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Title: Cytoskeletal organization in microtentacles

Abstract

Microtentacles are thin, flexible cell protrusions that have recently been described and whose presence enhances efficient attachment of circulating cells. They are found on circulating tumor cells and can be induced on a wide range of breast cancer cell lines, where they are promoted by factors that either stabilize microtubules or destabilize the actin cytoskeleton. Evidence suggests that they are relevant to the metastatic spread of cancer, so understanding their structure and formation may lead to useful therapies. Microtentacles are formed by microtubules and contain vimentin intermediate filaments, but beyond this, there is little information about their ultrastructure. We have used electron microscopy of high pressure frozen sections and tomography of cryo-prepared intact cells, along with super resolution fluorescence microscopy, to provide the first ultrastructural insights into microtubule and intermediate filament arrangement within microtentacles. By scanning electron microscopy it was seen that microtentacles form within minutes of addition of drugs that stabilize microtubules and destabilize actin filaments. Mature microtentacles were found to be well below one micrometer in diameter, tapering gradually to below 100 nm at the distal ends. They also contained frequent branches and bulges suggestive of heterogeneous internal structure. Super resolution fluorescence microscopy and examination of sectioned samplesmore » showed that the microtubules and intermediate filaments can occupy different areas within the microtentacles, rather than interacting intimately as had been expected. Cryo-electron tomography of thin regions of microtentacles revealed densely packed microtubules and absence of intermediate filaments. The number of microtubules ranged from several dozen in some areas to just a few in the thinnest regions, with none of the regular arrangement found in axonemes. Improved understanding of the mechanism of microtentacle formation, as well as the resultant structure, will be valuable in developing therapies against metastasis, if the hypothesized role of microtentacles in metastasis is confirmed. This work provides a significant step in this direction. - Highlights: • Microtentacles can form within minutes of the addition of inducing drugs. Microtentacle diameters are in the range of a tenth of a micrometer. • Microtubules are clearly resolved in microtentacles by cryo-electron tomography. • There is little microtubule-intermediate filament interaction in drug-induced microtentacles.« less

Authors:
 [1];  [1];  [1];  [1];  [2];  [1];  [3];  [1]
  1. Molecular Biophysics and Integrated Biomaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720 (United States)
  2. Department of Chemistry, University of California Berkeley, CA 94720 (United States)
  3. (United States)
Publication Date:
OSTI Identifier:
22738147
Resource Type:
Journal Article
Journal Name:
Experimental Cell Research
Additional Journal Information:
Journal Volume: 357; Journal Issue: 2; Other Information: Copyright (c) 2017 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 0014-4827
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; DRUGS; FILAMENTS; MAMMARY GLANDS; METASTASES; MICROTUBULES; NEOPLASMS; SCANNING ELECTRON MICROSCOPY; TOMOGRAPHY; TUMOR CELLS

Citation Formats

Killilea, Alison N., E-mail: ankillilea@berkeley.edu, Csencsits, Roseann, Le, Emily Bao Ngoc Thien, E-mail: emilybntle@gmail.com, Patel, Anand M., E-mail: anandpatel@lbl.gov, Kenny, Samuel J., E-mail: sjkenny1@berkeley.edu, Xu, Ke, Department of Chemistry, University of California Berkeley, CA 94720, and Downing, Kenneth H., E-mail: khdowning@lbl.gov. Cytoskeletal organization in microtentacles. United States: N. p., 2017. Web. doi:10.1016/J.YEXCR.2017.05.024.
Killilea, Alison N., E-mail: ankillilea@berkeley.edu, Csencsits, Roseann, Le, Emily Bao Ngoc Thien, E-mail: emilybntle@gmail.com, Patel, Anand M., E-mail: anandpatel@lbl.gov, Kenny, Samuel J., E-mail: sjkenny1@berkeley.edu, Xu, Ke, Department of Chemistry, University of California Berkeley, CA 94720, & Downing, Kenneth H., E-mail: khdowning@lbl.gov. Cytoskeletal organization in microtentacles. United States. doi:10.1016/J.YEXCR.2017.05.024.
Killilea, Alison N., E-mail: ankillilea@berkeley.edu, Csencsits, Roseann, Le, Emily Bao Ngoc Thien, E-mail: emilybntle@gmail.com, Patel, Anand M., E-mail: anandpatel@lbl.gov, Kenny, Samuel J., E-mail: sjkenny1@berkeley.edu, Xu, Ke, Department of Chemistry, University of California Berkeley, CA 94720, and Downing, Kenneth H., E-mail: khdowning@lbl.gov. Tue . "Cytoskeletal organization in microtentacles". United States. doi:10.1016/J.YEXCR.2017.05.024.
@article{osti_22738147,
title = {Cytoskeletal organization in microtentacles},
author = {Killilea, Alison N., E-mail: ankillilea@berkeley.edu and Csencsits, Roseann and Le, Emily Bao Ngoc Thien, E-mail: emilybntle@gmail.com and Patel, Anand M., E-mail: anandpatel@lbl.gov and Kenny, Samuel J., E-mail: sjkenny1@berkeley.edu and Xu, Ke and Department of Chemistry, University of California Berkeley, CA 94720 and Downing, Kenneth H., E-mail: khdowning@lbl.gov},
abstractNote = {Microtentacles are thin, flexible cell protrusions that have recently been described and whose presence enhances efficient attachment of circulating cells. They are found on circulating tumor cells and can be induced on a wide range of breast cancer cell lines, where they are promoted by factors that either stabilize microtubules or destabilize the actin cytoskeleton. Evidence suggests that they are relevant to the metastatic spread of cancer, so understanding their structure and formation may lead to useful therapies. Microtentacles are formed by microtubules and contain vimentin intermediate filaments, but beyond this, there is little information about their ultrastructure. We have used electron microscopy of high pressure frozen sections and tomography of cryo-prepared intact cells, along with super resolution fluorescence microscopy, to provide the first ultrastructural insights into microtubule and intermediate filament arrangement within microtentacles. By scanning electron microscopy it was seen that microtentacles form within minutes of addition of drugs that stabilize microtubules and destabilize actin filaments. Mature microtentacles were found to be well below one micrometer in diameter, tapering gradually to below 100 nm at the distal ends. They also contained frequent branches and bulges suggestive of heterogeneous internal structure. Super resolution fluorescence microscopy and examination of sectioned samples showed that the microtubules and intermediate filaments can occupy different areas within the microtentacles, rather than interacting intimately as had been expected. Cryo-electron tomography of thin regions of microtentacles revealed densely packed microtubules and absence of intermediate filaments. The number of microtubules ranged from several dozen in some areas to just a few in the thinnest regions, with none of the regular arrangement found in axonemes. Improved understanding of the mechanism of microtentacle formation, as well as the resultant structure, will be valuable in developing therapies against metastasis, if the hypothesized role of microtentacles in metastasis is confirmed. This work provides a significant step in this direction. - Highlights: • Microtentacles can form within minutes of the addition of inducing drugs. Microtentacle diameters are in the range of a tenth of a micrometer. • Microtubules are clearly resolved in microtentacles by cryo-electron tomography. • There is little microtubule-intermediate filament interaction in drug-induced microtentacles.},
doi = {10.1016/J.YEXCR.2017.05.024},
journal = {Experimental Cell Research},
issn = {0014-4827},
number = 2,
volume = 357,
place = {United States},
year = {2017},
month = {8}
}