RNA-dependent RNA polymerase

RNA-dependent RNA polymerase (RdRP, RDR) or RNA replicase is an enzyme that catalyzes the replication of RNA from an RNA template. This is in contrast to a typical DNA-dependent RNA polymerase, which catalyzes the transcription of RNA from a DNA template.

RNA-dependent RNA polymerase
Stalled HCV RNA replicase (NS5B), in complex with sofosbuvir (PDB 4WTG).
Identifiers
EC number2.7.7.48
CAS number9026-28-2
Databases
IntEnzIntEnz view
BRENDABRENDA entry
ExPASyNiceZyme view
KEGGKEGG entry
MetaCycmetabolic pathway
PRIAMprofile
PDB structuresRCSB PDB PDBe PDBsum
Gene OntologyAmiGO / QuickGO
RNA dependent RNA polymerase[lower-alpha 1]
Identifiers
SymbolRdRP_1
PfamPF00680
Pfam clanCL0027
InterProIPR001205
SCOPe2jlg / SUPFAM
Bunyavirus RNA replicase[lower-alpha 2]
Identifiers
SymbolBunya_RdRp
PfamPF04196
InterProIPR007322
RNA-dependent RNA polymerase, eukaryotic-type
Identifiers
SymbolRdRP_euk
PfamPF05183
InterProIPR007855

RdRP is an essential protein encoded in the genomes of all RNA-containing viruses with no DNA stage, i.e. of the RNA viruses.[1][2] It catalyses synthesis of the RNA strand complementary to a given RNA template. The RNA replication process is a two-step mechanism. First, the initiation step of RNA synthesis begins at or near the 3' end of the RNA template by means of a primer-independent (de novo), or a primer-dependent mechanism that utilizes a viral protein genome-linked (VPg) primer. The de novo initiation consists in the addition of a nucleoside triphosphate (NTP) to the 3'-OH of the first initiating NTP. During the following so-called elongation phase, this nucleotidyl transfer reaction is repeated with subsequent NTPs to generate the complementary RNA product.[3][4]

History

Viral RdRPs were discovered in the early 1960s from studies on mengovirus and polio virus when it was observed that these viruses were not sensitive to actinomycin D, a drug that inhibits cellular DNA-directed RNA synthesis. This lack of sensitivity suggested that there is a virus-specific enzyme that could copy RNA from an RNA template and not from a DNA template.

Distribution

RdRPs are highly conserved throughout viruses and is even related to telomerase, though the reason for this is an ongoing question as of 2009.[5] The similarity has led to speculation that viral RdRps are ancestral to human telomerase.

The most famous example of RdRP is that of the polio virus. The viral genome is composed of RNA, which enters the cell through receptor-mediated endocytosis. From there, the RNA is able to act as a template for complementary RNA synthesis, immediately. The complementary strand is then, itself, able to act as a template for the production of new viral genomes that are further packaged and released from the cell ready to infect more host cells. The advantage of this method of replication is that there is no DNA stage; replication is quick and easy. The disadvantage is that there is no 'back-up' DNA copy.

Many RdRPs are associated tightly with membranes and are, therefore, difficult to study. The best-known RdRPs are polioviral 3Dpol, vesicular stomatitis virus L,[6] and hepatitis C virus NS5B protein.

Many eukaryotes also have RdRPs involved in RNA interference; these amplify microRNAs and small temporal RNAs and produce double-stranded RNA using small interfering RNAs as primers.[7] In fact these same RdRPs that are used in the defense mechanisms can be usurped by RNA viruses for their benefit. Their evolutionary history has been reviewed.[8]

Structure

Viral/prokaryotic RNA-directed RNA polymerases, along with many single-subunit DNA-directed polymerases, employ a fold whose organization has been likened to the shape of a right hand with three subdomains termed fingers, palm, and thumb.[9] Only the palm subdomain, composed of a four-stranded antiparallel beta sheet with two alpha helices, is well conserved among all of these enzymes. In RdRP, the palm subdomain comprises three well-conserved motifs (A, B, and C). Motif A (D-x(4,5)-D) and motif C (GDD) are spatially juxtaposed; the aspartic acid residues of these motifs are implied in the binding of Mg2+ and/or Mn2+. The asparagine residue of motif B is involved in selection of ribonucleoside triphosphates over dNTPs and, thus, determines whether RNA rather than DNA is synthesized.[10] The domain organization[11] and the 3D structure of the catalytic centre of a wide range of RdRPs, even those with a low overall sequence homology, are conserved. The catalytic centre is formed by several motifs containing a number of conserved amino acid residues.

Eukaryotic RNA interference requires a cellular RNA-dependent RNA polymerase (cRdRP). Unlike the "hand" polymerases, they resemble simplified multi-subunit DdRPs, specifically in the catalytic β/β' subunits, in that they use two sets of double-psi β-barrels in the active site. QDE1 in Neurospora crassa, which forms a homodimer, is an example of such an enzyme.[12] Bacteriophage homologs, including yonO, appear to be closer to cRdRPs than DdRPs are.[13] yonO is a DNA-dependent RNA polymerase.[14]

In viruses

There are 4 superfamilies of viruses that cover all RNA-containing viruses with no DNA stage:

  • Viruses containing positive-strand RNA or double-strand RNA, except retroviruses and Birnaviridae
    • All positive-strand RNA eukaryotic viruses with no DNA stage
    • All RNA-containing bacteriophages; there are two families of RNA-containing bacteriophages: Leviviridae (positive ssRNA phages) and Cystoviridae (dsRNA phages)
    • dsRNA virus family Reoviridae, Totiviridae, Hypoviridae, Partitiviridae
  • Mononegavirales (negative-strand RNA viruses with non-segmented genomes; InterPro: IPR016269)
  • Negative-strand RNA viruses with segmented genomes (InterPro: IPR007099), such as orthomyxoviruses and bunyaviruses
  • dsRNA virus family Birnaviridae (InterPro: IPR007100)

RNA transcription is similar to but not the same as DNA replication.

Flaviviruses produce a polyprotein from the ssRNA genome. The polyprotein is cleaved to a number of products, one of which is NS5, an RNA-dependent RNA polymerase. This RNA-directed RNA polymerase possesses a number of short regions and motifs homologous to other RNA-directed RNA polymerases.[15]

RNA replicase found in positive-strand ssRNA viruses are related to each other, forming three large superfamilies.[16] Birnaviral RNA replicase is unique in that it lacks motif C (GDD) in the palm.[17] Mononegaviral RdRP (PDB 5A22) has been automatically classified as similar to (+)-ssRNA RdRPs, specifically one from Pestivirus and one from Leviviridae.[18] Bunyaviral RdRP monomer (PDB 5AMQ) resembles the heterotrimeric complex of Orthomyxoviral (Influenza; PDB 4WSB) RdRP.[19]

Since it is a protein universal to RNA-containing viruses, RdRP is a useful marker for understanding their evolution.[20] The overall structural evolution of viral RdRPs has been reviewed.[21][22]

Recombination

When replicating its (+)ssRNA genome, the poliovirus RdRP is able to carry out recombination. Recombination appears to occur by a copy choice mechanism in which the RdRP switches (+)ssRNA templates during negative strand synthesis.[23] Recombination frequency is determined in part by the fidelity of RdRP replication.[24] RdRP variants with high replication fidelity show reduced recombination, and low fidelity RdRps exhibit increased recombination.[24] Recombination by RdRP strand switching also occurs frequently during replication in the (+)ssRNA plant carmoviruses and tombusviruses.[25]

See also
  • Spiegelman Monster
  • NS5B inhibitor

Notes
  1. See Pfam clan for other (+)ssRNA/dsRNA families.
  2. A (-)ssRNA polymerase.

References
  1. Koonin EV, Gorbalenya AE, Chumakov KM (July 1989). "Tentative identification of RNA-dependent RNA polymerases of dsRNA viruses and their relationship to positive strand RNA viral polymerases". FEBS Letters. 252 (1–2): 42–6. doi:10.1016/0014-5793(89)80886-5. PMID 2759231.
  2. Zanotto PM, Gibbs MJ, Gould EA, Holmes EC (September 1996). "A reevaluation of the higher taxonomy of viruses based on RNA polymerases". Journal of Virology. 70 (9): 6083–96. doi:10.1128/JVI.70.9.6083-6096.1996. PMC 190630. PMID 8709232.
  3. Jin Z, Leveque V, Ma H, Johnson KA, Klumpp K (March 2012). "Assembly, purification, and pre-steady-state kinetic analysis of active RNA-dependent RNA polymerase elongation complex". The Journal of Biological Chemistry. 287 (13): 10674–83. doi:10.1074/jbc.M111.325530. PMC 3323022. PMID 22303022.
  4. Kao CC, Singh P, Ecker DJ (September 2001). "De novo initiation of viral RNA-dependent RNA synthesis". Virology. 287 (2): 251–60. doi:10.1006/viro.2001.1039. PMID 11531403.
  5. Suttle CA (September 2005). "Viruses in the sea". Nature. 437 (7057): 356–61. Bibcode:2005Natur.437..356S. doi:10.1038/nature04160. PMID 16163346.
  6. Timm C, Gupta A, Yin J (August 2015). "Robust kinetics of an RNA virus: Transcription rates are set by genome levels". Biotechnology and Bioengineering. 112 (8): 1655–62. doi:10.1002/bit.25578. PMC 5653219. PMID 25726926.
  7. Iyer LM, Koonin EV, Aravind L (January 2003). "Evolutionary connection between the catalytic subunits of DNA-dependent RNA polymerases and eukaryotic RNA-dependent RNA polymerases and the origin of RNA polymerases". BMC Structural Biology. 3: 1. doi:10.1186/1472-6807-3-1. PMC 151600. PMID 12553882.
  8. Zong J, Yao X, Yin J, Zhang D, Ma H (November 2009). "Evolution of the RNA-dependent RNA polymerase (RdRP) genes: duplications and possible losses before and after the divergence of major eukaryotic groups". Gene. 447 (1): 29–39. doi:10.1016/j.gene.2009.07.004. PMID 19616606.
  9. Hansen JL, Long AM, Schultz SC (August 1997). "Structure of the RNA-dependent RNA polymerase of poliovirus". Structure. 5 (8): 1109–22. doi:10.1016/S0969-2126(97)00261-X. PMID 9309225.
  10. Gohara DW, Crotty S, Arnold JJ, Yoder JD, Andino R, Cameron CE (August 2000). "Poliovirus RNA-dependent RNA polymerase (3Dpol): structural, biochemical, and biological analysis of conserved structural motifs A and B". The Journal of Biological Chemistry. 275 (33): 25523–32. doi:10.1074/jbc.M002671200. PMID 10827187.
  11. O'Reilly EK, Kao CC (December 1998). "Analysis of RNA-dependent RNA polymerase structure and function as guided by known polymerase structures and computer predictions of secondary structure". Virology. 252 (2): 287–303. doi:10.1006/viro.1998.9463. PMID 9878607.
  12. Werner F, Grohmann D (February 2011). "Evolution of multisubunit RNA polymerases in the three domains of life". Nature Reviews. Microbiology. 9 (2): 85–98. doi:10.1038/nrmicro2507. PMID 21233849.
  13. Iyer LM, Koonin EV, Aravind L (January 2003). "Evolutionary connection between the catalytic subunits of DNA-dependent RNA polymerases and eukaryotic RNA-dependent RNA polymerases and the origin of RNA polymerases". BMC Structural Biology. 3: 1. doi:10.1186/1472-6807-3-1. PMC 151600. PMID 12553882.
  14. Forrest D, James K, Yuzenkova Y, Zenkin N (June 2017). "Single-peptide DNA-dependent RNA polymerase homologous to multi-subunit RNA polymerase". Nature Communications. 8: 15774. Bibcode:2017NatCo...815774F. doi:10.1038/ncomms15774. PMC 5467207. PMID 28585540.
  15. Tan BH, Fu J, Sugrue RJ, Yap EH, Chan YC, Tan YH (February 1996). "Recombinant dengue type 1 virus NS5 protein expressed in Escherichia coli exhibits RNA-dependent RNA polymerase activity". Virology. 216 (2): 317–25. doi:10.1006/viro.1996.0067. PMID 8607261.
  16. Koonin EV (September 1991). "The phylogeny of RNA-dependent RNA polymerases of positive-strand RNA viruses" (PDF). The Journal of General Virology. 72 ( Pt 9) (9): 2197–206. doi:10.1099/0022-1317-72-9-2197. PMID 1895057.
  17. Shwed PS, Dobos P, Cameron LA, Vakharia VN, Duncan R (May 2002). "Birnavirus VP1 proteins form a distinct subgroup of RNA-dependent RNA polymerases lacking a GDD motif". Virology. 296 (2): 241–50. doi:10.1006/viro.2001.1334. PMID 12069523.
  18. Structural Similarities for the Entities in PDB 5A22.
  19. Gerlach P, Malet H, Cusack S, Reguera J (June 2015). "Structural Insights into Bunyavirus Replication and Its Regulation by the vRNA Promoter". Cell. 161 (6): 1267–79. doi:10.1016/j.cell.2015.05.006. PMC 4459711. PMID 26004069.
  20. Wolf YI, Kazlauskas D, Iranzo J, Lucía-Sanz A, Kuhn JH, Krupovic M, Dolja VV, Koonin EV (November 2018). "Origins and Evolution of the Global RNA Virome". mBio. 9 (6). doi:10.1128/mBio.02329-18. PMC 6282212. PMID 30482837.
  21. Venkataraman S, Prasad BV, Selvarajan R (February 2018). "RNA Dependent RNA Polymerases: Insights from Structure, Function and Evolution". Viruses. 10 (2): 76. doi:10.3390/v10020076. PMC 5850383. PMID 29439438.
  22. Černý J, Černá Bolfíková B, Valdés JJ, Grubhoffer L, Růžek D (2014). "Evolution of tertiary structure of viral RNA dependent polymerases". PLOS One. 9 (5): e96070. Bibcode:2014PLoSO...996070C. doi:10.1371/journal.pone.0096070. PMC 4015915. PMID 24816789.
  23. Kirkegaard K, Baltimore D (November 1986). "The mechanism of RNA recombination in poliovirus". Cell. 47 (3): 433–43. doi:10.1016/0092-8674(86)90600-8. PMC 7133339. PMID 3021340.
  24. Woodman A, Arnold JJ, Cameron CE, Evans DJ (August 2016). "Biochemical and genetic analysis of the role of the viral polymerase in enterovirus recombination". Nucleic Acids Research. 44 (14): 6883–95. doi:10.1093/nar/gkw567. PMC 5001610. PMID 27317698.
  25. Cheng CP, Nagy PD (November 2003). "Mechanism of RNA recombination in carmo- and tombusviruses: evidence for template switching by the RNA-dependent RNA polymerase in vitro". Journal of Virology. 77 (22): 12033–47. doi:10.1128/jvi.77.22.12033-12047.2003. PMC 254248. PMID 14581540.
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