Riboswitch

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A 3D representation of the lysine riboswitch (PDB code:3DIL, orange and blue tubes) bound to lysine (shown as grey, red and blue spheres in the upper middle of the structure) Lys ribosw 1.jpg
A 3D representation of the lysine riboswitch (PDB code:3DIL, orange and blue tubes) bound to lysine (shown as grey, red and blue spheres in the upper middle of the structure)

In molecular biology, a riboswitch is a regulatory segment of a messenger RNA molecule that binds a small molecule, resulting in a change in production of the proteins encoded by the mRNA. [1] [2] [3] [4] Thus, an mRNA that contains a riboswitch is directly involved in regulating its own activity, in response to the concentrations of its effector molecule. The discovery that modern organisms use RNA to bind small molecules, and discriminate against closely related analogs, expanded the known natural capabilities of RNA beyond its ability to code for proteins, catalyze reactions, or to bind other RNA or protein macromolecules.

Contents

The original definition of the term "riboswitch" specified that they directly sense small-molecule metabolite concentrations. [5] Although this definition remains in common use, some biologists have used a broader definition that includes other cis-regulatory RNAs. However, this article will discuss only metabolite-binding riboswitches.

Most known riboswitches occur in bacteria, but functional riboswitches of one type (the TPP riboswitch) have been discovered in archaea, plants and certain fungi. TPP riboswitches have also been predicted in archaea, [6] but have not been experimentally tested.

Examples of three riboswitches with their natural ligands. From left to right: the SAM riboswitch (PDB code: 3gx5), the FMN riboswitch (2yie) and the lysine riboswitch (PDB ID: 3d0u) Examples of some riboswitches with natural ligands.png
Examples of three riboswitches with their natural ligands. From left to right: the SAM riboswitch (PDB code: 3gx5), the FMN riboswitch (2yie) and the lysine riboswitch (PDB ID: 3d0u)

History and discovery

Prior to the discovery of riboswitches, the mechanism by which some genes involved in multiple metabolic pathways were regulated remained mysterious. Accumulating evidence increasingly suggested the then-unprecedented idea that the mRNAs involved might bind metabolites directly, to affect their own regulation. These data included conserved RNA secondary structures often found in the untranslated regions (UTRs) of the relevant genes and the success of procedures to create artificial small molecule-binding RNAs called aptamers. [7] [8] [9] [10] [11] In 2002, the first comprehensive proofs of multiple classes of riboswitches were published, including protein-free binding assays, and metabolite-binding riboswitches were established as a new mechanism of gene regulation. [5] [12] [13] [14]

Many of the earliest riboswitches to be discovered corresponded to conserved sequence "motifs" (patterns) in 5' UTRs that appeared to correspond to a structured RNA. For example, comparative analysis of upstream regions of several genes expected to be co-regulated led to the description of the S-box [15] (now the SAM-I riboswitch), the THI-box [9] (a region within the TPP riboswitch), the RFN element [8] (now the FMN riboswitch) and the B12-box [16] (a part of the cobalamin riboswitch), and in some cases experimental demonstrations that they were involved in gene regulation via an unknown mechanism. Bioinformatics has played a role in more recent discoveries, with increasing automation of the basic comparative genomics strategy. Barrick et al. (2004) [17] used BLAST to find UTRs homologous to all UTRs in Bacillus subtilis . Some of these homologous sets were inspected for conserved structure, resulting in 10 RNA-like motifs. Three of these were later experimentally confirmed as the glmS, glycine and PreQ1-I riboswitches (see below). Subsequent comparative genomics efforts using additional taxa of bacteria and improved computer algorithms have identified further riboswitches that are experimentally confirmed, as well as conserved RNA structures that are hypothesized to function as riboswitches. [18] [19] [20]

Mechanisms

Riboswitches are often conceptually divided into two parts: an aptamer and an expression platform. The aptamer directly binds the small molecule, and the expression platform undergoes structural changes in response to the changes in the aptamer. The expression platform is what regulates gene expression.

Expression platforms typically turn off gene expression in response to the small molecule, but some turn it on. The following riboswitch mechanisms have been experimentally demonstrated.

Types

Secondary structure of a purine riboswitch from Bacillus subtilis RF00167.jpg
Secondary structure of a purine riboswitch from Bacillus subtilis

The following is a list of experimentally validated riboswitches, organized by ligand.

Presumed riboswitches:

Candidate metabolite-binding riboswitches have been identified using bioinformatics, and have moderately complex secondary structures and several highly conserved nucleotide positions, as these features are typical of riboswitches that must specifically bind a small molecule. Riboswitch candidates are also consistently located in the 5' UTRs of protein-coding genes, and these genes are suggestive of metabolite binding, as these are also features of most known riboswitches. Hypothesized riboswitch candidates highly consistent with the preceding criteria are as follows: crcB RNA Motif, manA RNA motif, pfl RNA motif, ydaO/yuaA leader, yjdF RNA motif, ykkC-yxkD leader (and related ykkC-III RNA motif) and the yybP-ykoY leader. The functions of these hypothetical riboswitches remain unknown.

Computational models

Riboswitches have been also investigated using in-silico approaches. [29] [30] [31] In particular, solutions for riboswitch prediction can be divided into two wide categories:

The SwiSpot tool [39] somehow covers both the groups, as it uses conformational predictions to assess the presence of riboswitches.

Other computational investigations look into the druggability properties of riboswitch binding sites, assessing their potency as potential novel drug targets. [40] [41] [42]

The RNA world hypothesis

Riboswitches demonstrate that naturally occurring RNA can bind small molecules specifically, a capability that many previously believed was the domain of proteins or artificially constructed RNAs called aptamers. The existence of riboswitches in all domains of life therefore adds some support to the RNA world hypothesis, which holds that life originally existed using only RNA, and proteins came later; this hypothesis requires that all critical functions performed by proteins (including small molecule binding) could be performed by RNA. It has been suggested that some riboswitches might represent ancient regulatory systems, or even remnants of RNA-world ribozymes whose bindings domains are conserved. [13] [18] [43]

As antibiotic targets

RNA is already an emerging drug target. [44] Multiple factors contribute to riboswitches as targets for novel antibiotics: [45] [46] [47]

Indeed, some antibiotics whose mechanism of action was unknown for decades have been shown to operate by targeting riboswitches. [49] For example, when the antibiotic pyrithiamine enters the cell, it is metabolized into pyrithiamine pyrophosphate. Pyrithiamine pyrophosphate has been shown to bind and activate the TPP riboswitch, causing the cell to cease the synthesis and import of TPP. Because pyrithiamine pyrophosphate does not substitute for TPP as a coenzyme, the cell dies.

Engineered riboswitches

Since riboswitches are an effective method of controlling gene expression in natural organisms, there has been interest in engineering artificial riboswitches [50] [51] [52] [53] for industrial and medical applications such as gene therapy. [54] [55]

See also

References

  1. Nudler E, Mironov AS (January 2004). "The riboswitch control of bacterial metabolism". Trends in Biochemical Sciences. 29 (1): 11–17. doi:10.1016/j.tibs.2003.11.004. PMID   14729327.
  2. Tucker BJ, Breaker RR (June 2005). "Riboswitches as versatile gene control elements". Current Opinion in Structural Biology. 15 (3): 342–348. doi:10.1016/j.sbi.2005.05.003. PMID   15919195.
  3. Vitreschak AG, Rodionov DA, Mironov AA, Gelfand MS (January 2004). "Riboswitches: the oldest mechanism for the regulation of gene expression?". Trends in Genetics. 20 (1): 44–50. CiteSeerX   10.1.1.312.9100 . doi:10.1016/j.tig.2003.11.008. PMID   14698618.
  4. Batey RT (June 2006). "Structures of regulatory elements in mRNAs". Current Opinion in Structural Biology. 16 (3): 299–306. doi:10.1016/j.sbi.2006.05.001. PMID   16707260.
  5. 1 2 Nahvi A, Sudarsan N, Ebert MS, Zou X, Brown KL, Breaker RR (September 2002). "Genetic control by a metabolite binding mRNA". Chemistry & Biology. 9 (9): 1043–1049. doi: 10.1016/S1074-5521(02)00224-7 . PMID   12323379.
  6. Sudarsan N, Barrick JE, Breaker RR (June 2003). "Metabolite-binding RNA domains are present in the genes of eukaryotes". RNA. 9 (6): 644–647. doi:10.1261/rna.5090103. PMC   1370431 . PMID   12756322.
  7. Nou X, Kadner RJ (June 2000). "Adenosylcobalamin inhibits ribosome binding to btuB RNA". Proceedings of the National Academy of Sciences of the United States of America. 97 (13): 7190–7195. Bibcode:2000PNAS...97.7190N. doi: 10.1073/pnas.130013897 . PMC   16521 . PMID   10852957.
  8. 1 2 Gelfand MS, Mironov AA, Jomantas J, Kozlov YI, Perumov DA (November 1999). "A conserved RNA structure element involved in the regulation of bacterial riboflavin synthesis genes". Trends in Genetics. 15 (11): 439–442. doi:10.1016/S0168-9525(99)01856-9. PMID   10529804.
  9. 1 2 Miranda-Ríos J, Navarro M, Soberón M (August 2001). "A conserved RNA structure (thi box) is involved in regulation of thiamin biosynthetic gene expression in bacteria". Proceedings of the National Academy of Sciences of the United States of America. 98 (17): 9736–9741. doi: 10.1073/pnas.161168098 . PMC   55522 . PMID   11470904.
  10. Stormo GD, Ji Y (August 2001). "Do mRNAs act as direct sensors of small molecules to control their expression?". Proceedings of the National Academy of Sciences of the United States of America. 98 (17): 9465–9467. Bibcode:2001PNAS...98.9465S. doi: 10.1073/pnas.181334498 . PMC   55472 . PMID   11504932.
  11. Gold L, Brown D, He Y, Shtatland T, Singer BS, Wu Y (January 1997). "From oligonucleotide shapes to genomic SELEX: novel biological regulatory loops". Proceedings of the National Academy of Sciences of the United States of America. 94 (1): 59–64. Bibcode:1997PNAS...94...59G. doi: 10.1073/pnas.94.1.59 . PMC   19236 . PMID   8990161.
  12. Mironov AS, Gusarov I, Rafikov R, Lopez LE, Shatalin K, Kreneva RA, Perumov DA, Nudler E (November 2002). "Sensing small molecules by nascent RNA: a mechanism to control transcription in bacteria". Cell. 111 (5): 747–756. doi: 10.1016/S0092-8674(02)01134-0 . PMID   12464185.
  13. 1 2 Winkler W, Nahvi A, Breaker RR (October 2002). "Thiamine derivatives bind messenger RNAs directly to regulate bacterial gene expression". Nature. 419 (6910): 952–956. Bibcode:2002Natur.419..952W. doi:10.1038/nature01145. PMID   12410317. S2CID   4408592.
  14. Winkler WC, Cohen-Chalamish S, Breaker RR (December 2002). "An mRNA structure that controls gene expression by binding FMN". Proceedings of the National Academy of Sciences of the United States of America. 99 (25): 15908–15913. Bibcode:2002PNAS...9915908W. doi: 10.1073/pnas.212628899 . PMC   138538 . PMID   12456892.
  15. Grundy FJ, Henkin TM (November 1998). "The S box regulon: a new global transcription termination control system for methionine and cysteine biosynthesis genes in gram-positive bacteria". Molecular Microbiology. 30 (4): 737–749. doi: 10.1046/j.1365-2958.1998.01105.x . PMID   10094622.
  16. Franklund CV, Kadner RJ (June 1997). "Multiple transcribed elements control expression of the Escherichia coli btuB gene". Journal of Bacteriology. 179 (12): 4039–4042. doi:10.1128/jb.179.12.4039-4042.1997. PMC   179215 . PMID   9190822.
  17. Barrick JE, Corbino KA, Winkler WC, Nahvi A, Mandal M, Collins J, Lee M, Roth A, Sudarsan N, Jona I, Wickiser JK, Breaker RR (April 2004). "New RNA motifs suggest an expanded scope for riboswitches in bacterial genetic control". Proceedings of the National Academy of Sciences of the United States of America. 101 (17): 6421–6426. Bibcode:2004PNAS..101.6421B. doi: 10.1073/pnas.0308014101 . PMC   404060 . PMID   15096624.
  18. 1 2 Corbino KA, Barrick JE, Lim J, Welz R, Tucker BJ, Puskarz I, Mandal M, Rudnick ND, Breaker RR (2005). "Evidence for a second class of S-adenosylmethionine riboswitches and other regulatory RNA motifs in alpha-proteobacteria". Genome Biology. 6 (8) R70. doi: 10.1186/gb-2005-6-8-r70 . PMC   1273637 . PMID   16086852.
  19. Weinberg Z, Barrick JE, Yao Z, Roth A, Kim JN, Gore J, Wang JX, Lee ER, Block KF, Sudarsan N, Neph S, Tompa M, Ruzzo WL, Breaker RR (2007). "Identification of 22 candidate structured RNAs in bacteria using the CMfinder comparative genomics pipeline". Nucleic Acids Research. 35 (14): 4809–4819. doi:10.1093/nar/gkm487. PMC   1950547 . PMID   17621584.
  20. Weinberg Z, Wang JX, Bogue J, Yang J, Corbino K, Moy RH, Breaker RR (March 2010). "Comparative genomics reveals 104 candidate structured RNAs from bacteria, archaea, and their metagenomes". Genome Biology. 11 (3) R31. doi: 10.1186/gb-2010-11-3-r31 . PMC   2864571 . PMID   20230605.
  21. Cheah MT, Wachter A, Sudarsan N, Breaker RR (May 2007). "Control of alternative RNA splicing and gene expression by eukaryotic riboswitches". Nature. 447 (7143): 497–500. Bibcode:2007Natur.447..497C. doi:10.1038/nature05769. PMID   17468745. S2CID   4393918.
  22. Wachter A, Tunc-Ozdemir M, Grove BC, Green PJ, Shintani DK, Breaker RR (November 2007). "Riboswitch control of gene expression in plants by splicing and alternative 3' end processing of mRNAs". The Plant Cell. 19 (11): 3437–3450. doi:10.1105/tpc.107.053645. PMC   2174889 . PMID   17993623.
  23. Bocobza S, Adato A, Mandel T, Shapira M, Nudler E, Aharoni A (November 2007). "Riboswitch-dependent gene regulation and its evolution in the plant kingdom". Genes & Development. 21 (22): 2874–2879. doi:10.1101/gad.443907. PMC   2049190 . PMID   18006684.
  24. André G, Even S, Putzer H, Burguière P, Croux C, Danchin A, Martin-Verstraete I, Soutourina O (October 2008). "S-box and T-box riboswitches and antisense RNA control a sulfur metabolic operon of Clostridium acetobutylicum". Nucleic Acids Research. 36 (18): 5955–5969. doi:10.1093/nar/gkn601. PMC   2566862 . PMID   18812398.
  25. Loh E, Dussurget O, Gripenland J, Vaitkevicius K, Tiensuu T, Mandin P, Repoila F, Buchrieser C, Cossart P, Johansson J (November 2009). "A trans-acting riboswitch controls expression of the virulence regulator PrfA in Listeria monocytogenes". Cell. 139 (4): 770–779. doi: 10.1016/j.cell.2009.08.046 . PMID   19914169.
  26. "Team:BYU Provo/Results - 2011.igem.org". 2011.igem.org.
  27. Sherman EM, Esquiaqui J, Elsayed G, Ye JD (March 2012). "An energetically beneficial leader-linker interaction abolishes ligand-binding cooperativity in glycine riboswitches". RNA. 18 (3): 496–507. doi:10.1261/rna.031286.111. PMC   3285937 . PMID   22279151.
  28. Bocobza SE, Aharoni A (October 2008). "Switching the light on plant riboswitches". Trends Plant Sci. 13 (10): 526–33. Bibcode:2008TPS....13..526B. doi:10.1016/j.tplants.2008.07.004. PMID   18778966.
  29. Barrick JE (2009). "Predicting Riboswitch Regulation on a Genomic Scale". Riboswitches. Methods in Molecular Biology. Vol. 540. pp. 1–13. doi:10.1007/978-1-59745-558-9_1. ISBN   978-1-934115-88-6. PMID   19381548.
  30. Barash D, Gabdank I (January 2010). "Energy minimization applied to riboswitches: a perspective and challenges". RNA Biology. 7 (1): 90–97. doi: 10.4161/rna.7.1.10657 . PMID   20061789.
  31. Chen, Shi-Jie; Burke, Donald H; Adamiak, R W (2015). Computational methods for understanding riboswitches / Methods in Enzymology, vol 553. Academic Press. ISBN   978-0-12-801618-3.
  32. Nawrocki EP, Kolbe DL, Eddy SR (May 2009). "Infernal 1.0: inference of RNA alignments". Bioinformatics. 25 (10): 1335–1337. doi:10.1093/bioinformatics/btp157. PMC   2732312 . PMID   19307242.
  33. Abreu-Goodger C, Merino E (July 2005). "RibEx: a web server for locating riboswitches and other conserved bacterial regulatory elements". Nucleic Acids Research. 33 (Web Server issue): W690-2. doi:10.1093/nar/gki445. PMC   1160206 . PMID   15980564.
  34. Chang TH, Huang HD, Wu LC, Yeh CT, Liu BJ, Horng JT (July 2009). "Computational identification of riboswitches based on RNA conserved functional sequences and conformations". RNA. 15 (7): 1426–1430. doi:10.1261/rna.1623809. PMC   2704089 . PMID   19460868.
  35. Voss B, Meyer C, Giegerich R (July 2004). "Evaluating the predictability of conformational switching in RNA". Bioinformatics. 20 (10): 1573–1582. doi: 10.1093/bioinformatics/bth129 . PMID   14962925.
  36. Janssen S, Giegerich R (February 2015). "The RNA shapes studio". Bioinformatics. 31 (3): 423–425. doi:10.1093/bioinformatics/btu649. PMC   4308662 . PMID   25273103.
  37. Freyhult E, Moulton V, Clote P (August 2007). "Boltzmann probability of RNA structural neighbors and riboswitch detection". Bioinformatics. 23 (16): 2054–2062. doi: 10.1093/bioinformatics/btm314 . PMID   17573364.
  38. Clote P, Lou F, Lorenz WA (April 2012). "Maximum expected accuracy structural neighbors of an RNA secondary structure". BMC Bioinformatics. 13 (Suppl 5) S6. doi: 10.1186/1471-2105-13-S5-S6 . PMC   3358666 . PMID   22537010.
  39. Barsacchi M, Novoa EM, Kellis M, Bechini A (November 2016). "SwiSpot: modeling riboswitches by spotting out switching sequences". Bioinformatics. 32 (21): 3252–3259. doi: 10.1093/bioinformatics/btw401 . hdl: 11568/817190 . PMID   27378291.
  40. Rekand, Illimar Hugo; Brenk, Ruth (2021-08-23). "DrugPred_RNA—A Tool for Structure-Based Druggability Predictions for RNA Binding Sites". Journal of Chemical Information and Modeling. 61 (8): 4068–4081. doi:10.1021/acs.jcim.1c00155. ISSN   1549-9596. PMC   8389535 . PMID   34286972 . Retrieved 2024-03-12.
  41. Ramaswamy Krishnan, Sowmya; Roy, Arijit; Wong, Limsoon; Gromiha, M Michael (2025-04-11). "DRLiPS: a novel method for prediction of druggable RNA-small molecule binding pockets using machine learning". Nucleic Acids Research. 53 (6): 239. doi:10.1093/nar/gkaf239. ISSN   1362-4962. PMC   11963762 . PMID   40173014 . Retrieved 2025-06-10.
  42. Xie, Jingru; Frank, Aaron T. (2021-06-10). "Mining for Ligandable Cavities in RNA". ACS Medicinal Chemistry Letters. 12 (6): 928–934. doi:10.1021/acsmedchemlett.1c00068. PMC   8201482 . PMID   34141071 . Retrieved 2025-06-10.
  43. Cochrane JC, Strobel SA (June 2008). "Riboswitch effectors as protein enzyme cofactors". RNA. 14 (6): 993–1002. doi:10.1261/rna.908408. PMC   2390802 . PMID   18430893.
  44. Yu, Ai-Ming; Choi, Young Hee; Tu, Mei-Juan (2020). "RNA Drugs and RNA Targets for Small Molecules: Principles, Progress, and Challenges". Pharmacological Reviews. 72 (4): 862–898. doi:10.1124/pr.120.019554. ISSN   1521-0081. PMC   7495341 . PMID   32929000.
  45. Panchal, Vipul; Brenk, Ruth (January 2021). "Riboswitches as Drug Targets for Antibiotics". Antibiotics. 10 (1): 45. doi: 10.3390/antibiotics10010045 . PMC   7824784 . PMID   33466288.
  46. Hewitt, William M.; Calabrese, David R.; Schneekloth, John S. (2019-06-01). "Evidence for ligandable sites in structured RNA throughout the Protein Data Bank". Bioorganic & Medicinal Chemistry. 27 (11): 2253–2260. doi:10.1016/j.bmc.2019.04.010. ISSN   0968-0896. PMC   8283815 . PMID   30982658.
  47. Rekand, Illimar Hugo; Brenk, Ruth (2021-08-23). "DrugPred_RNA—A Tool for Structure-Based Druggability Predictions for RNA Binding Sites". Journal of Chemical Information and Modeling. 61 (8): 4068–4081. doi:10.1021/acs.jcim.1c00155. ISSN   1549-9596. PMC   8389535 . PMID   34286972 . Retrieved 2024-03-12.
  48. Rekand, Illimar; Brenk, Ruth (2017). "Design of riboswitch ligands, an emerging target class for novel antibiotics". Future Med Chem. 9 (14): 1649–1662. doi:10.4155/fmc-2017-0063. PMID   28925284.
  49. Blount KF, Breaker RR (December 2006). "Riboswitches as antibacterial drug targets". Nature Biotechnology. 24 (12): 1558–1564. doi:10.1038/nbt1268. PMID   17160062. S2CID   34398395.
  50. Bauer G, Suess B (June 2006). "Engineered riboswitches as novel tools in molecular biology". Journal of Biotechnology. 124 (1): 4–11. doi:10.1016/j.jbiotec.2005.12.006. PMID   16442180.
  51. Dixon N, Duncan JN, Geerlings T, Dunstan MS, McCarthy JE, Leys D, Micklefield J (February 2010). "Reengineering orthogonally selective riboswitches". Proceedings of the National Academy of Sciences of the United States of America. 107 (7): 2830–2835. Bibcode:2010PNAS..107.2830D. doi: 10.1073/pnas.0911209107 . PMC   2840279 . PMID   20133756.
  52. Verhounig A, Karcher D, Bock R (April 2010). "Inducible gene expression from the plastid genome by a synthetic riboswitch". Proceedings of the National Academy of Sciences of the United States of America. 107 (14): 6204–6209. Bibcode:2010PNAS..107.6204V. doi: 10.1073/pnas.0914423107 . PMC   2852001 . PMID   20308585.
  53. Fernandez-de-Cossio-Diaz, Jorge; Hardouin, Pierre; Moutier, Francois-Xavier Lyonnet du; Gioacchino, Andrea Di; Marchand, Bertrand; Ponty, Yann; Sargueil, Bruno; Monasson, Rémi; Cocco, Simona (2024-04-20), Designing Molecular RNA Switches with Restricted Boltzmann Machines, doi:10.1101/2023.05.10.540155 , retrieved 2024-11-10
  54. Ketzer P, Kaufmann JK, Engelhardt S, Bossow S, von Kalle C, Hartig JS, Ungerechts G, Nettelbeck DM (February 2014). "Artificial riboswitches for gene expression and replication control of DNA and RNA viruses". Proceedings of the National Academy of Sciences of the United States of America. 111 (5): E554–562. Bibcode:2014PNAS..111E.554K. doi: 10.1073/pnas.1318563111 . PMC   3918795 . PMID   24449891.
  55. Strobel B, Klauser B, Hartig JS, Lamla T, Gantner F, Kreuz S (October 2015). "Riboswitch-mediated Attenuation of Transgene Cytotoxicity Increases Adeno-associated Virus Vector Yields in HEK-293 Cells". Molecular Therapy. 23 (10): 1582–1591. doi:10.1038/mt.2015.123. PMC   4817922 . PMID   26137851.

Further reading