| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
(Received for publication, September 22, 1995; and in revised form, January 10, 1996) From the
Activation of CD28 on T lymphocytes initiates a cascade of
intracellular events, which in concert with activation of the T cell
receptor, culminates in production of cytokines and a functional immune
response. One of the earliest biochemical changes observed following
stimulation of CD28 is tyrosine phosphorylation. We have demonstrated
that both the LCK and the EMT/ITK/TSK (EMT) intracellular tyrosine
kinases are activated following cross-linking of CD28. Utilizing
somatic cell mutants lacking LCK, we demonstrate that functional LCK is
required for CD28-induced activation of EMT as evidenced by increased
tyrosine phosphorylation and kinase activity. In support of a role for
LCK in EMT activation, reconstitution of a LCK-negative Jurkat T cell
line by transfection with normal LCK recreates CD28-mediated EMT
activation. Furthermore, co-transfection of LCK and EMT into COS-7
cells showed that EMT becomes phosphorylated in the presence of LCK. In
addition, increases in EMT association with CD28 were eliminated in a
LCK-negative Jurkat cell line, but were restored following transfection
of wild type LCK. The data are most compatible with a model in which
LCK, either directly or indirectly, initiates EMT activation and
association with CD28 following ligation of CD28.
Co-stimulation of T lymphocytes requires the cooperation of two
signals delivered by antigen presenting cells: one stimulatory signal
derived from interaction of the T cell receptor (TCR) ( Upon activation of CD28, there is a rapid and immediate increase in
tyrosine phosphorylation of a number of specific
substrates(8, 9, 10, 11, 12) .
However, because CD28 does not contain an intrinsic kinase domain, it
must activate intracellular tyrosine kinases. In addition,
cross-linking of CD28 leads to the induction of a number of early
signaling events, including increases in cytosolic free
calcium(7, 8) , activation of RAS(13) ,
activation of mitogen-activated protein kinase (13) ,
activation of phosphatidylinositol
3`-kinase(14, 15, 16, 17, 18) ,
activation of JNK kinase (also known as stress-activated kinase) (19) and activation of RAF kinase(20) . Although it is
clear that cross-linking of CD28 can induce a number of early signals,
the role that activation of these biochemical changes plays in the
ability of CD28 to synergize with the TCR to induce a functional T cell
response remains unclear. Furthermore, the mechanisms leading to
activation of these enzymes by CD28 and in particular the order of
activation of each of these enymes is unknown. We have demonstrated
recently that EMT/ITK/TSK (EMT), a TEC family protein tyrosine kinase,
becomes activated after CD28 cross-linking, as evidenced by a transient
increase in tyrosine phosphorylation and kinase activity. In addition,
stimulation of CD28 results in a rapid increase in the association of
EMT with CD28(21) . Thus EMT has the potential to play a role
in CD28 signal transduction. LCK is also activated after stimulation of
CD28, suggesting that this kinase may have a signaling role through
CD28 in addition to its dual role downstream of both the TCR and
CD4/8(21, 22) . The TEC family of intracellular
kinases currently consists of members that contain SH2 and SH3 SRC
homology (SH) domains but lack the negative regulatory tyrosine present
at the carboxyl terminus and the myristoylation site found at the amino
terminus of SRC family members. Thus the TEC family of tyrosine kinases
must be regulated in a different manner from the SRC family of tyrosine
kinases. BTK and EMT contain, in addition to the SH2 and SH3 domains, a
pleckstrin homology (PH) domain. The exact function of this domain is
currently unknown, but it may play a role in the ability of BTK and EMT
to associate with other molecules such as protein kinase
C(23, 24) . Both BTK and EMT have restricted patterns
of expression; BTK is expressed mainly in mast cells and B cells and
EMT is expressed primarily in mast cells and T
cells(25, 26) . BTK is involved in B cell signal
transduction; mutations in BTK have been causally linked to X-linked
agammaglobulinemia, a severe human B cell
immunodeficiency(27) . In addition, since cross-linking mouse
Fc Herein, we demonstrate
that CD28-mediated EMT activation and EMT association with CD28 is
greatly decreased in cells lacking functional LCK. Reconstitution of
LCK kinase activity by enforced expression of the normal human LCK
reconstituted ligand-induced EMT activation and increased EMT
association with CD28, confirming a role for LCK in EMT activation. In
addition co-expression of EMT and LCK in COS-7 cells lead to tyrosine
phosphorylation of EMT. Thus, EMT appears to be located downstream of
LCK in the signaling pathways activated by CD28.
Figure 1:
CD28-induced tyrosine phosphorylation
of EMT requires functional LCK. Jurkat T cells were stimulated by
cross-linking CD28 with 10 µg/ml anti-CD28 (9.3) antibody and 10
µg/ml RAM for the indicated times. Similar results were observed in
the absence of RAM (see (18) and data not presented). The
cells were then lysed as described under ``Materials and
Methods.'' A, EMT was immunoprecipitated with anti-EMT
serum as described under ``Materials and Methods.'' The
precipitates were loaded on a 10% SDS-PAGE gel, transferred to
Immobilon, and Western-blotted with anti-phosphotyrosine antibody
(4G10). B, the blot was stripped with 1% SDS and reprobed with
anti-EMT antibody. C, total phosphotyrosine-containing
proteins were immunoprecipitated with polyclonal anti-phosphotyrosine
antibodies prepared in this laboratory, loaded on SDS-PAGE, and blotted
with anti-phosphotyrosine antibody (4G10). Immunoreactivity was
detected by ECL in all cases. Parental Jurkat (panel a), JCaM
1.6 (panel b), and LCK transfected JCaM 1.6 (panel c)
were treated similarly; the result shown represents one of three
similar experiments. In panel d, data from panels
a-d were converted to relative levels by densitometry and
normalized to EMT protein levels determined by densitometry. The
relative level of tyrosine phosphorylation is presented as the -fold
increase over basal levels of EMT tyrosine phosphorylation.
Figure 2:
Activation-induced increase in EMT
tyrosine kinase activity. The relative increase in kinase activity of
EMT was determined after stimulation of Jurkat cells as described in
the legend to Fig. 1. EMT was immunoprecipitated and kinase
activity was determined utilizing the SRC peptide as substrate as
described under ``Materials and Methods.'' Kinase activity is
presented as -fold increase over that observed in unstimulated cells to
allow comparison between experiments. The results represent mean and
standard error of the mean of three repeats of three independent
experiments.
Figure 3:
EMT
association with CD28 following CD28 activation. EMT association with
CD28 was determined by lysing JCaM1.6 cells (a) or JCaM1.6
cells transfected with LCK in Nonidet P-40 lysing buffer after
cross-linking CD28 for the indicated time periods (b). CD28
was immunoprecipitated from cell lysates with anti-CD28 (9.3, 10
µg/ml) and RAM (10 µg/ml) and Western-blotted with anti-EMT as
described under ``Materials and Methods.'' Activation of the
parental Jurkat cell line with anti-CD28 for 5 min provides a positive
control for CD28-induced EMT association with CD28. The data represent
one of three similar experiments. ipt,
immunoprecipitation.
To confirm that the decreased activation
of EMT and reduced association of EMT with CD28 in the JCaM 1.6 line
was due to a lack of functional LCK, we determined whether the response
to CD28 cross-linking was restored in a JCaM 1.6 cell line transfected
to express normal human LCK. Previous studies had demonstrated that
expression of LCK in JCaM 1.6 restores anti-TCR-mediated signaling and
interleukin-2 production (29) . In JCaM 1.6 LCK transfectants,
CD28-induced tyrosine phosphorylation, and specifically, tyrosine
phosphorylation of EMT induced by cross-linking CD28 was completely
restored (Fig. 1, panels c and d). Similarly,
anti-CD28 mediated increases in EMT kinase activity were restored in
the JCaM 1.6 LCK transfectants (Fig. 2). In addition, the
increase in association of EMT with CD28 was restored (Fig. 3b). The ability of transfected LCK to
reconstitute CD28-mediated EMT activation confirms that the defect in
EMT activation in JCaM 1.6 was indeed a consequence of lack of
functional LCK.
Figure 4:
LCK tyrosine phosphorylation of EMT in
COS-7 cells. COS-7 cells were transfected with expression vectors
encoding EMT/ITK in the presence or absence of LCK as described under
``Materials and Methods.'' Cells were harvested and EMT was
immunoprecipitated and separated on a 10% SDS-PAGE gel and transferred
to Immobilon membrane, which was then probed with anti-phosphotyrosine
antibodies (top). The blot was then stripped and reprobed with
anti-EMT antibodies (bottom).
The data presented are most consistent with a model wherein
CD28-induced activation of the LCK tyrosine kinase is proximal in a
cascade leading to CD28-induced activation of EMT. In support of this
possibility, we have demonstrated that LCK kinase activity is increased
following stimulation of CD28 (21, 22) and that LCK
can lead to tyrosine phosphorylation of EMT in COS-7 cells (Fig. 4). Whether LCK directly phosphorylates EMT or EMT
activation is a consequence of LCK-mediated phosphorylation of an
intermediary molecule following CD28 activation is currently unknown.
As Jurkat cells express both SRC and FYN(32) , these Src family
tyrosine kinases, in contrast to LCK, are either not sufficient for
CD28-mediated EMT activation or are not stimulated following CD28
ligation. Regulation of EMT kinase activity may well be at the level
of tyrosine phosphorylation, since tyrosine phosphorylation of EMT and
EMT kinase activity demonstrated concurrent changes following
cross-linking of CD28 (Fig. 1d and Fig. 2). EMT
contains a tyrosine in a conserved internal site which becomes
autophosphorylated in Src family tyrosine kinases and likely positively
regulates kinase activity(26) . However, kinase assays revealed
that EMT is very inefficient at autophosphorylation as compared to Src
family tyrosine kinases(21) . Since EMT is inefficient in
autophosphorylation (21) and LCK has the ability to induce
tyrosine phosphorylation of EMT (Fig. 4), this phosphorylation
site in EMT could be a direct target for other kinases such as LCK.
Indeed a number of different tyrosine kinases are both positively and
negatively regulated by other tyrosine kinases in activation cascades.
For example, phosphorylation of the negative regulatory site of SRC
family kinases seems to be dependent on the action of CSK family
kinases(33, 34) . In turn, a number of different SRC
family kinases have been demonstrated to regulate non-SRC family
kinases. This is well documented for LYN and SYK in B
cells(35) , SRC and FAK in fibroblasts(36) , and FYN or
LCK and ZAP70 in T cells(2) . ZAP70 activation is, however, not
detectable after CD28 ligation. ( After CD28 activation, the receptor becomes
tyrosine-phosphorylated (15, 18) . Signaling proteins
such as phosphatidylinositol 3`-kinase and Grb2 bind to these
phosphotyrosine residues through their SH2 domains (14, 15, 16, 17, 18, 38, 39) .
As demonstrated previously(21) , EMT binds to CD28
constitutively and after activation the association is up-regulated
presumably by SH2 domain interactions. This increase in association was
greatly reduced in the JCaM 1.6 cell line and restored in the JCaM 1.6
cell line transfected with LCK. This suggests that functional LCK is
required for CD28 cross-linking induced increases in EMT association
with CD28. This may be a consequence of LCK either directly or
indirectly tyrosine phosphorylating CD28. Recent results, using
transfection into Spodoptera cells, supports LCK as the
primary mediator of CD28 phosphorylation(38) . Furthermore,
EMT, phosphatidylinositol 3`-kinase, and GRB2 association with CD28 in
transfected Spodoptera cells required co-expression of LCK (38) . Although the PH domain of both BTK and EMT associate
with protein kinase C isoymes(24) , limited data are present to
date on the interactions between TEC family kinases and other tyrosine
kinases. Although EMT and BTK appear to associate with SRC family
kinases (including LCK and FYN) in the yeast two-hybrid system and when
expressed as bacterial fusion
proteins(40, 41, 42) , it has been difficult
to demonstrate interactions in intact
cells(40, 41, 42) . Indeed, under conditions
(using Nonidet P-40 buffers) in which we can readily demonstrate
association of EMT with CD28 (Fig. 3, (21) ), we have
been unable to demonstrate association of EMT with LCK, FYN, SRC, or
TTK (all of which are expressed by Jurkat T cells; 10, 32, 43, 44) as
indicated by co-immunoprecipitation (data not shown). In summary, we
have demonstrated that the ability of CD28 to optimally activate EMT is
dependent on the presence of functional LCK. The data are most
compatible with a model of CD28 signaling in which activated LCK
mediates phosphorylation and activation of EMT and mediates EMT
association with CD28. Thus EMT activation seems to be located
downstream of LCK in a kinase cascade stimulated by CD28.
Volume 271,
Number 12,
Issue of March 22, 1996 pp. 7079-7083
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
FOOTNOTES
ACKNOWLEDGEMENTS
REFERENCES
)complex with antigen in the context of the major
histocompatibility complex and a second co-stimulatory signal from the
ligation of accessory molecules(1) . In the absence of the
co-stimulatory signal, T cells fail to undergo clonal expansion and
instead ultimately enter an abortive pathway characterized by antigen
desensitization, anergy, or programmed cell
death(1, 2, 3, 4) . The interaction
of CD28 on T lymphocytes with B7.1 (CD80) or B7.2 (CD86) on
antigen-presenting cells is the most potent identified co-stimulatory
signal. Indeed, cross-linking of CD28 can prevent activation-induced
desensitization, anergy, and programmed cell
death(4, 5, 6, 7, 8) . RI leads to activation of BTK (28) , this kinase may
also play a role in mast cell activation.
Antibodies
Monoclonal anti-CD28 antibody 9.3
(IgG2a) was a kind gift of J. Ledbetter (Bristol-Myers Squibb Research
Institute, Seattle). Anti-phosphotyrosine antibody (4G10, IgG1) was
purchased from Upstate Biotechnology, Inc. (Lake Placid, NY). The
production and specificity of the anti-EMT serum used in these studies
was described previously(26) .Cell Lines
COS-7 cells, the parental human Jurkat
leukemic cell line E6.1 and the JCaM1.6 Jurkat clone (which is
LCK-negative, (29) ) were from the American Type Culture
Collection (ATCC, Rockville, MD). JCaM 1.6 transfected with LCK (JCaM
1.6-LCK) was obtained from Art Weiss (University of California, San
Francisco, (40) ). By Western blotting the protein level of LCK
in JCaM1.6-LCK was comparable with the parental Jurkat cell line (data
not shown) and was not detected in the JCaM1.6 cell line. Similar
levels of EMT were present, as assessed by Western blotting, in all the
Jurkat T cell lines studied (data not shown). CD28 expression was the
same for all Jurkat cell lines studied (data not shown). Both Jurkat
E6.1 and JCaM1.6 have been demonstrated to lack functional
SYK(30) .Cell Culture, Stimulation, Transfection, and
Lysis
Jurkat cells were cultured and starved as described
previously(21, 26) . Anti-CD28 antibodies were added
at 1 µg/5 
10
cells at 37 °C as indicated.
Rabbit anti-mouse (RAM) antibodies (10 µg/ml) were added 1 min
after addition of anti-CD28 antibodies, in order to induce
cross-linking. After activation, the cells were pelleted and
immediately incubated with lysis buffer (26) for 15 min at 4
°C. COS-7 cells were transfected with plasmids pA1068 (hlck in
pMEXneo) and/or pCMVEMTneo (hEMT in pCMVneo) by calcium phosphate
precipitation and lysed 2 days later.Immunoprecipitation and Western Blotting
Cell
lysates were centrifuged at 14,000 g for 15 min at 4
°C. After centrifugation the supernatant was immunoprecipitated and
Western-blotted as described previously(21, 26) . The
blots were blocked overnight in either 5% bovine serum albumin for the
detection of phosphotyrosine residues or 5% non-fat milk for detection
of EMT. Anti-phosphotyrosine antibodies (4G10, 1:2500 dilution) or EMT
antibodies (1:1000) were added for 1 h. Membranes were washed,
incubated with the appropriate secondary antibody, and protein was
detected by enhanced chemiluminescence (ECL). Where indicated, the
blots were stripped with 1% SDS, reprobed with anti-EMT antibodies
(1:1000 dilution), and visualized by ECL.In Vitro Kinase Assay
Tyrosine kinase activity of
EMT was assayed by immunoprecipitation of lysates as described above.
The immunoprecipitates were then washed once in kinase wash buffer (150
mM NaCl, 10 mM Tris, pH 7.2, 1 mM phenylmethylsulfonyl fluoride, and 1 mM sodium vanadate).
The precipitates were then incubated in 45 µl of kinase buffer (10
mM manganese chloride, 10 mM HEPES, pH 7.0, 1 mM phenylmethylsulfonyl fluoride, and 0.1 mM sodium
orthovanadate) containing 5 µCi of
[-
P]ATP and 5 µg of a peptide
(Arg-Arg-Leu-Ile-Glu-Asp-Ala-Glu-Tyr-Ala-Ala-Arg-Gly) (Sigma) derived
from the sequence surrounding the SRC tyrosine kinase
autophosphorylation site(31) . This mixture was incubated for
15 min at room temperature which was found to be within linear range of
the assay. The mixture was then blotted onto phosphocellulose paper and
washed six times with phosphoric acid. The amount of
-
P incorporated was determined by scintillation
counting.
Optimal CD28-induced EMT Activation Requires
LCK
Activation of CD28 initiates an intracellular kinase cascade
that involves the LCK kinase(21, 22) . To test whether
LCK expression is required for optimal activation of EMT by CD28, we
measured the level of EMT activation, as assessed by tyrosine
phosphorylation and kinase activity, after CD28 cross-linking in the
JCaM 1.6 Jurkat T cell line that does not express LCK. In contrast to
results in the parental Jurkat T cell line, both basal and CD28-induced
total tyrosine phosphorylation was markedly decreased in the JCaM 1.6
line in at least three similar experiments (Fig. 1, panels b and d, and data not presented). Strikingly, although
CD28-induced tyrosine phosphorylation of EMT was readily detectable in
parental Jurkat cells (Fig. 1, panels a and d)(21) , no CD28-induced increase in EMT
phosphorylation was detected in JCaM 1.6 cells (Fig. 1, panels b and d). In parallel with the tyrosine
phosphorylation data, incubation of JCaM 1.6 with anti-CD28 antibodies
did not induce EMT kinase activity (Fig. 2). Furthermore,
increased EMT association with CD28 was not detected in the JCaM 1.6
cell line as compared with the parental Jurkat cell line (Fig. 3a).
,
cell line: Jurkat, stimulation: CD28, readout: EMT Tyr(P); 
, cell
line: JCaM1, stimulation: CD28, readout, EMT Tyr(P); , cell line:
JCaM1 LCK, stimulation: CD28, readout: EMT
Tyr(P).
, cell line: Jurkat, stimulation: CD28, readout:
EMT immunoprecipitation kinase; , cell line: Jcam1, stimulation:
CD28, readout: EMT immunoprecipitation kinase; , cell line: Jcam1
LCK, stimulation: CD28, readout, EMT immunoprecipitation
kinase.
Co-transfection of LCK and EMT into COS-7 Cells Leads to
EMT Phosphorylation
CD28-mediated EMT activation seems to
require functional LCK in Jurkat cells. This suggests that EMT could be
a target for tyrosine phosphorylation by LCK. In support of this
possibility, both EMT and LCK were transfected in COS-7 cell either
alone or together and the extent of EMT phosphorylation determined. As
shown in Fig. 4, transfection of vector alone or EMT alone did
not result in detectable tyrosine phosphorylation of EMT. However,
concurrent transfection of both LCK and EMT in COS-7 cells resulted in
tyrosine phosphorylation of a unique 72-kDa band consistent with LCK
phosphorylating EMT.
)Further support for a
model in which tyrosine kinases are coordinately regulated in an
activation cascade is provided by the demonstration that functional
signaling by the platelet-derived growth factor receptor is dependent
on the presence of functional SRC family kinases(37) . It is
important to note, in terms of potential kinase activation cascades,
that the experiments presented herein, although demonstrating that LCK
is required for CD28-mediated EMT activation, do not address the
question of whether LCK is reciprocally regulated either negatively or
positively by EMT.
))
We thank Dr. A. Weiss for the kind gift of cell lines
that made these experiments possible and Dr. J. Ledbetter for the gift
of anti-CD28 antibody (9.3) along with his expert advice.
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  K. Nika, L. Tautz, Y. Arimura, T. Vang, S. Williams, and T. Mustelin A Weak Lck Tail Bite Is Necessary for Lck Function in T Cell Antigen Receptor Signaling J. Biol. Chem., December 7, 2007; 282(49): 36000 - 36009. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Q. Qi, N. Sahu, and A. August Tec Kinase Itk Forms Membrane Clusters Specifically in the Vicinity of Recruiting Receptors J. Biol. Chem., December 15, 2006; 281(50): 38529 - 38534. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C.-R. Li and L. J. Berg Cutting Edge: Itk Is Not Essential for CD28 Signaling in Naive T Cells J. Immunol., April 15, 2005; 174(8): 4475 - 4479. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Rahmouni, T. Vang, A. Alonso, S. Williams, M. van Stipdonk, C. Soncini, M. Moutschen, S. P. Schoenberger, and T. Mustelin Removal of C-Terminal Src Kinase from the Immune Synapse by a New Binding Protein Mol. Cell. Biol., March 15, 2005; 25(6): 2227 - 2241. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Tavano, G. Gri, B. Molon, B. Marinari, C. E. Rudd, L. Tuosto, and A. Viola CD28 and Lipid Rafts Coordinate Recruitment of Lck to the Immunological Synapse of Human T Lymphocytes J. Immunol., November 1, 2004; 173(9): 5392 - 5397. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. H. Ellis, C. Ashman, M. N. Burden, K. E. Kilpatrick, M. A. Morse, and P. A. Hamblin GRID: A Novel Grb-2-Related Adapter Protein That Interacts with the Activated T Cell Costimulatory Receptor CD28 J. Immunol., June 1, 2000; 164(11): 5805 - 5814. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J.-G. Nemorin and P. Duplay Evidence That Lck-mediated Phosphorylation of p56dok and p62dok May Play a Role in CD2 Signaling J. Biol. Chem., May 5, 2000; 275(19): 14590 - 14597. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. J. Perez-Villar and S. B. Kanner Regulated Association Between the Tyrosine Kinase Emt/Itk/Tsk and Phospholipase-C{gamma}1 in Human T Lymphocytes J. Immunol., December 15, 1999; 163(12): 6435 - 6441. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. L. Sommers, R. L. Rabin, A. Grinberg, H. C. Tsay, J. Farber, and P. E. Love A Role for the Tec Family Tyrosine Kinase Txk in T Cell Activation and Thymocyte Selection J. Exp. Med., November 15, 1999; 190(10): 1427 - 1438. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. L. Leung, L. Haughn, A. Veillette, R. G. Hawley, R. Rottapel, and M. Julius TCR{alpha}{beta}-Independent CD28 Signaling and Costimulation Require Non-CD4-Associated Lck J. Immunol., August 1, 1999; 163(3): 1334 - 1341. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Chuang, K.-M. Lee, M. D. Robbins, J. M. Duerr, M.-L. Alegre, J. E. Hambor, M. J. Neveu, J. A. Bluestone, and C. B. Thompson Regulation of Cytotoxic T Lymphocyte-Associated Molecule-4 by Src Kinases J. Immunol., February 1, 1999; 162(3): 1270 - 1277. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W.-C. Yang, M. Ghiotto, B. Barbarat, and D. Olive The Role of Tec Protein-tyrosine Kinase in T Cell Signaling J. Biol. Chem., January 8, 1999; 274(2): 607 - 617. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Michel, L. Grimaud, L. Tuosto, and O. Acuto Fyn and ZAP-70 Are Required for Vav Phosphorylation in T Cells Stimulated by Antigen-presenting Cells J. Biol. Chem., November 27, 1998; 273(48): 31932 - 31938. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Lu, B. Cuevas, S. Gibson, H. Khan, R. LaPushin, J. Imboden, and G. B. Mills Phosphatidylinositol 3-Kinase Is Required for CD28 But Not CD3 Regulation of the TEC Family Tyrosine Kinase EMT/ITK/TSK: Functional and Physical Interaction of EMT with Phosphatidylinositol 3-Kinase J. Immunol., November 15, 1998; 161(10): 5404 - 5412. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Okkenhaug and R. Rottapel Grb2 Forms an Inducible Protein Complex with CD28 through a Src Homology 3 Domain-Proline Interaction J. Biol. Chem., August 14, 1998; 273(33): 21194 - 21202. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K.-Q. Liu, S. C. Bunnell, C. B. Gurniak, and L. J. Berg T Cell Receptor-initiated Calcium Release Is Uncoupled from Capacitative Calcium Entry in Itk-deficient T Cells J. Exp. Med., May 18, 1998; 187(10): 1721 - 1727. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Wong, D. Straus, and A. C. Chan Genetic Evidence of a Role for Lck in T-Cell Receptor Function Independent or Downstream of ZAP-70/Syk Protein Tyrosine Kinases Mol. Cell. Biol., May 1, 1998; 18(5): 2855 - 2866. [Abstract] [Full Text]
|
|
|
|
|
|
H.-H. Kim, M. Tharayil, and C. E. Rudd Growth Factor Receptor-bound Protein 2 SH2/SH3 Domain Binding to CD28 and Its Role in Co-signaling J. Biol. Chem., January 2, 1998; 273(1): 296 - 301. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. J. Phipps, S. Yousefi, and D. R. Branch Increased Enzymatic Activity of the T-Cell Antigen Receptor-Associated Fyn Protein Tyrosine Kinase in Asymptomatic Patients Infected With the Human Immunodeficiency Virus Blood, November 1, 1997; 90(9): 3603 - 3612. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Hanada, L. Lin, K. G. Chandy, S. S. Oh, and A. H. Chishti Human Homologue of the Drosophila Discs Large Tumor Suppressor Binds to p56lck Tyrosine Kinase and Shaker Type Kv1.3 Potassium Channel in T Lymphocytes J. Biol. Chem., October 24, 1997; 272(43): 26899 - 26904. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. August, A. Sadra, B. Dupont, and H. Hanafusa Src-induced activation of inducible T cell kinase (ITK) requires phosphatidylinositol 3-kinase activity and the Pleckstrin homology domain of inducible T cell kinase PNAS, October 14, 1997; 94(21): 11227 - 11232. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. D. Heyeck, H. M. Wilcox, S. C. Bunnell, and L. J. Berg Lck Phosphorylates the Activation Loop Tyrosine of the Itk Kinase Domain and Activates Itk Kinase Activity J. Biol. Chem., October 3, 1997; 272(40): 25401 - 25408. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
X. C. Liao, S. Fournier, N. Killeen, A. Weiss, J. P. Allison, and D. R. Littman Itk Negatively Regulates Induction of T Cell Proliferation by CD28 Costimulation J. Exp. Med., July 21, 1997; 186(2): 221 - 228. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
