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Self-organization of an acentrosomal microtubule network at the basal cortex of polarized epithelial cells.

Reilein A, Yamada S, Nelson WJ - J. Cell Biol. (2005)

Bottom Line: Microtubules undergoing dynamic instability without any stabilization points continuously remodel their organization without reaching a steady-state network.However, the addition of increased microtubule stabilization at microtubule-microtubule and microtubule-cortex interactions results in the rapid assembly of a steady-state microtubule network in silico that is remarkably similar to networks formed in situ.These results define minimal parameters for the self-organization of an acentrosomal microtubule network.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Beckman Center for Molecular and Genetic Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.

ABSTRACT
Mechanisms underlying the organization of centrosome-derived microtubule arrays are well understood, but less is known about how acentrosomal microtubule networks are formed. The basal cortex of polarized epithelial cells contains a microtubule network of mixed polarity. We examined how this network is organized by imaging microtubule dynamics in acentrosomal basal cytoplasts derived from these cells. We show that the steady-state microtubule network appears to form by a combination of microtubule-microtubule and microtubule-cortex interactions, both of which increase microtubule stability. We used computational modeling to determine whether these microtubule parameters are sufficient to generate a steady-state acentrosomal microtubule network. Microtubules undergoing dynamic instability without any stabilization points continuously remodel their organization without reaching a steady-state network. However, the addition of increased microtubule stabilization at microtubule-microtubule and microtubule-cortex interactions results in the rapid assembly of a steady-state microtubule network in silico that is remarkably similar to networks formed in situ. These results define minimal parameters for the self-organization of an acentrosomal microtubule network.

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Microtubule organization in polarized epithelial cells shows microtubule–microtubules intersections in the basal network. (A) GFP-tubulin–expressing MDCK cells reconstructed from images spaced 0.2 μm apart in the z axis. (B) Image rotated with Volocity software to provide a view looking from the bottom of the cells upwards to show the microtubule network on the basal surface. (C) Deconvolution microscopy of tubulin immunofluorescence at representative basal, middle, and apical sections of polarized MDCK cells shows the webbed microtubule network apical and basal to the nucleus and microtubule bundles running alongside the nucleus parallel to the lateral membrane. Bar, 10 μm. (D) Close-up views of the basal network of individual cells show intersections formed by microtubules (arrows) and the variable weave of the microtubule network. (E) Microtubules at the base of a Caco-2 cell are organized into a similar intersecting network. (D and E) Bars, 5 μm.
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fig1: Microtubule organization in polarized epithelial cells shows microtubule–microtubules intersections in the basal network. (A) GFP-tubulin–expressing MDCK cells reconstructed from images spaced 0.2 μm apart in the z axis. (B) Image rotated with Volocity software to provide a view looking from the bottom of the cells upwards to show the microtubule network on the basal surface. (C) Deconvolution microscopy of tubulin immunofluorescence at representative basal, middle, and apical sections of polarized MDCK cells shows the webbed microtubule network apical and basal to the nucleus and microtubule bundles running alongside the nucleus parallel to the lateral membrane. Bar, 10 μm. (D) Close-up views of the basal network of individual cells show intersections formed by microtubules (arrows) and the variable weave of the microtubule network. (E) Microtubules at the base of a Caco-2 cell are organized into a similar intersecting network. (D and E) Bars, 5 μm.

Mentions: Microtubules in columnar epithelial cells are organized in the apico–basal axis of the cell into parallel bundles with their plus ends oriented toward the basal membrane and weblike networks on the apical and basal cortex (Fig. 1, A–C; Bacallao et al., 1989; Gilbert et al., 1991; Meads and Schroer, 1995; Eaton et al., 1996). High resolution imaging of microtubules on the basal cortex reveals a network of microtubules formed by intersections that arise from the termination of one microtubule on the side of another or from two microtubules crossing over one another (Fig. 1, D and E). A similar microtubule network is found on the basal cortex of different types of polarized epithelial cells, including Caco-2 cells derived from colonic epithelium (Figs. 1 E and 2 D) and EpH4 cells derived from mammary epithelium (Fig. 2 E).


Self-organization of an acentrosomal microtubule network at the basal cortex of polarized epithelial cells.

Reilein A, Yamada S, Nelson WJ - J. Cell Biol. (2005)

Microtubule organization in polarized epithelial cells shows microtubule–microtubules intersections in the basal network. (A) GFP-tubulin–expressing MDCK cells reconstructed from images spaced 0.2 μm apart in the z axis. (B) Image rotated with Volocity software to provide a view looking from the bottom of the cells upwards to show the microtubule network on the basal surface. (C) Deconvolution microscopy of tubulin immunofluorescence at representative basal, middle, and apical sections of polarized MDCK cells shows the webbed microtubule network apical and basal to the nucleus and microtubule bundles running alongside the nucleus parallel to the lateral membrane. Bar, 10 μm. (D) Close-up views of the basal network of individual cells show intersections formed by microtubules (arrows) and the variable weave of the microtubule network. (E) Microtubules at the base of a Caco-2 cell are organized into a similar intersecting network. (D and E) Bars, 5 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171299&req=5

fig1: Microtubule organization in polarized epithelial cells shows microtubule–microtubules intersections in the basal network. (A) GFP-tubulin–expressing MDCK cells reconstructed from images spaced 0.2 μm apart in the z axis. (B) Image rotated with Volocity software to provide a view looking from the bottom of the cells upwards to show the microtubule network on the basal surface. (C) Deconvolution microscopy of tubulin immunofluorescence at representative basal, middle, and apical sections of polarized MDCK cells shows the webbed microtubule network apical and basal to the nucleus and microtubule bundles running alongside the nucleus parallel to the lateral membrane. Bar, 10 μm. (D) Close-up views of the basal network of individual cells show intersections formed by microtubules (arrows) and the variable weave of the microtubule network. (E) Microtubules at the base of a Caco-2 cell are organized into a similar intersecting network. (D and E) Bars, 5 μm.
Mentions: Microtubules in columnar epithelial cells are organized in the apico–basal axis of the cell into parallel bundles with their plus ends oriented toward the basal membrane and weblike networks on the apical and basal cortex (Fig. 1, A–C; Bacallao et al., 1989; Gilbert et al., 1991; Meads and Schroer, 1995; Eaton et al., 1996). High resolution imaging of microtubules on the basal cortex reveals a network of microtubules formed by intersections that arise from the termination of one microtubule on the side of another or from two microtubules crossing over one another (Fig. 1, D and E). A similar microtubule network is found on the basal cortex of different types of polarized epithelial cells, including Caco-2 cells derived from colonic epithelium (Figs. 1 E and 2 D) and EpH4 cells derived from mammary epithelium (Fig. 2 E).

Bottom Line: Microtubules undergoing dynamic instability without any stabilization points continuously remodel their organization without reaching a steady-state network.However, the addition of increased microtubule stabilization at microtubule-microtubule and microtubule-cortex interactions results in the rapid assembly of a steady-state microtubule network in silico that is remarkably similar to networks formed in situ.These results define minimal parameters for the self-organization of an acentrosomal microtubule network.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Beckman Center for Molecular and Genetic Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.

ABSTRACT
Mechanisms underlying the organization of centrosome-derived microtubule arrays are well understood, but less is known about how acentrosomal microtubule networks are formed. The basal cortex of polarized epithelial cells contains a microtubule network of mixed polarity. We examined how this network is organized by imaging microtubule dynamics in acentrosomal basal cytoplasts derived from these cells. We show that the steady-state microtubule network appears to form by a combination of microtubule-microtubule and microtubule-cortex interactions, both of which increase microtubule stability. We used computational modeling to determine whether these microtubule parameters are sufficient to generate a steady-state acentrosomal microtubule network. Microtubules undergoing dynamic instability without any stabilization points continuously remodel their organization without reaching a steady-state network. However, the addition of increased microtubule stabilization at microtubule-microtubule and microtubule-cortex interactions results in the rapid assembly of a steady-state microtubule network in silico that is remarkably similar to networks formed in situ. These results define minimal parameters for the self-organization of an acentrosomal microtubule network.

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