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doi: 10.15389/agrobiology.2021.3.465eng

UDC: 35.64:573.6.086.83:577.21]:58

Acknowledgements:
Supported financially by the Russian Science Foundation (grant No. 16-16-10043)

 

PRODUCTION AND ANALYSIS OF COMPOSITE TOMATO PLANTS Solanum lycopersicum L. CARRYING PEA GENES ENCODING THE RECEPTORS TO RHIZOBIAL SIGNAL MOLECULES

E.S. Rudaya, E.A. Dolgikh

All-Russian Research Institute for Agricultural Microbiology, Federal Agency for Scientific Organizations, 3, sh. Podbel’skogo, St. Petersburg, 196608 Russia, e-mailrudaya.s.e@gmail.com, dol2helen@yahoo.com (corresponding author ✉)

ORCID:
Rudaya E.S. orcid.org/0000-0002-3081-9880
Dolgikh E.A. orcid.org/0000-0002-5375-0943

Received December 30, 2020

The development of legume-rhizobial symbiosis is based on signal exchange between partners, which ensures their mutual recognition and activation of the infection process and the program of nodule organogenesis. In this regard, it is of great interest to study the possibility of acquisition by non-legume plants of the ability to perceive lipochito-oligosaccharide signal molecules of rhizobia, the Nod factors, and subsequent activation of signal transduction pathway. To study this possibility in our work, we carried out the transfer of the genes encoding receptors to Nod factors of legume plant pea Pisum sativum L. into tomato Solanum lycopersicum L. (Carmello cultivar) using the transformation with Agrobacterium rhizogenes. In pea, two receptor kinases, SYM10 and K1, were previously identified, which are necessary for the recognition of Nod factors during the initiation of symbiosis with rhizobia. Upon reception of Nod factors, a complex is formed between these two receptor kinases, which leads to signal transduction. In the present work, we carried out the transfer of two genes encoding LysM-RLK SYM10 and K1 in pea P. sativum into tomato plants S. lycopersicum using agrobacterial transformation. In composite plants transformed with PsSym10 or PsK1 genes, the possibility of expression activation of introduced receptor genes in response to inoculation with a typical rhizobial strain Rhizobium leguminosarum bv. viciae CIAM1026 was shown. It was also shown that, under the influence of receptors in genetically transformed roots of composite plants, the expression of genes is increased, which can be regulated by components of the “common” signal pathway. The aim of this work was to study the possibility of acquiring the ability of S. lycopersicum plants to recognize signal molecules of rhizobia after transfer of the genes encoding receptors for Nod factors in the legume plant P. sativum. Two types of constructs in the pKm43GW vector were obtained and used, in which the PsSym10 or PsK1 genes encoding receptors were cloned under the pSlEXT1 promotor of tomato extensin gene —pSlEXT1::PsSym10-3xFLAG::T35S and pSlEXT1::PsK1-RFP::T35S. Young tomato seedlings of S. lycopersicum cv. Carmello were transformed with the Agrobacterium rhizogenes Arqua 1 strain. The transformed seedlings were placed on Murashige-Skoog (MS) agar medium without sucrose in Petri dishes and cultured in an upright position in a phytotron until callus is appeared. After that, the plants were transferred to MS medium with 3 % sucrose containing 0.3 mg/ml of the antibiotic cefotaxime and incubated until transgenic roots are appeared. Composite plants were transferred into vermiculite poured with 0.5× Fahreus medium and incubated under high humidity conditions for 2-3 days. The plants were then inoculated with R. leguminosarum bv. viciae CIAM1026 containing the uidA glucuronidase gene (GUS). For the analysis we used transformed roots of composite tomato plants without rhizobial inoculation (control, 7 days), as well as transformed roots at 7 and 21 days after inoculation. The analysis of gene expression was performed by quantitative PCR combined with reverse transcription (RT-PCR). In genetically transformed roots of tomato plants the expression of both PsSym10 and PsK1 genes was observed under the pSlEXT1 promoter, moreover the expression was enhanced under the influence of rhizobial inoculation. A significant (approximately 2.0-2.5-fold) increase in the expression of the PsSym10 gene was shown in response to inoculation with rhizobia both at 7 and 21 days. The level of PsK1 expression was found to be the highest 7 days after inoculation in the transformed roots of composite tomato plants as compared to the control. To determine whether the components of the “common” signal pathway will be activated under the influence of transferred receptors in composite tomato plants, the changes in the expression of S. lycopersicum SlD27, SlNSP2, SlRAM1, and SlMAPK6 genes were assessed. These genes encode carotenoid isomerase (DWARF27) which regulates the synthesis of the hormones strigolactones, transcription factors NSP2 and RAM1, and mitogen-activated protein kinase (MAPK6). Activation of the expression of two genes, the SlNSP2 and SlMAPK6 in response to inoculation may indicate the effect of the introduced pea K1 gene on the susceptibility of tomato plants to rhizobial inoculation.

Keywords: legume-rhizobial symbiosis, receptor-like kinases, Nod factors, composite plants, gene expression.

 

REFERENCES

  1. van Velzen R., Holmer R., Bu F., Rutten L., van Zeijl A., Liu W., Santuari L., Cao Q., Sharma T., Shen D., Roswanjaya Y., Wardhani T.A.K., Kalhor M.S., Jansen J., van den Hoogen J., Güngör B., Hartog M., Hontelez J., Verver J., Yang W.C., Schijlen E., Repin R., Schilthuizen M., Schranz M.E., Heidstra R., Miyata K., Fedorova E., Kohlen W., Bisseling T., Smit S., Geurts R. Comparative genomics of the nonlegume Parasponia reveals insights into evolution of nitrogen-fixing rhizobium symbioses. Proceedings of the National Academy of Sciences, 2018, 115(20): E4700-E4709 CrossRef
  2. Murray J.D. Invasion by invitation: rhizobial infection in legumes. Molecular Plant-Microbe Interactions, 2011, 24(6): 631-639 CrossRef
  3. Cullimore J.V., Ranjeva R., Bono J.-J. Perception of lipo-chitooligosaccharidic Nod factors in legumes. Trends in Plant Science, 2001, 6(1): 24-30 CrossRef
  4. Martin F.M., Uroz S., Barker D.G. Ancestral alliances: plant mutualistic symbioses with fungi and bacteria. Science, 2017, 356(6340): eaad4501 CrossRef
  5. de Bruijn F.J. The common symbiotic signaling pathway (CSSP or SYM). In: The model legume Medicago truncatula. F.J. de Bruijn (ed.). John Wiley & Sons, New Jersey, 2020 CrossRef
  6. Diédhiou I., Diouf D. Transcription factors network in root endosymbiosis establishment and development. World Journal of Microbiology and Biotechnology, 2018, 34(3): 37 CrossRef
  7. Xue L., Klinnawee L., Zhou Y., Saridis G., Vijayakumar V., Brands M., Dörmann P., Gigolashvili T., Turck F., Bucher M. AP2 transcription factor CBX1 with a specific function in symbiotic exchange of nutrients in mycorrhizal Lotus japonicus. Proceedings of the National Academy of Sciences, 2018, 115(39): E9239-E9246 CrossRef
  8. Broghammer A., Krusell L., Blaise M., Sauer J., Sullivan J.T., Maolanon N., Vinther M., Lorentzen A., Madsen E.B., Jensen K.J., Roepstorff P., Thirup S., Ronson C.W., Thygesen M.B., Stougaard J. Legume receptors perceive the rhizobial lipochitin oligosaccharide signal molecules by direct binding. Proceedings of the National Academy of Sciences, 2012, 109(34): 13859-13864 CrossRef
  9. Fliegmann J., Canova S., Lachaud C., Uhlenbroich S., Gasciolli V., Pichereaux C., Rossignol M., Rosenberg C., Cumener M., Pitorre D., Lefebvre B., Gough C., Samain E., Fort S., Driguez H., Vauzeilles B., Beau J.M., Nurisso A., Imberty A., Cullimore J., Bono J.J. Lipo-chitooligosaccharidic symbiotic signals are recognized by LysM receptor-like kinase LYR3 in the legume Medicago truncatula. ACS Chemical Biology, 2013, 8(9): 1900-1906 CrossRef
  10. Bozsoki Z., Gysel K., Hansen S.B., Lironi D., Krönauer C., Feng F., de Jong N., Vinther M., Kamble M., Thygesen M.B., Engholm E., Kofoed C., Fort S., Sullivan J.T., Ronson C.W., Jensen K.J., Blaise M., Oldroyd G., Stougaard J., Andersen K.R., Radutoiu S. Ligand-recognizing motifs in plant LysM receptors are major determinants of specificity. Science, 2020, 369(6504): 663-670 CrossRef
  11. Bozsoki Z., Cheng J., Feng F., Gysel K., Vinther M., Andersen K.R., Oldroyd G., Blaise M., Radutoiu S., Stougaard J. Receptor-mediated chitin perception in legume roots is functionally separable from Nod factor perception. Proceedings of the National Academy of Sciences, 2017, 114(38): E8118-E8127 CrossRef
  12. Leppyanen I.V., Shakhnazarova V.Y., Shtark O.Y., Vishnevskaya N.A., Tikhonovich I.A., Dolgikh E.A. Receptor-like kinase LYK9 in Pisum sativum L. is the CERK1-like receptor that controls both plant immunity and AM symbiosis development. International Journal of Molecular Sciences, 2017, 19(1): 8 CrossRef
  13. Bisseling T., Geurts R. Specificity in legume nodule symbiosis. Science, 2020, 369(6504): 620-621 CrossRef
  14. He J., Zhang C., Dai H., Liu H., Zhang X., Yang J., Chen X., Zhu Y., Wang D., Qi X., Li W., Wang Z., An G., Yu N., He Z., Wang Y.-F., Xiao Y., Zhang P., Wang E. A LysM receptor heteromer mediates perception of arbuscular mycorrhizal symbiotic signal in rice. Molecular Plant, 2019, 12(12): 1561-1576 CrossRef
  15. Den Camp R.O., Streng A., De Mita S., Cao Q., Polone E., Liu W., Ammiraju J.S.S., Kudrna D., Wing R., Untergasser A., Bisseling T., Geurts R. LysM-type mycorrhizal receptor recruited for rhizobium symbiosis in nonlegume Parasponia. Science, 2011, 331(6019): 909-912 CrossRef
  16. Buendia L., Wang T., Girardin A., Lefebvre B. The LysM receptor-like kinase SlLYK10 regulates the arbuscular mycorrhizal symbiosis in tomato. New Phytologist, 2016, 210(1): 184-195 CrossRef
  17. Liao D., Sun X., Wang N., Song F., Liang Y. Tomato LysM receptor-like kinase SlLYK12 is involved in arbuscular mycorrhizal symbiosis. Frontiers in Plant Science, 2018, 9: 1004 CrossRef
  18. Girardin A., Wang T., Ding Y., Keller J., Buendia L., Gaston M., Ribeyre C., Gasciolli V., Auriac M.C., Vernié T., Bendahmane A., Ried M.K., Parniske M., Morel P., Vandenbussche M., Schorderet M., Reinhardt D., Delaux P.M., Bono J.J., Lefebvre B. LCO receptors involved in arbuscular mycorrhiza are functional for rhizobia perception in legumes. Current Biology, 2019, 29(24): 4249-4259.e5 CrossRef
  19. Staehelin C., Granado J., Müller J., Wiemken A., Mellor R.B., Felix G., Regenass M., Broughton W.J., Boller T. Perception of Rhizobium nodulation factors by tomato cells and inactivation by root chitinases. Proceedings of the National Academy of Sciences, 1994, 91(6): 2196-2200 CrossRef
  20. Kirienko A.N., Porozov Y.B., Malkov N. V., Akhtemova G.A., Le Signor C., Thompson R., Saffray C., Dalmais M., Bendahmane A., Tikhonovich I.A., Dolgikh E.A. Role of a receptor-like kinase K1 in pea Rhizobium symbiosis development. Planta, 2018, 248(5): 1101-1120 CrossRef
  21. Madsen E.B., Madsen L.H., Radutoiu S., Olbryt M., Rakwalska M., Szczyglowski K., Sato S., Kaneko T., Tabata S., Sandal N., Stougaard J. A receptor kinase gene of the LysM type is involved in legume perception of rhizobial signals. Nature, 2003, 425(6958): 637-640 CrossRef
  22. Murashige T., Skoog F. A revised medium for rapid growth and bio-assays with tobacco tissue cultures. Physiologia Plantarum, 1962, 15(3): 473-497 CrossRef
  23. Rival P., de Billy F., Bono J.J., Gough C., Rosenberg C., Bensmihen S. Epidermal and cortical roles of NFP and DMI3 in coordinating early steps of nodulation in Medicago truncatula. Development, 2012, 139(18): 3383-3391 CrossRef
  24. Ligero F., Lluch C., Olivares J. Evolution of ethylene from roots of Medicago sativa plants inoculated with Rhizobium meliloti. Journal of Plant Physiology, 1986, 125(3-4): 361-365 CrossRef
  25. Chiu C.H., Paszkowski U. Receptor-like kinases sustain symbiotic scrutiny. Plant Physiology, 2020, 182(4): 1597-1612 CrossRef

 

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