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Effect of rosR Gene Overexpression on Biofilm Formation by Rhizobium leguminosarum

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Abstract—

The regulatory protein encoded by the rosR gene is involved in the processes of adaptation of root nodule bacteria Rhizobium leguminosarum to changes in environmental conditions. It affects the expression of a number of genes associated with the synthesis of exopolysaccharides, which play a critical role in formation of bacterial biofilms on various surfaces. The goal of this work was to study the effect of overexpression of the rosR gene on formation of R. leguminosarum biofilms on inert surfaces and roots of heterologous plant hosts by obtaining recombinant strains with an additional copy of the rosR gene under the control of the Pm promoter. Our analysis of recombinant strains showed that additional expressed copies of the rosR gene allowed rhizobia to overcome the inhibitory effect on biofilm formation caused by low calcium concentrations in the medium, the effect of proteases and detergents, and to form biofilms more efficiently on the surface of tomato and clover roots. The results obtained open up prospects for using rosR as a tool both for increasing the efficiency of endosymbiosis of rhizobia with legumes, and for the formation of stable associative interactions with other agricultural crops.

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REFERENCES

  1. Baymiev, A.Kh., Yamidanov, R.S., Matniyazov, R.T., Blagova, D.K., and Chemeris, A.V., Preparation of fluorescent labeled nodule bacteria strains of wild legumes for their detection in vivo and in vitro, Mol. Biol. (Moscow), 2011, vol. 45, pp. 904–910.

    Article  CAS  Google Scholar 

  2. Bittinger, M.A., Milner, J.L., Saville, B.J., and Handelsman, J., RosR, a determinant of nodulation competitiveness in Rhizobium etli, Mol. Plant-Microbe Interact., 1997, vol. 10, pp. 180–186.

    Article  CAS  Google Scholar 

  3. Blatny, J.M., Brautaset, T., Winther-Larsen, H.C., Karunakaran, P., and Valla, S., Improved broad-host-range RK2 vectors useful for high and low regulated gene expression levels in gram-negative bacteria, Plasmid, 1997, vol. 38, pp. 35–51.

    Article  CAS  Google Scholar 

  4. Chou, A.Y., Archdeacon, J., and Kado, C.I., Agrobacterium transcriptional regulator Ros is a prokaryotic zinc finger protein that regulates the plant oncogene ipt, Proc. Natl. Acad. Sci. USA, 1998, vol. 95, pp. 5293–5298.

    Article  CAS  Google Scholar 

  5. D’Souza-Ault, M.R., Cooley, M.B., and Kado, C.I., Analysis of the Ros repressor of Agrobacterium virC and virD operons: molecular intercommunication between plasmid and chromosomal genes, J. Bacteriol., 1993, vol. 175, pp. 3486–3490.

    Article  Google Scholar 

  6. Flemming, H.C., Wingender, J., Griegbe, T., and Mayer, C., Physico-chemical properties of biofilms, in Biofilms: Recent Advances in Their Study and Control, Evans, L.V., Ed., Amsterdam: Harwood Academic Publishers, 2000, pp. 19–34.

    Book  Google Scholar 

  7. Fujishige, N.A., Kapadia, N.N., DeHoff, P.L., and Hirsch, A.M., Investigations of Rhizobium biofilm formation, FEMS Microbiol. Ecol., 2006, vol. 56, pp. 195–206.

    Article  CAS  Google Scholar 

  8. González, V., Santamaría, R.I., Bustos, P., Hernández-Gonzalez, I., Medrano-Soto, A., Moreno-Hagelsieb, G., Chandra Janga, S., Ramírez, M.A., Jiménez-Jacinto, V., Collado-Vides, J., and Dávila, G., The partitioned Rhizobium etli genome: genetic and metabolic redundancy in seven interacting replicons, Proc. Natl. Acad. Sci. USA, 2006, vol. 103, pp. 3834–3839.

    Article  Google Scholar 

  9. Janczarek, M., Krol, J., and Skorupska, A., The pssB gene product of Rhizobium leguminosarum bv. trifolii is homologous to a family of inositol monophosphatases, FEMS Microbiol. Lett., 1999, vol. 173, pp. 319–325.

    Article  CAS  Google Scholar 

  10. Janczarek, M. and Skorupska, A., The Rhizobium leguminosarum bv. trifolii RosR: transcriptional regulator involved in exopolysaccharide production, Mol. Plant-Microbe Interact., 2007, vol. 20, pp. 867–881.

    Article  CAS  Google Scholar 

  11. Janczarek, M., Jaroszuk-Ściseł, J., and Skorupska, A., Multiple copies of rosR and pssA genes enhance exopolysaccharide production, symbiotic competitiveness and clover nodulation in Rhizobium leguminosarum bv. trifolii, Antonie van Leeuwenhoek, 2009, vol. 96, pp. 471–486.

    Article  CAS  Google Scholar 

  12. Janczarek, M., Kutkowska, J., Piersiak, T., and Skorupska, A., Rhizobium leguminosarum bv. trifolii rosR is required for interaction with clover, biofilm formation and adaptation to the environment, BMC Microbiol., 2010, vol. 10, art. 284.

    Article  Google Scholar 

  13. Janczarek, M. and Skorupska, A., Modulation of rosR expression and exopolysaccharide production in Rhizobium leguminosarum bv. Trifolii by phosphate and clover root exudates, Int. J. Mol. Sci., 2011, vol. 12, pp. 4132–4155.

    Article  CAS  Google Scholar 

  14. Jaszek, M., Janczarek, M., Kuczyński, K., Piersiak, T., and Grzywnowicz, K., The response of the Rhizobium leguminosarum bv. trifolii wild-type and exopolysaccharide-deficient mutants to oxidative stress, Plant and Soil, 2014, vol. 376, pp. 75–94.

    Article  CAS  Google Scholar 

  15. Keller, M., Roxlau, A., Weng, W.M., Schmidt, M., Quand-t, J., Niehaus, K., Jording, D., Arnold, W., and Pühler, A., Molecular analysis of the Rhizobium meliloti mucR gene regulating the biosynthesis of the exopolysaccharides succinoglycan and galactoglucan, Mol. Plant-Microbe Interact., 1995, vol. 8, pp. 267–277.

    Article  CAS  Google Scholar 

  16. Kopycińska, M., Lipa, P., Cieś la, J., Kozieł, M., and Janczarek, M., Extracellular polysaccharide protects Rhizobium leguminosarum cells against zinc stress in vitro and during symbiosis with clover, Environ. Microbiol. Rep., 2018, vol. 10, pp. 355–368.

    Article  Google Scholar 

  17. Livak, K.J. and Schmittgen, T.D., Analysis of relative gene expression data using real-time quantitative PCR and the 2‑ΔΔCT method, Methods, 2001, vol. 25, pp. 402–408.

    Article  CAS  Google Scholar 

  18. Murashige, T. and Skoog, F., A revised medium for rapid growth and bioassays with tobacco tissue cultures, Physiol. Plant, 1962, vol. 15, pp. 473–497.

    Article  CAS  Google Scholar 

  19. O’Toole, G.A. and Kolter, R., Initiation of biofilm formation in Pseudomonas fluorescens WCS365 proceeds via multiple, convergent signalling pathways: a genetic analysis, Mol. Microbiol., 1998, vol. 28. pp. 449–461.

    Article  Google Scholar 

  20. Rachwał, K., Matczyńska, E., and Janczarek, M., Transcriptome profiling of a Rhizobium leguminosarum bv. trifolii rosR mutant reveals the role of the transcriptional regulator RosR in motility, synthesis of cell-surface components, and other cellular processes, BMC Genomics, 2015, vol. 16, art. 1111. https://doi.org/10.1186/s12864-015-2332-4

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Rachwał, K., Boguszewska, A., Kopcińska, J., Karaś, M., Tchórzewski, M., and Janczarek, M., The regulatory protein RosR affects Rhizobium leguminosarum bv. trifolii protein profiles, cell surface properties, and symbiosis with clover, Front. Microbiol., 2016, vol. 7, art. 1302, pp. 1–21.

  22. Rachwał, K., Lipa, P., Wojda, I., Vinardell, J.M., and Janczarek, M., Regulatory elements located in the upstream region of the Rhizobium leguminosarum rosR global regulator are essential for its transcription and mRNA stability, Genes, 2017, vol. 8, art. 388.

    Article  Google Scholar 

  23. Reeve, W.G., Dilworth, M.J., Tiwari, R.P., and Glenn, A.R., Regulation of exopolysaccharide production in Rhizobium leguminosarum biovar viciae WSM710 involves exoR, Microbiology (SGM), 1996, vol. 143, pp. 1951–1958.

    Article  Google Scholar 

  24. Rinaudi, L., Fujishige, N.A., Hirsch, A.M., Banchio, E., Zorreguieta, A., and Giordano, W., Effects of nutritional and environmental conditions on Sinorhizobium meliloti biofilm formation, Res. Microbiol., 2006, vol. 157, pp. 867–875.

    Article  CAS  Google Scholar 

  25. Rinaudi, L.V. and Gonzalez, J.E., The low-molecular-weight fraction of exopolysaccharide II from Sinorhizobium meliloti is a crucial determinant of biofilm formation, J. Bacteriol., 2009, vol. 191, pp. 7216–7224.

    Article  CAS  Google Scholar 

  26. Sambrook, J., Fritsch, E., and Maniatis, T., Molecular Cloning: A Laboratory Manual, N.Y.: Cold Spring Harbor Lab. Press, 1989.

    Google Scholar 

  27. Santos, R., Herouart, D., Sigaud, S., Touati, D., and Puppo, A., Oxidative burst in alfalfa-Sinorhizobium meliloti symbiotic interaction, Mol. Plant-Microbe Interact., 2001, vol. 14, pp. 86–89.

    Article  CAS  Google Scholar 

  28. Smith, R.J., Calcium and bacteria, Adv. Microb. Physiol., 1995, vol. 37, pp. 83–133.

    Article  CAS  Google Scholar 

  29. Vershinina, Z.R., Blagova, D.K., Nigmatullina, L.R., Lavina, A.M., Baymiev, Al.Kh., and Chemeris, A.V., Associative symbiosis of transgenic tomatoes with rhizobia increases plant resistance to Fusarium oxysporum f. sp. lycopersici, Biotekhnologiya, 2015, vol. 31, no. 3. pp. 42–53.

    Article  Google Scholar 

  30. Vershinina, Z.R., Khakimova, L.R., Lavina, A.M., Karimova, L.R., Fedyaev, V.V., Baymiev, An.Kh., and Baymiev, Al.Kh., Interaction of tomato (Solanum lycopersicum L.) transformed with rapAl with bacteria Pseudomonas sp. 102 resistant to high cadmium concentrations as a basis of an efficient symbiotic phytoremediation system, Biotekhnologiya, 2019, vol. 35, no. 2, pp. 38–48.

    Article  Google Scholar 

  31. Vershinina, Z.R., Lavina, A.M., and Chubukova, O.V., Exopolysaccharides of Rhizobium leguminosarum: a brief review, Biomika, 2020, vol. 12, pp. 27–49.

    Google Scholar 

  32. Vincent, J.M., A Manual for Practical Study of Root Nodule Bacteria, IBP Handbook No. 15, Oxford: Blackwell Scientific Publishers, 1970.

  33. Wimpenny, J.W.T. and Colasanti, R.A., A unifying hypothesis for the structure of microbial films based on cellular automation models, FEMS Microbiol. Ecol., 1997, vol. 22. pp. 1–16.

    Article  CAS  Google Scholar 

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Funding

This work was performed using the equipment of the Regional Center of Shared Use “Agidel” (Ufa Federal Research Center of the Russian Academy of Sciences) as a part of State Assignment no. AAAA-A16-116020350028-4; it was financially supported by the Russian Foundation for Basic Research, project no. 18-34-20004 mol_a_ved.

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Authors and Affiliations

  1. Institute of Biochemistry and Genetics, Subdivision of the Ufa Federal Research Center of the Russian Academy of Sciences, 450098, Ufa, Russia

    Z. R. Vershinina, O. V. Chubukova, Yu. M. Nikonorov, L. R. Khakimova, A. M. Lavina, L. R. Karimova, An. Kh. Baymiev & Al. Kh. Baymiev

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  1. Z. R. Vershinina
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  2. O. V. Chubukova
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  3. Yu. M. Nikonorov
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  4. L. R. Khakimova
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  5. A. M. Lavina
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  6. L. R. Karimova
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  7. An. Kh. Baymiev
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  8. Al. Kh. Baymiev
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Correspondence to Z. R. Vershinina.

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Translated by D. Timchenko

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Vershinina, Z.R., Chubukova, O.V., Nikonorov, Y.M. et al. Effect of rosR Gene Overexpression on Biofilm Formation by Rhizobium leguminosarum . Microbiology 90, 198–209 (2021). https://doi.org/10.1134/S0026261721020144

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