- Email: [email protected]
SOS Repair
SOS Repair V Verbenko, Russian Academy of Sciences, Gatchina, St. Petersburg, Russia
© 2013 Elsevier Inc. All rights reserved.
This article is a revision of the previous edition article by BA Bridges, volume 4, pp 1853–1855, © 2001, Elsevier Inc.
Homologous recombination A housekeeping process involved in the maintenance of chromosome integrity and generation of genetic variability in which nucleotide sequences are exchanged between two similar or identical molecules of DNA.
Prophage A latent form of a bacteriophage inserted and integrated into the circular bacterial DNA chromosome. Regulon A network of genes or operons under regulation by the same regulatory protein. W-reactivation Inducible (Weigle) reactivation of irradiated bacteriophages plated on preliminary irradiated cells.
Bacterial cells permanently undergo the action of exogenic and endogenic factors damaging DNA. This damage being not repaired can lead to cell death or mutation. A large part of DNA breaks and nucleotide modifications is fixed by basal activity of repair enzymes. If damage interfering with DNA replication persists, expression of set of genes is induced. In order to survive in various environmental conditions, cells have a repertoire of genes that can express or silence according to their needs. Among this vast collection of genetically controlled stress networks, the SOS repair is an inducible DNA repair system that allows bacteria to survive sudden increases in DNA damage. The importance of the SOS system is under scored by the observation that this regulatory network is widely present in bacteria, reflecting the need to maintain the integrity of their genomes. Numerous experiments preceded SOS hypothesis. Observation of W-reactivation of ultraviolet (UV)-irradiated phage lambda, induction of prophage lambda and lysis of UV-irradiated lysogenic bacteria, filamentous growth of Escherichia coli B cells in response to UV irradiation, all sug gested a relation between the arrest of cell division, the mechanisms of lambda prophage induction, and UV-induced mutagenesis. These data led Miroslaw Radman to conclude to an unpublished letter in 1970 that in E. coli there is a DNA repair system dependent on the LexA and RecA proteins that is induced when DNA is severely damaged and its synthesis is arrested and the induction of this system is connected with induction of mutations. This coordinated cellular response to DNA damage was named ‘SOS response’. Two proteins play key roles in the regulation of the SOS response: a repressor named LexA (lexA is a locus for X-ray sensitivity A) and an inducer named the RecA filament (recA is a locus for recombinase A). The SOS response in E. coli can be induced by stalled replica tion forks, unrepaired defects following recombination or chromosome segregation, and DNA damage caused by irradia tion, radiomimetic, chemical mutagens, or metabolic intermediates in well-fed or starved cells. LexA protein is the repressor, which during normal bacterial growth downregu lates its own expression and, in E. coli, the expression of at least 43 unlinked genes. The RecA protein is the inducer, which, in response to DNA damage, binds to appeared single-stranded DNA (ssDNA) to form a filament. The ssDNA-RecA filament interacts with LexA as RecA* coprotease and activates a self-cleaving activity in LexA. Upon
self-cleavage, LexA dissociates from its DNA targets, causing the induction of the SOS regulon. Subsequently, as DNA damage is repaired, the coprotease activity of the RecA protein disappears and this allows functional LexA to reaccumulate and to bind to target sites to prevent expression of the SOS genes. Each of the SOS-induced damage-inducible (din) or SOS genes has near its promoter site a specific 20-nucleotide-long ‘SOS box’ (also named, LexA-box) to which the LexA repressor protein is bound, preventing RNA polymerase binding and gene expression. By comparing the sequences of SOS boxes from din genes, the consensus SOS box sequence was estab lished and it was a perfect palindrome, TACTG(TA)5CAGTA. The LexA repressor binds to SOS box as a dimer. RecA-induced self-cleavage of LexA is more rapid when LexA is dimeric, so LexA bound with promoters degrades first of all. The role of the RecA* coprotease in SOS-induced cells is not only to assist in the cleavage of LexA protein at the Ala84–Gly85 site, but also to cleave the CI repressor of lambda phage, which transforms the phage from a lysogenic to a lytic form and to process UmuD–UmuD′ by nicking UmuD, which is a prerequisite for the assembly of the SOS-induced mutagenic DNA polymerase V (PolV) consisting of UmuD′2C. Generally, only one SOS box is present in one operon. The sequences of the SOS boxes are different. The level, timing, and duration of induction of dif ferent LexA-regulated genes differ significantly, depending on the strength of the specific SOS boxes, their location relative to the target promoter and promoter strength. Since LexA binds some operators more weakly than others, selective derepression of certain genes might occur in response to even minor endo genous DNA damage. In contrast, some genes may be expressed only upon drastic DNA damage and a persistent inducing signal. Substantial expressions of some SOS genes are supported due to imperfect SOS box or alternative promo ters. Regulation of genes, which SOS boxes have low affinity to LexA repressor are achieved by high concentration of protein or two SOS boxes in genes lexA, and ColE, and three SOS boxes in recN. A lot is known of mutations in the lexA and recA genes affecting on steady state of the SOS regulon. The self-cleavage also triggers LexA degradation. After induction, dormant pro tease recognition signals are exposed in the cleaved LexA N-terminal and C-terminal fragments, resulting in degradation by the ClpXP protease. This process is important, since accu mulation of the LexA DNA binding N-terminal domain, which retains some repressor function, might be deleterious after
Glossary
Brenner’s Encyclopedia of Genetics, 2nd edition, Volume 6
doi:10.1016/B978-0-12-374984-0.01447-9
489
490
SOS Repair
DNA damage. Degradation of cleaved C-terminal domain is facilitated by the Lon protease. The lon gene is a member of both the SOS system and the heat-shock protein network. More recently, microarrays were used to measure the timing and the amplitude of the induction in bacterial populations. Induction of the SOS response proceeds until 45–60 min after treatment of bacteria with SOS-inducing agents and then abruptly ceases. Within this time, most of the lesions have been repaired. Induction of SOS gene expression is synchro nized with the DNA repair process. The first genes to be induced are uvrA, uvrB, and uvrD. These proteins, together with the endonuclease UvrC, catalyze nucleotide excision repair (NER), a reaction that excises the damaged nucleotides from double-stranded DNA. As a second defense against DNA lesions, expression of recA, recN, and other homologous recombination functions increase more slowly, about 10-fold. Homologous recombination allows the repair of lesions that occur on ssDNA regions at replication forks by rendering them double-stranded (and hence a substrate for NER) and also restores disintegrated replication forks. Depending on the nat ure of the DNA damage, the RecFOR or RecBCD complexes facilitate RecA filament assembly on 3′-ssDNA ends. In this case, RecA filament plays its role in recombinational repair. It invades an intact homologous double-stranded DNA molecule forming a synaptic complex. The division inhibitor SfiA is also induced to give the bacterium time to complete the repairs. Finally, about 40 min after DNA damage (and if the damage was not fully repaired by NER and homologous recombina tion), the mutagenic DNA repair polymerase PolV is induced. This response also allows bacteria to render DNA lesions double-stranded but at the expense of introducing errors into the genome. If this arrangement does not restore DNA replica tion, continuing SOS induction arouses colicin production and cryptic prophage induction followed by cell lysis. Induction of lysis by this way is an example of ‘bacterial apoptosis’. SOS induction was measured in bulk cultures until fluores cent microscopy techniques became available that allowed the direct measurement of gene expression in individual cells. An additional level of regulation beyond repression by LexA exists, and the expression of SOS genes is not simply induced until DNA damage is repaired and then turned off. The E. coli SOS network is turned on in a pattern of discrete activation pulses and the number of pulses, but not their amplitude, increases with the level of DNA damage. The UmuC and UmuD proteins appear to be key factors in maintaining this pattern, by some how modulating the activities of SOS gene promoters. Other proteins also play a role in regulating the SOS response. The DinI and RecX proteins, respectively, stabilize or destabilize active RecA filaments. More sophisticated regulation should be mentioned. The main protein of another stress system, the cold-shock protein CspA, activates synthesis of RecA after shift to low temperature. Beyond being a model of a DNA repair regulatory network, the SOS system has played an important role in shaping the bacterial world. This is mainly because it increases mutations and genetic exchanges. Induction of the E. coli SOS regulon involves three DNA polymerases, PolII (polB), PolIV (dinA), and PolV (umuDC), that operate in a poorly processive and error-prone manner, bypassing ‘irreparable’ DNA lesions that block replication. Not only do the polymerases fulfill transle sion synthesis, but also they enable bacteria to increase their
mutation rate. For example, when wild-type E. coli cells are placed in the presence of a carbon source that they cannot use, a substrate for PolIV arises in their chromosomes. Due to the mutagenic action of this DNA polymerase, a subpopulation of cells acquires the capacity to use the carbon source and propagates. The SOS response is widespread among bacteria and exhi bits considerable variation in its composition and regulation. Many different factors can trigger induction of the SOS response. Thus, when strains of E. coli pass into warm-blooded animals, they encounter many chemicals that can induce the SOS response. The production of antimicrobial molecules such as hydrogen peroxide by neutrophils and nitric oxide inside macrophages results in DNA damage and contributes to patho genesis in enterohemorrhagic E. coli and Salmonella. The discovery that LexA directly regulates the expression of different colicins clearly shows that members of the LexA regulon are not solely concerned with the upkeep of the genome. In some well-characterized pathogens, induction of the SOS response modulates the evolution and dissemination of drug resistance, as well as synthesis, secretion, and dissemination of the viru lence. Sublethal doses of some commonly used antibiotics induce the SOS response and the synthesis of error-prone DNA polymerases. Hence, antibiotics can speed up mutagen esis by the acquisition of point mutations that result in the drug’s inactivation or efflux. SOS-inducing antibiotics can also trigger the self-catalytic cleavage of phage repressors, leading to the horizontal spread of temperate phage and associated pathogenicity islands. Recent progress in understanding the molecular details of specific LexA binding at targets and how it is cleaved, together with genomic information on the SOS regulon in different organisms, make a possibility the LexA repressor be considered as a drug target. The SOS response has become a paradigm for the field of DNA repair. During the past 40 years, many laboratories have addressed questions concerning the function of the SOS genes and mechanisms of their regulation. Work on the SOS response illustrates fundamental and applied results of bacterial studies as SOS repair is both an irreplaceable source of concepts for the understanding of DNA repair regulation networks, and a mod ulator of bacterial propagation during pathogenicity. Several important issues remain to be addressed. More than a dozen SOS-induced genes encode proteins of unknown function. The identification of their physiological role may reveal new means of regulation of the SOS response, links, and overlap with other cellular stress systems, and unsuspected consequences of the SOS induction.
See also: Mutation; Repair Mechanisms; Recombination Pathways.
Further Reading Butala M, Zgur-Bertok D, and Busby SJW (2009) The bacterial LexA transcriptional repressor. Cellular and Molecular Life Sciences 66: 82–93. Dwyer DJ, Kohanski MA, and Collins JJ (2009) Role of reactive oxygen species in antibiotic action and resistance. Current Opinion in Microbiology 12(5): 482–489. Janion C (2008) Inducible SOS response system of DNA repair and mutagenesis in Escherichia coli. International Journal of Biological Sciences 4(6): 338–344.
SOS Repair Patel M, Jiang Q, Woodgate R, Cox MM, and Goodman MF (2010) A new model for SOS-induced mutagenesis: How RecA protein activates DNA polymerase V. Critical Reviews in Biochemistry and Molecular Biology 45(3): 171–184. Phadtare S and Severinov K (2010) RNA remodeling and gene regulation by cold shock proteins. RNA Biology 7(6): 788–795. Pruteanua M and Bakera TA (2009) Proteolysis in the SOS response and metal homeostasis in Escherichia coli. Research in Microbiology 160(9): 677–683.
Relevant Websites http://ecocyc.org – EcoCyc. http://www.ecogene.org – EcoGene. http://ecoliwiki.net – EcoliWiki.
491
© 2013 Elsevier Inc. All rights reserved.
This article is a revision of the previous edition article by BA Bridges, volume 4, pp 1853–1855, © 2001, Elsevier Inc.
Homologous recombination A housekeeping process involved in the maintenance of chromosome integrity and generation of genetic variability in which nucleotide sequences are exchanged between two similar or identical molecules of DNA.
Prophage A latent form of a bacteriophage inserted and integrated into the circular bacterial DNA chromosome. Regulon A network of genes or operons under regulation by the same regulatory protein. W-reactivation Inducible (Weigle) reactivation of irradiated bacteriophages plated on preliminary irradiated cells.
Bacterial cells permanently undergo the action of exogenic and endogenic factors damaging DNA. This damage being not repaired can lead to cell death or mutation. A large part of DNA breaks and nucleotide modifications is fixed by basal activity of repair enzymes. If damage interfering with DNA replication persists, expression of set of genes is induced. In order to survive in various environmental conditions, cells have a repertoire of genes that can express or silence according to their needs. Among this vast collection of genetically controlled stress networks, the SOS repair is an inducible DNA repair system that allows bacteria to survive sudden increases in DNA damage. The importance of the SOS system is under scored by the observation that this regulatory network is widely present in bacteria, reflecting the need to maintain the integrity of their genomes. Numerous experiments preceded SOS hypothesis. Observation of W-reactivation of ultraviolet (UV)-irradiated phage lambda, induction of prophage lambda and lysis of UV-irradiated lysogenic bacteria, filamentous growth of Escherichia coli B cells in response to UV irradiation, all sug gested a relation between the arrest of cell division, the mechanisms of lambda prophage induction, and UV-induced mutagenesis. These data led Miroslaw Radman to conclude to an unpublished letter in 1970 that in E. coli there is a DNA repair system dependent on the LexA and RecA proteins that is induced when DNA is severely damaged and its synthesis is arrested and the induction of this system is connected with induction of mutations. This coordinated cellular response to DNA damage was named ‘SOS response’. Two proteins play key roles in the regulation of the SOS response: a repressor named LexA (lexA is a locus for X-ray sensitivity A) and an inducer named the RecA filament (recA is a locus for recombinase A). The SOS response in E. coli can be induced by stalled replica tion forks, unrepaired defects following recombination or chromosome segregation, and DNA damage caused by irradia tion, radiomimetic, chemical mutagens, or metabolic intermediates in well-fed or starved cells. LexA protein is the repressor, which during normal bacterial growth downregu lates its own expression and, in E. coli, the expression of at least 43 unlinked genes. The RecA protein is the inducer, which, in response to DNA damage, binds to appeared single-stranded DNA (ssDNA) to form a filament. The ssDNA-RecA filament interacts with LexA as RecA* coprotease and activates a self-cleaving activity in LexA. Upon
self-cleavage, LexA dissociates from its DNA targets, causing the induction of the SOS regulon. Subsequently, as DNA damage is repaired, the coprotease activity of the RecA protein disappears and this allows functional LexA to reaccumulate and to bind to target sites to prevent expression of the SOS genes. Each of the SOS-induced damage-inducible (din) or SOS genes has near its promoter site a specific 20-nucleotide-long ‘SOS box’ (also named, LexA-box) to which the LexA repressor protein is bound, preventing RNA polymerase binding and gene expression. By comparing the sequences of SOS boxes from din genes, the consensus SOS box sequence was estab lished and it was a perfect palindrome, TACTG(TA)5CAGTA. The LexA repressor binds to SOS box as a dimer. RecA-induced self-cleavage of LexA is more rapid when LexA is dimeric, so LexA bound with promoters degrades first of all. The role of the RecA* coprotease in SOS-induced cells is not only to assist in the cleavage of LexA protein at the Ala84–Gly85 site, but also to cleave the CI repressor of lambda phage, which transforms the phage from a lysogenic to a lytic form and to process UmuD–UmuD′ by nicking UmuD, which is a prerequisite for the assembly of the SOS-induced mutagenic DNA polymerase V (PolV) consisting of UmuD′2C. Generally, only one SOS box is present in one operon. The sequences of the SOS boxes are different. The level, timing, and duration of induction of dif ferent LexA-regulated genes differ significantly, depending on the strength of the specific SOS boxes, their location relative to the target promoter and promoter strength. Since LexA binds some operators more weakly than others, selective derepression of certain genes might occur in response to even minor endo genous DNA damage. In contrast, some genes may be expressed only upon drastic DNA damage and a persistent inducing signal. Substantial expressions of some SOS genes are supported due to imperfect SOS box or alternative promo ters. Regulation of genes, which SOS boxes have low affinity to LexA repressor are achieved by high concentration of protein or two SOS boxes in genes lexA, and ColE, and three SOS boxes in recN. A lot is known of mutations in the lexA and recA genes affecting on steady state of the SOS regulon. The self-cleavage also triggers LexA degradation. After induction, dormant pro tease recognition signals are exposed in the cleaved LexA N-terminal and C-terminal fragments, resulting in degradation by the ClpXP protease. This process is important, since accu mulation of the LexA DNA binding N-terminal domain, which retains some repressor function, might be deleterious after
Glossary
Brenner’s Encyclopedia of Genetics, 2nd edition, Volume 6
doi:10.1016/B978-0-12-374984-0.01447-9
489
490
SOS Repair
DNA damage. Degradation of cleaved C-terminal domain is facilitated by the Lon protease. The lon gene is a member of both the SOS system and the heat-shock protein network. More recently, microarrays were used to measure the timing and the amplitude of the induction in bacterial populations. Induction of the SOS response proceeds until 45–60 min after treatment of bacteria with SOS-inducing agents and then abruptly ceases. Within this time, most of the lesions have been repaired. Induction of SOS gene expression is synchro nized with the DNA repair process. The first genes to be induced are uvrA, uvrB, and uvrD. These proteins, together with the endonuclease UvrC, catalyze nucleotide excision repair (NER), a reaction that excises the damaged nucleotides from double-stranded DNA. As a second defense against DNA lesions, expression of recA, recN, and other homologous recombination functions increase more slowly, about 10-fold. Homologous recombination allows the repair of lesions that occur on ssDNA regions at replication forks by rendering them double-stranded (and hence a substrate for NER) and also restores disintegrated replication forks. Depending on the nat ure of the DNA damage, the RecFOR or RecBCD complexes facilitate RecA filament assembly on 3′-ssDNA ends. In this case, RecA filament plays its role in recombinational repair. It invades an intact homologous double-stranded DNA molecule forming a synaptic complex. The division inhibitor SfiA is also induced to give the bacterium time to complete the repairs. Finally, about 40 min after DNA damage (and if the damage was not fully repaired by NER and homologous recombina tion), the mutagenic DNA repair polymerase PolV is induced. This response also allows bacteria to render DNA lesions double-stranded but at the expense of introducing errors into the genome. If this arrangement does not restore DNA replica tion, continuing SOS induction arouses colicin production and cryptic prophage induction followed by cell lysis. Induction of lysis by this way is an example of ‘bacterial apoptosis’. SOS induction was measured in bulk cultures until fluores cent microscopy techniques became available that allowed the direct measurement of gene expression in individual cells. An additional level of regulation beyond repression by LexA exists, and the expression of SOS genes is not simply induced until DNA damage is repaired and then turned off. The E. coli SOS network is turned on in a pattern of discrete activation pulses and the number of pulses, but not their amplitude, increases with the level of DNA damage. The UmuC and UmuD proteins appear to be key factors in maintaining this pattern, by some how modulating the activities of SOS gene promoters. Other proteins also play a role in regulating the SOS response. The DinI and RecX proteins, respectively, stabilize or destabilize active RecA filaments. More sophisticated regulation should be mentioned. The main protein of another stress system, the cold-shock protein CspA, activates synthesis of RecA after shift to low temperature. Beyond being a model of a DNA repair regulatory network, the SOS system has played an important role in shaping the bacterial world. This is mainly because it increases mutations and genetic exchanges. Induction of the E. coli SOS regulon involves three DNA polymerases, PolII (polB), PolIV (dinA), and PolV (umuDC), that operate in a poorly processive and error-prone manner, bypassing ‘irreparable’ DNA lesions that block replication. Not only do the polymerases fulfill transle sion synthesis, but also they enable bacteria to increase their
mutation rate. For example, when wild-type E. coli cells are placed in the presence of a carbon source that they cannot use, a substrate for PolIV arises in their chromosomes. Due to the mutagenic action of this DNA polymerase, a subpopulation of cells acquires the capacity to use the carbon source and propagates. The SOS response is widespread among bacteria and exhi bits considerable variation in its composition and regulation. Many different factors can trigger induction of the SOS response. Thus, when strains of E. coli pass into warm-blooded animals, they encounter many chemicals that can induce the SOS response. The production of antimicrobial molecules such as hydrogen peroxide by neutrophils and nitric oxide inside macrophages results in DNA damage and contributes to patho genesis in enterohemorrhagic E. coli and Salmonella. The discovery that LexA directly regulates the expression of different colicins clearly shows that members of the LexA regulon are not solely concerned with the upkeep of the genome. In some well-characterized pathogens, induction of the SOS response modulates the evolution and dissemination of drug resistance, as well as synthesis, secretion, and dissemination of the viru lence. Sublethal doses of some commonly used antibiotics induce the SOS response and the synthesis of error-prone DNA polymerases. Hence, antibiotics can speed up mutagen esis by the acquisition of point mutations that result in the drug’s inactivation or efflux. SOS-inducing antibiotics can also trigger the self-catalytic cleavage of phage repressors, leading to the horizontal spread of temperate phage and associated pathogenicity islands. Recent progress in understanding the molecular details of specific LexA binding at targets and how it is cleaved, together with genomic information on the SOS regulon in different organisms, make a possibility the LexA repressor be considered as a drug target. The SOS response has become a paradigm for the field of DNA repair. During the past 40 years, many laboratories have addressed questions concerning the function of the SOS genes and mechanisms of their regulation. Work on the SOS response illustrates fundamental and applied results of bacterial studies as SOS repair is both an irreplaceable source of concepts for the understanding of DNA repair regulation networks, and a mod ulator of bacterial propagation during pathogenicity. Several important issues remain to be addressed. More than a dozen SOS-induced genes encode proteins of unknown function. The identification of their physiological role may reveal new means of regulation of the SOS response, links, and overlap with other cellular stress systems, and unsuspected consequences of the SOS induction.
See also: Mutation; Repair Mechanisms; Recombination Pathways.
Further Reading Butala M, Zgur-Bertok D, and Busby SJW (2009) The bacterial LexA transcriptional repressor. Cellular and Molecular Life Sciences 66: 82–93. Dwyer DJ, Kohanski MA, and Collins JJ (2009) Role of reactive oxygen species in antibiotic action and resistance. Current Opinion in Microbiology 12(5): 482–489. Janion C (2008) Inducible SOS response system of DNA repair and mutagenesis in Escherichia coli. International Journal of Biological Sciences 4(6): 338–344.
SOS Repair Patel M, Jiang Q, Woodgate R, Cox MM, and Goodman MF (2010) A new model for SOS-induced mutagenesis: How RecA protein activates DNA polymerase V. Critical Reviews in Biochemistry and Molecular Biology 45(3): 171–184. Phadtare S and Severinov K (2010) RNA remodeling and gene regulation by cold shock proteins. RNA Biology 7(6): 788–795. Pruteanua M and Bakera TA (2009) Proteolysis in the SOS response and metal homeostasis in Escherichia coli. Research in Microbiology 160(9): 677–683.
Relevant Websites http://ecocyc.org – EcoCyc. http://www.ecogene.org – EcoGene. http://ecoliwiki.net – EcoliWiki.
491