1 Department of Biochemistry, University of Washington, Seattle, WA 98195,
USA
2 Molecular and Cellular Biology Program, University of Washington, Seattle, WA
98195, USA
3 State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell
Biology, Shanghai Institute for Biological Sciences, Shanghai, China
4 Department of Anatomy, University of California, San Francisco, CA 94143 and
Lawrence Berkeley National Laboratory, University of California, Berkeley, CA
94720, USA
5 Department of Genetics and Developmental Biology, University of Connecticut,
Farmington, CN 06030, USA
Author for correspondence (e-mail:
kimelman{at}u.washington.edu)
Accepted 23 July 2003
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SUMMARY |
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Key words: Frat, Wnt pathway, Axis specification, Cortical rotation, Microtubules
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Introduction |
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Although the molecular identity of the dorsal determinants is not clear, it
is known that their translocation leads to the dorsal accumulation of
ß-catenin, which then activates the expression of dorsal organizer genes
at the onset of zygotic transcription
(Schneider et al., 1996;
Larabell et al., 1997
;
Rowning et al., 1997
).
Cytoplasmic transplant experiments using ß-catenin-depleted embryos have
shown that ß-catenin is not the endogenous dorsalizing activity, but that
instead this activity probably consists of proteins involved in ß-catenin
stabilization (Marikawa and Elinson,
1999
). ß-catenin is normally phosphorylated by the
serine-threonine kinase glycogen synthase kinase 3 (GSK3) within a protein
complex that also includes Axin and the adenomatous polyposis coli gene
product (APC), and this phosphorylation targets ß-catenin for degradation
by the ubiquitin-proteosome pathway (reviewed by
Moon and Kimelman, 1998
;
Bienz, 1999
;
Polakis, 2000
). Work from many
laboratories has led to a model in which the localized inhibition of GSK3 in
the dorsal region causes the dorsal accumulation of ß-catenin
(He et al., 1995
;
Pierce and Kimelman, 1995
;
Yost et al., 1996
;
Yost et al., 1998
;
Dominguez and Green, 2000
;
Salic et al., 2000
). How GSK3
becomes locally inhibited by the dorsal determinants, however, is still an
open question.
A strong candidate component of the translocating dorsalizing activity is
GBP, a vertebrate-specific GSK3-binding protein
(Yost et al., 1998). Depletion
of endogenous GBP from the embryo with antisense oligonucleotides causes a
loss of dorsal axial structures, showing that GBP is required for dorsal axis
formation (Yost et al., 1998
).
GBP inhibits GSK3 activity by preventing its binding to Axin, thus preventing
GSK3 from phosphorylating ß-catenin
(Farr III et al., 2000
;
Salic et al., 2000
). When
microinjected ventrally, GBP mimics the endogenous dorsal signal and induces
the formation of a secondary dorsal axis
(Yost et al., 1998
), and
overexpression of GBP also leads to GSK3 degradation in the cortical shear
zone (Dominguez and Green,
2000
). In addition to binding GSK3, GBP also binds directly to Dsh
(Li et al., 1999
;
Salic et al., 2000
;
Lee et al., 2001
;
Hino et al., 2003
), a positive
effector of the canonical Wnt signaling pathway
(Klingensmith et al., 1994
;
Yanagawa et al., 1995
;
Sokol, 1996
;
Rothbacher et al., 2000
).
Together, these two proteins potently synergize to stabilize ß-catenin
(Li et al., 1999
;
Salic et al., 2000
;
Hino et al., 2003
).
Thus, both GBP and its binding partner Dsh have characteristics that
strongly suggest that they are part of the endogenous dorsalizing activity.
Furthermore, Dsh-GFP has been shown to form particles in the shear zone that
exhibit directed movement on microtubules, and endogenous Dsh accumulates
dorsally by the end of cortical rotation
(Miller et al., 1999).
However, no direct molecular link has yet been established between either GBP
or Dsh and the microtubule array. In this study, we demonstrate that GBP binds
kinesin light chain (KLC), a component of the plus end-directed microtubule
motor kinesin. Like Dsh, GBP-GFP and KLC-GFP form particles that exhibit fast,
directional translocation in the shear zone during the period of cortical
rotation. Our results suggest a model in which GBP acts initially as a link
between the transport apparatus and the dorsalizing activity, and subsequently
as an inhibitor of GSK3 in the ß-catenin degradation complex.
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Materials and methods |
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Two-hybrid and Xenopus cDNA screens
A ProQuest mouse brain cDNA library (Life Technologies) was screened with
full-length mouse Frat1 as bait using the ProQuest Two-Hybrid System
(Life Technologies) according to the manufacturer's instructions. One of the
KLC clones obtained was cloned into the pGluCMV vector
(Li et al., 1999) and used to
screen a Xenopus oocyte cDNA library in
GT10
(Rebagliati et al., 1985
),
from which a full-length KLC clone was obtained.
Egg manipulations and confocal microscopy
Oocytes were surgically removed from Xenopus females. Stage VI
oocytes were manually defolliculated and injected with 50-100 ng of synthetic
RNA, as was done previously (Miller et
al., 1999). After injection, oocytes were incubated at 17-18°C
for 9-14 hours, then matured in Oocyte Culture Medium (OCM; 60% Leibowitz L-15
medium with 0.4% BSA, 1 mM L-glutamine, 5 µg/ml gentamycin) [modified from
Kloc et al. (Kloc et al.,
1996
)] with 0.5-5 µM progesterone for 12-15 hours. After
germinal vesicle breakdown, mature eggs were placed in 0.3x Modified
Ringers (MR; 100 mM NaCl, 2 mM KCl, 2 mM CaCl2, 1 mM
MgCl2, 5 mM HEPES, pH 7.5) with 6% Ficoll and pricked in the top
third of the animal hemisphere with a fine glass needle to induce activation
and cortical rotation. Some eggs were incubated in 1 µg/ml Nile Red prior
to pricking to stain yolk platelets.
To monitor the behavior of GBP-GFP particles, images were captured with a scanning laser confocal microscope (model 1024, BioRad Laboratories, Hercules, CA, USA). A 60x PlanApo oil immersion objective lens, 1.4 NA, was used to capture a field of view at a focal plane 4-8 µm from the vegetal surface of the egg. Images were collected at 1.5-3.3-second intervals and typically several 3- to 5-minute-long movies were collected for each egg during the period of cortical rotation. Image shown was collected with an electronic zoom of 2. For XKLC4-GFP particles, images were collected with a Zeiss LSM510 NLO microscope equipped with an inverted microscope. A 40x Apochromat water immersion lens, 1.2 NA, was used to capture images as for GBP-GFP at 1.5-second intervals.
To image wild-type and mutant GBP-GFP particles in fixed eggs, eggs were prepared as for live imaging and fixed 60 minutes post-activation during peak cortical rotation in room temperature fix solution (4% paraformaldehyde, 0.1% glutaraldehyde, 0.1% Triton X-100, 100 mM KCl, 3 mM MgCl2, 5 mM HEPES, 150 mM sucrose pH 7.4). Images of the vegetal shear zone were collected using a Leica TCS SP/MP scanning confocal microscope with a 40x PlanApo oil immersion lens, 1.25 N.A. Images shown were collected with an electronic zoom of 2.
For whole-mount immunocytochemistry of eggs expressing GBP-GFP and XKLC4-HA, eggs were injected with approximately 50 ng of each RNA and prepared as for live imaging. Eggs were immersed in room temperature fix solution 60 minutes post-activation as described for GBP-GFP alone and incubated for 1 hour at room temperature, then incubated overnight at 4°C. Blocking of nonspecific binding was performed in Super Block (Pierce) with 0.2% Triton X-100. Eggs were incubated overnight at 4°C with anti-HA antibodies (1:10,000 dilution; Covance) in Super Block/0.2% Triton X-100 followed by three washes in Super Block/0.2% Triton X-100, then incubated overnight in Alexa Fluor 568 goat anti-mouse IgG (Molecular Probes) secondary antibodies (1:600 dilution). Eggs were imaged as described for GBP-GFP.
Embryos and microinjection
Embryos were microinjected (Moon and
Christian, 1989) with RNA synthesized from CS2+-derived constructs
linearized with Asp718 or NotI using the Sp6 mMessage mMachine kit (Ambion)
and purified away from unincorporated nucleotides with Microcon 100s
(Millipore). Wild-type (1-1.5 ng) or mutant XKLC4-HA RNA, 0.5-1 ng WT
or mutant GBP-myc RNA, and 1.5 ng WT or kinase-dead GSK3
(kdGSK3)-myc RNA were injected per embryo.
Immunoprecipitation and western blotting
Immunoprecipitations from COS-7 cells were performed as described
(Li et al., 1999).
Xenopus embryos were lysed 4-5 hours after RNA injection in 1% Triton
X-100 lysis buffer (Rubinfeld et al.,
1993
) supplemented with Complete protease inhibitors (Roche).
Embryo lysates were cleared by centrifugation in a microfuge at 14,926
g and half of the clear cytoplasmic layer retained for
analysis. Antibody for immunoprecipitation (anti-HA; Covance) was used at 0.2
µl per 100 µl of lysate. Immunocomplexes were collected on Protein G
beads (Amersham/Pharmacia) and washed with 1% NP-40, 50 mM Tris pH 8, 150 mM
NaCl. Proteins were detected on anti-HA, anti-myc (Covance) Western blots
using a goat anti-mouse IgG HRP secondary antibody (Zymed) developed with
enhanced chemiluminescence (NEN).
For analysis of expression of the WT GBP-GFP, C-III-GFP,
N-III-GFP,
-II-GFP and GFP constructs in eggs, stage VI oocytes
were injected with equivalent amounts of each RNA, incubated and matured in
vitro as for the live confocal imaging experiments. Eggs were then processed
for SDS-PAGE as in the immunoprecipitation experiments and Western blots on
the total lysates were performed with an anti-GFP antibody (Covance). In
Fig. 7G oocytes were injected
with 92 ng of RNA each, in Fig.
7H oocytes received 88 ng of RNA each, and in both cases
approximately 0.4 oocyte equivalent was loaded in each lane.
|
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Results |
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As was seen for Dsh-GFP (Miller et al.,
1999), GBP-GFP localized to bright particles that were enriched in
the 4-8-µm-deep subcortical shear zone relative to the deeper vegetal core
cytoplasm. During rotation, a subset of the GBP-GFP particles exhibited fast,
unidirectional movement opposite the rotation of the core
(Fig. 1B) (see also Movie 1 at
http://dev.biologists.org/supplemental/),
as was demonstrated for Dsh-GFP (Miller et
al., 1999
). The GBP-GFP particles moved with a wide range of
velocities (15-127 µm/minute) relative to the inner core. Velocities were
fairly evenly distributed across this range, with a mean velocity of
47±27 µm/minute (n=31 particles from 3 eggs). These
particles often appeared to stream one after the other and moved in a
saltatory fashion, interspersing periods of fast transport with pauses. In all
cases, we also observed a population of non-translocating particles that moved
in the reverse direction at a slower rate, accompanying the rotation of the
core (4-11 µm/minute). These results indicate that, like Dsh, GBP is
capable of undergoing fast, directed transport away from the vegetal pole of
the egg during cortical rotation.
KLC associates with GBP/Frat
In order to find new binding partners for GBP and its mammalian ortholog
Frat (Jonkers et al., 1997;
Yost et al., 1998
), we
performed a yeast two-hybrid screen using mouse Frat1 and a mouse
brain cDNA library. From 107 transformants we obtained 172 His+,
ß-gal+ positives of which 23 were identified as GSK3, the second
most abundant type of clone. Intriguingly, the most highly represented gene
among the positive clones was a member of the KLC family, mouse KLC1,
obtained in 31 of the 172 positives.
To determine whether the interaction between Frat1 and KLC1 seen in yeast occurs in mammalian cells, co-immunoprecipitations were performed from COS7 cells transfected with FLAG-tagged Frat1 along with one of the KLC1 clones from the two-hybrid screen fused to a Glu-Glu (GG) tag. KLC1-GG was co-immunoprecipitated by anti-FLAG antibody in the presence of Frat1-FLAG (Fig. 2A, lane 8), but not in its absence (Fig. 2A, lane 7). Conversely, Frat1-FLAG was co-immunoprecipitated by the anti-GG antibody in the presence of KLC1-GG (Fig. 2A, lane 4) at a level much greater than background (Fig. 2A, lane 3).
|
|
The heptad repeat region of XKLC4 is required for GBP binding
Kinesin light chains are subunits of kinesin, a heterotetrameric,
plus-end-directed microtubule motor protein. The two highly conserved domains
in KLC have independent functions; the heptad repeat region binds kinesin
heavy chain, the motor-containing subunit of kinesin, whereas the TPR region
has recently been shown to associate with proteins involved in transporting
cargo (reviewed by Kamal and Goldstein,
2002) (see also the Kinesin Home Page at
http://www.blocks.fhcrc.org/~kinesin/).
To determine whether either of these conserved domains are important for the
GBP/XKLC4 interaction, we first made constructs that consisted of the
N-terminal third of XKLC4, which includes the heptad repeats (XKLC4-N) or the
C-terminal two-thirds of XKLC4, which includes the TPR repeats (XKLC4-TPR),
fused to an HA tag (Fig. 4A).
Whereas full-length XKLC4 and XKLC4-N co-immunoprecipitated GBP-myc from
Xenopus embryos, XKLC4-TPR failed to interact with GBP
(Fig. 4B). This result is
consistent with our observation that some of the KLC1 clones obtained in the
two-hybrid screen, including the KLC1 clone used in the co-immunoprecipitation
experiment with Frat1 (Fig.
2A), contained all of the heptad repeats but none of the TPR
repeats. These results indicate that the GBP-binding region resides in the
N-terminal third of XKLC4.
|
XKLC4 translocates during cortical rotation
Although our results demonstrated that XKLC4 could bind GBP, it was
important to determine whether XKLC4 could also form particles and translocate
like GBP and Dsh. As with GBP, we used time-lapse confocal microscopy to
observe the localization of XKLC4-GFP in the vegetal region of immobilized,
prick-activated eggs during cortical rotation. Nile Red was used to label yolk
platelets to assess the direction and velocity of core rotation. Like GBP-GFP,
XKLC4-GFP formed bright particles in the 4-8-µm-deep shear zone between the
cortex and the inner core cytoplasm. Just prior to the initiation of cortical
rotation, XKLC4-GFP particles began to exhibit random saltations (see Movie 2
at
http://dev.biologists.org/supplemental/).
Once rotation was underway these saltations became organized into directional
translocation opposite the direction of the core rotation
(Fig. 5A,B) (Movie 3 at
http://dev.biologists.org/supplemental/),
with a range of velocities similar to that observed for GBP-GFP (18-89
µm/minute, with an average velocity of 48±18 µm/minute). Late in
rotation, the velocity of core rotation decreased, as did the frequency of
directed particle translocation (Movie 4 at
http://dev.biologists.org/supplemental/).
As with GBP-GFP, a population of XKLC4-GFP particles in each egg did not
actively translocate, but rather moved slowly with the core in the opposite
direction (7-9 µm/minute). These observations indicate that, like GBP,
XKLC4 is capable of undergoing fast, directional transport in the vegetal
shear zone of the egg during cortical rotation.
|
Domain III of GBP is required for binding to XKLC4
GBP contains three domains, I, II and III, that are conserved between the
Xenopus and mammalian proteins
(Yost et al., 1998). We
previously identified domain III as the site of GSK3 binding
(Yost et al., 1998
). No
binding partners have yet been identified for domains I or II. In order to
determine the binding site for KLC, we generated a series of mutant GBP
constructs containing 8-15 amino acid deletions, focusing on regions within
the three conserved domains (Fig.
6A). Xenopus embryos expressing XKLC4-HA and either a
myc-tagged mutant GBP or WT GBP-myc were lysed and immunoprecipitated with the
anti-HA antibody. Deletion of domain II (
-II) had no effect on the
ability of GBP-myc to associate with XKLC4-HA, nor did deletion of the
N-terminal half of domain I (
N-I;
Fig. 6B). Deletion of the
C-terminal half of domain I (
C-I) slightly but consistently diminished
the interaction (Fig. 6C). The
most severe effects on binding were achieved by removing either the N-terminal
(
N-III) or C-terminal (
C-III) half of domain III. With either of
these mutations, the amount of co-precipitating GBP-myc was diminished to
almost background levels (Fig.
6C; see also Fig.
6B). We conclude that domain III of GBP is required for its
interaction with XKLC4, with a more minor role played by a region in the
C-terminus of domain I.
|
Domain III of GBP is required for normal particle formation
We predicted that a GBP mutant incapable of binding KLC should also be
compromised in its ability to form particles and translocate on microtubules
during cortical rotation. We therefore compared the localization of GFP-tagged
C-III, which lacks 10 amino acids in domain III
(Fig. 6A), with that of GBP-GFP
in prick-activated eggs. In some experiments, eggs were fixed during cortical
rotation and imaged later. In obvious contrast to the punctate localization of
WT GBP-GFP,
C-III-GFP appeared primarily as a bright, diffuse green
glow throughout the shear zone, with very few particles forming
(Fig. 7B). This distribution is
different from what we observed for WT GBP-GFP, which also exhibited a
background glow but in most cases formed hundreds of discrete particles in a
single egg (Fig. 7A; see also
Fig. 1B). In the live eggs, the
few
C-III-GFP particles moved slowly in the same direction as the core,
with the exception of one particle in one egg (n=14 eggs) seen to
quickly dart opposite the direction of core rotation (data not shown).
Analysis of the expression of WT GBP-GFP and
C-III-GFP by Western blot
demonstrated that they were expressed at the same levels in eggs
(Fig. 7G).
To further investigate the correspondence between the ability of GBP to
interact with KLC and its ability to form particles, we assessed the
localization of the other deletion mutant in domain III, N-III-GFP, in
the shear zone during cortical rotation. Like
C-III-GFP,
N-III-GFP was present as a bright, diffuse glow in the vegetal region
of the egg, forming very few particles
(Fig. 7E). GFP alone, as
previously reported, also localized as a diffuse glow
(Fig. 7C)
(Miller et al., 1999
). In
contrast, the localization of a GBP deletion mutant that retained KLC binding
in the co-immunoprecipitation assay,
-II, was indistinguishable from
that of WT GBP-GFP (Fig. 7F).
Again, analysis of the expression of the GBP-GFP constructs by Western blot
demonstrated that they were expressed at equivalent levels in eggs
(Fig. 7H).
We conclude that domain III of GBP is important for both in vivo association with KLC and for proper localization and translocation of GBP during cortical rotation. The residual ability of a domain III deletion mutant to form particles and translocate may be because of its ability to make contacts with other components of a larger complex of proteins, allowing some association with endogenous particles even in the absence of KLC binding. However, in general we observed a good correlation between the ability of GBP constructs to associate with KLC and their ability to form particles in the shear zone, suggesting that KLC binding is a key factor in GBP particle formation.
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Discussion |
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Evidence for involvement of kinesin-related proteins in the
translocation of dorsal determinants
Several lines of evidence point to the involvement of an endogenous
kinesin-related protein in the active transport of the dorsalizing activity to
the dorsal region from the vegetal pole. First, cytoplasmic transfer
experiments have demonstrated that the endogenous dorsal determinants move
from the vegetal region toward the prospective dorsal side of the embryo
beyond the extent of cortical displacement due to rotation, indicating that
the dorsalizing activity does not just move with the cortex
(Yuge et al., 1990;
Fujisue et al., 1993
;
Holowacz and Elinson, 1993
;
Kikkawa et al., 1996
;
Sakai, 1996
;
Kageura, 1997
). Second, hook
assays have shown that approximately 90% of the subcortical microtubule
bundles that make up the parallel array are aligned with their plus ends
toward the future dorsal side, suggesting that active transport in this
direction is performed by a plus end-directed motor
(Houliston and Elinson,
1991b
). Third, endogenous vesicles in the shear zone move dorsally
from the vegetal pole with a velocity and saltatory behavior suggestive of
kinesin-mediated transport (Rowning et
al., 1997
). Fourth, Dsh-GFP, GBP-GFP and XKLC4-GFP
(Miller et al., 1999
; this
work) move with velocities that are in the range in which kinesin and
kinesin-like proteins have been measured to move in vitro and in vivo
(Howard et al., 1989
;
Kawaguchi and Ishiwata, 2000
;
Zhou et al., 2001
;
Yajima et al., 2002
).
Together, these observations support our evidence that a motor protein in the
kinesin family is involved in translocating dorsal determinants along the
subcortical microtubule array.
Is GBP a component of the dorsalizing activity?
GBP is known to be required for dorsal axis formation
(Yost et al., 1998), but its
relationship to the dorsalizing activity that translocates during the first
cell cycle has been unclear. However, evidence is mounting that GBP is
probably an important component of this activity. Dominguez and Green
demonstrated that translocation of the dorsalizing activity leads to a
decrease in GSK3 protein levels in the dorsal shear zone
(Dominguez and Green, 2000
),
and showed that overexpressed GBP, unlike other candidate dorsalizing
proteins, was able to reproduce this activity in the ventral shear zone. Our
observations that GBP is capable of binding a microtubule motor and undergoing
directed translocation during cortical rotation further support the role of
GBP as a component of the translocating dorsalizing activity. However, work by
Marikawa and Elinson suggests that the dorsalizing activity is not composed
solely of GBP (Marikawa and Elinson,
1999
). Before cortical rotation, the region of the shear zone
where the dorsal determinants reside is called the vegetal cortical cytoplasm,
or VCC. Marikawa and Elinson addressed the nature of the dorsal determinants
by investigating the ability of the VCC to activate dorsal markers when
transplanted into blastomeres overexpressing various ventralizing molecules.
Their results suggested that neither Dsh nor GBP alone is the endogenous
dorsal determinant in the VCC. Salic et al.
(Salic et al., 2000
)
subsequently demonstrated that GBP and Dsh, when expressed together,
powerfully synergize to stabilize ß-catenin levels (see also
Li et al., 1999
;
Hino et al., 2003
). It
therefore seems probable that neither GBP nor Dsh act alone as the endogenous
dorsalizing activity, but rather act in combination with each other and
perhaps other ß-catenin stabilizing proteins as well. It will therefore
be of great importance to determine which other proteins are present in the
particles formed by GBP-GFP and XKLC4-GFP. In particular, although we do not
yet know whether Dsh joins these particles, the fact that Dsh binds GBP
(Li et al., 1999
;
Salic et al., 2000
) and
exhibits directed translocation (Miller et
al., 1999
) suggests that it is likely to join KLC and GBP in a
multimeric translocating complex. Future studies will investigate this
issue.
A revised model for translocation of dorsal determinants
The results of this study contribute to a growing body of work that aims to
describe the molecular components of the dorsalizing activity in
Xenopus and the mechanism by which they travel to the dorsal region
of the embryo in order to stabilize ß-catenin. We propose a model in
which a complex of proteins including KLC, GBP and Dsh is assembled during
oogenesis at the vegetal pole (Fig.
8A). Upon fertilization, the sperm aster introduces a minus-end
microtubule organizing center into the egg, initiating microtubule assembly.
Kinesin-like proteins embedded in the cortex begin to move on the
microtubules, thus helping to align the forming microtubule array from the
point of sperm entry (minus end) toward the future dorsal side (plus end),
analogous to a comb moving through hair. As the array lines up, the
GBP/KLC/Dsh particles move toward the plus end of the microtubules using a
conventional kinesin motor (Fig.
8B). When the particles approach the equator of the embryo, Dsh
binds to Axin and recruits GBP to the ß-catenin destruction complex,
causing GBP to dissociate from KLC in favor of binding to GSK3. Because GBP
competes with Axin for GSK3 binding (Farr
et al., 2000), this removes GSK3 from the Axin/APC/ß-catenin
complex, thus preventing the phosphorylation and degradation of ß-catenin
on the dorsal side of the embryo (Fig.
8C). Locally stabilized ß-catenin then activates the
expression of dorsal genes at the onset of zygotic transcription.
|
It will be interesting to determine the relevance of the interaction
between GBP and KLC in other biological contexts. The interaction between
GBP/Frat and KLC was observed in both Xenopus and mammalian cells,
and the high degree of conservation within their mutual binding domains
suggests that the interaction will probably occur in other vertebrate species
as well. Although cortical rotation appears to be a process specific to
amphibians, the translocation of signaling molecules by molecular motors is a
very general phenomenon. Fertilization in zebrafish causes the formation of a
parallel array of microtubules that is required for the translocation of
dorsal determinants from the vegetal region of the yolk into the embryonic
blastomeres, resulting in the stabilization of ß-catenin
(Jesuthasan and Stahle, 1997).
Kinesin light chains have been shown to bind the scaffolding proteins Sunday
driver/JIP3, JIP1 and JIP2 as cargo (Bowman
et al., 2000
; Verhey et al.,
2001
), which in mammalian neurons allows the kinesin-mediated
transport of entire JNK signaling cassettes from the cell body to neurite
tips. An interesting possibility is that components of the Wnt pathway may
also be able to form multimeric signaling complexes that are transported to
different subcellular locations in various cell types by virtue of the
interaction between GBP and kinesin.
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ACKNOWLEDGMENTS |
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![]() |
Footnotes |
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* These authors contributed equally to this work
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