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The distributional changes and role of microtubules in Nod factor-challenged Medicago sativa root hairs

Planta volume 218, pages 276–287 (2003)Cite this article

Abstract

The normal tip-growing pattern exhibited by root hairs of legumes is disrupted when the hair is exposed to Nod factors generated by compatible bacteria capable of inducing nodule formation. Since microtubules (MTs) play an important role in regulating directionality and stability of apical growth in root hairs [T.N. Bibikova et al. (1999) Plant J 17:657–665], we examined the possibility that Nod factors might affect the MT distribution patterns in root hairs of Medicago sativa L. We observed that Nod factor application caused rapid changes in the pattern of MTs starting as early as 3 min after perfusion. Within 3 to 10 min after Nod factor application, first endoplasmic and then cortical MTs depolymerised, initially at the proximal ends of cells. Twenty minutes after exposure to Nod factors, a transverse band of microtubules was seen behind the tip, while almost all other MTs had depolymerised. By 30 min, very few MTs remained in the root hair and yet by 1 h the MT cytoskeleton re-formed. When Nod factors were applied in the presence of 10 μM oryzalin or 5 μM taxol, the MTs appeared disintegrated while the morphological effects, such as bulging and branching, became enhanced. Compared to the treatments with oryzalin or taxol alone, the combinatory treatments exhibited higher growth rates. Since microtubule reorganization is one of the earliest measurable events following Nod factor application we conclude that microtubules have an important role in the early phases of the signalling cascade. Microtubule involvement could be direct or a consequence of Nod factor-induced changes in ion levels.

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Fig. 1a–c.
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Fig. 3a–d.
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Fig. 8a–h.
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Fig. 10a–f.

Abbreviations

BNM:

buffered nodulation medium

CLSM:

confocal laser scanning microscopy

MT:

microtubule

References

  • Allen NS, Bennett MN (1996) Electro-optical imaging of F-actin and endoplasmic reticulum in living and fixed plant cells. In: Malecki M, Roomans G (eds) Science of specimen preparation for microscopy and microanalysis, vol 10. SMI Press, AMF, Chicago, pp 177–187

  • Allen NS, Bennett MN, Cox DN, Shipley A, Ehrhardt DW, Long SR (1994) Effects of Nod factors on alfalfa root hair Ca++ and H+ currents on cytoskeleton behavior. In: Daniels MJ, Downie JA, Osbourn AE (eds) Advances in molecular genetics of plant-microbe interactions, vol 3. Kluwer, Dordrecht, pp 107–114

  • Baluška F, Salaj J, Mathur J, Braun M, Jasper F, Šamaj J, Chua NH, Barlow PW, Volkmann D (2000) Root hair formation: F-actin-dependent tip growth is initiated by local assembly of profilin-supported F-actin meshworks accumulated within expansin-enriched bulges. Dev Biol 227:618–632

    PubMed  Google Scholar 

  • Bibikova TN, Zhigilei A, Gilroy S (1997) Root hair growth in Arabidopsis thaliana is directed by calcium and an endogenous polarity. Planta 203:495–505

    CAS  PubMed  Google Scholar 

  • Bibikova TN, Blancaflor EB, Gilroy S (1999) Microtubules regulate tip growth and orientation in root hairs of Arabidopsis thaliana. Plant J 17:657–665

    Article  CAS  PubMed  Google Scholar 

  • Cárdenas L, Vidali L, Domínguez J, Pérez H, Sánchez F, Hepler PK, Quinto C (1998) Rearrangement of actin microfilaments in plant root hairs responding to Rhizobium etli nodulation signals. Plant Physiol 116:871–877

    PubMed  Google Scholar 

  • Cárdenas L, Feijó JA, Kunkel JG, Sánchez F, Holdaway-Clarke T, Hepler PK, Quinto C (1999) Rhizobium Nod factors induce increases in intracellular free calcium and extracellular calcium influxes in bean root hairs. Plant J 19:347–352

    Article  PubMed  Google Scholar 

  • Catoira R, Galera C, de Billy F, Penmetsa VR, Juornet EP, Maillet F, Rosenberg C, Cook D, Gough C, Dénarié J (2000) Four genes of Medicago truncatula controlling components of a Nod factor transduction pathway. Plant Cell 12:1647–1665

    CAS  PubMed  Google Scholar 

  • Collings DA, Allen NS (2000) Cortical actin interacts with the plasma membrane and microtubules. In: Staiger CJ, Baluška F, Volkmann D, Barlow PW (eds) Actin: a dynamic framework for multiple plant cell functions. Kluwer, Dordrecht, pp 145–164

  • Cormack RGH (1949) The development of root hairs in angiosperms. Bot Rev 15:583–612

    Google Scholar 

  • Cyr RJ (1991) Calcium calmodulin affects microtubule stability in lysed protoplasts. J Cell Sci 100:311–317

    CAS  Google Scholar 

  • Cyr RJ (1994) Microtubules in plant morphogenesis: role of the cortical array. Annu Rev Cell Biol 10:153–180

    CAS  PubMed  Google Scholar 

  • de Ruijter NCA, Rook MB, Bisseling T, Emons AMC (1998) Lipochitooligosaccharides re-initiate root hair tip growth in Vicia sativa with high calcium and spectrin like antigen at the tip. Plant J 13:341–350

    Article  Google Scholar 

  • de Ruijter NCA, Bisseling T, Emons AMC (1999) Rhizobium Nod factors induce an increase in sub-apical fine bundles of actin filaments in Vicia sativa root hairs within minutes. Mol Plant Microbe Interact 12:829–832

    Google Scholar 

  • Dénarié J, Debellé F, Promé JC (1996) Rhizobium lipo-chitooligosaccharide nodulation factors: signaling molecules mediating recognition and morphogenesis. Annu Rev Biochem 65:503–535

    PubMed  Google Scholar 

  • Doonan JH, Cove DJ, Lloyd CW (1988) Microtubules and microfilaments in tip growth — evidence that microtubules impose polarity on protonemal growth in Physcomitrella patens. J Cell Sci 89:533–540

    Google Scholar 

  • Downie JA, Walker S (1999) Plant responses to Nodulation factors. Curr Opin Plant Biol 2:483–489

    CAS  PubMed  Google Scholar 

  • Ehrhardt DW, Atkinson EM, Long SR (1992) Depolarization of alfalfa root hair membrane potential by Rhizobium meliloti Nod factors. Science 256:998–1000

    CAS  PubMed  Google Scholar 

  • Ehrhardt DW, Wais R, Long SR (1996) Calcium spiking in plant root hairs responding to Rhizobium nodulation signals. Cell 85:673–681

    PubMed  Google Scholar 

  • Emons AMC (1989) Helicoidal microfibril deposition in a tip-growing cell and microtubule alignment during tip morphogenesis — a dry-cleaving and freeze-substitution study. Can J Bot 67:2401–2408

    Google Scholar 

  • Emons AMC, Derksen J (1986) Microfibrils, microtubules and microfilaments of the trichoblast of Equisetum hyemale. Acta Bot Neerl 35:311–320

    Google Scholar 

  • Emons AMC, de Ruijter NCA (2000) Actin: a target for signal transduction in root hairs. In: Staiger CJ, Baluška F, Volkmann D, Barlow PW (eds) Actin: a dynamic framework for multiple plant cell functions. Kluwer, Dordrecht, pp 373–390

  • Felle HH, Kondorosi E, Kondorosi A, Schultze M (1995) Nod signal induced plasma membrane potential changes in alfalfa root hairs are differentially sensitive to structural modifications of the lipochitooligosaccharides. Plant J 7:939–947

    Article  CAS  Google Scholar 

  • Felle HH, Kondorosi E, Kondorosi A, Schultze M (1996) Rapid alkalinization in alfalfa root hairs in response to rhizobial lipochitooligosaccharides signals. Plant J 10:295–301

    Article  CAS  Google Scholar 

  • Felle HH, Kondorosi E, Kondorosi A, Schultze M (1998) The role of ion fluxes in Nod factor signaling in Medicago sativa. Plant J 13:453–463

    Article  Google Scholar 

  • Felle HH, Kondorosi E, Kondorosi A, Schultze M (1999a) Nod factors modulate the concentration of cytosolic free calcium differently in growing and non-growing root hairs of Medicago sativa L. Planta 209:207–212

    CAS  PubMed  Google Scholar 

  • Felle HH, Kondorosi E, Kondorosi A, Schultze M (1999b) Elevation of the cytosolic free [Ca2+] is indispensable for the transduction of the Nod factor signal in alfalfa. Plant Physiol 121:273–280

    Article  CAS  PubMed  Google Scholar 

  • Fu Y, Wu G, Yang ZB (2001) Rop GTPase-dependent dynamics of tip-localized F-actin controls tip growth in pollen tubes. J Cell Biol 152:1019–1032

    CAS  PubMed  Google Scholar 

  • Gehring CA, Irving HR, Kabbara AA, Parish RW, Boukli NM, Broughton WJ (1997) Rapid, plateau-like increases in intracellular calcium are associated with Nod factor induced root hair deformation. Mol Plant Microbe Interact 10:791–802

    CAS  Google Scholar 

  • Geitmann A, Emons AMC (2000) The cytoskeleton in plant and fungal cell tip growth. J Microsc 198:218–245

    Article  CAS  Google Scholar 

  • Gianì S, Campanoni P, Breviaro D (2002) A dual effect on protein synthesis and degradation modulates the tubulin level in rice cells treated with oryzalin. Planta 214:837–847

    PubMed  Google Scholar 

  • Gilroy S, Jones DL (2000) Through form to function: root hair development and nutrient uptake. Trends Plant Sci 5:56–60

    CAS  PubMed  Google Scholar 

  • Gundersen GG, Cook TA (1999) Microtubules and signal transduction. Curr Opin Cell Biol 11:81–94

    CAS  PubMed  Google Scholar 

  • Heidstra R, Bisseling T (1996) Nod factor induced host responses and mechanisms of Nod factor perception. New Phytol 133:25–43

    CAS  Google Scholar 

  • Horvath B, Heidstra R, Lados M, Moerman M, Spaink HP, Prome JC, Van Kammen A, Bisseling T (1993) Lipo-oligosaccharides of Rhizobium induce infection-related early nodulin gene expression in pea root hairs. Plant J 4:727–733

    Article  CAS  PubMed  Google Scholar 

  • Igarashi H, Orii H, Mori H, Shimmen T, Sonobe S (2000) Isolation of a novel protein from tobacco BY-2 cells: possible involvement in the interaction between actin filaments and microtubules Plant Cell Physiol 41:920–931

    Google Scholar 

  • Journet EP, Pichon M, Dedieu A, De Billy F, Truchet G, Barker DG (1994) Rhizobium meliloti Nod factors elicit cell specific transcription of ENOD12 gene in transgenic alfalfa. Plant J 6:241–249

    Article  CAS  PubMed  Google Scholar 

  • Kadota A, Wada M (1992) The circular arrangement of cortical microtubules around the subapex of tip-growing fern protonemata is sensitive to cytochalasin B. Plant Cell Physiol 33:99–102

    CAS  Google Scholar 

  • Kropf DL, Bisgrove SR, Hable WE (1998) Cytoskeletal control of polar growth in plant cells. Curr Opin Plant Biol 10:117–122

    CAS  Google Scholar 

  • Kurkdjian AC (1995) Role of differentiation of root epidermal cells in Nod factor (from Rhizobium meliloti)-induced root hair depolarization of Medicago sativa. Plant Physiol 107:783–790

    CAS  PubMed  Google Scholar 

  • Lhuissier FGP, de Ruijter NCA, Sieberer BJ, Esseling JJ, Emons AMC (2001) Time course of cell biological events evoked in legume root hairs by Rhizobium Nod factors: state of the art. Ann Bot 87:289–302

    Article  CAS  Google Scholar 

  • Lloyd CW, Pearce KJ, Rawlins DJ, Ridge RW, Shaw PJ (1987) Endoplasmic microtubules connect the advancing nucleus to the tip of legume root hairs, but F-actin is involved in basipetal migration. Cell Motil Cytoskel 8:27–36

    Google Scholar 

  • Long SR (1996) Rhizobium symbiosis: Nod factors in perspective. Plant Cell 8:1885–1898

    CAS  PubMed  Google Scholar 

  • Meske V, Ruppert V, Hartmann E (1996) Structural basis for the red light induced repolarization of tip growth in caulonema cells of Ceratodon purpureus. Protoplasma 192:189–198

    Google Scholar 

  • Miller DD, de Ruijter NCA, Emons AMC (1997) From signal to form: aspects of the cytoskeleton plasma membrane continuum in root hair tips. J Exp Bot 48:1881–1896

    Article  CAS  Google Scholar 

  • Ridge RW (1992) A model of legume root hair growth and Rhizobium infection. Symbiosis 14:359–373

    Google Scholar 

  • Schultze M, Kondorosi A (1998) Regulation of symbiotic root nodule development. Annu Rev Genet 32:33–57

    CAS  PubMed  Google Scholar 

  • Schwuchow J, Sack FD, Hartmann E (1990) Microtubule distribution in gravitropic protonemata of the moss Ceratodon. Protoplasma 159:60–69

    CAS  PubMed  Google Scholar 

  • Sieberer BJ, Timmers ACJ, Lhuissier FGP, Emons AMC (2002) Endoplasmic microtubules configure the subapical cytoplasm and are required for fast growth of Medicago truncatula root hairs. Plant Physiol 130:977–988

    CAS  PubMed  Google Scholar 

  • Staiger CJ (2000) Signaling to the actin cytoskeleton in plants. Annu Rev Plant Phys Plant Mol Biol 51:257–288

    CAS  Google Scholar 

  • Terasaka O, Niitsu T (1994) Differential roles of microtubules and actin-myosin cytoskeleton in the growth of Pinus pollen tubes. Sex Plant Reprod 7:264–272

    Google Scholar 

  • That TC, Rossier C-T, Barja F, Turian G, Roos UP (1988) Induction of multiple germ tubes in Neurospora crassa by anti-tubulin agents. Eur J Cell Biol 46:68–79

    CAS  PubMed  Google Scholar 

  • Thion L, Mazars C, Thuleau P, Graziana A, Rossignol M, Moreau M, Ranjeva R (1996) Activation of plasma membrane voltage-dependent calcium-permeable channels by disruption of microtubules in carrot cells. FEBS Lett 393:13–18

    CAS  PubMed  Google Scholar 

  • Thion L, Mazars C, Nacry P, Bouchez D, Moreau M, Ranjeva R, Thuleau P (1998) Plasma membrane depolarization-activated calcium channels, stimulated by microtubule depolymerizing drugs in wild-type Arabidopsis thaliana protoplasts, display constitutively large activities and longer half life in ton2 mutant cells affected in the organization of cortical microtubules. Plant J 13:603–610

    CAS  PubMed  Google Scholar 

  • Timmers ACJ, Auriac M-C, de Billy F, Truchet G (1998) Nod factor internalization and microtubular cytoskeleton changes occur concomitantly during nodule differentiation in alfalfa. Development 125:339–349

    CAS  PubMed  Google Scholar 

  • Timmers ACJ, Auriac M-C, Truchet G (1999) Refined analysis of early symbiotic steps of the Rhizobium–Medicago interaction in relationship with microtubular cytoskeleton rearrangements. Development 126:3617–3628

    CAS  PubMed  Google Scholar 

  • Tominaga M, Morita K, Sonobe S, Yokota E, Shimmen T (1997) Microtubules regulate the organization of actin filaments at the cortical region in root hair cells of Hydrocharis. Protoplasma 199:83–92

    CAS  Google Scholar 

  • Truchet G, Roche P, Lerouge P, Vasse J, Camut S, de Billy F, Promé JC, Dénarié J (1991) Sulfated lipo-oligosaccharide signals of Rhizobium meliloti elicit root nodule organogenesis in alfalfa. Nature 351:670–673

    CAS  Google Scholar 

  • Van Brussel AAN, Bakhuizen R, Van Spronsen PC, Spaink HP, Tak T, Lugtenberg BJJ, Kijne JW (1992) Induction of pre-infection thread structures in the leguminous host plant by mitogenic lipooligosaccharides of Rhizobium. Science 257:70–72

    Google Scholar 

  • Wacker I, Quader H, Schnepf E (1988) Influence of the herbicide oryzalin on cytoskeleton and growth of Funaria hygrometrica protonemata. Protoplasma 142:55–67

    Google Scholar 

  • Wais RJ, Galera C, Oldroyd G, Catoira R, Penmetsa VR, Cook D, Gough C, Dénarié J, Long S (2000) Genetic analysis of calcium spiking responses in nodulation mutants of Medicago truncatula. Proc Natl Acad Sci USA 97:13407–13412

    CAS  PubMed  Google Scholar 

  • Whitehead LF, Day DA, Hardham AR (1998) Cytoskeletal arrays in the cells of soybean root nodules: the role of actin microfilaments in the organization of symbiosomes. Protoplasma 203:194–205

    Google Scholar 

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Acknowledgements

Financial support was provided by the North Carolina Agricultural Research Station (NC AgResStat 407050, NS Allen) and the National Science Foundation (NSF DBI0077503, D Bird).

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Authors and Affiliations

  1. Department of Botany, North Carolina State University, Box 7612, Raleigh, NC 27695-7612, USA

    Ravisha R. Weerasinghe, Eva Johannes & Nina Strömgren Allen

  2. Plant Cell Biology Group, Research School of Biological Sciences, Australian National University, GPO Box 475, Canberra ACT 2601, Australia

    David A. Collings

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  1. Ravisha R. Weerasinghe
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  2. David A. Collings
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  3. Eva Johannes
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  4. Nina Strömgren Allen
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Correspondence to Nina Strömgren Allen.

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Weerasinghe, R.R., Collings, D.A., Johannes, E. et al. The distributional changes and role of microtubules in Nod factor-challenged Medicago sativa root hairs. Planta 218, 276–287 (2003). https://doi.org/10.1007/s00425-003-1097-1

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Keywords

  • Nodulation
  • Nod factor
  • Microtubule
  • Medicago
  • Rhizobium