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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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New microtubules arise and are integrated into the existing network. Ovals represent points of stabilization on the cortex. Circles indicate the growth of microtubules from the sides of other microtubules. Microtubules are referred to by arrow color. Note that most microtubules grow distances of 5 μm or more. (A) Depolymerization to a fixed location; corresponds to Video 3 (available at http://www.jcb.org/cgi/content/full/jcb.200505071/DC1). The red microtubule arises (red arrow and red line in schematic drawing) apparently from the side of the blue microtubule. After 15 s, the minus end of the red microtubule shrinks to a nearby location that is traversed by the orange microtubule and remains at this location for the 25 min of imaging. The orange microtubule depolymerizes. The end of the blue microtubule has formed a connection with the red microtubule. (B) Growth to a fixed location. The yellow microtubule grows upward to the location where the green microtubule begins and then continues to grow at a different angle. After a few minutes, the yellow microtubule depolymerizes from its minus end and pauses at this same point (see drawing). The green microtubule depolymerizes as well. Later, the pink microtubule arises and grows to this same location (yellow arrow is carried through to mark the location). (C) Depolymerization to a fixed location; corresponds to Video 4. The yellow arrows show a microtubule that arises and whose minus end quickly shrinks and is stabilized nearby. This microtubule curves as it grows, interacts with other microtubules, and remains in the network for the 23 min of imaging. The pink microtubule arises and, after 7 min, depolymerizes from its minus end, pausing at points on the cortex and at another microtubule during depolymerization. The orange microtubule arises and persists for the duration of imaging (15 min). (D) Stabilization of the plus end at a fixed location. The light blue microtubule grows to and becomes stabilized at a point on the basal cortex (see drawing). (E) Growth over a fixed location; corresponds to Video 5. The blue microtubule grows over the same point (yellow arrow) where another microtubule ends and interacts with this microtubule (it bends slightly). The plus end of the blue microtubule continues to grow, pausing at different microtubules along its path (bottom, open arrows; see life history plot to the right; asterisks indicate points of pause). At the end of imaging, the minus end of the blue microtubule has depolymerized a short distance and becomes stabilized at a point along its path (last panel). The microtubule had paused at this same location while growing. Bars, 5 μm.
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fig4: New microtubules arise and are integrated into the existing network. Ovals represent points of stabilization on the cortex. Circles indicate the growth of microtubules from the sides of other microtubules. Microtubules are referred to by arrow color. Note that most microtubules grow distances of 5 μm or more. (A) Depolymerization to a fixed location; corresponds to Video 3 (available at http://www.jcb.org/cgi/content/full/jcb.200505071/DC1). The red microtubule arises (red arrow and red line in schematic drawing) apparently from the side of the blue microtubule. After 15 s, the minus end of the red microtubule shrinks to a nearby location that is traversed by the orange microtubule and remains at this location for the 25 min of imaging. The orange microtubule depolymerizes. The end of the blue microtubule has formed a connection with the red microtubule. (B) Growth to a fixed location. The yellow microtubule grows upward to the location where the green microtubule begins and then continues to grow at a different angle. After a few minutes, the yellow microtubule depolymerizes from its minus end and pauses at this same point (see drawing). The green microtubule depolymerizes as well. Later, the pink microtubule arises and grows to this same location (yellow arrow is carried through to mark the location). (C) Depolymerization to a fixed location; corresponds to Video 4. The yellow arrows show a microtubule that arises and whose minus end quickly shrinks and is stabilized nearby. This microtubule curves as it grows, interacts with other microtubules, and remains in the network for the 23 min of imaging. The pink microtubule arises and, after 7 min, depolymerizes from its minus end, pausing at points on the cortex and at another microtubule during depolymerization. The orange microtubule arises and persists for the duration of imaging (15 min). (D) Stabilization of the plus end at a fixed location. The light blue microtubule grows to and becomes stabilized at a point on the basal cortex (see drawing). (E) Growth over a fixed location; corresponds to Video 5. The blue microtubule grows over the same point (yellow arrow) where another microtubule ends and interacts with this microtubule (it bends slightly). The plus end of the blue microtubule continues to grow, pausing at different microtubules along its path (bottom, open arrows; see life history plot to the right; asterisks indicate points of pause). At the end of imaging, the minus end of the blue microtubule has depolymerized a short distance and becomes stabilized at a point along its path (last panel). The microtubule had paused at this same location while growing. Bars, 5 μm.

Mentions: Microtubule dynamics were directly imaged in basal membrane cytoplasts that were prepared from polarized MDCK cells expressing GFP-tubulin. The majority of basal cytoplasts contained a steady-state network of microtubules in which the overall dimensions and organization of microtubules remained stable (Fig. 4). Generally, the network consists of many short (5.2 ± 2.2 μm) microtubules that arrange themselves into a network as they encounter one another through growth and shrinkage and form connections by adhering to one another end-to-side and side-to-side. Microtubules also bind to the basal cortex (Reilein and Nelson, 2005). The striking feature of the basal microtubule network is that it appears to be organized predominantly by stable interactions between individual microtubules and between microtubules and the basal cortex. Note, however, that the network is also undergoing slow, subtle remodeling by dynamic instability and de novo growth of a few microtubules.


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)

New microtubules arise and are integrated into the existing network. Ovals represent points of stabilization on the cortex. Circles indicate the growth of microtubules from the sides of other microtubules. Microtubules are referred to by arrow color. Note that most microtubules grow distances of 5 μm or more. (A) Depolymerization to a fixed location; corresponds to Video 3 (available at http://www.jcb.org/cgi/content/full/jcb.200505071/DC1). The red microtubule arises (red arrow and red line in schematic drawing) apparently from the side of the blue microtubule. After 15 s, the minus end of the red microtubule shrinks to a nearby location that is traversed by the orange microtubule and remains at this location for the 25 min of imaging. The orange microtubule depolymerizes. The end of the blue microtubule has formed a connection with the red microtubule. (B) Growth to a fixed location. The yellow microtubule grows upward to the location where the green microtubule begins and then continues to grow at a different angle. After a few minutes, the yellow microtubule depolymerizes from its minus end and pauses at this same point (see drawing). The green microtubule depolymerizes as well. Later, the pink microtubule arises and grows to this same location (yellow arrow is carried through to mark the location). (C) Depolymerization to a fixed location; corresponds to Video 4. The yellow arrows show a microtubule that arises and whose minus end quickly shrinks and is stabilized nearby. This microtubule curves as it grows, interacts with other microtubules, and remains in the network for the 23 min of imaging. The pink microtubule arises and, after 7 min, depolymerizes from its minus end, pausing at points on the cortex and at another microtubule during depolymerization. The orange microtubule arises and persists for the duration of imaging (15 min). (D) Stabilization of the plus end at a fixed location. The light blue microtubule grows to and becomes stabilized at a point on the basal cortex (see drawing). (E) Growth over a fixed location; corresponds to Video 5. The blue microtubule grows over the same point (yellow arrow) where another microtubule ends and interacts with this microtubule (it bends slightly). The plus end of the blue microtubule continues to grow, pausing at different microtubules along its path (bottom, open arrows; see life history plot to the right; asterisks indicate points of pause). At the end of imaging, the minus end of the blue microtubule has depolymerized a short distance and becomes stabilized at a point along its path (last panel). The microtubule had paused at this same location while growing. Bars, 5 μm.
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fig4: New microtubules arise and are integrated into the existing network. Ovals represent points of stabilization on the cortex. Circles indicate the growth of microtubules from the sides of other microtubules. Microtubules are referred to by arrow color. Note that most microtubules grow distances of 5 μm or more. (A) Depolymerization to a fixed location; corresponds to Video 3 (available at http://www.jcb.org/cgi/content/full/jcb.200505071/DC1). The red microtubule arises (red arrow and red line in schematic drawing) apparently from the side of the blue microtubule. After 15 s, the minus end of the red microtubule shrinks to a nearby location that is traversed by the orange microtubule and remains at this location for the 25 min of imaging. The orange microtubule depolymerizes. The end of the blue microtubule has formed a connection with the red microtubule. (B) Growth to a fixed location. The yellow microtubule grows upward to the location where the green microtubule begins and then continues to grow at a different angle. After a few minutes, the yellow microtubule depolymerizes from its minus end and pauses at this same point (see drawing). The green microtubule depolymerizes as well. Later, the pink microtubule arises and grows to this same location (yellow arrow is carried through to mark the location). (C) Depolymerization to a fixed location; corresponds to Video 4. The yellow arrows show a microtubule that arises and whose minus end quickly shrinks and is stabilized nearby. This microtubule curves as it grows, interacts with other microtubules, and remains in the network for the 23 min of imaging. The pink microtubule arises and, after 7 min, depolymerizes from its minus end, pausing at points on the cortex and at another microtubule during depolymerization. The orange microtubule arises and persists for the duration of imaging (15 min). (D) Stabilization of the plus end at a fixed location. The light blue microtubule grows to and becomes stabilized at a point on the basal cortex (see drawing). (E) Growth over a fixed location; corresponds to Video 5. The blue microtubule grows over the same point (yellow arrow) where another microtubule ends and interacts with this microtubule (it bends slightly). The plus end of the blue microtubule continues to grow, pausing at different microtubules along its path (bottom, open arrows; see life history plot to the right; asterisks indicate points of pause). At the end of imaging, the minus end of the blue microtubule has depolymerized a short distance and becomes stabilized at a point along its path (last panel). The microtubule had paused at this same location while growing. Bars, 5 μm.
Mentions: Microtubule dynamics were directly imaged in basal membrane cytoplasts that were prepared from polarized MDCK cells expressing GFP-tubulin. The majority of basal cytoplasts contained a steady-state network of microtubules in which the overall dimensions and organization of microtubules remained stable (Fig. 4). Generally, the network consists of many short (5.2 ± 2.2 μm) microtubules that arrange themselves into a network as they encounter one another through growth and shrinkage and form connections by adhering to one another end-to-side and side-to-side. Microtubules also bind to the basal cortex (Reilein and Nelson, 2005). The striking feature of the basal microtubule network is that it appears to be organized predominantly by stable interactions between individual microtubules and between microtubules and the basal cortex. Note, however, that the network is also undergoing slow, subtle remodeling by dynamic instability and de novo growth of a few microtubules.

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.

Show MeSH
Related in: MedlinePlus