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| {{Short description|Fluoride-binding RNA structure}} | |||
| ] | |||
| {{Infobox rfam | |||
| The ''' ''crcB'' RNA motif''' is a conserved ] structure identified by ] in a wide variety of ] and ].<ref name="Weinberg2010">{{cite journal |author=Weinberg Z, Wang JX, Bogue J, ''et al.'' |title=Comparative genomics reveals 104 candidate structured RNAs from bacteria, archaea and their metagenomes |journal=Genome Biol |volume=11 |issue=3 |pages=R31 |year=2010 |month=March |pmid=20230605 |doi=10.1186/gb-2010-11-3-r31 |url=}}</ref> The motif exhibits the rigid or almost perfect conservation of the nucleotide identity at several positions (colored red in the diagram) across two ] of life. Such conservation is consistent with a biochemical requirement to bind a small molecule that does not evolve (like ]s or other ]s would). Given this observation, and the fact that ''crcB'' RNAs are typically located in possible ]s (5' UTRs) of protein-coding ]s, it was proposed that ''crcB'' RNAs likely function as ]es. | |||
| | Name = crcB RNA motif | |||
| | image = CrcB-RNA.svg | |||
| | width = | |||
| | caption = Consensus secondary structure of ''crcB'' RNAs | |||
| | Symbol = crcB RNA | |||
| | AltSymbols = | |||
| | Rfam = RF01734 | |||
| | miRBase = | |||
| | miRBase_family = | |||
| | RNA_type = ]; ] | |||
| | Tax_domain =] | |||
| | GO = | |||
| | SO = | |||
| | CAS_number = | |||
| | EntrezGene = | |||
| | HGNCid = | |||
| | OMIM = | |||
| | PDB = | |||
| | RefSeq = | |||
| | Chromosome = | |||
| | Arm = | |||
| | Band = | |||
| | LocusSupplementaryData = | |||
| }} | |||
| The ''' fluoride riboswitch''' (formerly called the '''''crcB'' RNA motif''') is a conserved ] structure identified by ] in a wide variety of ] and ].<ref name="Weinberg2010">{{cite journal |vauthors=Weinberg Z, Wang JX, Bogue J |title=Comparative genomics reveals 104 candidate structured RNAs from bacteria, archaea and their metagenomes |journal=Genome Biol |volume=11 |issue=3 |pages=R31 |date=March 2010 |pmid=20230605 |doi=10.1186/gb-2010-11-3-r31 |pmc=2864571|display-authors=etal |doi-access=free }}</ref> These RNAs were later shown to function as ]es that sense ] ]s.<ref name="Baker2012">{{cite journal |vauthors=Baker JL, Sudarsan N, Weinberg Z |title=Widespread genetic switches and toxicity resistance proteins for fluoride |journal=Science |volume=335 |issue=6065 |pages=233–235 |date=January 2012 |pmid=22194412 |doi=10.1126/science.1215063 |pmc=4140402|display-authors=etal}}</ref> These "fluoride riboswitches" increase expression of downstream genes when fluoride levels are elevated, and the genes are proposed to help mitigate the toxic effects of very high levels of fluoride. | |||
| Many genes are presumed to be regulated by ''crcB'' RNAs. The most common are ''mutS'', which is functions within DNA repair; the ''crcB'' gene that is thought to play a role in ] condensation; transporters of ], ] or ] ]s; the ''nifU'' iron-sulfur protein; and genes that encode ]s. Many ''crcB'' RNAs overlap predicted ] ]s. Assuming that ''crcB'' RNAs are riboswitches whose conserved structure is stabilized by the interaction with their small molecule ligand, high concentrations of this ligand should destabilize the transcription terminators, and thereby lead to increased gene expression. | |||
| Many genes are presumed to be regulated by these fluoride riboswitches. Two of the most common encode proteins that are proposed to function by removing fluoride from the cell. These proteins are CrcB proteins and a fluoride-specific subtype of chloride channels referred to as EriC<sup>F</sup> or ClC<sup>F</sup>. ClC<sup>F</sup> proteins have been shown to function as fluoride-specific fluoride/] antiporters.<ref name="Stockbridge2012"/> | |||
| Only one other class of riboswitch, the ], is found in organisms other than bacteria. Although several classes of RNAs are present in more than one domain of life (e.g., ] RNAs, ]s, ]s, ] RNAs and ]s), these RNAs are a small minority compared to classes of RNAs restricted to a single domain. Moreover, few of the highly widespread RNAs were initially discovered recently. The biological role played by ''crcB'' RNAs, although currently unknown, is apparently relevant to the physiology of both bacteria and archaea. | |||
| The three-dimensional structure of a fluoride riboswitch has been solved at atomic resolution by X-ray crystallography.<ref>{{cite journal |vauthors=Ren A, Rajashankar KR, Patel DJ |title=Fluoride ion encapsulation by Mg2+ ions and phosphates in a fluoride riboswitch |journal=Nature |volume=486 |issue=7401 |pages=85–89 |date=June 2012 |pmid=22678284 |doi=10.1038/nature11152 |pmc=3744881}}</ref> | |||
| Fluoride riboswitches are found in many organisms within the domains ] and ], indicating that many of these organisms sometimes encounter elevated levels of fluoride. Of particular interest is '']'', a major cause of ]. It has been shown that sodium fluoride has inhibited the growth rate of ''S. mutans'' using glucose as an energy and carbon source.<ref>{{cite journal|last=Yost|first=K G|author2=VanDemark, P J|title=Growth inhibition of Streptococcus mutans and Leuconostoc mesenteroides by sodium fluoride and ionic tin|journal=Applied and Environmental Microbiology|date=May 1978|volume=35|issue=5|pages=920–924|doi=10.1128/aem.35.5.920-924.1978|pmid=655708|pmc=242953}}</ref> However, it is also noteworthy that many organisms that do not encounter fluoride in the human mouth carry fluoride riboswitches or resistance genes. | |||
| == Discovery of the fluoride riboswitch == | |||
| The identity of fluoride as the riboswitch ligand was accidentally discovered when a compound contaminated with fluoride caused significant conformational changes to the non-coding ''crcB'' RNA motif during an in-line probing experiment.<ref name="Baker2012" /> In-line probing was used to illuminate the secondary structure of the ''crcB'' RNA motif and structural changes associated with possible binding to specific ] or ions.<ref name="Regulski">{{Cite book|last=Regulski|first=EE|author2=Breaker RR|chapter=In-Line Probing Analysis of Riboswitches |title=Post-Transcriptional Gene Regulation|year=2008|volume=419|pages=|doi=10.1007/978-1-59745-033-1_4|pmid=18369975|series=Methods in Molecular Biology|isbn=978-1-58829-783-9|chapter-url-access=registration|chapter-url=https://archive.org/details/posttranscriptio00wilu/page/53}}</ref> The results of the probing showed the addition of increasing fluoride ion concentrations suppressed certain regions of spontaneous RNA cleavage and heightening other regions. These nucleotide regions in the ''crcB'' RNA motif play important roles in the ] binding region for fluoride.<ref name="Baker2012"/> | |||
| Upon binding fluoride ions, the fluoride riboswitch showed regulation of downstream gene transcription.<ref name="Baker2012" /> These downstream genes transcribe fluoride sensitive enzymes <ref name="Baker2012" /> such as ], ], the presumed fluoride exporter CrcB and a superfamily of CLC membrane proteins called Eric<sup>F</sup> proteins.<ref name="Stockbridge2012">{{cite journal|last=Stockbridge|first=RB|author2=Lim HH |author3=Otten R |author4=Williams C |author5=Shane T |author6=Weinberg Z |author7=Miller C |title=Fluoride resistance and transport by riboswitch-controlled CLC antiporters|journal=Proc Natl Acad Sci U S A|date=18 September 2012|volume=109|issue=38|pages=15289–15294|pmid=22949689|doi=10.1073/pnas.1210896109 |pmc=3458365|doi-access=free}}</ref> The CLC<sup>F</sup> proteins have been shown to function as fluoride transporters against fluoride toxicity.<ref name="Stockbridge2012"/> The ''eric<sup>F</sup>'' gene is a mutant version of the chloride channel gene that is less common in bacteria than ]-specific ]s, but is nonetheless found in the genome of ].<ref>{{cite journal|last=Breaker|first=R.R.|title=New Insight on the Response of Bacteria to Fluoride|journal=Caries Research|date=10 February 2012|volume=46|issue=1|pages=78–81|doi=10.1159/000336397|pmid=22327376|url= |pmc=3331882}}</ref> The Eric<sup>F</sup> protein in particular carries specific amino acids in their channels that targets fluoride anions whereas the regular Eric protein favored chloride over fluoride ions.<ref name="Baker2012" /> | |||
| == Fluoride riboswitch structure == | |||
| ] The discovery of the fluoride riboswitch was surprising as both fluoride ions and the ''crcB'' RNA phosphate groups are negatively charged and should not be able to bind to one another.<ref name="Baker2012"/> Previous research came across this question in elucidating the cofactor thiamine pyrophosphate (TPP) riboswitch. The ] structure showed the assistance of two hydrated Mg<sup>2+</sup> ions that help stabilize the connection between the phosphates of TPP and guanine bases of the RNA.<ref name="Serganov">{{cite journal|last=Serganov|first=A|author2=Polonskaia A |author3=Phan AT |author4=Breaker RR |author5=Patel DJ |title=Structural basis for gene regulation by a thiamine pyrophosphate-sensing riboswitch|journal=Nature|date=29 June 2006|volume=441|issue=7097|pages=1167–1171|pmid=16728979|doi=10.1038/nature04740|pmc=4689313}}</ref><ref name="Thore">{{cite journal|last=Thore|first=S|author2=Leibundgut M |author3=Ban N |title=Structure of the eukaryotic thiamine pyrophosphate riboswitch with its regulatory ligand|journal=Science|date=26 May 2006|volume=312|issue=5777|pages=1208–1211|pmid=16675665|doi=10.1126/science.1128451|s2cid=32389251|doi-access=free}}</ref> This guiding research help characterize the fluoride riboswitch's own interactions with fluoride and its structure. Through in-line probing and mutational studies the fluoride riboswitch of the organism Thermotoga petrophila is recognized to have two helical stems adjoined by a helical loop with the capacity to become a ].<ref name="Ren">{{cite journal|last=Ren|first=A|author2=Rajashankar KR |author3=Patel DJ |title=Fluoride ion encapsulation by Mg<sup>2+</sup> ions and phosphates in a fluoride riboswitch|journal=Nature|date=13 May 2012|volume=486|issue=7401|pages=85–89|pmid=22678284|doi=10.1038/nature11152|pmc=3744881}}</ref> The bound fluoride ligand is found to be located within the center of the riboswitch fold, enclosed by three Mg<sup>2+</sup> ions. The Mg<sup>2+</sup> ions are octahedrally coordinated with five outer backbone phosphates and water molecules making a metabolite specific pocket for coordinating the fluoride ligand to bind. The placement of the Mg<sup>2+</sup> ions positions the fluoride ion into the negatively charged ''crcB'' RNA scaffold.<ref name="Ren"/> | |||
| == Biological significance == | |||
| ] | |||
| In the ], fluoride is the 13th most abundant element.<ref name="Baker2012"/> It is commonly used in oral healthcare products and water.<ref name="Baker2012"/> The fluoride acts as a hardening agent with the ] base on teeth, remineralizing and protecting them from harsh acids and bacteria in the oral cavity.<ref name="Wolfgang">{{cite journal|last=Wolfgang|first=Arnold|author2=Andreas Dorow |author3=Stephanie Langenhorst |author4=Zeno Gintner |author5=Jolan Banoczy |author6=Peter Gaengler |title=Effect of fluoride toothpastes on enamel demineralization|journal=BMC Oral Health|date=15 June 2006|volume=6|issue=8|pages=8|doi=10.1186/1472-6831-6-8|pmid=16776820 |pmc=1543617 |doi-access=free }}</ref> Additionally, its significance lies in the effect of the toxicity of fluoride at high concentrations to bacteria, especially those that cause ]. It has long been known that many species encapsulate a sensor system for toxic metals such as cadmium and silver.<ref name="Baker2012"/> However, a sensor system against fluoride remained unknown. The fluoride riboswitch elucidates the bacterial defense mechanism in counteracting against the toxicity of high concentrations of fluoride by regulating downstream genes of the riboswitch upon binding the fluoride ligand.<ref name="Baker2012"/> Further elucidating the mechanism of how bacteria protect themselves from fluoride toxicity can help modify the mechanism to make smaller concentrations of fluoride even more lethal to bacteria. Additionally, the fluoride riboswitch and the downstream regulated genes can be potential targets for drug development in the future. Overall, these advancements will help towards making fluoride and future drugs strong protectors against oral health disease. | |||
| ==References== | ==References== | ||
| {{reflist}} | |||
| <references/> | |||
| ] | ] | ||
| {{molecular-cell-biology-stub}} | |||
Latest revision as of 00:34, 4 December 2023
Fluoride-binding RNA structure RNA family| crcB RNA motif | |
|---|---|
 Consensus secondary structure of crcB RNAs | |
| Identifiers | |
| Symbol | crcB RNA |
| Rfam | RF01734 |
| Other data | |
| RNA type | Cis-reg; riboswitch |
| Domain(s) | Prokaryota |
| PDB structures | PDBe |
The fluoride riboswitch (formerly called the crcB RNA motif) is a conserved RNA structure identified by bioinformatics in a wide variety of bacteria and archaea. These RNAs were later shown to function as riboswitches that sense fluoride ions. These "fluoride riboswitches" increase expression of downstream genes when fluoride levels are elevated, and the genes are proposed to help mitigate the toxic effects of very high levels of fluoride.
Many genes are presumed to be regulated by these fluoride riboswitches. Two of the most common encode proteins that are proposed to function by removing fluoride from the cell. These proteins are CrcB proteins and a fluoride-specific subtype of chloride channels referred to as EriC or ClC. ClC proteins have been shown to function as fluoride-specific fluoride/proton antiporters.
The three-dimensional structure of a fluoride riboswitch has been solved at atomic resolution by X-ray crystallography.
Fluoride riboswitches are found in many organisms within the domains bacteria and archaea, indicating that many of these organisms sometimes encounter elevated levels of fluoride. Of particular interest is Streptococcus mutans, a major cause of dental caries. It has been shown that sodium fluoride has inhibited the growth rate of S. mutans using glucose as an energy and carbon source. However, it is also noteworthy that many organisms that do not encounter fluoride in the human mouth carry fluoride riboswitches or resistance genes.
Discovery of the fluoride riboswitch
The identity of fluoride as the riboswitch ligand was accidentally discovered when a compound contaminated with fluoride caused significant conformational changes to the non-coding crcB RNA motif during an in-line probing experiment. In-line probing was used to illuminate the secondary structure of the crcB RNA motif and structural changes associated with possible binding to specific metabolites or ions. The results of the probing showed the addition of increasing fluoride ion concentrations suppressed certain regions of spontaneous RNA cleavage and heightening other regions. These nucleotide regions in the crcB RNA motif play important roles in the aptamer binding region for fluoride.
Upon binding fluoride ions, the fluoride riboswitch showed regulation of downstream gene transcription. These downstream genes transcribe fluoride sensitive enzymes such as enolase, pyrophosphatase, the presumed fluoride exporter CrcB and a superfamily of CLC membrane proteins called Eric proteins. The CLC proteins have been shown to function as fluoride transporters against fluoride toxicity. The eric gene is a mutant version of the chloride channel gene that is less common in bacteria than chloride-specific homologs, but is nonetheless found in the genome of Streptococcus mutans. The Eric protein in particular carries specific amino acids in their channels that targets fluoride anions whereas the regular Eric protein favored chloride over fluoride ions.
Fluoride riboswitch structure
The discovery of the fluoride riboswitch was surprising as both fluoride ions and the crcB RNA phosphate groups are negatively charged and should not be able to bind to one another. Previous research came across this question in elucidating the cofactor thiamine pyrophosphate (TPP) riboswitch. The TPP riboswitch structure showed the assistance of two hydrated Mg ions that help stabilize the connection between the phosphates of TPP and guanine bases of the RNA. This guiding research help characterize the fluoride riboswitch's own interactions with fluoride and its structure. Through in-line probing and mutational studies the fluoride riboswitch of the organism Thermotoga petrophila is recognized to have two helical stems adjoined by a helical loop with the capacity to become a pseudoknot. The bound fluoride ligand is found to be located within the center of the riboswitch fold, enclosed by three Mg ions. The Mg ions are octahedrally coordinated with five outer backbone phosphates and water molecules making a metabolite specific pocket for coordinating the fluoride ligand to bind. The placement of the Mg ions positions the fluoride ion into the negatively charged crcB RNA scaffold.
Biological significance
In the Earth's crust, fluoride is the 13th most abundant element. It is commonly used in oral healthcare products and water. The fluoride acts as a hardening agent with the enamel base on teeth, remineralizing and protecting them from harsh acids and bacteria in the oral cavity. Additionally, its significance lies in the effect of the toxicity of fluoride at high concentrations to bacteria, especially those that cause dental caries. It has long been known that many species encapsulate a sensor system for toxic metals such as cadmium and silver. However, a sensor system against fluoride remained unknown. The fluoride riboswitch elucidates the bacterial defense mechanism in counteracting against the toxicity of high concentrations of fluoride by regulating downstream genes of the riboswitch upon binding the fluoride ligand. Further elucidating the mechanism of how bacteria protect themselves from fluoride toxicity can help modify the mechanism to make smaller concentrations of fluoride even more lethal to bacteria. Additionally, the fluoride riboswitch and the downstream regulated genes can be potential targets for drug development in the future. Overall, these advancements will help towards making fluoride and future drugs strong protectors against oral health disease.
References
- Weinberg Z, Wang JX, Bogue J, et al. (March 2010). "Comparative genomics reveals 104 candidate structured RNAs from bacteria, archaea and their metagenomes". Genome Biol. 11 (3): R31. doi:10.1186/gb-2010-11-3-r31. PMC 2864571. PMID 20230605.
- ^ Baker JL, Sudarsan N, Weinberg Z, et al. (January 2012). "Widespread genetic switches and toxicity resistance proteins for fluoride". Science. 335 (6065): 233–235. doi:10.1126/science.1215063. PMC 4140402. PMID 22194412.
- ^ Stockbridge, RB; Lim HH; Otten R; Williams C; Shane T; Weinberg Z; Miller C (18 September 2012). "Fluoride resistance and transport by riboswitch-controlled CLC antiporters". Proc Natl Acad Sci U S A. 109 (38): 15289–15294. doi:10.1073/pnas.1210896109. PMC 3458365. PMID 22949689.
- Ren A, Rajashankar KR, Patel DJ (June 2012). "Fluoride ion encapsulation by Mg2+ ions and phosphates in a fluoride riboswitch". Nature. 486 (7401): 85–89. doi:10.1038/nature11152. PMC 3744881. PMID 22678284.
- Yost, K G; VanDemark, P J (May 1978). "Growth inhibition of Streptococcus mutans and Leuconostoc mesenteroides by sodium fluoride and ionic tin". Applied and Environmental Microbiology. 35 (5): 920–924. doi:10.1128/aem.35.5.920-924.1978. PMC 242953. PMID 655708.
- Regulski, EE; Breaker RR (2008). "In-Line Probing Analysis of Riboswitches". Post-Transcriptional Gene Regulation. Methods in Molecular Biology. Vol. 419. pp. 53–67. doi:10.1007/978-1-59745-033-1_4. ISBN 978-1-58829-783-9. PMID 18369975.
- Breaker, R.R. (10 February 2012). "New Insight on the Response of Bacteria to Fluoride". Caries Research. 46 (1): 78–81. doi:10.1159/000336397. PMC 3331882. PMID 22327376.
- Serganov, A; Polonskaia A; Phan AT; Breaker RR; Patel DJ (29 June 2006). "Structural basis for gene regulation by a thiamine pyrophosphate-sensing riboswitch". Nature. 441 (7097): 1167–1171. doi:10.1038/nature04740. PMC 4689313. PMID 16728979.
- Thore, S; Leibundgut M; Ban N (26 May 2006). "Structure of the eukaryotic thiamine pyrophosphate riboswitch with its regulatory ligand". Science. 312 (5777): 1208–1211. doi:10.1126/science.1128451. PMID 16675665. S2CID 32389251.
- ^ Ren, A; Rajashankar KR; Patel DJ (13 May 2012). "Fluoride ion encapsulation by Mg ions and phosphates in a fluoride riboswitch". Nature. 486 (7401): 85–89. doi:10.1038/nature11152. PMC 3744881. PMID 22678284.
- Wolfgang, Arnold; Andreas Dorow; Stephanie Langenhorst; Zeno Gintner; Jolan Banoczy; Peter Gaengler (15 June 2006). "Effect of fluoride toothpastes on enamel demineralization". BMC Oral Health. 6 (8): 8. doi:10.1186/1472-6831-6-8. PMC 1543617. PMID 16776820.