, Volume 159, Issue 1, pp 44–59 | Cite as

The effects of microtubule and microfilament disrupting agents on cytoskeletal arrays and wall deposition in developing cotton fibers

  • R. W. Seagull


The effects of various cytoskeletal disrupting agents (cholchicine, oryzalin, trifluralin, taxol, cytochalasins B and D) on microtubules, microfilaments and wall microfibril deposition were monitored in developing cotton fibers, using immunocytochemical and fluorescence techniques. Treatment with 10−4 M colchicine, 10−6 M trifluralin or 10−6 M oryzalin resulted in a reduction in the number of microtubules, however, the “drug-stable” microtubules still appear to influence wall deposition. Treatment with 10−5 M taxol increased the numbers of microtubules present within 15 minutes of application. New microtubules were aligned parallel to the existing ones, however, some evidence of random arrays was observed. Microtubules stabilized with taxol appeared to function in wall organization but do not undergo normal re-orientations during development. Microtubule disrupting agent had no detectable affect on the microfilament population. Exposure to either 4×10−5 M cytochalasin B or 2×10−6M cytochalasin D resulted in a disruption of microfilaments and a re-organization of microtubule arrays. Treatment with either cytochalasin caused a premature shift in the orientation of microtubules in young fibers, whereas in older fibers the microtubule arrays became randomly organized. These observations indicate that microtubule populations during interphase are heterogeneous, differing at least in their susceptibility to disruption by depolymerizing agents. Changes in microtubule orientation (induced by cytochalasin) indicate that microfilaments may be involved in regulating microtubule orientation during development.


Microtubules Microfilaments Wall microfibrils Cotton fiber Cytoskeletal disruption 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Beasley CA, Ting IP (1973) The effects of plant growth substances on in vitro fiber development in fertilized cotton ovules. Amer J Bot 60: 130–139Google Scholar
  2. Bajer AS, Mole-Bajer J (1986) Drugs with colchicine-like effects that specifically disassemble plant but not animal microtubules. Ann NY Acad Sci 466: 767–784Google Scholar
  3. Cyr RJ, Palevitz BA (1989) Microtubule-binding proteins from carrot. I. Initial characterization and microtubule bundling. Planta 177: 245–260Google Scholar
  4. —, Bustos MM, Guiltinan MJ, Fosket DE (1987) Developmental modulation of tubulin protein and mRNA levels during somatic embryogenesis in cultured carrot cells. Planta 171: 365–376Google Scholar
  5. Dawson PJ, Lloyd CW (1985) Identification of multiple tubulins in taxol microtubules purified from carrot suspension cells. EMBO J 4: 2451–45Google Scholar
  6. Derksen J, Traas JA (1984) Growth of tobacco pollen in vitro: effects of drugs interfering with the cytoskeleton. In: Willemse MTM, Went JL (eds) Proceedings 8th International Symposium Sexual Reproduction in Seed Plants, Ferns, and Mosses. Pudoc, Wageningen, pp 64–67Google Scholar
  7. — — Oosterdorp T (1986) Distribution of actin filaments in differentiating cells ofEquisetum hyemale root tips. Plant Sci 43: 77–81Google Scholar
  8. Falconer MM, Seagull RW (1985 a) Immunofluorescent and calcofluor white staining of developing tracheary elements inZinnia elegans L. suspension cultures. Protoplasma 125: 190–198Google Scholar
  9. — — (1985 b) Xylogenesis in tissue culture: taxol effects on microtubule reorientation and lateral association in differentiating cells. Protoplasma 128: 157–166Google Scholar
  10. — — (1987) Amiprophos-methyl (APM): a rapid, reversible, antimicrotubule agent for plant cell cultures. Protoplasma 136: 118–124Google Scholar
  11. Farrell KW, Wilson L (1980) Proposed mechanism for colchicine poisoning of microtubules reassembled in vitro fromStongylocintrotus purpatus sperm tail outer doublet tubulin. Biochemistry 9: 3466–3472Google Scholar
  12. Fukuda H, Kobayashi H (1989) Dynamic organization of the cytoskeleton during tracheary-element differentiation. Dev Growth Different 31: 9–16Google Scholar
  13. Galatis B (1977) Differentiation of stomatal meristemoids and guard cell mother cells into guard-like cells inVigna sinensis leaves after colchicine treatment. Planta 136: 103–114Google Scholar
  14. Gunning BES, Hardham AR (1982) Microtubules. Annu Rev Plant Physiol 33: 651–698Google Scholar
  15. Hardham AR, McCully ME (1982) Preprogramming of cells following wounding in pea (Pisum sativum L.) root. II. The effects of caffeine and colchicine on the development of new vascular elements. Protoplasma 112: 152–166Google Scholar
  16. —, Gunning BES (1979) Interpolation of microtubules into cortical arrays during cell elongation and differentiation in roots ofAzolla pinnata. J Cell Sci 37: 411–442Google Scholar
  17. Hartwig JH, Stossel TP (1979) Cytochalasin B and the structure of actin gels. J Mol Biol 134: 539–553Google Scholar
  18. Heath IB, Seagull RW (1982) Oriented cellulose microfibrils and the cytoskeleton: a critical comparison of models. In: Lloyd CW (ed) The cytoskeleton in plant growth and development. Academic Press, London, pp 162–182Google Scholar
  19. Hensel W (1986) Cytodifferentiation in polar plant cells. Use of antimicro tubular agents during the differentiation of statocytes from cress roots (Lepidium sativum L.). Planta 169: 293–303Google Scholar
  20. Hepler PK, Palevitz BA (1974) Microtubules and microfilaments. Annu Rev Plant Physiol 25: 309–362Google Scholar
  21. Hess FD, Bayer DE (1977) Binding of the herbicide trifluralin toChlamydomonas flagellar tubulin. J Cell Sci 24: 351–360Google Scholar
  22. Hussey PJ, Gull K (1985) Multiple isotypes of A and B tubulin in the plantPhaseolus vulgaris. FEBS Lett 181: 113–118Google Scholar
  23. —, Traas JA, Gull K, Lloyd CW (1987) Isolation of cytoskeletons from synchronized plant cells: the interphase microtubule array utilizes multiple tubulin isotypes. J Cell Sci 88: 225–230Google Scholar
  24. Juniper BE, Lawton JR (1979) The effect of caffeine, different fixation regimes and low temperature on microtubules in the cells of higher plants. Planta 145: 411–416Google Scholar
  25. Kakimoto T, Shibaoka H (1987) Actin filaments and microtubules in the pre-prophase band and phragmoplast of tobacco cells. Protoplasma 140: 151–156Google Scholar
  26. Katsuta J, Shibaoka H (1988) The roles of the cytoskeleton and the cell wall in nuclear positioning in tobacco BY-2 cells. Plant Cell Physiol 29: 403–413Google Scholar
  27. Kloth R (1989) Changes in the level of tubulin subunits during development of cotton (Gossypium hirsutum) fiber. Physiol Plant 76: 37–41Google Scholar
  28. Kobayashi H, Fukuda H, Shibaoka H (1989) Interrelation between the spatial disposition of actin filaments and microtubules during the differentiation of tracheary elements in culturedZinnia elegans. Protoplasma 143: 29–37Google Scholar
  29. Lancelle SA, Hepler PK (1987) Ultrastructure of the cytoskeleton in freeze-substituted pollen tubes ofNicotiana alata. Protoplasma 140: 141–150Google Scholar
  30. Lloyd CW (1984) Toward a dynamic helical model for the influence of microtubules on wall patterns in plants. Int Rev Cytol 86: 1–51Google Scholar
  31. MacLean-Fletcher S, Pollard TD (1980) Mechanism of action of cytochalasin B on actin. Cell 20: 329–341Google Scholar
  32. Margolis RL, Wilson L (1977) Addition of colchicine-tubulin complex to microtubule ends: the mechanism of substoichiometric colchicine poisoning. Proc Natl Acad Sci USA 74: 3466–3470Google Scholar
  33. Meyer Y, Herth W (1978) Chemical inhibition of cell wall formation and cytokinesis, but not nuclear division, in protoplasts inNicotiana tabacum L. cultivated in vitro. Planta 142: 253–262Google Scholar
  34. Morejohn LC, Fosket DE (1984) Inhibition of plant microtubule polymerization in vitro by the phosphoric amide herbicide Amiprophos-methyl. Science 224: 874–876Google Scholar
  35. —, Bureau TE, Tocchi LP, Fosket DE (1984) Tubulins from different higher plant species are immunologically nonidentical and bind colchicine differentially. Proc Natl Acad Sci USA 81: 1440–1444Google Scholar
  36. — —, Mole-Bajer J, Bajer AS, Fosket DE (1987) Oryzalin, a dinitroaniline herbicide, binds to plant tubulin and inhibits microtubule polymerization in vitro. Planta 172: 252–264Google Scholar
  37. Nelms BJ, Preston RD, Ashworth D (1973) A possible function of microtubules suggested by their distribution in rubbery wood. J Cell Sci 37: 411–424Google Scholar
  38. Palevitz BA (1980) Comparative effects of phalloidin and cytochalasin B on motility and morphogenesis inAllium. Can J Bot 58: 772–785Google Scholar
  39. — (1987 a) Actin in the preprophase band ofAllium cepa. J Cell Biol 104: 1515–1519Google Scholar
  40. — (1987 b) Accumulation of F-actin during cytokinesis inAllium. Correlation with microtubule disruption and the effects of drugs. Protoplasma 141: 24–32Google Scholar
  41. — (1988) Cytochalasin-induced reorganization of actin inAllium root cells. Cell Motil Cytoskeleton 9: 283–298Google Scholar
  42. Parthasarathy MV, Perdue TD, Witztum A, Alvernaz J (1985) Aetin network as a normal component of the cytoskeleton in many vascular plant cells. Amer J Bot 72: 1318–1323Google Scholar
  43. Pickett-Heaps JD (1967) The effects of colchicine on the ultrastructure of dividing plant cells, xylem wall differentiation and distribution of cytoplasmic microtubules. Dev Biol 15: 206–236Google Scholar
  44. Pierson ES (1988) Rhodamine-phalloidin staining of F-actin in pollen after dimethylsulfoxide permeabilization. Sex Plant Reprod 1: 83–87Google Scholar
  45. Quader H, Filner P (1980) The action of antimitotic herbicides on flagellar regeneration inChlamydomonas reinhardtii: a comparison with the action of colchicine. Eur J Cell Biol 21: 301–304Google Scholar
  46. Ridge R (1988) Freeze-substitution improves the ultrastructural preservation of legume root hairs. Bot Mag Tokyo 101: 427–441Google Scholar
  47. Schiff PB, Fant J Horowitz SB (1979) Promotion of microtubule assembly in vitro by taxol. Nature 277: 665–667Google Scholar
  48. Schiff PB, Horowitz SB (1980) Taxol stabilizes microtubules in mouse fibroblast cells. Proc Natl Acad Sci USA 77: 1561–1565Google Scholar
  49. Schmit A-C, Lambert A-M (1988) Plant actin filament and microtubule interactions during anaphase-telophase transition: effects of antagonist drugs. Biol Cell 64: 309–319Google Scholar
  50. Seagull RW (1986) Changes in microtubule organization and microfibril orientation during in vitro cotton fiber development: an immunofluorescent study. Can J Bot 64: 1373–1381Google Scholar
  51. — (1989 a) The plant cytoskeleton. CRC Crit Rev Plant Sci 8: 131–167Google Scholar
  52. - (1989 b) Changes in microtubule arrays during cotton fiber development. In: Bailey GW (ed) Proceedings of the 47th Annual Meeting of the Electron Microscopy Society of America, San Antonio, Texas, pp 760–761Google Scholar
  53. — (1990) The role of the cytoskeleton in cellulose synthesis. In: Haigler C, Weimer PO (eds) Biogenesis and biodégradation of cellulose and cellulosic materials. Marcel Dekker, New York (in press)Google Scholar
  54. —, Heath IB (1980 a) The differential effects of cytochalasin B on microfilament populations and cytoplasmic streaming. Protoplasma 103: 231–240Google Scholar
  55. — (1980b) The organization of cortical microtubule arrays in the radish root hair. Protoplasma 103: 205–229Google Scholar
  56. —, Falconer MM, Weerdenberg CA (1987) Microfilaments: dynamic arrays in higher plant cells. J Cell Biol 104: 995–1004Google Scholar
  57. —, Holland S, Gibson DM (1989) Isolation of cytoskeletal proteins from cotton (Gossypium hirsutum) suspension culture cells. J Cell Biol 109: 78 aGoogle Scholar
  58. Sheterline P (1983) Mechanisms of cell motility: molecular aspects of contractility. Academic Press, London, pp 167–175Google Scholar
  59. Simmonds DH, Seagull RW, Setterfield G (1985) Evaluation of techniques for immunofluorescent staining of microtubules in cultured plant cells. J Histochem Cytochem 33: 345–352Google Scholar
  60. Tang X, Lancelle SA, Hepler PK (1989) Fluorescence microscopic localization of actin in pollen tubes: comparison of actin antibody and phalloidin staining. Cell Motil Cytoskeleton 12: 216–224Google Scholar
  61. Tewinkel M, Kruse S, Quader H, Volkmann D, Sievers I (1989) Visualization of actin filament pattern in plant cells without prefixation: a comparison of differently modified phallotoxins. Protoplasma 149: 178–182Google Scholar
  62. Tiwari CC, Gunning BES (1986) Colchicine inhibits plasmodium formation and disrupts pathway of sporopollenin secretion in the anther tapetum ofTradescantia virginiana L. Protoplasma 114: 115–128Google Scholar
  63. —, Polito VS (1988) Organization of the cytoskeleton in pollen tubes ofPyrus communis: a study employing conventional and freezesubstitution electron microscopy, immunofluorescence and rhodamine-phalloidin. Protoplasma 147: 100–112Google Scholar
  64. Traas JA, Doonan JH, Rawlins DJ, Shaw PJ, Watts J, Lloyd CW (1987) An actin network is present in the cytoplasm throughout the cell cycle of carrot cells and associates with the dividing nucleus. J Cell Biol 105: 387–395Google Scholar
  65. Vaughn MA, Vaughn KC (1987) Effects of microfilament disrupters on microfilament distribution and morphology in maize root cells. Histochemistry 87: 129–137Google Scholar
  66. Volkmann D, Czaja AWP (1981) Reversible inhibition of secretion in root cap cells of cress after treatment with cytochalasin B. Exp Cell Res 135: 229–236Google Scholar
  67. Waterkyn L (1985) Light microscopy of the cotton fiber. Tech Monogr Belgian Cotton Res Council. International Institute of Cotton, Manchester, UKGoogle Scholar
  68. Weerdenberg CA, Falconer MM, Setterfield G, Seagull RW (1986) Effects of taxol on microtubule arrays in cultured higher plant cells. Cell Motil Cytoskeleton 6: 469–478Google Scholar
  69. Yatsu LY, Jacks TJ (1981) An ultrastructural study of the relationship between microtubules and microfibrils in cotton (Gossypium hirsutum L.) cell wall reversals. Amer J Bot 68: 771–777Google Scholar

Copyright information

© Springer-Verlag 1990

Authors and Affiliations

  • R. W. Seagull
    • 1
  1. 1.Southern Regional Research CenterUSDA/ARSNew OrleansUSA

Personalised recommendations