COMMUNICATION
Interaction of Heterotrimeric G Protein G
o
with Purkinje Cell Protein-2
EVIDENCE FOR A NOVEL NUCLEOTIDE EXCHANGE FACTOR*
Yuan
Luo
and
Bradley M.
Denker§
From the Renal Division, Department of Medicine, Brigham and
Women's Hospital and Harvard Medical School,
Boston, Massachusetts 02115
 |
ABSTRACT |
The heterotrimeric G protein
G
o is ubiquitously expressed throughout the
central nervous system, but many of its functions remain to be defined.
To search for novel proteins that interact with G
o, a
mouse brain library was screened using the yeast two-hybrid interaction
system. Pcp2 (Purkinje cell
protein-2) was identified as a partner for
G
o in this system. Pcp2 is expressed in cerebellar Purkinje cells and retinal bipolar neurons, two locations where G
o is also expressed. Pcp2 was first identified as a
candidate gene to explain Purkinje cell degeneration in pcd
mice (Nordquist, D. T., Kozak, C. A., and Orr, H. T. (1988) J. Neurosci. 8, 4780-4789), but its function
remains unknown as Pcp2 knockout mice are normal (Mohn, A. R.,
Feddersen, R. M., Nguyen, M. S., and Koller, B. H. (1997) Mol. Cell. Neurosci. 9, 63-76). G
o
and Pcp2 binding was confirmed in vitro using glutathione
S-transferase-Pcp2 fusion proteins and in vitro
translated [35S]methionine-labeled G
o. In
addition, when G
o and Pcp2 were cotransfected into COS
cells, G
o was detected in immunoprecipitates of Pcp2. To
determine whether Pcp2 could modulate G
o function, kinetic constants kcat and
koff of bovine brain G
o were
determined in the presence and absence of Pcp2. Pcp2 stimulates GDP
release from G
o more than 5-fold without affecting
kcat. These findings define a novel nucleotide
exchange function for Pcp2 and suggest that the interaction between
Pcp2 and G
o is important to Purkinje cell function.
 |
INTRODUCTION |
Cell surface receptors coupled to G
proteins1 enable cells to
respond to a wide range of extracellular signals. G proteins, composed
of G
and G
subunits, associate at the plasma membrane in a
complex with a serpentine transmembrane receptor. Agonist-liganded receptors activate G
by inducing a change in conformation that leads
to GDP release and GTP-binding. GTP-liganded G
dissociates from
G
, and both subunits can interact with a variety of intracellular effectors. G
and G
remain activated until the intrinsic GTPase activity of G
hydrolyzes GTP to GDP. The mechanisms utilized by
cells to respond to specific signals in a precise manner is not well
understood. In reconstituted systems there is ample evidence that
multiple G proteins can couple to the same sets of receptors and
effectors (reviewed in Ref. 1), and in transfected cells a single G
subunit can couple to at least three different effector pathways (2).
One important mechanism that contributes to regulation of signaling
pathways is the existence of proteins that modulate particular points
in the pathway. For example, G protein receptor kinases desensitize
receptors (such as
-adrenergic receptor kinase, (reviewed in Ref. 3)
and RGS (regulators of G protein
signaling) proteins turn off effector responses by
accelerating the GTPase activity of G
subunits (reviewed in Ref. 4).
Since some G protein family members are predominantly expressed in
specific tissues, cell type-specific modulators of G protein signaling are likely to exist.
G
o is a member of the pertussis toxin family of G
subunits and is predominantly expressed in the central nervous system and heart. Although G
o comprises 0.2-0.5% of brain
particulate protein (5), many of its functions are yet to be defined.
G
o couples to several well characterized receptors in
the brain and can regulate both N-type Ca+2 channels as
well as some K+ channels (see Ref. 6 and references
therein). In addition, G
o can be regulated by
neuromodulin (GAP43 (growth cone-associated protein)) in developing
neurites (7). Knockout of neuromodulin in mice causes significant
abnormalities in neuronal pathfinding (8), but G
o
knockout mice have anatomically normal brains. Despite the normal
central nervous system anatomy in the G
o knockout mice,
they develop a spectrum of neurologic abnormalities (tremor and
impairments of motor control and behavior) and shortened survival (6,
9). We utilized the yeast two-hybrid interaction system to search for
unique modulators of G
o that are expressed in the central nervous system. An interaction between G
o and
Pcp2 (Purkinje cell
protein-2), a protein of unknown function
expressed in Purkinje cells and retinal bipolar neurons was identified.
Although G
o and Pcp2 have not yet been definitively
colocalized in cerebellar Purkinje cells, this interaction was
confirmed in vitro and by coimmunoprecipitation from
transfected cells. Furthermore, Pcp2 can function as a nucleotide
exchange factor by stimulating GDP release from G
o.
 |
EXPERIMENTAL PROCEDURES |
Yeast Two-hybrid Screening--
G
o cDNA in
pBS (previously described in Ref. 10) was cloned into
EcoRI/SalI sites of PAS2-1
(CLONTECH), a vector that encodes GAL4 DNA-binding
domain. G
o-PAS2-1 was used as a "bait" to screen a
mouse brain library in pACT (CLONTECH). Both
G
o-PAS2-1 and the mouse brain cDNA libraries were
cotransformed into Y190, a yeast lacZ/HIS3 reporter strain,
using standard methods. The transformed mix was screened for growth on
plates containing selective medium (synthetic complete medium lacking
tryptophan, leucine, and histidine in the presence of 25 mM
3-aminotriazole) and incubated at 30 °C for 8-12 days.
His+ colonies were screened for
-galactosidase activity
using a filter lift assay, and positive "blue" colonies
(
-galactosidase-positive) were further confirmed by a yeast mating
assay. Individual plasmids were transformed into Escherichia
coli by electroporation and plasmids analyzed by restriction
analysis and dideoxynucleotide sequencing. GenBankTM data
bases were screened with each sequence using BLAST analysis.
In Vitro Binding Assay--
Pcp2 was excised from pACT2 using
EcoRI/XhoI and cloned into pGEX4-2 (Amersham
Pharmacia Biotech). The resulting Pcp2-pgex cDNA and pGEX4-2
without insert (GST alone) were transformed into E. coli,
and the expressed proteins were purified from bacterial pellets after
induction with isopropyl-1-thio-
-D-galactopyranoside. Bacterial lysates were incubated with glutathione-agarose beads (2 ml)
and rotated at 4 °C for 30 min. After centrifugation, GST-Pcp2 and
GST alone were eluted from beads using glutathione elution buffer for
10 min at room temperature (Amersham Pharmacia Biotech). Samples were
recentrifuged, and the resulting supernatant containing the fusion
proteins was analyzed according to the Bradford method to determine
protein concentration. G
o (subtype 1) and
G
i2 in pBS (G
i2 plasmid construction
described in Ref. 11) and G
s in pcDNAI (from ATCC,
Manassas, VA) were used for in vitro translation. Labeled
G
subunits were made using 1 µg of cDNA, appropriate RNA
polymerase in a coupled rabbit reticulocyte translation system (TNT
system, Promega, Madison WI) plus [35S]methionine (NEN
Life Science Products; 20 µCi/reaction) as described previously (11).
[35S]Methionine-labeled G
subunits were analyzed by
SDS-PAGE and autoradiography, and the amounts were normalized by
densitometric analysis of translated products (NIH Image 1.61/fat,
Wayne Rasband, NIH, Bethesda, MD). Equivalent amounts of fusion
proteins (GST or GST- Pcp2) were incubated with glutathione-agarose
beads for 30 min at room temperature, followed by incubation with
equivalent amounts of in vitro translated
[35S]methionine-labeled G
o,
G
s, or G
i2 in PBS with 0.05% Triton X-100 overnight at 4 °C with rocking. Beads were centrifuged, washed
with PBS with 0.05% Triton X-100 three times, eluted with SDS sample
buffer, and analyzed by SDS-PAGE and autoradiography.
Coimmunoprecipitation from Transiently Transfected COS
Cells--
To preserve the HA epitope located at the N terminus of
Pcp2 in pACT2, the plasmid was cut with BglII and filled
with Klenow to generate a blunt end. Following XhoI digest,
the fragment was cloned into the EcoRV/XhoI sites
of pcDNA3 (Invitrogen). G
o in pcDNA3 (12) was
transfected into COS-7 cells alone or in combination with Pcp2 using
LipofectAMINETM (Life Technologies, Inc.) according to the
manufacturer's protocol. At 72 h after the transfection, cells
were washed with ice-cold PBS and lysed for 30 min in lysis buffer (50 mM Hepes, pH 7.5, 6 mM MgCl2, 1 mM EDTA, 75 mM sucrose, 2.5 mM
benzamidine, 1 mM dithiothreitol, and 1% Triton X-100).
Lysates were cleared by low speed centrifugation, and the supernatant
was incubated with the HA-specific monoclonal antibody 12CA5 (1:100)
overnight at 4 °C. Protein A-Sepharose (Sigma) was added for 1 h, and the samples were rocked at 4 °C. Samples were then
centrifuged, and the pellets were washed three times with PBS with
0.05% Triton X-100. The precipitated proteins were eluted with SDS
sample buffer and analyzed by SDS-PAGE and Western blotting using a
rabbit polyclonal anti-G
o antibody (5), and bands were
visualized by chemiluminescence (Pierce).
Determination of kcat and
koff--
Bovine brain G
o was kindly
provided by E. Neer (Harvard Medical School) and used for kinetic
analysis in the presence and absence of Pcp2. Pcp2 was prepared from
GST fusion proteins by cleavage with thrombin at 2.5 units/mg protein
for 1 h at room temperature and separated from GST by incubation
with glutathione-agarose beads. Samples were concentrated and protein
concentration determined (Bradford). Single turnover GTP hydrolysis
(kcat) was determined by incubating
G
o (50 nM) with 1 µM
[
-32P]GTP (5500 cpm/pmol) for 20 min in the presence
(1:1 molar ratio) or absence of Pcp2 in buffer A (50 mM
Tris, pH 7.6, 5 mM EDTA, 1 mM dithiothreitol,
and 0.1% Triton X-100). The hydrolysis reaction was started by
addition of MgCl2 (final concentration, 10 mM) and 100 µM GTP. Aliquots were diluted into 1 ml of 5%
(w/v) trichloracetic acid in 5% charcoal and counted as described
previously (13). The amount of
-32P released at each
time point was fit to an exponential function using GraphPad Prism. For
determining koff, G
o (2 pmol) in
the presence and absence of Pcp2 or GST (1:1 ratio with
G
o) was incubated in buffer A with 10 mM
MgCl2, 1 µM GTP [
-32P]GTP
(specific activity 1.1 × 105 cpm/pmol), and 10 µg/ml bovine serum albumin for 20 min at room temperature. The
stoichiometry of binding was approximately 60%, and the amount bound
prior to initiating nucleotide exchange was set at 100%. GDP
dissociation from G
o was initiated by the addition of 1 mM GDP, and aliquots were filtered onto nitrocellulose,
washed, and counted. The data were fit to a one phase exponential decay using GraphPad Prism (San Diego, CA).
 |
RESULTS AND DISCUSSION |
The yeast two-hybrid interaction system detects low affinity
protein interactions and has been successfully used for finding novel
proteins that interact with G protein
subunits (14). The initial
screen yielded 34
-galactosidase-positive clones, and this number
was reduced to six positive clones after yeast mating controls. These
six clones were sequenced and searched in GenBankTM (BLAST)
data bases. Three of these cDNAs had no sequence homology to known
genes, and one was identified as lactate dehydrogenase. One of the
remaining genes was a previously identified human mosaic protein, LGN,
that was discovered in a yeast two-hybrid screen using
G
i2 as bait (15). The other remaining gene was a
full-length clone of PCD5 (now called Pcp2) (16) that had been
initially identified as a candidate gene important for Purkinje cell
development. In pcd mice, Purkinje cells develop normally
but then begin to degenerate at 15-18 days after birth leading to the
development of ataxia (17).
To determine whether G
o could interact with Pcp2 in an
independent assay, GST pull-down experiments were performed. G
subunits were translated and [35S]methionine-labeled
in vitro and then incubated with equivalent amounts of GST
or GST-Pcp2 fusion proteins. Fig. 1 shows
that in vitro translated G
o binds to GST-Pcp2
(lane 2), whereas no significant interaction was seen with
the GST control protein (black arrow, lane 1). We
consistently detected approximately 10% of in vitro
translated G
o coprecipitating with GST-Pcp2. Nonspecific
interactions of G
o with GST were less than 1%. We next
asked whether the conformation of G
o affected the
interaction with GST-Pcp2. G
o was preincubated with
GTP
S (nonhydrolyzable GTP analogue) or GDP prior to binding. As
shown in Fig. 1, the amount of G
o that associates with
Pcp2 is similar irrespective of the nucleotide bound to
G
o. To address the issue of which G
subunits could
interact with Pcp2, two other G
subunits expressed in the central
nervous system were studied. G
i2 (pertussis toxin family
member; ~70% amino acid identity to G
o) and
G
s (cholera toxin family member; ~40% amino acid
identity to G
o) were characterized in pull-down
experiments with GST-Pcp2. Equal amounts of 35S-labeled
G
i2 and G
s were translated in
vitro and used for binding to GST proteins, and the amount of
nonspecific binding between G
i2 or G
s
with GST was similar to that seen with G
o (not shown). Fig. 1 shows that in vitro translated G
i2
interacts with GST-Pcp2, but G
s is barely detectable in
GST-Pcp2 precipitates (lane 6, 52 kDa, open
arrow). Taken together, these results suggest that the related
pertussis toxin family members (G
o and
G
i2) interact in vitro with Pcp2 but that
G
s subunits do not.

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Fig. 1.
Interaction of in vitro
translated G subunits with GST fusion
proteins. Equivalent amounts of GST or the fusion protein GST-Pcp2
were incubated with glutathione-agarose for 30 min and then overnight
with equal amounts of [35S]methionine-labeled
G o, G i2, or G s translated
in vitro. Samples were washed and eluted with SDS sample
buffer and analyzed by SDS-PAGE followed by autoradiography.
Nonspecific binding of in vitro translated G o
with GST is shown in lane 1 and was similar for
G i2 and G s (not shown). A representative
interaction of G o with GST-Pcp2 is shown in lane
2 and was consistently seen in four independent experiments
performed in duplicate. Preincubating G o with GTP S
(100 µM, lane 3) or GDP (100 µM,
lane 4) for 10 min at 30 °C did not affect binding. The
binding of GST-Pcp2 to G i2 is seen in lane 5,
and there was minimal interaction of GST-Pcp2 with G s
(lane 6). The closed arrow is the size of
G o (39 kDa), and the open arrow is the
expected size of G s (52 kDa). The exposure time was
16 h.
|
|
To look for evidence that G
o and Pcp2 could interact in
cells, cotransfection studies were performed in COS cells. COS cells were transfected with G
o, Pcp2, empty expression vector
pcDNA3 (PC) alone or in combination. Cell lysates were
immunoprecipitated using the 12CA5 antibody directed toward the
hemagglutinin epitope on the N terminus of Pcp2 and then analyzed by
Western with anti-G
o antibody (Fig.
2). COS cells do not normally express
G
o, and in cells transfected with vector (PC), Pcp2, or
G
o and then immunoprecipitated with 12CA5 there is no
detectable G
o immunoreactivity (Fig. 2, arrow). However, when both G
o and Pcp2 are
cotransfected into the same cells, a fraction of G
o is
found in association with Pcp2 (Fig. 2, lane 4). We were
unable to detect Pcp2 in immunoprecipitates of G
o,
presumably due to disruption of the association with G
o by the stringent detergent conditions necessary to immunoprecipitate G
o (18). Attempts to detect G
i2 in Pcp2
immunoprecipitates were unsuccessful, although the level of
G
i2 expression was much lower than for
G
o. The observation that G
o and Pcp2
interact in an intact cell raises the possibility that Pcp2 could be a modulator of G
o.

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Fig. 2.
Coimmunoprecipitation of
G o and PCP2 from transfected COS
cells. COS cells were transfected with pcDNA 3 (PC), G o, Pcp2, or G o plus
Pcp2 and immunoprecipitated (IP) with HA-specific monoclonal
antibody 12CA5 directed toward the N terminus of Pcp2. 72 h after
the transfection, cellular lysates were immunoprecipitated and eluted
with SDS sample buffer followed by SDS-PAGE and Western blotting with
anti-G o antibody. The migration of G o in
cell lysates of COS cells transfected with G o is shown
in lane 1. The arrow denotes the size of
G o at 39 kDa. Lanes 1-3 are from a different
gel (and divided by white line) than lanes 4-6
but represent a single experiment. Similar results were seen in three
independent experiments. Immunoprecipitation of Pcp2 was confirmed in
independent experiments (not shown). In cells cotransfected with both
G o and Pcp2, a fraction of G o can be
detected in association with precipitated Pcp2 (lane 4,
arrow). In cells transfected with vector alone
(PC), G o alone, or Pcp2 alone (lanes
2, 3, 5, and 6) and analyzed in
parallel with G o + Pcp2, no G o (39 kDa,
arrow) is detectable.
|
|
There are several possible mechanisms for regulation of G
subunits
by accessory proteins, including effects on nucleotide binding or GTP
hydrolysis. To address the possibility that Pcp2 modulates the
enzymatic properties of G
o, we measured
kcat and koff of bovine
brain purified G
o (5) in the presence and absence of
Pcp2. Fig. 3 shows a representative
experiment, and the results are summarized in Table
I. The kcat and
koff values obtained for G
o are
similar to literature values (19), and there was no significant
difference in kcat of G
o in
presence of Pcp2 (Fig. 3a and Table I). However, as seen in
Fig. 3b and Table I, there is significant stimulation of
koff in the presence of Pcp2. In experiments
simultaneously comparing koff of
G
o in the presence and absence of Pcp2 or GST, there was
5.2 ± 0.5-fold stimulation of koff in the
presence of Pcp2 (n = 7). Incubating G
o
with GST had no effect on koff (Fig.
3b and Table I), and including bovine serum albumin in the
binding buffer also had no effect. The increase in
koff of G
o in the presence of
Pcp2 is similar to the observed increases seen with G
subunits
reconstituted with receptors plus agonists (4-6-fold increases in
koff) (20). This finding suggests that Pcp2
could mimic a receptor by stimulating GDP release. A family of proteins
that promote nucleotide exchange have been described for monomeric G
proteins such as Ras (21), but only neuromodulin has been shown to
affect nucleotide binding to G
o. This mechanism of
activation is likely to be distinct from Pcp2 because neuromodulin
stimulates GTP
S binding through its N-terminal domain, which is
homologous to the cytoplasmic tail of G protein-coupled receptors
(7).

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Fig. 3.
Determination of
kcat and koff
for G o in the presence and absence
of Pcp2. a, single turnover GTPase activity for
G o was determined in the presence ( ) and absence
( ) of PCP2 (1:1 molar ratio). The fraction of GTP hydrolyzed was
measured for bovine brain purified G o in the presence of
[ -32P]GTP, and kcat was
determined by fitting the data to a single exponential association
function (GraphPad Prism). The results are summarized in Table I.
b, the rate constant for GDP release
(koff) from G o was determined
alone ( ) or in the presence of PCP2 ( ) or an equivalent amount of
GST (×). The percentage of GDP remaining bound to G o
was determined by equilibrating 1 µM
[ -32P]GTP for 20 min and initiating nucleotide
exchange with 1 mM GDP. The fraction of GDP bound to
G o was measured over time. The data were fit to a single
exponential dissociation, and the results are summarized in Table I.
The fold increase in koff for this experiment
was 5.9 (the mean of all experiments (n = 7) was
5.2 ± 0.5).
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Table I
Summary of G o rate constants
Results are expressed as the means of individual experiments ± S.E. (n) where n = the number of
experiments. There is no significant difference in any values except in
koff for G o + Pcp2 versus
G o alone or G o + GST (p < 0.006).
|
|
The gene for Pcp2 is located on mouse chromosome 8 and encodes a
cytosolic 99-amino acid protein without significant homology to other
proteins (including G protein-coupled receptors). There is some amino
acid sequence similarity to the c-sis/PDGF2 gene, but the
implications of this are unknown (22). The expression profile of Pcp2
is consistent with a role in Purkinje cell development (23), but in
mice without Pcp2 expression, Purkinje cells develop normally
(presumably through other compensatory mechanisms) (24, 25). The
function of Pcp2 and the mechanism(s) for increased Purkinje cell
apoptosis in pcd mice remains unknown. Several genes in
pcd mice have altered expression patterns including
up-regulation of the early response genes fos and
jun and down-regulation of the anti-apoptosis gene
bcl-2 (26). Several G protein
subunits trigger apoptosis
(27), and G
o may be involved in apoptosis triggered by a
mutated amyloid precursor protein in Alzheimer's disease (28). The
results described in these studies suggest that Pcp2 may be involved in
a signal transduction pathway involving G
o, but it
remains to be determined whether this pathway relates to apoptosis. In
addition, our results do not exclude important interactions of Pcp2
with other G
subunits, particularly G
i1, which is
highly expressed in the brain. Increasingly, novel functions (and
locations) for G proteins are being identified (such as in the Golgi,
in the endoplasmic reticulum, and on intracellular vesicles) where they
regulate protein processing and vesicular targeting (29-31). Because G
protein-coupled transmembrane receptors have not been identified in
many of these locations, G
subunits may be activated by nucleotide
exchange factors. Pcp2 may be such a factor for G
o in
cerebellar Purkinje cells, and future studies will define the
functional consequences of this interaction.
 |
FOOTNOTES |
*
This work was supported by the National Institutes of Health
and by a pilot award from the Harvard Digestive Disease Center (to
B. M. D.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Present address: Dept. of Biological Sciences, University of
Southern Mississippi, Hattiesburg, MS 39406.
§
To whom correspondence should be addressed: Harvard Inst. of
Medicine, 77 Ave. Louis Pasteur, Boston, MA 02115. Tel.: 617-525-5809; Fax: 617-525-5830; E-mail: bdenker{at}rics.bwh.harvard.edu.
 |
ABBREVIATIONS |
The abbreviations used are:
G proteins, guanine
nucleotide-binding proteins;
GTP
S, guanosine
5'-(
-thio)triphosphate;
PAGE, polyacrylamide gel electrophoresis;
GST, glutathione S-transferase;
PBS, phosphate-buffered
saline;
HA, hemagglutinin.
 |
REFERENCES |
-
Neer, E. J.
(1995)
Cell
80,
249-257[Medline]
[Order article via Infotrieve]
-
Hunt, T. W.,
Carroll, R. C.,
and Peralta, E. G.
(1994)
J. Biol. Chem.
269,
29565- 29570[Abstract/Free Full Text]
-
Lefkowitz, R. J.
(1998)
J. Biol. Chem.
273,
18677-18680[Free Full Text]
-
Berman, D. M.,
and Gilman, A. G.
(1998)
J. Biol. Chem.
273,
1269-1272[Free Full Text]
-
Huff, R. M.,
Axton, J. M.,
and Neer, E. J.
(1985)
J. Biol. Chem.
260,
10864-10871[Abstract/Free Full Text]
-
Valenzuela, D.,
Han, X.,
Mende, U.,
Fankhauser, C.,
Mashimo, H.,
Huang, P.,
Pfeffer, J.,
Neer, E. J.,
and Fishman, M. C.
(1997)
Proc. Natl. Acad. Sci. U. S. A.
94,
1727-1732[Abstract/Free Full Text]
-
Strittmatter, S. M.,
Valenzuela, D.,
Kennedy, T. E.,
Neer, E. J.,
and Fishman, M. C.
(1990)
Nature
344,
836-841[CrossRef][Medline]
[Order article via Infotrieve]
-
Strittmatter, S. M.,
Fankhauser, C.,
Huang, P. L.,
Mashimo, H.,
and Fishman, M. C.
(1995)
Cell
80,
445-452[Medline]
[Order article via Infotrieve]
-
Jiang, M.,
Gold, M. S.,
Boulay, G.,
Spicher, K.,
Peyton, M.,
Brabet, P.,
Srinivasan, Y.,
Rudolph, U.,
Ellison, G.,
and Birnbaumer, L.
(1998)
Proc. Natl. Acad. Sci. U. S. A.
95,
3269-3274[Abstract/Free Full Text]
-
Denker, B. M.,
Neer, E. J.,
and Schmidt, C. J.
(1992)
J. Biol. Chem.
267,
6272-6277[Abstract/Free Full Text]
-
Denker, B. M.,
Boutin, P. M.,
and Neer, E. J.
(1995)
Biochemistry
34,
5544-5553[Medline]
[Order article via Infotrieve]
-
Denker, B. M.,
Saha, C.,
Khawaja, S.,
and Nigam, S. J.
(1996)
J. Biol. Chem.
271,
25750-25753[Abstract/Free Full Text]
-
Denker, B. M.,
Tempst, P.,
and Neer, E. J.
(1991)
Biochem. J.
278,
341-345[Medline]
[Order article via Infotrieve]
-
De Vries, L.,
Mousli, M.,
Wurmser, A.,
and Farquhar, M. G.
(1995)
Proc. Natl. Acad. Sci. U. S. A.
92,
11916-11920[Abstract]
-
Mochizuki, N.,
Cho, G.,
Wen, B.,
and Insel, P. A.
(1996)
Gene (Amst.)
181,
39-43[CrossRef][Medline]
[Order article via Infotrieve]
-
Nordquist, D. T.,
Kozak, C. A.,
and Orr, H. T.
(1988)
J. Neurosci.
8,
4780-4789[Abstract]
-
Mullen, R. J.,
Eicher, E. M.,
and Sidman, R. L.
(1976)
Proc. Natl. Acad. Sci. U. S. A.
73,
208-212[Abstract]
-
Busconi, L,
and Denker, B. M.
(1997)
Biochem. J.
328,
23-31[Medline]
[Order article via Infotrieve]
-
Fields, T. A,
and Casey, P. J.
(1997)
Biochem. J.
321,
561-571[Medline]
[Order article via Infotrieve]
-
Senogles, S. E.,
Spiegel, A. M.,
Padrell, E.,
Iyengar, R. I.,
and Caron, M. G.
(1990)
J. Biol. Chem.
265,
4507-4514[Abstract/Free Full Text]
-
Shou, C.,
Farnsworth, C. L.,
Neel, B. G.,
and Feig, L. A.
(1992)
Nature
358,
351-354[CrossRef][Medline]
[Order article via Infotrieve]
-
Oberdick, J.,
Levinthal, F.,
and Levinthal, C.
(1988)
Neuron
1,
367-376[Medline]
[Order article via Infotrieve]
-
Vandaele, S.,
Nordquist, D. T.,
Feddersen, R. M.,
Tretjakoff, I.,
Peterson, A. C.,
and Orr, H. T.
(1991)
Genes Dev.
5,
1136-1148[Abstract]
-
Mohn, A. R.,
Feddersen, R. M.,
Nguyen, M. S.,
and Koller, B. H.
(1997)
Mol. Cell. Neurosci.
9,
63-76[CrossRef][Medline]
[Order article via Infotrieve]
-
Vassileva, G.,
Smeyne, R. J.,
and Morgan, J. I.
(1997)
Mol. Brain Res.
46,
333-337[CrossRef][Medline]
[Order article via Infotrieve]
-
Gillardon, F.,
Baurle, J.,
Wickert, H.,
Grusser-Cornehls, U.,
and Zimmermann, M.
(1995)
J. Neurosci. Res.
41,
708-715[Medline]
[Order article via Infotrieve]
-
Althoefer, H.,
Eversole-Cire, P.,
and Simon, M. I.
(1997)
J. Biol. Chem.
272,
24380-24386[Abstract/Free Full Text]
-
Yamatsuji, T.,
Matsui, T.,
Okamoto, T.,
Komatsuzaki, K.,
Takeda, S.,
Fukumoto, H.,
Iwatsubo, T.,
Suzuki, N.,
Asami-Odaka, A.,
Ireland, S.,
Kinane, T. B.,
Giambarella, U.,
and Nishimoto, I.
(1996)
Science
272,
1349-1352[Abstract]
-
Denker, S. P.,
McCaffery, J. M.,
Palade, G. E.,
Insel, P. A.,
and Farquhar, M. G.
(1996)
J. Cell Biol.
133,
1027-1040[Abstract]
-
Stow, J. L.,
de Almeida, J. B.,
Narula, N.,
Holtzman, E.,
Ercolani, L.,
and Ausiello, D. A.
(1991)
J. Cell Biol.
114,
1113-1124[Abstract]
-
Pimplikar, S. W,
and Simons, K.
(1993)
Nature
362,
456-458[CrossRef][Medline]
[Order article via Infotrieve]
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