Abstract
Exploration of molecular structure of β-tubulin is key to understand mechanism of action of carbendazim since its activity depends on strong binding to β-tubulin. Resistance against the fungicide is often associated with mutation in β-tubulin gene. A full-length (1619 bp) β-tubulin gene has been cloned and sequenced from a carbendazim resistant and a sensitive isolates of F. solani isolated from agricultural fields of Murshidabad (24.23 °N, 88.25 °E), West Bengal, India. Phylogenetic position of the isolates was confirmed using internal transcribed spacer and β-tubulin gene sequences. In the β-tubulin based phylogenetic tree, Fusarium species with available data were clustered in nine species complexes and members of both F. solani species complex and F. fujikuroi species complex were distributed into three clades each. The β-tubulin gene of F. solani was found to be shortest due to least number of non-coding sequences indicating its primitiveness among the Fusarium species. The coding region (G + C 58.54%) was organized into five exons. The protein has 446 amino acid, 49.834 KD molecular weight and 4.64 isoelectric point. Amino acid sequence of the resistant and the sensitive isolates were identical, suggesting that the mechanism of carbendazim resistance in the F. solani isolate was not due to point mutation in β-tubulin gene. The secondary and tertiary structure of β-tubulin were similar in all the species except F. oxysporum f.sp. cubense. The identification of binding sites for GDP, carbendazim and α-tubulin would resolve how carbendazim prevents tubulin polymerization. All the data are useful to design tubulin-targeted fungicide with better performance.
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References
Albertini C, Gredt M, Leroux P (1999) Mutations of the β-tubulin gene associated with different phenotypes of benzimidazole resistance in the cereal eyespot fungi Tapesia yallundae and Tapesia acuformis. Pestic Biochem Physiol 64:17–31. https://doi.org/10.1006/pest.1999.2406
Al-Hatmi AMS, Ahmed SA, van Diepeningen AD, Drogari-Apiranthitou M, Verweij PE, Meis JF, de Hoog GS (2018) Fusarium metavorans sp. nov.: the frequent opportunist “FSSC6.” Med Mycol 56:S144–S152. https://doi.org/10.1093/mmy/myx107
Andrade AC, Del Sorbo G, Van Nistelrooy JGM, De Waard MA (2000) The ABC transporter AtrB from Aspergillus nidulans mediates resistance to all major classes of fungicides and some natural toxic compounds. Microbiology 146:1987–1997. https://doi.org/10.1099/00221287-146-8-1987
Aoki T, O’Donnell K, Geiser DM (2014) Systematics of key phytopathogenic Fusarium species: current status and future challenges. J Gen Plant Pathol 80:189–201. https://doi.org/10.1007/s10327-014-0509-3
Avenot HF, Morgan DP, Quattrini J, Michailides TJ (2020) Resistance to thiophanate-methyl in Botrytis cinerea isolates from Californian vineyards and Pistachio and Pomegranate Orchards. Plant Dis 104:1069–1075. https://doi.org/10.1094/PDIS-02-19-0353-RE
Begerow D, John B, Oberwinkler F (2004) Evolutionary relationships among β-tubulin gene sequences of basidiomycetous fungi. Mycol Res 108:1257–1263. https://doi.org/10.1017/S0953756204001066
Benkert P, Künzli M, Schwede T (2009) QMEAN server for protein model quality estimation. Nucleic Acids Res 37:510–514. https://doi.org/10.1093/nar/gkp322
Bogale M, Steenkamp ET, Wingfield MJ, Wingfield BD (2009) Diverse Fusarium solani isolates colonise agricultural environments in Ethiopia. Eur J Plant Pathol 124:369–378
Chen C, Wang J, Luo Q, Yuan S, Zhou M (2007) Characterization and fitness of carbendazim-resistant strains of Fusarium graminearum (wheat scab). Pest Manag Sci 63:1201–1207. https://doi.org/10.1002/ps.1449
Chen Z, Gao T, Liang S et al (2014) Molecular mechanism of resistance of Fusarium fujikuroi to benzimidazole fungicides. FEMS Microbiol Lett 357:77–84. https://doi.org/10.1111/1574-6968.12504
Chou PY, Fasman GD (1974) Conformational parameters for amino acids in helical, β-sheet, and random coil regions calculated from proteins. Biochemistry 13:211–222. https://doi.org/10.1021/bi00699a001
Chou PY, Fasman GD (1978) Empirical predictions of protein conformation. Annu Rev Biochem 47:251–276. https://doi.org/10.1146/annurev.bi.47.070178.001343
Coleman JJ (2016) The Fusarium solani species complex: ubiquitous pathogens of agricultural importance. Mol Plant Pathol 17:146–158. https://doi.org/10.1111/mpp.12289
D’Addazio V, dos Santos RAA, Leitao ASB et al (2016) Evaluation of in vitro inhibition of mycelial growth of Fusarium solani f. sp. piperis by different products in Brazil. Afric J Microbiol Res 10(47):1992–1998. https://doi.org/10.58897/AJMR2016.8292
De Vries SJ, Van Dijk M, Bonvin AMJJ (2010) The HADDOCK web server for data-driven biomolecular docking. Nat Protoc 5:883–897. https://doi.org/10.1038/nprot.2010.32
Deber CM, Glibowicka M, Woolley GA (1990) Conformations of proline residues in membrane environments. Biopolymers 29:149–157. https://doi.org/10.1002/bip.360290120
Dominguez C, Boelens R, Bonvin AMJJ (2003) HADDOCK: a protein-protein docking approach based on biochemical or biophysical information. J Am Chem Soc 125:1731–1737. https://doi.org/10.1021/ja026939x
Dorugade SP, Madhale SV, Sardesai VA, Kamble SS (2021) Survival ability of benomyl resistant isolate of Fusarium solani causing dry rot of elephant foot yam. Bioinfolet 18(1B):197–198
Dutta S, Tarafder M, Islam R, Datta B (2018) Characterization of cellulolytic enzymes of Fusarium soil Isolates. Biocatal Agric Biotechnol 14:279–285
Einax E, Voigt K (2003) Oligonucleotide primers for universal amplification of β-tubulin genes facilitate phylogenetic analyses in the regnum Fungi. Org Divers Evol 3(3):185–194
Geiser DM, Al-Hatmi A, Aoki T, et al (2020) Phylogenomic analysis of a 55.1 kb 19-gene dataset resolves a monophyletic Fusarium that includes the Fusarium solani Species Complex. Phytopathology® 1–64. https://doi.org/10.1094/phyto-08-20-0330-le
Grosdidier A, Zoete V, Michielin O (2007) EADock: docking of small molecules into protein active sites with a multiobjective evolutionary optimization. Proteins 67:1010–1025. https://doi.org/10.1002/prot.21367
Gupta PK, Singh SK, Shikha S (2020) In vitro efficacy of different fungicides against Fusarium solani isolate causing root rot of papaya (Carica papaya L.). Int J Chem Stud 8(3):221–224. https://doi.org/10.22271/chemi.2020.v8.i3c.9229
Herron DA, Wingfield MJ, Wingfield BD et al (2015) Novel taxa in the Fusarium fujikuroi species complex from Pinus spp. Stud Mycol 80:131–150. https://doi.org/10.1016/j.simyco.2014.12.001
Horio T, Murata T, Murata T (2014) The role of dynamic instability in microtubule organization. Front Plant Sci 5:1–11. https://doi.org/10.3389/fpls.2014.00511
Hurley JH, Mason DA, Matthews BW (1992) Flexible-geometry conformational energy maps for the amino acid residue preceding a proline. Biopolymers 32:1443–1446. https://doi.org/10.1002/bip.360321104
Kawchuk LM, Hutchison LJ, Verhaeghe CA et al (2002) Isolation of the β-tubulin gene and characterization of thiabendazole resistance in Gibberella pulicaris. Can J Plant Pathol 24:233–238. https://doi.org/10.1080/07060660309507001
Kolattukudy PE, Gamble DL (1995) Nectria haematococca: pathogenesis and host specificity in plant diseases. In: Kohmoto K, Singh US, Singh RP (eds) Pathogenesis and host specificity in plant pathogenic fungi and nematodes, vol 2. Eukaryotes Pergamon, Oxford, pp 83–102
Kozakov D, Hall DR, Xia B et al (2017) The ClusPro web server for protein-protein docking. Nat Protoc 12:255–278. https://doi.org/10.1038/nprot.2016.169
Kumar S, Stecher G, Li M et al (2018) MEGA X: Molecular evolutionary genetics analysis across computing platforms. Mol Biol Evol 35:1547–1549. https://doi.org/10.1093/molbev/msy096
Leslie JF, Summerell BA (2008) The Fusarium laboratory manual. Wiley, New York
Liu S, Liu J, Fu L et al (2020) Baseline sensitivity of Fusarium graminearum from wheat fields in Henan, China, to metconazole and analysis of cross resistance with carbendazim and phenamacril. J Phytopathol 168:156–161. https://doi.org/10.1111/jph.12874
Ma Z, Michailides TJ (2005) Advances in understanding molecular mechanisms of fungicide resistance and molecular detection of resistant genotypes in phytopathogenic fungi. Crop Prot 24:853–863. https://doi.org/10.1016/j.cropro.2005.01.011
MacArthur MW, Thornton JM (1991) Influence of proline residues on protein conformation. J Mol Biol 218:397–412. https://doi.org/10.1016/0022-2836(91)90721-H
Msiska Z, Morton JB (2009) Isolation and sequence analysis of a beta-tubulin gene from arbuscular mycorrhizal fungi. Mycorrhiza 19:501–513. https://doi.org/10.1007/s00572-009-0248-z
Nakaune R, Nakano M (2007) Benomyl resistance of Colletotrichum acutatum is caused by enhanced expression of β-tubulin 1 gene regulated by putative leucine zipper protein CaBEN1. Fungal Genet Biol 44:1324–1335. https://doi.org/10.1016/j.fgb.2007.03.007
Nakaune R, Adachi K, Nawata O et al (1998) A novel ATP-binding cassette transporter involved in multidrug resistance in the phytopathogenic fungus Penicillium digitatum. Appl Environ Microbiol 64:3983–3988. https://doi.org/10.1128/aem.64.10.3983-3988.1998
Narayanan P et al (2015) Efficacy of bio-control agents and fungicides in management of mulberry wilt caused by Fusarium solani. http://ischolar.info/index.php/jbc/article/view/79116
Nogales E, Downing KH, Amos LA, Lowe J (1998) Tubulin and FtsZ form a distinct family of GTPases. Nat Struct Mol Biol 5:451–458. https://doi.org/10.1038/nsb0698-45
O’Donnell K (2000) Molecular phylogeny of the Nectria haematococca-Fusarium solani species complex. Mycologia 92:919–938. https://doi.org/10.1080/00275514.2000.12061237
O’Donnell K, Cigelnik E (1997) Two divergent intragenomic rDNA ITS2 types within a monophyletic lineage of the fungus fusarium are nonorthologous. Mol Phylogenet Evol 7:103–116. https://doi.org/10.1006/mpev.1996.0376
O’Donnell K, Rooney AP, Proctor RH et al (2013) Phylogenetic analyses of RPB1 and RPB2 support a middle Cretaceous origin for a clade comprising all agriculturally and medically important fusaria. Fungal Genet Biol 52:20–31. https://doi.org/10.1016/j.fgb.2012.12.004
O’Donnell K, Al-Hatmi AMS, Aoki T et al (2020) No to Neocosmospora: phylogenomic and practical reasons for continued inclusion of the Fusarium solani species complex in the genus Fusarium. mSphere. https://doi.org/10.1128/msphere.00810-20
O’Donnell K, Cigelnik E, Nirenberg HI (1998) Molecular systematics and phylogeography of the Gibberella fujikuroi species complex. Mycologia 90:465–493. https://doi.org/10.1080/00275514.1998.12026933
Padvi SA, Hingole DG, Khaire PB (2018) In vitro efficacy of fungicides against Fusarium solani incited by dry root rot of sweet orange. J Pharmaconosy Phytochem 7(4):3270–3273
Petkar A, Langston DB, Buck JW et al (2017) Sensitivity of Fusarium oxysporum f. sp. niveum to prothioconazole and thiophanate-methyl and gene mutation conferring resistance to thiophanate-methyl. Plant Dis 101:366–371. https://doi.org/10.1094/PDIS-09-16-1236-RE
Pettersen EF, Goddard TD, Huang CC et al (2004) UCSF Chimera—a visualization system for exploratory research and analysis. J Comput Chem 25:1605–1612. https://doi.org/10.1002/jcc.20084
Porollo AA, Adamczak R, Meller J (2004) POLYVIEW: a flexible visualization tool for structural and functional annotations of proteins. Bioinformatics 20:2460–2462. https://doi.org/10.1093/bioinformatics/bth248
Qiu JB, Xu JQ, Yu JJ et al (2011) Localisation of the benzimidazole fungicide binding site of Gibberella zeae β2-tubulin studied by site-directed mutagenesis. Pest Manag Sci 67:191–198. https://doi.org/10.1002/ps.2050
Richards KL, Anders KR, Nogales E et al (2000) Structure-function relationships in yeast tubulins. Mol Biol Cell 11:1887–1903. https://doi.org/10.1091/mbc.11.5.1887
Richardson JS, Richardson DC (1988) Amino acid preferences for specific locations at the ends of α helices. Science 240(4859):1648–1652. https://doi.org/10.1126/science.242.4886.1624-b
Richardson JS, Richardson DC (1989) Principles and patterns of protein conformation. Predict Protein Struct Princ Protein Conform. https://doi.org/10.1007/978-1-4613-1571-1_1
Schroers HJ, Samuels GJ, Zhang N, Short DP, Juba J, Geiser DM (2016) Epitypification of Fusisporium (Fusarium) solani and its assignment to a common phylogenetic species in the Fusarium solani species complex. Mycologia 108:806–819
Sevastos A, Labrou NE, Flouri F, Malandrakis A (2017) Glutathione transferase-mediated benzimidazole-resistance in Fusarium graminearum. Pestic Biochem Physiol 141:23–28. https://doi.org/10.1016/j.pestbp.2016.11.002
Smith JA, Pease LG, Kopple KD (1980) Reverse turns in peptides and protein. Crit Rev Biochem Mol Biol 8:315–399. https://doi.org/10.3109/10409238009105470
Suga H, Nakajima T, Kageyama K, Hyakumachi M (2011) The genetic profile and molecular diagnosis of thiophanate-methyl resistant strains of Fusarium asiaticum in Japan. Fungal Biol 115:1244–1250. https://doi.org/10.1016/j.funbio.2011.08.009
Tamura K, Nei M (1993) Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Mol Biol Evol 10:512–526. https://doi.org/10.1093/oxfordjournals.molbev.a040023
Tarafder M, Islam R, Datta B (2019) Carbendazim sensitivity of Fusarium solani soil isolates. Indian Phytopathol 72:705–710. https://doi.org/10.1007/s42360-019-00151-8
Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680. https://doi.org/10.1093/nar/22.22.4673
Vajda S, Yueh C, Beglov D et al (2017) New additions to the ClusPro server motivated by CAPRI. Proteins Struct Funct Bioinforma 85:435–444. https://doi.org/10.1002/prot.25219
Vela-Corcia D, Romero D, De Vicente A, Perez-Garcia A (2018) Analysis of β-tubulin-carbendazim interaction reveals that binding site for MBC fungicides does not include residues involved in fungicide resistance. Sci Rep 8:1–12. https://doi.org/10.1038/s41598-018-25336-5
Víglaš J, Olejníkova P (2021) An update on ABC transporters of filamentous fungi—from physiological substrates to xenobiotics. Microbiol Res 246:126684. https://doi.org/10.1016/j.micres.2020.126684
Wiederstein M, Sippl MJ (2007) ProSA-web: Interactive web service for the recognition of errors in three-dimensional structures of proteins. Nucleic Acids Res 35:407–410. https://doi.org/10.1093/nar/gkm290
Williams KA, Deber CM (1991) Proline residues in transmembrane helices: structural or dynamic role? Biochemistry 30:8919–8923. https://doi.org/10.1021/bi00101a001
Xu S, Wang J, Wang H et al (2019) Molecular characterization of carbendazim resistance of Fusarium species complex that causes sugarcane pokkah boeng disease. BMC Genom 20:1–14. https://doi.org/10.1186/s12864-019-5479-6
Yan K, Dickman MB (1996) Isolation of a β-tubulin gene from Fusarium moniliforme that confers cold-sensitive benomyl resistance. Appl Environ Microbiol 62:3053–3056. https://doi.org/10.1128/aem.62.8.3053-3056.1996
Yang Y, Li MX, Duan YB et al (2018) A new point mutation in β2-tubulin confers resistance to carbendazim in Fusarium asiaticum. Pestic Biochem Physiol 145:15–21. https://doi.org/10.1016/j.pestbp.2017.12.006
Zhang H, Brankovics B, van der Lee TAJ et al (2016) A single-nucleotide-polymorphism-based genotyping assay for simultaneous detection of different carbendazim-resistant genotypes in the Fusarium graminearum species complex. PeerJ. https://doi.org/10.7717/peerj.2609
Zhao Z, Liu H, Luo Y et al (2014) Molecular evolution and functional divergence of tubulin superfamily in the fungal tree of life. Sci Rep 4:1–13. https://doi.org/10.1038/srep06746
Zhou Y, Xu J, Zhu Y et al (2016) Mechanism of action of the benzimidazole fungicide on Fusarium graminearum: interfering with polymerization of monomeric tubulin but not polymerized microtubule. Phytopathology 106:807–813. https://doi.org/10.1094/PHYTO-08-15-0186-R
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The work has been supported by the funds received from University of Kalyani and DST-FIST and DST-PURSE programme II, Govt. of India
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Tarafder, M., Datta, B. Deciphering β-tubulin gene of carbendazim resistant Fusarium solani isolate and its comparison with other Fusarium species. Curr Genet 68, 429–447 (2022). https://doi.org/10.1007/s00294-022-01238-y
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DOI: https://doi.org/10.1007/s00294-022-01238-y