Plant Molecular Biology

, Volume 96, Issue 4–5, pp 339–351 | Cite as

Mitogen activated protein kinase 6 and MAP kinase phosphatase 1 are involved in the response of Arabidopsis roots to l-glutamate

  • Jesús Salvador López-Bucio
  • Javier Raya-González
  • Gustavo Ravelo-Ortega
  • León Francisco Ruiz-Herrera
  • Maricela Ramos-Vega
  • Patricia León
  • José López-Bucio
  • Ángel Arturo Guevara-García


Key message

The function and components of l-glutamate signaling pathways in plants have just begun to be elucidated. Here, using a combination of genetic and biochemical strategies, we demonstrated that a MAPK module is involved in the control of root developmental responses to this amino acid.


Root system architecture plays an essential role in plant adaptation to biotic and abiotic factors via adjusting signal transduction and gene expression. l-Glutamate (l-Glu), an amino acid with neurotransmitter functions in animals, inhibits root growth, but the underlying genetic mechanisms are poorly understood. Through a combination of genetic analysis, in-gel kinase assays, detailed cell elongation and division measurements and confocal analysis of expression of auxin, quiescent center and stem cell niche related genes, the critical roles of l-Glu in primary root growth acting through the mitogen-activated protein kinase 6 (MPK6) and the dual specificity serine–threonine–tyrosine phosphatase MKP1 could be revealed. In-gel phosphorylation assays revealed a rapid and dose-dependent induction of MPK6 and MPK3 activities in wild-type Arabidopsis seedlings in response to l-Glu. Mutations in MPK6 or MKP1 reduced or increased root cell division and elongation in response to l-Glu, possibly modulating auxin transport and/or response, but in a PLETHORA1 and 2 independent manner. Our data highlight MPK6 and MKP1 as components of an l-Glu pathway linking the auxin response, and cell division for primary root growth.


Arabidopsis l-Glutamate MPK6 MKP1 Auxins Root development 



We thank Patricia Jarillo for technical support. We appreciate the kind donation of seeds by Drs. Ben Scheres, Alfredo Cruz-Ramírez, Shuqun Zhang, Scot C. Peck and Marina A. González Basteiro. This work was supported by Universidad Nacional Autónoma de México (UNAM)-Dirección General de Asuntos del Personal Académico (DGAPA)-Programa de Apoyo a Proyectos de Investigación e Innovación Tecnológica (PAPIIT) (Grants: IN207014 & IN210917 to AGG) and Consejo Nacional de Ciencia y Tecnología (CONACYT)-México (Grants: CB2015-251848 to AGG and CB-177775 to JLB).

Author contributions

JSL-B, JR-G, GR-O, FR-H, MR-V: planning and execution of experiments and analysis of data/results. PL: funding, discussion of results and editing of the manuscript. JL-B: experiment planning, data/results analysis and writing/editing of the manuscript. AAG-G: funding, experiment planning, data/results analysis and writing/editing of the manuscript.

Supplementary material

11103_2018_699_MOESM1_ESM.pdf (42.6 mb)
Supplementary material 1 (PDF 43619 KB)


  1. Aida M, Beis D, Heidstra R, Willemsen V, Blilou I, Galinha C, Nussaume L, Noh YS, Amasino R, Scheres B (2004) The PLETHORA genes mediate patterning of the Arabidopsis root stem cell niche. Cell 119:109–120CrossRefPubMedGoogle Scholar
  2. Alonso JM, Stepanova AN, Leisse TJ, Kim CJ, Chen H, Shinn P, Stevenson DK, Zimmerman J, Barajas P, Cheuk R et al (2003) Genome-wide insertional mutagenesis of Arabidopsis thaliana. Science 301:653–657CrossRefPubMedGoogle Scholar
  3. Andreasson E, Ellis B (2009) Convergence and specificity in the Arabidopsis MAPK nexus. Trends Plant Sci 15:106–113CrossRefGoogle Scholar
  4. Asai T, Tena G, Plotnikova J, Willmann MR, Chiu WL, Gomez-Gomez L, Boller T, Ausubel FM, Sheen J (2002) MAP kinase signalling cascade in Arabidopsis innate immunity. Nature 415:977–983CrossRefPubMedGoogle Scholar
  5. Bartels S, Anderson JC, Gonzalez Besteiro MA, Carreri A, Hirt H, Buchala A, Metraux JP, Peck SC, Ulm R (2009) MAP kinase phosphatase1 and protein tyrosine phosphatase1 are repressors of salicylic acid synthesis and SNC1-mediated responses in Arabidopsis. Plant Cell 21:2884–2897CrossRefPubMedPubMedCentralGoogle Scholar
  6. Bartels S, González Besteiro MA, Lang D, Ulm R (2010) Emerging functions for plant MAP kinase phosphatases. Trends Plant Sci 15:322–329CrossRefPubMedGoogle Scholar
  7. Benková E, Michniewicz M, Sauer M, Teichmann T, Seifertov D Jürgens G, Friml J (2003) Local, efflux-dependent auxin gradients as a common module for plant organ formation. Cell 115:591–602CrossRefPubMedGoogle Scholar
  8. Biancucci M, Mattioli R, Moubayidin L, Sabatini S, Constantino P, Trovato M (2015) Proline affects the size of the root meristematic zone in Arabidopsis. BMC Plant Biol 15:263CrossRefPubMedPubMedCentralGoogle Scholar
  9. Blilou I, Xu J, Wildwater M, Willemsen V, Paponov I, Friml J, Heidstra R, Scheres B (2005) The PIN auxin efflux facilitator network controls growth and patterning in Arabidopsis roots. Nature 443:39–44CrossRefGoogle Scholar
  10. Bush SM, Krysan PJ (2007) Mutational evidence that the Arabidopsis MAP kinase MPK6 is involved in anther, inflorescence, and embryo development. J Exp Bot 58:2181–2191CrossRefPubMedGoogle Scholar
  11. Casimiro I, Marchant A, Bhalerao R, Beeckman T, Dhooge S, Swarup R, Graham N, Inzé D, Sandberg G, Casero P, Bennett M (2001) Auxin transport promotes Arabidopsis lateral root initiation. Plant Cell 13:843–852CrossRefPubMedPubMedCentralGoogle Scholar
  12. Cederholm HM, Iyer-Pascuzzi AS, Benfey P (2012) Patterning the primary root in Arabidopsis. Dev Biol 1:675–691Google Scholar
  13. Chapman E, Estelle M (2009) Mechanisms of auxin-regulated gene expression in plants. Annu Rev Genet 43:265–285CrossRefPubMedGoogle Scholar
  14. Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743CrossRefPubMedGoogle Scholar
  15. Colcombet J, Hirt H (2008) Arabidopsis MAPKs: a complex signalling network involved in multiple biological processes. Biochem J 413:217–226CrossRefPubMedGoogle Scholar
  16. Colón-Carmona A, You R, Haimovitch-Gal T, Doerner P (1999) Spatio-temporal analysis of mitotic activity with a labile cyclin-GUS fusion protein. Plant J 20:503–508CrossRefPubMedGoogle Scholar
  17. Contreras-Cornejo HA, López-Bucio JS, Méndez-Bravo A, Macías-Rodríguez L, Ramos-Vega M, Guevara-García A, López-Bucio J (2015) Mitogen-activated protein kinase 6 and ethylene and auxin signaling pathways are involved in Arabidopsis root-system architecture alterations by Trichoderma atroviride. Mol Plant Microbe Interact 28:701–710CrossRefPubMedGoogle Scholar
  18. Ding Z, Friml J (2010) Auxin regulates distal stem cell differentiation in Arabidopsis roots. Proc Natl Acad Sci USA 107:12046–12051CrossRefPubMedPubMedCentralGoogle Scholar
  19. Djamei A, Pitzschke A, Nakagami H, Rajh I, Hirt H (2007) Trojan horse strategy in Agrobacterium transformation: abusing MAPK defense signaling. Science 318:453–456CrossRefPubMedGoogle Scholar
  20. Dolan L, Janmaat K, Willemsen V, Linstead P, Poethig S, Roberts K, Scheres B (1993) Cellular organization of the Arabidopsis thaliana root. Development 119:71–84PubMedGoogle Scholar
  21. Fiil BK, Petersen K, Petersen M, Mundy J (2009) Gene regulation by MAP kinase cascades. Curr Opin Plant Biol 12:615–621CrossRefPubMedGoogle Scholar
  22. Forde BG (2014) Glutamate signalling in roots. J Exp Bot 65:779–787CrossRefPubMedGoogle Scholar
  23. Forde BG, Roberts MR (2014) Glutamate receptor-like channels in plants: a role as amino acid sensors in plant defence? F1000 Prime Rep 6 (37):7Google Scholar
  24. Forde BG, Cutler SR, Zaman N, Krysan PJ (2013) Glutamate signalling via a MEKK1 kinase-dependent pathway induces changes in Arabidopsis root architecture. Plant J 75:1–10CrossRefPubMedPubMedCentralGoogle Scholar
  25. Franklin KA, Lee SH, Patel D, Kumar SV, Spartz AK, Gu C, Ye S, Yu P, Breen G, Cohen JD et al (2011) Phytochrome-interacting factor 4 (PIF4) regulates auxin biosynthesis at high temperature. Proc Natl Acad Sci USA 108:20231–20235CrossRefPubMedPubMedCentralGoogle Scholar
  26. Fukaki H, Tasaka M (2009) Hormone interactions during lateral root formation. Plant Mol Biol 69:437–449CrossRefPubMedGoogle Scholar
  27. Galinha C, Hofhuis H, Luijten M, Willemsen V, Blilou I, Heidstra R, Scheres B (2007) PLETHORA proteins as dose-dependent master regulators of Arabidopsis root development. Nature 449:1053–1057CrossRefPubMedGoogle Scholar
  28. Gray WM, Ostin A, Sandberg G, Romano CP, Estelle M (1998) High temperature promotes auxin-mediated hypocotyl elongation in Arabidopsis. Proc Natl Acad Sci USA 95:7197–7202CrossRefPubMedPubMedCentralGoogle Scholar
  29. Gupta R, Luan S (2003) Redox control of protein tyrosine phosphatases and mitogen-activated protein kinases in plants. Plant Physiol 132:1149–1152CrossRefPubMedPubMedCentralGoogle Scholar
  30. Kan CC, Chung TY, Wu HY, Juo YA, Hsieh MH (2017) Exogenous glutamate rapidly induces the expression of genes involved in metabolism and defense responses in rice roots. BMC Genom 18:186. CrossRefGoogle Scholar
  31. Kanno S, Arrighi JF, Chiarenza S, Bayle V, Berthomé R, Péret B, Javot H, Delannoy E, Marin E, Nakanishi TM, Thibaud MC, Nussaume L (2016) A novel role for the root cap in phosphate uptake and homeostasis. eLife 5:e14577CrossRefPubMedPubMedCentralGoogle Scholar
  32. Keyse SM (2008) The regulation of stress-activated MAP kinase signalling by protein phosphatases. In: Posas F, R., NA (eds) Topics in current genetics: stress-activated protein kinases, vol 20. Spinger, Berlin Heidelberg, pp 33–49CrossRefGoogle Scholar
  33. Kong D, Hu HC, Okuma E, Lee Y, Lee H, Munemasa S, Cho D, Ju C, Pedoeim L, Rodriguez B et al (2016) L-Met activates Arabidopsis GLR Ca2+ channels upstream of ROS production and regulates stomatal movement. Cell Rep 17:2553–2561CrossRefPubMedGoogle Scholar
  34. Křeček P, Skúpa P, Libus J, Naramoto S, Tejos R, Friml J, Zažimalová E (2009) The PIN-FORMED (PIN) protein family of auxin transporters. Genome Biol 10:249–249.11CrossRefPubMedPubMedCentralGoogle Scholar
  35. Lam HM, Chiu J, Hsieh MH, Meisel L, Olivera IC, Shin M, Coruzzi G (1998) Glutamate-receptor genes in plants. Nature 396:125–126CrossRefPubMedGoogle Scholar
  36. Lampard GR, Macalister CA, Bergmann DC (2008) Arabidopsis stomatal initiation is controlled by MAPK-mediated regulation of the bHLH SPEECHLESS. Science 322:1113–1116CrossRefPubMedGoogle Scholar
  37. Li Y, Yang S, Yang H, Hua J (2007) The TIR-NB-LRR gene SNC1 is regulated at the transcript level by multiple factors. Mol Plant Microbe Interact 20:1449–1456CrossRefPubMedGoogle Scholar
  38. Liu Y, Zhang S (2004) Phosphorylation of 1-aminocyclopropane-1-carboxylic acid synthase by MPK6, a stress-responsive mitogen-activated protein kinase, induces ethylene biosynthesis in Arabidopsis. Plant Cell 16:3386–3399CrossRefPubMedPubMedCentralGoogle Scholar
  39. López-Bucio J, Cruz-Ramírez A, Herrera-Estrella L (2003) The role of nutrient availability in regulating root architecture. Curr Opin Plant Biol 6:280–287CrossRefPubMedGoogle Scholar
  40. López-Bucio JS, Dubrovsky JG, Raya-González J, Ugartechea-Chirino Y, López-Bucio J, de Luna-Valdés LA, Ramos-Vega M, León P, Guevara-García AA (2014) Arabidopsis thaliana mitogen-activated protein kinase 6 is involved in seed formation and modulation of primary and lateral root development. J Exp Bot 65:169–183CrossRefPubMedGoogle Scholar
  41. Malamy JE (2005) Intrinsic and environmental response pathways that regulate root system architecture. Plant Cell Environ 28:67–77CrossRefPubMedGoogle Scholar
  42. MAPK Group (2002) Mitogen-activated protein kinase cascades in plants: a new nomenclature. Trends Plant Sci 7: 301–308CrossRefGoogle Scholar
  43. Meyerhoff O, Müller K, Roelfsema MR, Latz A, Lacombe B, Hedrich R, Dietrich P, Becker D (2005) AtGLR3.4, a glutamate receptor channel like gene is sensitive to touch and cold. Planta 222:418–427CrossRefPubMedGoogle Scholar
  44. Michard E, Lima PT, Borges F, Silva AC, Portes MT, Carvalho JE, Gilliham M, Liu LH, Obermeyer G, Feijo JA (2011) Glutamate receptor-like genes form Ca2+ channels in pollen tubes and are regulated by pistil D-serine. Science 332:434–437CrossRefPubMedGoogle Scholar
  45. Moe LA (2013) Amino acids in the rhizosphere: from plants to microbes. Am J Bot 100:1692–1705CrossRefPubMedGoogle Scholar
  46. Mousavi SA, Chauvin A, Pascaud F, Kellenberger S, Farmer EE (2013) GLUTAMATE RECEPTOR-LIKE genes mediate leaf-to-leaf wound signalling. Nature 500:422–426CrossRefPubMedGoogle Scholar
  47. Okumoto S, Funck D, Trovato M, Forlani G (2016) Amino acids of the glutamate family: functions beyond primary metabolism. Front Plant Sci 7:318CrossRefPubMedPubMedCentralGoogle Scholar
  48. Ottenschläger I, Wolff P, Wolverton C, Bhalerao RP, Sandberg G, Ishikawa H, Evans M, Palme K (2003) Gravity-regulated differential auxin transport from columella to lateral root cap cells. Proc Natl Acad Sci USA 100:2987–2991CrossRefPubMedPubMedCentralGoogle Scholar
  49. Pi L, Aichinger E, van der Graaff E, Llavata-Peris CI, Weijers D, Hennig L, Groot E, Laux T (2015) Organizer-derived WOX5 signal maintains root columella stem cells through chromatin-mediated repression of CDF4 expression. Dev Cell 33:576–588CrossRefPubMedGoogle Scholar
  50. Qi Z, Stephens NR, Spalding EP (2006) Calcium entry mediated by GLR3.3, an Arabidopsis glutamate receptor with a broad agonist profile. Plant Physiol 142:963–971CrossRefPubMedPubMedCentralGoogle Scholar
  51. Scheres B, Benfey P, Dolan L (2002) Root development. In: The American Society of Plant Biologists. eds. The Arabidopsis Book, vol 1, p e0101Google Scholar
  52. Suárez-Rodriguez MC, Petersen M, Mundy J (2010) Mitogen-activated protein kinase signaling in plants. Annu Rev Plant Biol 61:621–649CrossRefGoogle Scholar
  53. Sun J, Qi L, Li Y, Chu J, Li C (2012) PIF4-mediated activation of YUCCA8 expression integrates temperature into the auxin pathway in regulating Arabidopsis hypocotyl growth. PLoS Genet 8:e1002594CrossRefPubMedPubMedCentralGoogle Scholar
  54. Ulm R, Revenkova E, di Sansebastiano GP, Bechtold N, Paszkowski J (2001) Mitogen-activated protein kinase phosphatase is required for genotoxic stress relief in Arabidopsis. Genes Dev 15:699–709CrossRefPubMedPubMedCentralGoogle Scholar
  55. Ulm R, Ichimura K, Mizoguchi T, Peck SC, Zhu T, Wang X, Shinozaki K, Paszkowski J (2002) Distinct regulation of salinity and genotoxic stress responses by Arabidopsis MAP kinase phosphatase 1. EMBO J 21:6483–6493CrossRefPubMedPubMedCentralGoogle Scholar
  56. Vieten A, Vanneste S, Wiśniewska J, Benková E, Benjamins R, Beeckman T, Luschnig C, Friml J (2005) Functional redundancy of PIN proteins is accompanied by auxin-dependent cross-regulation of PIN expression. Development 132:4521–4531CrossRefPubMedGoogle Scholar
  57. Vincill ED, Clarin AE, Molenda JN, Spalding EP (2013) Interacting glutamate receptor-like proteins in phloem regulate lateral root initiation in Arabidopsis. Plant Cell 25:1304–1313CrossRefPubMedPubMedCentralGoogle Scholar
  58. Walch-Liu P, Liu LH, Remans T, Tester M, Forde BG (2006) Evidence that L-glutamate can act as an exogenous signal to modulate root growth and branching in Arabidopsis thaliana. Plant Cell Physiol 47:1045–1057CrossRefPubMedGoogle Scholar
  59. Wang H, Ngwenyama N, Liu Y, Walker JC, Zhang S (2007a) Stomatal development and patterning are regulated by environmentally responsive mitogen activated protein kinases in Arabidopsis. Plant Cell 19:63–73CrossRefPubMedPubMedCentralGoogle Scholar
  60. Wang JQ, Fibuch EE, Mao L (2007b) Regulation of mitogen-activated protein kinases by glutamate receptors. J Neurochem 100:1–11CrossRefPubMedGoogle Scholar
  61. Xu J, Zhang S (2015) Mitogen-activated protein kinase cascades in signaling plant growth and development. Trends Plant Sci 20:56–64CrossRefPubMedGoogle Scholar
  62. Zhang S, Klessig DF (1997) Salicylic acid activates a 48-kD MAP kinase in tobacco. Plant Cell 9:809–824CrossRefPubMedPubMedCentralGoogle Scholar
  63. Zhang P, Luo Q, Wang R, Xu J (2017) Hydrogen sulfide toxicity inhibits primary root growth through the ROS-NO pathway. Sci Rep 7:868CrossRefPubMedPubMedCentralGoogle Scholar

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© Springer Science+Business Media B.V., part of Springer Nature 2018

Authors and Affiliations

  1. 1.Instituto de BiotecnologíaUniversidad Nacional Autónoma de MéxicoCuernavacaMexico
  2. 2.Instituto de Investigaciones Químico-BiológicasUniversidad Michoacana de San Nicolás de HidalgoMoreliaMexico
  3. 3.CONACYT-Instituto de Investigaciones Químico-BiológicasUniversidad Michoacana de San Nicolás de HidalgoMoreliaMexico

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