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(Received for publication, July 12,
1995; and in revised form, September 12, 1995) From the
A 120-kDa protein that is tyrosine-phosphorylated upon antigen
receptor ligation in B lymphocytes has been identified as the product
of the c-cbl protooncogene. Tyrosine phosphorylation of Cbl
depends on the efficient association of membrane immunoglobulin heavy
chains with the Ig
The antigen receptor on B lymphocytes is made up of membrane
immunoglobulins associated with the Ig Following B cell receptor ligation, cytoplasmic tyrosines in the
ITAMs (immunoreceptor tyrosine-associated motifs) of Ig An alternative pathway by
which antigen-selected B lymphocytes may be induced to proliferate is
mediated by T lymphocyte-derived cytokines and the triggering of CD40
on B cells by its ligand on activated T
cells(24, 25, 26) . Cross-linking of CD40 can
lead to the activation of tyrosine and serine/threonine kinases, the
activation of PI 3-kinase, the activation of phospholipase C Cbl is the cellular
homolog of the v-Cbl oncoprotein (27, 28) and is a
predominantly cytosolic protein, which contains 17 proline-rich motifs
potentially capable of binding a range of SH3 domains. Tyrosine
phosphorylation of Cbl has been described in response to receptor
occupancy of a number of receptors including the antigen receptor on T
cells, the erythropoietin receptor, the granulocyte/macrophage
colony-stimulating factor receptor, the Fc We
describe here the identification of Cbl as a prominent substrate for
tyrosine phosphorylation in antigen-receptor-ligated B lymphocytes.
This phosphorylation event requires the efficient interaction of
membrane immunoglobulin heavy chains with the Ig
Immunoprecipitations,
anti-phosphotyrosine immunoblots, and in vitro kinase assays
were performed as described previously(17, 34) .
Figure 1:
Cbl is the prominent
120-kDa protein that is tyrosine-phosphorylated following anti-IgM and
anti-IgG cross-linking. Upper panel, anti-phosphotyrosine
immunoblot. Lower panel, anti-Cbl immunoblot. WEHI 231 (IgM) B
cells and the 4JJ subclone of A20.25 (IgG; A20/4JJ) were either not
stimulated (N) or stimulated (S) with F(ab)2
anti-mouse IgM (WEHI 231)or anti-mouse IgG (A20/4JJ) for 5 min. Lysates
were either not precleared(-) or were precleared with anti-Cbl (Cbl) or with a control rabbit IgG (IgG). Total
lysate was then separated on SDS/PAGE transferred to an Immobilon-P
(Millipore) membrane and reacted sequentially with an
anti-phosphotyrosine monoclonal antibody and a peroxidase conjugated
rabbit anti-mouse antibody. The ECL system (Amersham Corp.) was used
for detection. Positions of molecular weight markers are indicated on
the left of each panel.
In Fig. 2A, Cbl tyrosine phosphorylation is
apparent from the 30-s time point. In a slightly longer exposure,
phosphorylation was apparent from the 10-s time point.
Tyrosine-phosphorylated proteins with a slightly slower mobility than
the major Cbl band probably represent Cbl species phosphorylated on
multiple sites. These bands were depleted by anti-Cbl as seen in Fig. 1and Fig. 2A. Similar bands were observed
on anti-Cbl immunoblots of lysates from activated B cells (data not
shown). The tyrosine-phosphorylated 70-kDa species seen both in Fig. 1and 2A, exactly comigrates with, and probably
represents, tyrosine-phosphorylated Syk. Phosphorylation of Cbl is
observed soon after the cross-linking of membrane immunoglobulins but
is not observed after cross-linking CD40 (Fig. 2B),
although both stimuli can independently contribute to the initiation of
similar signaling pathways.
Figure 2:
A, time course of tyrosine
phosphorylation following receptor ligation of BAL 17 B cells. Lysates
were prepared 10 s (10``), 30 s (30''), 1
min (1`), 3 min (3`), 5 min (5`), and 15 min (15`) after activation. Additional lysates at the 5-min time
point were precleared either with anti-Cbl or with preimmune IgG.
Details as in legend to Fig. 1. B, Cbl is
tyrosine-phosphorylated after antigen receptor ligation but not
following CD40 cross-linking. BAL 17 B cells were either not stimulated (N) or were stimulated with anti-IgM for 5 min (S1),
anti-CD40 for 30 s (S2), or anti-CD40 for 5 min (S3).
In leftmost four lanes, total lysates were separated on
SDS-PAGE. In rightmost four lanes, anti-Cbl immunoprecipitates
were separated. Proteins were visualized by an anti-phosphotyrosine
immunoblot.
Figure 3:
Cross-linking of a transmembrane IgM
mutant that associates poorly with the Ig
Figure 4:
Cbl is efficiently tyrosine-phosphorylated
following receptor ligation in Xid splenic B cells. Splenic B cells
were purified from CBA/CaHN-xid/J mice and were either not stimulated (N) or cross-linked with anti-IgM (S). Total lysates
and anti-Cbl immunoprecipitates were separated on SDS/PAGE and
tyrosine-phosphorylated proteins revealed by an immunoblot
assay.
We wished to ascertain whether the
formation of a complex between Shc, Grb2, and Cbl could be demonstrated
in activated B lymphocytes. We were able to confirm, as described
previously(41) , the formation of Shc-Grb2 complexes after B
cell activation (Fig. 5A) and also the association of
fusion proteins containing an intact N-terminal SH3 domain of Grb2 with
Cbl from B cell lysates (Fig. 5B). While these data
suggest that Shc-Grb2-Cbl complexes may well be formed in activated B
cells, in our hands we were able to observe an in vivo association between Grb2 and Cbl only in three murine pre-B cell
lines.
Figure 5:
Grb2
complexes with Shc in activated B cells. BAL 17 cells were either not
stimulated (N) or stimulated with anti-IgM (S) and
immunoprecipitated with anti-Grb2 or anti-Shc. Transferred proteins
were revealed on an immunoblot using anti-Grb2 (upper panel)
or anti-phosphotyrosine (lower panel) antibodies. B,
Cbl binds to the N-terminal SH3 domain of Grb2. Lysates from
non-stimulated (N) and anti-IgG-stimulated (S) A20.25
B cells were bound to a control glutathione S-transferase
affinity matrix (GST), to a wild type GST-Grb2 matrix (G2), or to a mutant GSTGrb2 matrix (K36; Trp
The band that has been labeled as
Shc in the lower panel of Fig. 5A is inferred
to be the phosphorylated form of p52 Shc. Anti-Shc immunoblots detect
the p52 and p46 species of Shc in B cells but fail to reveal p66.
Although the p46 form of Shc is also phosphorylated in activated B
cells, the band representing it is probably obscured in Fig. 5A by rabbit IgG detected by the second antibody.
Figure 6:
An 85-kDa protein is associated with Cbl
in activated B cells. WEHI 231 B cells were either not stimulated (N) or stimulated with anti-IgM (S),
immunoprecipitated with preimmune serum or anti-Cbl, and subjected to
an in vitro kinase assay using
[
Figure 7:
A, the p85 subunit of PI 3-kinase
associates with Cbl in receptor-ligated B cells. BAL 17 B cells were
either not stimulated (N) or cross-linked with anti-mouse IgM (S) and immunoprecipitated with anti-Cbl. The presence of
associated p85 was detected using an immunoblot assay. B,
Anti-Cbl-associated PI 3-kinase activity in receptor-ligated B cells.
BAL 17 B cells were either not stimulated (N) or cross-linked
with anti-IgM (S) and immunoprecipitated with preimmune serum,
anti-Cbl, or anti-phosphotyrosine antibodies. The presence of
associated PI 3-kinase activity was detected by incubation with
micellar PI and [
Since B lymphocyte antigen receptor cross-linking
leads to an increase in PI 3-kinase activity(42, 43) ,
we were keen to establish whether there was an increase in PI 3- kinase
activity associated with Cbl following anti-IgM exposure. As seen in Fig. 7B, a dramatic increase in Cbl-associated
phosphatidylinositol 3-kinase kinase activity was observed in activated
B cell lysates. Three different cell lines representing distinct
stages of differentiation were used in all the studies described above
(other than those examining membrane IgM mutants). Most experiments
were performed with all three cell lines, although in some only two of
the three lines were used. No functional differences between these cell
lines was observed in any of the above studies. Since the tyrosine phosphorylation of Cbl is an early event
downstream of the engagement of a number of mitogenic receptors, we
sought to identify proteins that specifically associate with Cbl after
the initiation of signal transduction. A number of proteins associate
constitutively with Cbl usually via SH3 domain-mediated interactions.
These proteins include Src family kinases, Grb2, and
Nck(29, 31, 44) . We have demonstrated here
that an 85-kDa protein specifically associates with Cbl following B
cell receptor ligation and that this 85-kDa protein is the p85 subunit
of PI 3-kinase. In addition we have also demonstrated a convincing
increase in PI 3-kinase activity associated with Cbl in activated B
cells, suggesting that both subunits of this enzyme associate with Cbl
and implying that Cbl might play a role in the activation of PI
3-kinase. It is quite likely that the major functional role of Cbl in
activated B cells is to serve as a docking/activation site for PI
3-kinase. PI 3-kinase is known to be activated when one or both of
the SH2 domains in the p85 subunit of this enzyme bind to
tyrosine-phosphorylated proteins that contain Y-X-X-M
motifs(45) . The activation of PI-3 kinase in response to
antigen receptor ligation in B cells has been described previously (42, 43) and association of PI-3 kinase with CD19
(which contains a Y-X-X-M motif in its cytoplasmic
tail) has also been demonstrated(46) . Given the ease with
which a Cbl-PI 3-kinase complex can be detected in vivo in
activated B cells, and the dramatic increase in Cbl-associated PI-3
kinase catalytic activity following antigen receptor ligation, we
postulate that the major functional consequence of Cbl tyrosine
phosphorylation is the catalytic activation of PI 3-kinase. The 110-kDa
catalytic subunit of this enzyme possesses both a lipid kinase activity
and a serine/threonine protein kinase activity. The lipid kinase
activity may, via PI intermediates phosphorylated in the 3-position of
inositol, contribute to the activation of the The tyrosine phosphorylation of
Cbl in response to antigen receptor cross-linking requires the
associated Ig
Volume 270,
Number 46,
Issue of November 27, 1995 pp. 27504-27509
©1995 by The American Society for Biochemistry and Molecular Biology, Inc.
in Antigen Receptor-ligated B Cells (*)
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
FOOTNOTES
ACKNOWLEDGEMENTS
REFERENCES
/
heterodimer but is unimpaired in splenic
B cells from the Xid mouse. Cross-linking of membrane IgM and membrane
IgG, but not of CD40, leads to the tyrosine phosphorylation of Cbl. In
receptor-ligated B lymphocytes, p120 associates
with an 85-kDa protein that has been identified as the 85-kDa subunit
of phosphatidylinositol 3-kinase.
/
heterodimer(1, 2) . This heterodimer serves to link
the receptor with associated Src family kinases such as Blk, Lyn, Fyn,
and Fgr(3, 4, 5, 6) , and also with
the Syk tyrosine kinase(7, 8) . Tyrosine
phosphorylation appears to be an obligatory event (9) in the
initiation of signal transduction pathways that promote B lymphocyte
proliferation and differentiation. A number of signaling events
downstream of this receptor depend on the interaction of membrane
immunoglobulin heavy chains with the associated Ig/
heterodimer(10, 11, 12) , while some are
initiated independently of these associated glycoproteins(13) .
Activated protein tyrosine kinases are presumed to participate in
triggering the Ras pathway, in activating PI
3-kinase, and
in initiating the hydrolysis of phosphatidylinositol 4,5-bisphosphate
by specific isoforms of phosphatidylinositol-specific phospholipase C. and
Ig are tyrosine-phosphorylated, presumably by activated Src family
kinases. The subsequent recruitment of Syk to phosphorylated
ITAMs(14, 15) , and possibly an interaction between
Src family kinases and Syk(16, 17, 18) ,
leads to the activation of the latter. Another protein tyrosine kinase
that is activated following receptor ligation and which plays a role in
antigen receptor mediated signaling is
Btk(19, 20, 21) . Splenic B cells in Xid
mice, which carry a point mutation in the PH domain of Btk, appear to
be identical to gene-targeted mice in which no Btk protein is
synthesized(22) . Btk appears to be critical for the entry into
S phase of B lymphocytes that have been stimulated by antigen receptor
cross-linking(22, 23) .
isoforms, and the nuclear translocation of the NF
B transcription
factor. While the molecular consequences of activating B lymphocytes
via the antigen receptor and by CD40-mediated signaling are similar,
the mechanisms by which these individual pathways are activated by
these different ligands may well be distinct. receptor, the
colony-stimulating factor-1 receptor, and the epidermal growth factor
receptor(29, 30, 31, 32) . Tyrosine
phosphorylation of Cbl has also been observed in v-src- and
v-abl-transformed cells (32, 33) , and this
modification may be a critical event in mitogenic signaling.
/
heterodimer, and is unimpaired in splenic B cells from the Xid mouse.
Following receptor ligation, Cbl is seen to associate with an 85-kDa
phosphoprotein, identified as the 85-kDa subunit of PI 3-kinase. Our
studies suggest that a major role of Cbl in B cells may be the
recruitment and activation of PI 3-kinase following antigen receptor
ligation.
Cells
Cell lines used included WEHI 231 (IgM,
immature B), BAL 17 (IgM, mature B), and A20.25 (IgG,
mature B), as well as transfectants of A20.25 described below. The
source for WEHI 231 cells was described earlier(34) . BAL 17
cells were kindly provided by Dr. W. E. Paul. A20.25 transfectants
expressing wild type human IgM and the YS/VV transmembrane human IgM
mutant were kindly provided by Drs. A. Abbas and R.
Mitchell(10) . Splenic B cells from CBA/CaHN-xid/J mice
(Jackson Laboratory, Bar Harbor, ME) were purified as described in (17) .
Antisera
Antibodies to Cbl (Santa Cruz), Shc
(Upstate Biotechnology Inc.), Grb2 (Santa Cruz), phosphotyrosine
(Zymed), and the p85 subunit of phosphatidylinositol 3-kinase (Santa
Cruz) were obtained from commercial sources. Anti-Btk was obtained as
described earlier(17) .Lymphocyte Activation
Anti-IgM and anti-IgG
stimulation of B cells was performed as described earlier(17) .
Anti-CD40 stimulation was performed incubating 2 
10
BAL 17 B cells in 200 µl of serum-free RPMI medium at 37
°C with anti-mouse CD40 at a concentration known to induce
proliferation (2 µg/ml; Serotec).GST-Grb2 Association with Cbl
Glutathione S-transferase fusion proteins including Grb2 and a mutant Grb2
protein in which the N-terminal SH3 domain is non-functional (a
tryptophan to lysine substitution in codon 36) were kindly provided by
Dr. Bruce Mayer. Lysates from non-stimulated and stimulated A20.25 B
cells were made using 1% Nonidet P-40 in 20 mM Tris, pH 7.5,
150 mM NaCl, and 2 mM phenylmethylsulfonyl fluoride.
Binding to immobilized GST fusion proteins, elution, and analysis on
polyacrylamide/SDS gels was performed as described
earlier(34) . Cbl associated with Grb2 was revealed by an
immunoblot assay.Assay for PI 3-Kinase Activity in Anti-Cbl,
Anti-phosphotyrosine, and Anti-p85 Immunoprecipitates
The assay
was performed essentially as described(35) . Briefly anti-Cbl,
anti-PY, and anti-p85 immunoprecipitates were incubated with micellar
phosphatidylinositol (0.2 mg/ml) in 25 mM MOPS, pH 7.0, 5
mM MgCl
, 1 mM EDTA, 150 µM unlabeled ATP, and 25 µCi of
[-
P]ATP in a total reaction volume of 50
µl. After a 20-min incubation at 37 °C, the reaction was
stopped using 100 µl of 1:1 CH
OH:1 N HCl.
Labeled phospholipid was extracted with 200 µl of CHCl
and was dried under N. Thin layer chromatography was
performed overnight with HO:glacial acetic
acid:n-propanol (34:1:65).
Cbl Is Tyrosine-phosphorylated following Membrane IgM
or Membrane IgG Cross-linking of B Cells
Membrane
immunoglobulins were cross-linked on an IgM-expressing B cell line
(WEHI 231) and an IgG-expressing B cell line (the 4JJ subclone of
A20.25, which also expresses a transfected human IgM mutant; (10) ) using anti-mouse IgM and IgG, respectively. Lysates from
stimulated cells contained a 120-kDa tyrosine-phosphorylated protein (Fig. 1, upper panel), which was depleted by
preclearing with anti-Cbl, but not by control antibodies. Depletion of
Cbl was confirmed by an immunoblot assay (lower panel).
Tyrosine phosphorylation of Cbl is seen as early as 10 s after
cross-linking and peaks at about 5 min. Similar results were observed
with the surface IgM-positive BAL 17 line and untransfected A20.25
cells.
Cross-linking of a Transmembrane IgM Mutant Fails to
Initiate the Tyrosine Phosphorylation of Cbl
Mutations in the
the transmembrane domain of membrane IgM have been demonstrated to
impair signal transduction. A Tyr/Ser Val/Val mutation (10) in human membrane IgM expressed in a murine B cell line
(A20.25) has been examined in a number of studies. While cross-linking
of a wild type human IgM transfectant leads to the effective initiation
of intracellular signaling, this mutant human membrane IgM associates
poorly with the murine Ig
/ heterodimer and is compromised in
terms of its ability to initiate a calcium flux (10) or to lead
to the tyrosine phosphorylation of a number of unidentified cellular
proteins (11, 12) . We immunoprecipitated Cbl
following cross-linking of the above A20.25 transfectants either with
anti-mouse IgG or anti-human IgM. As seen in Fig. 3(upper
panel), cross-linking of the YS/VV mutant human IgM does not lead
to the efficient tyrosine phosphorylation of Cbl.
/ heterodimer
compromises the tyrosine phosphorylation of Cbl. A20.25 cells
expressing transfected wild-type human IgM (A20 WT) or a
transmembrane YS/VV mutant (A20/4J) were individually either
not stimulated(-) or cross-linked with anti-human IgM
(µ) or anti-mouse IgG (). Lysates were
immunoprecipitated with anti-Cbl and were separated on SDS-PAGE,
followed by an immunoblot analysis using anti-phosphotyrosine antibody (upper panel). Immunoprecipitated Cbl was quantified (lower panel) by reprobing with
anti-Cbl.
Tyrosine Phosphorylation of Cbl Is Unimpaired upon
Cross-linking of Membrane IgM Expressed by Splenic B Cells from Xid
Mice
The catalytic activation of Src family kinases is known to
occur in the first minute after B cell antigen receptor
cross-linking(20) , while the activation of Syk (20) and Btk peaks a few minutes
later(19, 20) . The dependence of Cbl tyrosine
phosphorylation on the Ig/ heterodimer and the time course of
Cbl phosphorylation suggest that antigen receptor-associated Src family
kinases and Syk probably play a role in this tyrosine phosphorylation
event. Xid mice harbor a point mutation in the PH domain of
Btk(36, 37) . In response to antigen receptor
ligation, splenic B cells from Xid mice appear to initiate signal
transduction in a manner similar to B cells from wild type mice, but
the Xid B cells, in contrast to wild type B cells, fail to enter S
phase following stimulation(23) . While Xid B cells are
defective in terms of T-independent responses, they respond normally to
protein antigens. Splenic B cells from gene-targeted mice, which fail
to synthesize any Btk, cannot be functionally distinguished from Xid B
cells(22) , suggesting that the Xid point mutation completely
disrupts Btk function. Receptor ligation of Xid B cells, however, leads
to the efficient tyrosine phosphorylation of Cbl (Fig. 4),
suggesting that the phosphorylation of Cbl occurs independently of Btk.
Shc-Grb2-Cbl Complexes in Lysates from Activated B
Lymphocytes
Cbl contains multiple proline-rich stretches, which
presumably constitute sites for SH3 interactions, and also contains a
number of tyrosine residues, which, if phosphorylated, could serve as
sites for SH2 binding. While the N-terminal SH3 domain of Grb2 has been
demonstrated to associate with Cbl in lysates from T
lymphocytes(29) , the significance of this association remains
unclear. One possibility could involve the recruitment of Cbl to the
antigen receptor via Shc and Grb2. While it has been suggested that
Shc-Grb2-SOS complexes contribute to the recruitment of SOS to the
membrane in activated T cells(38) , this remains an unresolved
issue. Recruitment of Grb2-SOS to the membrane may depend on its
association in activated T cells with a 36-kDa membrane-associated
phosphoprotein(39) . Complex formation between T cell
receptor-associated ITAMs and Shc may be relatively
inefficient(40) . Although we could detect phosphorylated Shc species
in anti-Cbl immunoprecipitates from lysates of activated B cells (data
not shown), we were unable, in any of the three activated B cell lines
tested, to demonstrate an interaction in vivo between Grb2 and
Cbl. However, Shc-Grb2-Cbl complexes are readily observed in activated
human B cell lines.
Lys mutation in codon 36, which abrogates binding in the N-terminal SH3
domain). Bound proteins were separated in parallel with proteins from
original lysate (Total). Presence of Cbl was monitored by an
immunoblot assay.
In Receptor-ligated B Cells, Cbl Associates with the
85-kDa Subunit of Phosphatidylinositol 3-Kinase
Since Cbl is
tyrosine-phosphorylated in response to receptor ligation, we wished to
ascertain if Cbl is specifically associated with any proteins after
antigen receptor cross-linking. Given the knowledge that Cbl associates
constitutively with Src family kinases via the SH3 domains of the
latter (29, 31) and with Syk by a poorly understood
mechanism(31) , one of the approaches that was used to identify
proteins associated with Cbl was an in vitro kinase assay on
Cbl immunoprecipitates of detergent lysates from non-activated and
activated B cells. As seen in Fig. 6, phosphorylation of Cbl as
revealed by this assay is increased after receptor ligation. In
addition, a prominent 85-kDa phosphoprotein was seen to be associated
specifically with Cbl immunoprecipitated from lysates of activated B
cells, but not from non-activated B cell lysates. Since this protein
was of the predicted size for the 85-kDa subunit of
phosphatidylinositol 3-kinase, we examined lysates of B cells before
and after activation for Cbl-associated PI-3 kinase as well as for Btk.
Lysates were immunoprecipitated from non-activated and activated B
cells with anti-Cbl antibodies and were analyzed for associated PI-3
kinase using antibodies specific for the p85 subunit in an immunoblot
assay. The same filter was also probed with anti-Btk but no significant
association of Btk with Cbl was observed (data not shown). As seen in Fig. 7A, p85 was seen to be associated with Cbl only in
activated B cells.
-
P]ATP.
-
P]ATP and analysis on
thin layer chromatography.
isoform of protein
kinase C(47) . Activation of Cbl presumably initiates a cascade
of events including the activation of S6 kinase and the induction of
proliferation(48, 49) . While this report was being
prepared, the association of PI 3-kinase with Cbl was reported in
receptor-ligated T cells(50) , and we are aware of similar to
be reported results from a second group(51) . It is likely that
the activation of PI-3 kinase by Cbl will prove to be a recurring theme
in mitogenic signal transduction.
/ complex and does not require functional Btk.
While CD40 cross-linking can also lead to the activation of PI-3 kinase
in B cells, this apparently occurs via a Cbl-independent mechanism. It
is unclear as to whether there is a functional significance to the
association of Cbl with Grb2 and the formation of complexes between
Shc, Grb2, and Cbl following B cell activation. While such a complex
could in theory promote the association of Cbl with the antigen
receptor and therefore with cellular membranes, given the apparently
low stoichiometry with which such complexes are generated, it is
unclear whether such an event is necessary for the activation of PI
3-kinase activity. It is worth noting that previous attempts to
demonstrate PI 3-kinase association in vivo with the
Ig/ heterodimer have proved unsuccessful(44) ,
suggesting that recruitment of PI 3-kinase via Shc, Grb2 and Cbl to the
antigen receptor does not occur readily. However it has also been
suggested, albeit based on indirect evidence, that membrane association
of PI 3-kinase might be necessary for catalytic
activation(52) . The association of Cbl with Golgi membranes
has been demonstrated in activated macrophages(32) . This
phenomenon, as well as the activation of PI 3-kinase, might be linked
to the formation of Grb2-Cbl or Shc-Grb2-Cbl complexes in cells that
have received a mitogenic stimulus.
)))
We thank Bruce Mayer for Grb2 fusion proteins, Abul
Abbas and Rick Mitchell for the human IgM transfectants of A20.25, Tom
Rothstein and Bill Paul for BAL17 cells, Ajay Rana for advice on lipid
kinase assays, and Hamid Band for sharing information prior to
publication.
©1995 by The American Society for Biochemistry and Molecular Biology, Inc.
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 M. Kato, R. T. Abraham, S. Okada, and H. Kita Ligation of the beta 2 Integrin Triggers Activation and Degranulation of Human Eosinophils Am. J. Respir. Cell Mol. Biol., May 1, 1998; 18(5): 675 - 686. [Abstract] [Full Text]
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M. Deckert, C. Elly, A. Altman, and Y.-C. Liu Coordinated Regulation of the Tyrosine Phosphorylation of Cbl by Fyn and Syk Tyrosine Kinases J. Biol. Chem., April 10, 1998; 273(15): 8867 - 8874. [Abstract] [Full Text] [PDF]
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E. A. Feshchenko, W. Y. Langdon, and A. Y. Tsygankov Fyn, Yes, and Syk Phosphorylation Sites in c-Cbl Map to the Same Tyrosine Residues That Become Phosphorylated in Activated T Cells J. Biol. Chem., April 3, 1998; 273(14): 8323 - 8331. [Abstract] [Full Text] [PDF]
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K. A. Reedquist and J. L. Bos Costimulation through CD28 Suppresses T Cell Receptor-dependent Activation of the Ras-like Small GTPase Rap1 in Human T Lymphocytes J. Biol. Chem., February 27, 1998; 273(9): 4944 - 4949. [Abstract] [Full Text] [PDF]
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V. Ribon, J. A. Printen, N. G. Hoffman, B. K. Kay, and A. R. Saltiel A Novel, Multifunctional c-Cbl Binding Protein in Insulin Receptor Signaling in 3T3-L1 Adipocytes Mol. Cell. Biol., February 1, 1998; 18(2): 872 - 879. [Abstract] [Full Text]
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B. L. Craddock and M. J. Welham Interleukin-3 Induces Association of the Protein-tyrosine Phosphatase SHP2 and Phosphatidylinositol 3-Kinase with a 100-kDa Tyrosine-phosphorylated Protein in Hemopoietic Cells J. Biol. Chem., November 14, 1997; 272(46): 29281 - 29289. [Abstract] [Full Text] [PDF]
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D. L. Barber, J. M. Mason, T. Fukazawa, K. A. Reedquist, B. J. Druker, H. Band, and A. D. D'Andrea Erythropoietin and Interleukin-3 Activate Tyrosine Phosphorylation of CBL and Association With CRK Adaptor Proteins Blood, May 1, 1997; 89(9): 3166 - 3174. [Abstract] [Full Text] [PDF]
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H. Odai, K. Sasaki, A. Iwamatsu, T. Nakamoto, H. Ueno, T. Yamagata, K. Mitani, Y. Yazaki, and H. Hirai Purification and Molecular Cloning of SH2- and SH3-Containing Inositol Polyphosphate-5-Phosphatase, Which Is Involved in the Signaling Pathway of Granulocyte-Macrophage Colony-Stimulating Factor, Erythropoietin, and Bcr-Abl Blood, April 15, 1997; 89(8): 2745 - 2756. [Abstract] [Full Text] [PDF]
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Y.-C. Liu, Y. Liu, C. Elly, H. Yoshida, S. Lipkowitz, and A. Altman Serine Phosphorylation of Cbl Induced by Phorbol Ester Enhances Its Association with 14-3-3Proteins in T Cells via a Novel Serine-rich 14-3-3-binding Motif J. Biol. Chem., April 11, 1997; 272(15): 9979 - 9985. [Abstract] [Full Text] [PDF]
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M. Ojaniemi, S. S. Martin, F. Dolfi, J. M. Olefsky, and K. Vuori The Proto-oncogene Product p120cbl Links c-Src and Phosphatidylinositol 3'-Kinase to the Integrin Signaling Pathway J. Biol. Chem., February 7, 1997; 272(6): 3780 - 3787. [Abstract] [Full Text] [PDF]
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Y.-C. Liu, C. Elly, W. Y. Langdon, and A. Altman Ras-dependent, Ca2+-stimulated Activation of Nuclear Factor of Activated T Cells by a Constitutively Active Cbl Mutant in T Cells J. Biol. Chem., January 3, 1997; 272(1): 168 - 173. [Abstract] [Full Text] [PDF]
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