Abstract
As sessile organisms, plants need to counteract different biotic and abiotic stresses to survive. RNA interference provides natural immunity against various plant pathogens, especially against viral infections via inhibition of viral genome replication or translation. In plants, DRB3, a multi-domain protein containing two N-terminal dsRNA binding domains (dsRBD), plays a vital role in RNA-directed DNA methylation of the geminiviral genome. Additionally, DRB3 arrests the replication of the viral genome in the viral replication complex of RNA viruses through a mechanism that has yet to be fully deciphered. Therefore, as a first step towards exploring the structural details of DRB3, we present a nearly complete backbone and side chain assignment of the two N-terminal dsRBD domains.
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Data availability
Backbone and sidechain chemical shift assignments data for DRB3 dsRBD1 and DRB3 dsRBD2 have been deposited in the BMRB database with accession No. 52133 and 52151, respectively.
References
Barton DA, Roovers EF, Gouil Q, Da Fonseca GC, Reis RS, Jackson C, Overall RL, Fusaro AF, Waterhouse PM (2017) Live cell imaging reveals the relocation of dsRNA binding proteins upon viral infection. Mol Plant Microbe Interact 30(6):435–443. https://doi.org/10.1094/MPMI-02-17-0035-R
Chiliveri SC, Deshmukh MV (2015) Chemical shift assignments of DRB4 (1–153), a dsRNA binding protein in A. thaliana RNAi pathway. Biomol NMR Assign 9(2):253–256. https://doi.org/10.1007/s12104-014-9585-8
Chiliveri SC, Aute R, Rai U, Deshmukh MV (2017) DRB4 dsRBD1 drives dsRNA recognition in Arabidopsis thaliana tasi/siRNA pathway. Nucleic Acids Res 45(14):8551–8563. https://doi.org/10.1093/nar/gkx481
Clavel M, Pélissier T, Montavon T, Tschopp MA, Pouch-Pélissier MN, Descombin J, Jean V, Dunoyer P, Bousquet-Antonelli C, Deragon JM (2016) Evolutionary history of double-stranded RNA binding proteins in plants: identification of new cofactors involved in easiRNA biogenesis. Plant Mol Biol 91(1–2):131–147. https://doi.org/10.1007/s11103-016-0448-9
Diercks T, Coles M, Kessler H (1999) An efficient strategy for assignment of cross-peaks in 3D heteronuclear NOESY experiments. J Biomol NMR 15(2):177–180. https://doi.org/10.1023/A:1008367912535
Eamens AL, Kim KW, Curtin SJ, Waterhouse PM (2012a) DRB2 is required for microRNA biogenesis in Arabidopsis thaliana. PLoS ONE 7(4):e35933. https://doi.org/10.1371/journal.pone.0035933
Eamens AL, Kim KW, Waterhouse PM (2012b) DRB2, DRB3 and DRB5 function in a non-canonical microRNA pathway in Arabidopsis thaliana. Plant Signal Behav 7(10):1224–1229. https://doi.org/10.4161/psb.21518
Gleghorn ML, Maquat LE (2014) “Black sheep” that don’t leave the dsRBD fold. Trends Biochem Sci 39(7):328–340. https://doi.org/10.1016/j.tibs.2014.05.003
Hafsa NE, Arndt D, Wishart DS (2015) CSI 3.0: a web server for identifying secondary and super-secondary structure in proteins using NMR chemical shifts. Nucleic Acids Res 43(W1):W370-377. https://doi.org/10.1093/nar/gkv494
Kay LE, Xu GY, Singer AU, Muhandiram DR, Formankay JD (1993) A gradient-enhanced HCCH-TOCSY experiment for recording side-chain 1H and 13C correlations in H2O samples of proteins. J Magn Reson Series B 101(3):333–337. https://doi.org/10.1006/jmrb.1993.1053
Kay LE, Xu GY, Yamazaki T (1994) Enhanced-sensitivity triple-resonance spectroscopy with minimal H2O saturation. J Magn Reson, Series A 109(1):129–133. https://doi.org/10.1006/jmra.1994.1145
Keller R 1966 (2004) The computer aided resonance assignment tutorial. ISBN 3–85600–112–3
Klukowski P, Riek R, Guntert P (2023) NMRtist: an online platform for automated biomolecular NMR spectra analysis. Bioinformatics. https://doi.org/10.1093/bioinformatics/btad066
Lee W, Tonelli M, Markley JL (2015) NMRFAM-SPARKY: enhanced software for biomolecular NMR spectroscopy. Bioinformatics 31(8):1325–1327. https://doi.org/10.1093/bioinformatics/btu830
Lee YY, Lee H, Kim H, Kim VN, Roh SH (2023) Structure of the human DICER–pre-miRNA complex in a dicing state. Nature 615(7951):331–338. https://doi.org/10.1038/s41586-023-05723-3
Masliah G, Barraud P, Allain FHT (2013) RNA recognition by double-stranded RNA binding domains: a matter of shape and sequence. Cell Mol Life Sci 70(11):1875–1895. https://doi.org/10.1007/s00018-012-1119-x
Montelione GT, Lyons BA, Emerson SD, Tashiro M (1992) An efficient triple resonance experiment using carbon-13 isotropic mixing for determining sequence-specific resonance assignments of isotopically-enriched proteins. J Am Chem Soc 114(27):10974–10975. https://doi.org/10.1021/ja00053a051
Muhandiram DR, Kay LE (1994) Gradient-enhanced triple-resonance three-dimensional nmr experiments with improved sensitivity. J Magn Reson Series B 103(3):203–216. https://doi.org/10.1006/jmrb.1994.1032
Muhandiram R, Kay LE (2007) Three-dimensional HMQC-NOESY, NOESY-HMQC, and NOESY-HSQC. Encycl Magn Reson. https://doi.org/10.1002/9780470034590.emrstm0563
Neri D, Szyperski T, Otting G, Senn H, Wüthrich K (1989) Stereospecific nuclear magnetic resonance assignments of the methyl groups of valine and leucine in the DNA-binding domain of the 434 repressor by biosynthetically directed fractional 13C labeling. Biochemistry 28(19):7510–7516. https://doi.org/10.1021/bi00445a003
Paturi S, Deshmukh MV (2021) A glimpse of “Dicer Biology” through the structural and functional perspective. Front Mol Biosci 8:1–18. https://doi.org/10.3389/fmolb.2021.643657
Raja P, Sanville BC, Buchmann RC, Bisaro DM (2008) Viral genome methylation as an epigenetic defense against geminiviruses. J Virol 82(18):8997–9007. https://doi.org/10.1128/jvi.00719-08
Raja P, Jackel JN, Li S, Heard IM, Bisaro DM (2014) Arabidopsis double-stranded rna binding protein DRB3 Participates in methylation-mediated defense against geminiviruses. J Virol 88(5):2611–2622. https://doi.org/10.1128/JVI.02305-13
Rasia RM, Mateos J, Bologna NG, Burdisso P, Imbert L, Palatnik JF, Boisbouvier J (2010) Structure and RNA interactions of the plant MicroRNA processing-associated protein HYL1. Biochemistry 49(38):8237–8239. https://doi.org/10.1021/bi100672x
Sawano H, Matsuzaki T, Usui T, Tabara M, Fukudome A, Kanaya A, Tanoue D, Hiraguri A, Horiguchi G, Ohtani M, Demura T, Kozaki T, Ishii K, Moriyama H, Fukuhara T (2017) Double-stranded RNA-binding protein DRB3 negatively regulates anthocyanin biosynthesis by modulating PAP1 expression in Arabidopsis thaliana. J Plant Res 130(1):45–55. https://doi.org/10.1007/s10265-016-0886-0
Shen Y, Bax A (2013) Protein backbone and sidechain torsion angles predicted from NMR chemical shifts using artificial neural networks. J Biomol NMR 56(3):227–241. https://doi.org/10.1007/s10858-013-9741-y
Tants JN, Fesser S, Kern T, Stehle R, Geerlof A, Wunderlich C, Juen M, Hartlmüller C, Böttcher R, Kunzelmann S, Lange O, Kreutz C, Förstemann K, Sattler M (2017) Molecular basis for asymmetry sensing of siRNAs by the Drosophila Loqs-PD/Dcr-2 complex in RNA interference. Nucleic Acids Res 45(21):12536–12550. https://doi.org/10.1093/nar/gkx886
Vukovic L, Koh HR, Myong S, Schulten K (2014) Substrate recognition and specificity of double-stranded RNA binding proteins. Biochemistry 53(21):3457–3466. https://doi.org/10.1021/bi500352s
Wilson RC, Schneider CP, Correspondence JAD, Tambe A, Kidwell MA, Noland CL, Doudna JA (2015) Dicer-TRBP complex formation ensures accurate Mammalian microRNA biogenesis article dicer-TRBP complex formation ensures accurate Mammalian microRNA biogenesis. Mol Cell 57(3):397–407. https://doi.org/10.1016/j.molcel.2014.11.030
Wishart DS, Bigam CG, Yao J, Abildgaard F, Dyson HJ, Oldfield E, Markley JL, Sykes BD (1995) 1H, 13C and 15N chemical shift referencing in biomolecular NMR. J Biomol NMR 6(2):135–140. https://doi.org/10.1007/BF00211777
Acknowledgements
The authors acknowledge the initial assistance of Mr. Ramdas Aute, Dr. Sai Chaitanya Chiliveri, Dr. Upasana Rai, Dr. Jaishri Rubina Das, and Ms. Richa Garg in cloning DRB3 domains from drb3 full-length gene.
Funding
This work was supported by the Council of Scientific and Industrial Research, Government of India, through a CSIR-FIRST grant (MLP161). JP acknowledges Ph.D. Fellowship (JRF and SRF) from the INSPIRE program of the Department of Science and Technology (DST), Government of India.
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MVD planned the study. MVD and JP designed and performed the experiments. Both MVD and JP wrote the manuscript.
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Paul, J., Deshmukh, M.V. Chemical shift assignment of dsRBD1 and dsRBD2 of Arabidopsis thaliana DRB3, an essential protein involved in RNAi-mediated antiviral defense. Biomol NMR Assign 18, 99–104 (2024). https://doi.org/10.1007/s12104-024-10174-6
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DOI: https://doi.org/10.1007/s12104-024-10174-6