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The characterization of plasma membrane-bound tubulin of cauliflower using Triton X-114 fractionation

Sonesson, Anders LU ; Berglund, M.; Staxén, Irina and Widell, Susanne LU (1997) In Plant Physiology 115(3). p.1001-1007
Abstract
The cortical microtubules determine how cellulose microfibrils are deposited in the plant cell wall and are thus important for the control of cell expansion. To understand how microtubules can control microfibril deposition, the components that link the microtubules to the plasma membrane (PM) of plant cells must be isolated. To obtain information on the properties of the tubulin-membrane associations, cauliflower (Brassica oleracea) PM was subjected to Triton X-114 fractionation, and the distribution of α- and β-tubulin was analyzed using immunoblotting. Approximately one-half of the PM-associated tubulin was solubilized by Triton X-114 and 10 to 15 % of both α- and β-tubulin was recovered in the detergent phase (indicative of hydrophobic... (More)
The cortical microtubules determine how cellulose microfibrils are deposited in the plant cell wall and are thus important for the control of cell expansion. To understand how microtubules can control microfibril deposition, the components that link the microtubules to the plasma membrane (PM) of plant cells must be isolated. To obtain information on the properties of the tubulin-membrane associations, cauliflower (Brassica oleracea) PM was subjected to Triton X-114 fractionation, and the distribution of α- and β-tubulin was analyzed using immunoblotting. Approximately one-half of the PM-associated tubulin was solubilized by Triton X-114 and 10 to 15 % of both α- and β-tubulin was recovered in the detergent phase (indicative of hydrophobic properties) and 30 to 40% was recovered in the aqueous phase. The hydrophobic tubulin could be released from the membrane by high pH extraction with preserved hydrophobicity. A large part of the PM-associated tubulin was found in the Triton-insoluble fraction. When this insoluble material was extracted a second time, a substantial amount of hydrophobic tubulin was released if the salt concentration was increased, suggesting that the hydrophobic tubulin was linked to a high-salt-sensitive protein aggregate that probably includes other components of the cytoskeleton. The hydrophobicity of a fraction of PM-associated tubulin could reflect a direct or indirect interaction of this tubulin with the lipid bilayer or with an integral membrane protein and may represent the anchoring of the cortical microtubules to the PM, a key element in the regulation of cell expansion. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Brassica, Cell Membrane, Detergents, Hydrogen-Ion Concentration, Membrane Proteins, Polyethylene Glycols, Tubulin
in
Plant Physiology
volume
115
issue
3
pages
1001 - 1007
publisher
American Society of Plant Biologists
external identifiers
  • scopus:0031277491
ISSN
1532-2548
DOI
10.1104/pp.115.3.1001
language
English
LU publication?
yes
id
ebcb8c12-5dca-4ab1-8818-20785efd130e (old id 4438059)
alternative location
http://www.jstor.org/stable/4277978
date added to LUP
2014-05-21 11:45:39
date last changed
2017-01-01 07:31:10
@article{ebcb8c12-5dca-4ab1-8818-20785efd130e,
  abstract     = {The cortical microtubules determine how cellulose microfibrils are deposited in the plant cell wall and are thus important for the control of cell expansion. To understand how microtubules can control microfibril deposition, the components that link the microtubules to the plasma membrane (PM) of plant cells must be isolated. To obtain information on the properties of the tubulin-membrane associations, cauliflower (Brassica oleracea) PM was subjected to Triton X-114 fractionation, and the distribution of α- and β-tubulin was analyzed using immunoblotting. Approximately one-half of the PM-associated tubulin was solubilized by Triton X-114 and 10 to 15 % of both α- and β-tubulin was recovered in the detergent phase (indicative of hydrophobic properties) and 30 to 40% was recovered in the aqueous phase. The hydrophobic tubulin could be released from the membrane by high pH extraction with preserved hydrophobicity. A large part of the PM-associated tubulin was found in the Triton-insoluble fraction. When this insoluble material was extracted a second time, a substantial amount of hydrophobic tubulin was released if the salt concentration was increased, suggesting that the hydrophobic tubulin was linked to a high-salt-sensitive protein aggregate that probably includes other components of the cytoskeleton. The hydrophobicity of a fraction of PM-associated tubulin could reflect a direct or indirect interaction of this tubulin with the lipid bilayer or with an integral membrane protein and may represent the anchoring of the cortical microtubules to the PM, a key element in the regulation of cell expansion.},
  author       = {Sonesson, Anders and Berglund, M. and Staxén, Irina and Widell, Susanne},
  issn         = {1532-2548},
  keyword      = {Brassica,Cell Membrane,Detergents,Hydrogen-Ion Concentration,Membrane Proteins,Polyethylene Glycols,Tubulin},
  language     = {eng},
  number       = {3},
  pages        = {1001--1007},
  publisher    = {American Society of Plant Biologists},
  series       = {Plant Physiology},
  title        = {The characterization of plasma membrane-bound tubulin of cauliflower using Triton X-114 fractionation},
  url          = {http://dx.doi.org/10.1104/pp.115.3.1001},
  volume       = {115},
  year         = {1997},
}