein synthesis. Transcriptional homeostasis is a fundamental mechanism that can regulate RNA transcription and protein expression and its use to mediate immune escape, control cellular differentiation, or other functions, would not be surprising. Finally, modification of RNA template by proteins (phenotype) will accelerate adaptation,
Figure 1 Transcriptional homeostasis. Starting in stable equilibrium A fi a1 fi a2 fi A1 . Excessive mutation (mutant proteins Emt ) translated (b1 ) by ribosomes (R) from mutant RNA1 result in competitive blockade (B) of RNA polymerase, changing conformation and functional properties of polymerase reducing increasing fidelity. The altered conformation results in more faithful transcription of RNA [2], with subsequent increased translation of wild type envelope (Ewt ) by ribosomes (R, b2 ), returning the system to the equilibrium point (A1 ). Generation of excessive wild type envelope Ewt from excessively homogeneous RNA3 (c1 ), leads to high affinity Ewt/pol complexes of Ewt to RNA pol (C). Altered polymerase conformation results in less faithful transcription of RNA to generate more mutated RNAs (RNA4 ), with subsequent increased translation of mutant envelope proteins by ribosomes (c2 ), returning the system to the equilibrium point (A1 ).
234 at least for RNA viruses, where the RNA template is the genotype. While introduction of mutations to most RNA genomes may not adversely influence the function of any specific protein function to any significant degree, transcriptional homeostasis ensures any RNA mutations that do arise, and result in a beneficial phenotype, would tend to be retained, accelerating adaptation. DNA-dependent DNA polymerases undergo similar conformational changes under the influence of accessory proteins [4]. In principle, transcriptional homeostasis could operate at the DNA level, at least in germ cells, where other mechanisms exist to minimise the likelihood deleterious structural mutations are retained, or in “junk” DNA sequences where transcriptional homeostasis would provide a mechanism to allow coding of memory in “hardhardcopy”, and thus an explanation of “instinct” or species memory.
Potential importance of transcriptional homeostasis Apart from its potential importance in maintaining the integrity of cellular protein synthesis, transcriptional homeostasis is potentially the mechanism that enables pure RNA viruses (and retroviruses) maintain quasispecies stability.
Sallie
Testing the hypothesis This hypothesis is readily testable in viral culture systems; if wild-type or mutant envelope proteins are added to a stably replicating viral culture (for example, HBV in HepG2 Cells) then alteration in the amount and fidelity, both easily measured, of viral RNAs generated by the system would be expected.
Acknowledgements I thank Profs. W.D. Reed, M.G. McCall, R.A. Joske, and Bill Musk for critical clinical and scientific guidance and S.J. Coleman, Matt and Tim for everything else.
References [1] Steinhauer DA, Domingo E, Holland JJ. Lack of evidence for proofreading mechanisms associated with an RNA virus polymerase. Gene 1992;122(2):281–8. [2] Preston BD, Poiesz BJ, Loeb LA. Fidelity of HIV-1 reverse transcriptase. Science 1988;242(4882):1168–11671. [3] Erie DA, Hajiseyedjavadi O, Young MC, von Hippel PH. Multiple RNA polymerase conformations and GreA: control of the fidelity of transcription. Science 1993;262(5135): 867–73. [4] Maga G, Frouin I, Spadari S, Hubscher U. Replication protein A as a “fidelity clamp” for DNA polymerase alpha. J Biol Chem 2001;276(21):18235–42.
http://intl.elsevierhealth.com/journals/mehy
Transcriptional homeostasis: a mechanism of protein quality control Richard Sallie* Western Gastroenterology, Department of Hepatology, St. John of God Hospital, Suite 35, 95 Monash Avenue, Nedlands, Perth, Western Australia 6009, Australia Received 5 February 2004; accepted 19 February 2004
Summary Sickle cell anaemia confirms that minor mutations in protein sequence can have catastrophic effects. However, RNA transcription depends on polymerases that have low fidelity and introduce errors at a rate of 10 5 substitutions/base synthesised. For many large proteins translation of these errors could result in significant loss of function, while for others (for example, cell surface signalling proteins) variability of protein expression could be advantageous. This paper outlines a mechanism that enables proteins to modulate error incorporation (variability) into RNAs sysnthesised by RNA polymerase – transcriptional homeostasis – thereby modulating, and error correcting, their own sysnthesis. Transcriptional homeostasis is a fundamental regulatory mechanism relevant to control of gene expression. c 2004 Published by Elsevier Ltd.
Background Proteins synthesis occurs once RNAs are synthesised by RNA polymerases, enzymes that lack fidelity or proofreading function [1]. Non-faithful template copying by RNApol results in introduction of mutations at a rate of 1 10 5 mutations/base RNA synthesised [2], a rate that could impact significantly on the function of large proteins or proteins with high turnover rates. Additionally, many cellular processes, including cell to cell signalling and control of proliferation by contact inhibition, may require proteins with similar, but subtly different, protein sequences or conformations to permit differential signalling *
Tel.: +61893897101; fax: +61893897103. E-mail addresses: [email protected], [email protected].
0306-9877/$ - see front matter c 2004 Published by Elsevier Ltd. doi:10.1016/j.mehy.2004.02.024
to occur. Stable maintenance of these differences requires control over the fidelity rate of RNA polymerases. Most cellular enzymes are under some form of kinetic control, usually by product inhibition. While simple negative feedback is sufficient to control the velocity of enzyme activity, it is insufficient to ensure the functional quality of any complex molecules synthesised, especially proteins. The functionality of RNApol , output is determined by the proteins translated from any RNAs it synthesises. Modulation of its fidelity would result in control over protein sequence. However, regulation of this adaptability requires that viral RNApol , function be dynamically modifiable and controllable and that requires a close nexus between the functional output of polymerases (i.e. the functionality of any resulting protein) and the fidelity of that polymerase.
Transcriptional homeostasis
The mechanism The mechanism of regulation proposed – transcriptional homeostasis – results from minor differential interactions of wild type (wt) and mutant (mt) proteins on RNApol in a series of feedback epicycles that link and modulate RNApol function, RNA replication and protein synthesis and is outlined schematically (Fig. 1). In principle, homeostatic systems depend on: (i) an efferent arm that produces a desired change in response to perturbations of equilibria; (ii) an afferent arm to monitor the systems response to that change; (iii) mechanism(s) by which (i) and (ii) can communicate. The mechanism described here to regulate protein expression requires the following: i) that translated proteins interact with RNApol ; (ii) that these interactions alter both polymerase processivity and fidelity; (iii) that wild-type protein/ RNApol interaction are more avid and replicate cognate RNAs more rapidly than mutant protein/ RNApol interactions. RNA polymerase function is
233 known to be responsive to and influenced by accessory proteins that induce conformational changes and modulate both processivity and fidelity [3], an experimental finding that provides the basis for the mechanism proposed. Transcriptional homeostasis is an adaptation that facilitates stable protein expression in cells and permits depends on, protein (phenotype) expression to inducing, and subtly influencing, rates of genetic mutation at an RNA level. If protein expression (i.e. phenotype) modulates RNApol properties in a manner contingent on the proteins functionality, and modulates the sequence of RNA templates RNApol synthesises, a subtle form of “quality control” is exerted over protein synthesis. Transcriptional homeostasis is a fundamental mechanism that can regulate RNA transcription and protein expression and its use to mediate immune escape, control cellular differentiation, or other functions, would not be surprising. Finally, modification of RNA template by proteins (phenotype) will accelerate adaptation,
Figure 1 Transcriptional homeostasis. Starting in stable equilibrium A fi a1 fi a2 fi A1 . Excessive mutation (mutant proteins Emt ) translated (b1 ) by ribosomes (R) from mutant RNA1 result in competitive blockade (B) of RNA polymerase, changing conformation and functional properties of polymerase reducing increasing fidelity. The altered conformation results in more faithful transcription of RNA [2], with subsequent increased translation of wild type envelope (Ewt ) by ribosomes (R, b2 ), returning the system to the equilibrium point (A1 ). Generation of excessive wild type envelope Ewt from excessively homogeneous RNA3 (c1 ), leads to high affinity Ewt/pol complexes of Ewt to RNA pol (C). Altered polymerase conformation results in less faithful transcription of RNA to generate more mutated RNAs (RNA4 ), with subsequent increased translation of mutant envelope proteins by ribosomes (c2 ), returning the system to the equilibrium point (A1 ).
234 at least for RNA viruses, where the RNA template is the genotype. While introduction of mutations to most RNA genomes may not adversely influence the function of any specific protein function to any significant degree, transcriptional homeostasis ensures any RNA mutations that do arise, and result in a beneficial phenotype, would tend to be retained, accelerating adaptation. DNA-dependent DNA polymerases undergo similar conformational changes under the influence of accessory proteins [4]. In principle, transcriptional homeostasis could operate at the DNA level, at least in germ cells, where other mechanisms exist to minimise the likelihood deleterious structural mutations are retained, or in “junk” DNA sequences where transcriptional homeostasis would provide a mechanism to allow coding of memory in “hardhardcopy”, and thus an explanation of “instinct” or species memory.
Potential importance of transcriptional homeostasis Apart from its potential importance in maintaining the integrity of cellular protein synthesis, transcriptional homeostasis is potentially the mechanism that enables pure RNA viruses (and retroviruses) maintain quasispecies stability.
Sallie
Testing the hypothesis This hypothesis is readily testable in viral culture systems; if wild-type or mutant envelope proteins are added to a stably replicating viral culture (for example, HBV in HepG2 Cells) then alteration in the amount and fidelity, both easily measured, of viral RNAs generated by the system would be expected.
Acknowledgements I thank Profs. W.D. Reed, M.G. McCall, R.A. Joske, and Bill Musk for critical clinical and scientific guidance and S.J. Coleman, Matt and Tim for everything else.
References [1] Steinhauer DA, Domingo E, Holland JJ. Lack of evidence for proofreading mechanisms associated with an RNA virus polymerase. Gene 1992;122(2):281–8. [2] Preston BD, Poiesz BJ, Loeb LA. Fidelity of HIV-1 reverse transcriptase. Science 1988;242(4882):1168–11671. [3] Erie DA, Hajiseyedjavadi O, Young MC, von Hippel PH. Multiple RNA polymerase conformations and GreA: control of the fidelity of transcription. Science 1993;262(5135): 867–73. [4] Maga G, Frouin I, Spadari S, Hubscher U. Replication protein A as a “fidelity clamp” for DNA polymerase alpha. J Biol Chem 2001;276(21):18235–42.