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29 CELL CYCLE AND GROWTH REGULATION (Full Edition)
23 A monomeric G protein controls spindle assembly
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The active form of the G protein Ran (Ran-GTP) causes importin dimers to release proteins that trigger microtubule nucleation.
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The Ran-activating protein RCC is localized on chromosomes, generating a high local concentration of Ran-GTP.
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The proteins released by the importins have several different functions that assist microtubule nucleation.
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The trigger for the reorganization of
microtubules from the interphase network into the spindle may be the
breakdown of the nuclear envelope, which exposes nuclear components to
cytoplasmic components. Indirect evidence has been available for some
time to indicate a connection, but only recently has a molecular
mechanism been suggested. The important point here is that the ability
of an MTOC to nucleate microtubules must be controlled,
so that it happens only in the right time and place.
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The critical component is a monomeric G
protein called Ran, which controls the direction of protein transport
through the nuclear envelope. Like all members of its class, Ran is
active when bound to GTP, and inactive when bound to GDP (see G proteins).
Conditions in the nucleus and cytosol differ so that typically there is
Ran-GTP in the nucleus, but there is Ran-GDP in the cytosol. Protein
export complexes are stable in the presence of Ran-GTP, whereas import
complexes are stable in the presence of Ran-GDP. So export complexes
are driven to form in the nucleus and dissociate in the cytosol,
whereas the reverse is true of import complexes (see Transport
receptors carry cargo proteins through the pore).
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Mutations in some proteins that bind to
Ran cause the spindle to malfunction, and overexpression of the protein
RanBPM (another Ran-binding protein) causes the formation of ectopic
asters — arrays of microtubules emanating from centrosomes. The usual
assay for these experiments is to inject demembranated sperm into Xenopus
eggs. The sperm centrioles assemble into centrosomes that nucleate
microtubule asters. Using this assay identifies proteins that can
stimulate nucleation. These include a mutant of Ran and the protein RCC
that maintains Ran in the GTP-bound active state (1592; 1593).
The most likely explanation is that the breakdown of the nuclear
envelope releases Ran-GTP, which then triggers microtubule nucleation
by centrosomes.
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Does Ran act directly or indirectly? One
of the targets for Ran in the nuclear transport process is the import
receptor importin-ß,
which (in combination with importin-a)
transports cargo proteins from the cytoplasm to the nucleus. It turns
out that the importin dimer binds to proteins that affect microtubules.
One of these proteins is Xklp2, which connects a motor (a protein that
moves other proteins) to microtubules at the poles; another is NuMA
which cross-links microtubules at the poles during mitosis. When the
complex of importins with either of these proteins is exposed to
Ran-GTP, it dissociates, releasing the cargo protein, which can then
act to trigger microtubule nucleation (1594; 1595).
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Figure 29.42
The Ran-activating protein RCC is localized on
chromosomes. Ran-GTP is high in the nucleus in the interphase cell.
When the nuclear envelope breaks down, RCC maintains a high level of
Ran-GTP in the vicinity of the chromosomes. This causes importin dimers
to release the proteins bound to them. These proteins cause microtubule
nucleation.
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How does the exposure of the importins to
Ran-GTP change at mitosis? Figure 29.42
shows that the situation in the cytoplasm of the interphase cell, and
then correspondingly in the spindle, is that Ran is predominantly in
the form of Ran-GDP, and therefore does not affect the importin
complex. But there are localized areas where Ran-GTP is formed. The
Ran-activating protein RCC is located on chromatin, so Ran-GTP forms in
the vicinity of the chromosomes. This releases the proteins that are
bound to the importin dimer, which activate the kinetochores to connect
to microtubules.
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Last Revised on May 20, 2004
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1592 Ohba, T., Nakamura, M., Nishitani, H., and Nishimoto, T.
(1999).
Self-organization of microtubule asters induced in Xenopus egg extracts by GTP-bound Ran.
Science 284, 1356-1358.
PubMed Journal
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1593 Wilde, A. and Zheng, Y.
(1999).
Stimulation of microtubule aster formation and spindle assembly by the small GTPase Ran.
Science 284, 1359-1362.
PubMed Journal
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1594 Gruss, O. J., Carazo-Salas, R. E., Schatz,
C. A., Guarguaglini, G., Kast, J., Wilm, M., Le Bot, N., Vernos, I.,
Karsenti, E., and Mattaj, I. W. (2001). Ran induces spindle
assembly by reversing the inhibitory effect of importin alpha on TPX2
activity. Cell 104, 83-93. PubMed Journal
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1595 Nachury, M. V., Maresca, T. J., Salmon, W. C., Waterman-Storer, C. M., Waterman-Storer, R., Heald, R., and Weis, K.
(2001).
Importin beta is a mitotic target of the small GTPase Ran in spindle assembly.
Cell 104, 95-106.
PubMed Journal
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© Jones and Bartlett Publishers (2007)
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