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CHEMOKINES
From the Department of Microbiology, Department of
Immunology and Medicine, and Department of Biochemistry and Molecular
Biology, Indiana University School of Medicine; and the Walther
Oncology Center, Indianapolis, IN 46202.
Chemokines play a pivotal role in regulating leukocyte migration as
well as other biological functions. CC chemokine receptor 9 (CCR9) is a
specific receptor for thymus-expressed CC chemokine (TECK). It
is shown here that engagement of CCR9 with TECK leads to
phosphorylation of Akt (protein kinase B), mitogen-activated protein kinases (MAPKs), glycogen synthase kinase-3 Chemokines refer to a group of cytokines whose
function was originally confined to regulation of leukocyte
trafficking.1 Chemokines have since been implicated in
other biological functions.2 These involve regulation of
hematopoiesis, T helper cell development, inflammation,
angiogenesis/angiostasis, regulation of cytokine production from
certain dendritic cells, and cell survival. These diverse functions of
chemokines result primarily from the interaction of chemokines with
their cognate receptors, collectively termed chemokine receptors.
Chemokine receptors are a subfamily of the G protein-coupled receptor
supergene family that uses heterotrimeric G proteins (G Upon binding of certain chemokines to chemokine receptors, signaling
events are noted; these include F-actin formation; transient calcium
mobilization; inhibition of intracellular cyclic adenosine monophosphate (cAMP) levels; activation of mitogen-activated protein kinases (MAPKs); protein kinase C (PKC); c-Jun-NH2 terminal
kinase; Pyk2; focal adhesion kinase; phosphoinositide-3 kinase
(PI-3 kinase); Akt (protein kinase B); the Jak/signal transducer and
activator of transcription pathway; and nuclear factor We and others identified CC chemokine receptor 9 (CCR9), previously
designated GPR-9-6, as a specific receptor for thymus-expressed chemokine (TECK).12,13 One interesting feature of CCR9 is
that it is expressed mainly in immature T cells such as double-positive (DP) T cells or gut-associated T cells.14 This makes CCR9
a valuable model system for understanding T-cell development and tissue-specific homing. Additionally, DP T cells undergo extensive apoptosis, thereby becoming single-positive T cells.14
Therefore, it may be possible that CCR9-mediated signaling is
associated with this apoptosis. Because several studies have shown that
chemokine receptor-mediated signaling activates a battery of protein
or lipid kinases that are generally thought to be involved in cell survival or cell proliferation,3-5 it is possible that
CCR9/TECK interaction provides antiapoptotic signaling to DP T cells.
Thus, we decided to evaluate signaling components activated following CCR9/TECK interaction. This led to the finding that this
receptor/ligand interaction induces activation of PI-3 kinase as well
as its downstream target Akt, MAPK, glycogen synthase kinase-3 Chemokines and reagents
Cell culture, fluorescence-activated cell-sorting analysis, and
transfection
Construction of expression plasmids and transfection The CCR9 expression vector has been previously described.12 Akt complementary DNA (cDNA) was obtained from MOLT4 cDNA library. Akt cDNA was cloned into pCEP4 (Invitrogen, Carlsbad, CA) that had been digested with XhoI and BamHI. The transcriptional unit encompassing the cytomegalovirus early promoter, Akt, and 3' polyadenylation site was released by SalI digestion and cloned into the CCR9 expression vector. This expression vector was then transfected into Cos7 cells by means of Fugene 6 (Roche). After 48 hours, the transfected cells were serum deprived for 2 hours and stimulated with TECK, and cell lysates were prepared. Activated Akt was detected by the phospho-specific Akt antibody.Calcium flux assay Receptor activation was assessed in chemokine receptor-expressing cell lines by real-time measurement of (Ca++)i changes by means of an MSIII fluorometer (Photon Technology International, S Brunswick, NJ). The cell preparation was made as described.11 Excitation scans between 300 and 400 nm were performed to determine whether fura-2AM was appropriately loaded. Excitation ratios were measured at 340 and 380 nm.Detection of phosphorylated kinases and downstream targets of Akt MOLT4 cells were serum starved overnight or for 2 hours and stimulated with TECK (1 µg/mL) for varying time points. Cell lysates were prepared as described elsewhere,15 and protein concentrations were determined by means of a Pierce chemical kit. Equal amounts of protein lysates were employed for Western blot analysis. Subsequently, protein blots were incubated with proper concentrations of phospho-specific antibodies or respective pan-antibodies. Target protein bands were detected by chemiluminescence.Chemotaxis assay A bare-filter transwell assay was performed as previously described.13 MOLT4 cells were washed once with chemotaxis assay buffer (RPMI, 20 mM Hepes, pH 7.4, 1% bovine serum albumin) and resuspended in the same buffer. We placed 5 × 105 cells in 100 µL buffer in the upper inset, and optimal concentrations of TECK in 600 µL buffer were placed in the lower chamber. Cell migration was allowed to take place at 37°C for 4 hours. Migrated cells were harvested and counted with a hemocytometer. For chemotaxis inhibition assay, MOLT4 cells were pretreated with varying concentrations of wortmannin, PD98059 (50 µM), and pertussis toxin (1 µg/mL) or vehicle for 1 hour, washed once with phosphate-buffered saline (PBS), and used for chemotaxis.Apoptosis assay We deprived 1 × 106 MOLT cells of serum overnight. These cells were preincubated with cycloheximide (CHX) at a final concentration of 2.5 µg/mL for 1 hour. Afterwards, cells were stimulated with CH-11 (0.5 µg/mL) and 1 µg/mL TECK for 6 hours. For inhibitor studies, CHX-treated cells were incubated with wortmannin (25 or 50 nM), PD98059 (50 µM), and rapamycin (10 µM) for 30 minutes and then stimulated with CH-11 and TECK for 6 hours. The treated cells were washed twice with cold PBS and stained with annexin V for 15 minutes. The apoptotic cell population that was annexin-V-positive and propidium iodide-negative was measured by means of flow cytometry.
PI-3 kinase-dependent migration of MOLT4 cells in response to TECK MOLT4 cells have been shown to express high levels of CCR9.13,14 CCR9 is a specific receptor for TECK.12,13 Binding of TECK to CCR9 induced a rapid and transient calcium flux (Figure 1A). In order to evaluate the chemotactic stimulating ability of TECK, bare-filter transwell migration assays were performed with MOLT4 cells. The maximal chemotactic responsiveness of MOLT4 cells to TECK was seen at 1 µg/mL, with migration desensitized at higher concentrations (Figure 1B). TECK-mediated migration of MOLT4 cells was completely abrogated by pertussis toxin (PT) (Figure 1B), suggesting that CCR9 is coupled to G i proteins in the cells.
PI-3 kinases and MAPK have been implicated in cell movement or cell
shape change.16,17 However, information on involvement of
PI-3 kinase or MAPK in chemokine-mediated chemotaxis is fairly limited.
It has been shown that CXCR4 or RANTES-receptor-mediated chemotaxis is
inhibited by PI-3 kinase inhibitors.6,18 The MAPK
inhibitor PD99059 has been shown to block monocyte and eosinophil chemotaxis in response to monocyte chemoattractant protein (MCP)-1 and
eotaxin-1, respectively.8,19 To determine whether these kinases were responsible for TECK-mediated chemotaxis, MOLT4 cells were
pretreated with varying concentrations of wortmannin, a potent PI-3-kinase inhibitor; PD98059, a MAPK inhibitor; or vehicle, DMSO.
Transwell migration assays were performed. As seen in Figure 2A, only wortmannin significantly
inhibited cell migration, an effect that was maximally 5- to 6-fold at
1 µM. PD98059 did not inhibit cell migration. Wortmannin at the same
concentrations did not affect the ability of TECK to induce transient
calcium mobilization (Figure 2B). These data suggests that PI-3 kinase, but not MAPK, plays an important role in TECK-mediated chemotaxis.
PI-3-kinase- and G protein-dependent activation of Akt by TECK A growing body of evidence has demonstrated that Akt is a downstream target of PI-3 kinase.20 Upon activation of growth factor receptors, Akt is phosphorylated and activated in the presence of PI-(3,4,5)P3 (PIP3).21 This lipid product appears to recruit the cytosolic Akt to the membrane compartment by interaction with the pleckstrin homology (PH) domain of Akt. To investigate whether TECK activates Akt, MOLT4 cells were stimulated with TECK, and activation of Akt was assessed at different times by means of phopho-specific Akt antibody. As shown in Figure 3A, an increase in Akt phosphorylation was observed within 10 minutes, and maximal phosphorylation was attained at 30 minutes. This was sustained for 1 hour (data not shown). Wortmannin or pertussis toxin completely abrogated this TECK-enhanced effect, suggesting that activation of Akt is both PI-3-kinase- and Gi-protein-dependent. In terms of loading controls, we found that
complete stripping of the phospho-specific Akt antibody at stringent
conditions rarely occurred, and this interfered with use of the pan-Akt
antibody. However, separate loading of equal amounts of cell lysates
gave uniform intensity of phosphorylation. See an example of this in
Figure 4. It has been recently suggested
that CCR11 is also a receptor for TECK.22 Therefore, we evaluated whether the CCR9/TECK interaction leads to
activation of Akt in heterologous cells. When CCR9 was ectopically expressed in TECK-stimulated Cos7 cells, mainly marginal
activation of endogenous Akt was detected. This low-level activation
was due to low levels of endogenous Akt in the Cos cells (Figure 3B). To enhance expression levels of Akt, an expression vector harboring CCR9 and human Akt together was constructed, and these were
overexpressed in Cos7 cells. When these cells were stimulated with
TECK, activation of Akt was observed in a few seconds and peaked at 20 minutes. A substantial difference in total Akt was noted between the
mock-transfected and the expression vector-transfected cells. At this
time, we do not know whether MOLT4 cells express functional
CCR11. Taken together, these data strongly suggest that
CCR9/TECK interaction leads to the activation of Akt in a PI-3 kinase-
and Gi protein-dependent manner.
CCR9/TECK interaction results in activation of multiple signaling components In the case of the -adrenergic receptor, the chemokine receptor
CXCR2, or the fMLPR receptor (single-letter amino acid codes), it has
been shown that PI-3 kinase  , a subclass of PI-3 kinase which is
believed to be coupled to G protein-coupled receptors including
chemokine receptors, is required for activation of MAPKs, Erk1/Erk2.23,24 Since we demonstrated that Akt is
activated via a PI-3-kinase-dependent pathway upon stimulation of
MOLT4 cells with TECK, this gave us an opportunity to test whether
Erk1/Erk2 is activated by the CCR9/TECK interaction. As seen in Figure
4, a robust activation of Erk1/Erk2 was noticed in 5 minutes, and the
activated state was sustained for 30 minutes. However, in contrast to
Akt, wortmannin did not block the activation of Erk1/Erk2 whereas
pertussis toxin effectively blocked the activation. This suggests that
unlike CXCR2 or the fMLPR in neutrophils, PI-3 kinase may not be
involved in CCR9-mediated activation of Erk1/Erk2 in MOLT4 cells. As in
activation of MAPK, p70 S6 kinase was phosphorylated. However, its
phosphorylation was not inhibited by wortmannin. Rather, it was
inhibited by pertussis toxin.
Activated Akt has been shown to phosphorylate multiple downstream
targets, most of which are involved in cell survival.20 These include Bad, GSK-3 The CCR9/TECK interaction blocks CHX-induced or Fas-mediated apoptosis In addition to the implication of Akt, MAPK, GSK-3, and FKHR in
cell survival signaling,20 it has also been shown that Akt
and MAPK are involved in resistance to Fas-mediated cell
death.25,26 MOLT4 cells express Fas on the cell surface
and are killed by agonistic anti-Fas, CH-11.27 Because Akt
and MAPK are activated via CCR9/TECK interaction, we evaluated whether
Fas-mediated cell death in these cells could be blocked by
CCR9-mediated signaling. MOLT4 cells were cultured in serum-free medium
overnight and stimulated with CH-11 alone, or along with TECK in
the presence of CHX. CHX is known to facilitate Fas-mediated
cell death or, by itself, to be able to kill some T
cells.28 The CXCR4/stromal-derived factor (SDF)-1
interaction was used for comparison. Apoptotic cells were measured by
annexin-V staining. CH-11 treatment was done in the absence of serum,
because MOLT4 cells were highly resistant to CH-11 in the presence of
serum (data not shown), and we hoped to maximize activities of the
signaling components described in Figure 4 upon stimulation of MOLT4
cells with TECK. The percentage listed under each flow analysis
reflects the percentage of apoptotic cells, as seen in the lower
right-hand quadrant. As seen in Figure 5
(an example of flow analysis), CHX (5 µg/mL) or CH-11 (0.5 µg/mL)
induced a rapid apoptosis of MOLT4 cells. Notably, TECK blocked
CHX-induced apoptosis by 80% to 90% and partially rescued the cells
from Fas-mediated cell death. As reported, CHX potentiated Fas-mediated
cell death, which was significantly attenuated by TECK. Because
Z-VAD-FMK, a potent caspase inhibitor, completely blocked both
CHX-induced and Fas-mediated apoptosis, it seemed likely that the
blocking activity displayed by TECK was due to the inhibition of
caspase activity. The in-depth analysis of 3 experiments is presented
in Figure 6A. TECK was capable of blocking CHX-induced apoptosis at varying concentrations of CHX (Figure
6B). The blocking activity was dependent on the dose of TECK. Efficient
blockade of CHX-induced cell death occurred at 1 µg/mL TECK, whereas
the efficiency rapidly declined at 100 ng/mL TECK (Figure 6C). This
blocking activity was not unique to TECK because SDF-1 also efficiently
blocked CHX-induced apoptosis in a dose-dependent manner. Half-maximal
blocking activity was seen even at 10 ng/mL SDF-1. However, SDF-1 did
not appear to effectively block Fas-mediated or CHX-potentiated
Fas-mediated apoptosis (data not shown).
CHX-mediated apoptosis of MOLT4 cells is c-FLIPL-independent, and down-regulation of caspase 3 may account for the antiapoptotic activity displayed by chemokines It has been suggested that CHX induced or potentiated apoptosis by down-regulation of c-FLIPL, an antiapoptotic protein.29,30 This prompted us to investigate whether CHX down-regulated c-FLIPL and, if so, whether TECK up-regulated c-FLIPL. As shown in Figure 7A, CHX-induced or CHX-potentiated Fas-mediated cleavage of PARP was significantly blocked by TECK. However, CHX did not alter expression levels of c-FLIPL. Down-regulation of c-FLIPL was not seen at higher concentrations of CHX with prolonged incubation in which the cleavage of PARP was evident but was not significantly inhibited by TECK (Figure 7B). Combined treatment of MOLT4 cells with CHX and CH-11 induced a cleavage of ICAD and Bcl-XL. Although cleavage of caspase 8 is not evident in this Figure, prolonged exposure revealed that CHX alone or along with CH-11 indeed cleaved caspase 8 and that the cleavage was blocked by TECK (data not shown). Regardless of treatments, TECK did not alter expression levels of Bcl-2 or Bcl-XL. We conclude that CHX may induce apoptosis of MOLT4 cells via a novel pathway, presumably independently of c-FLIPL, and that antiapoptotic Bcl-2 family members were not involved in CCR9- or CXCR4-mediated resistance to CHX-induced apoptosis. Some studies suggest that activation of NF- B
leads to antiapoptosis and that Akt is involved in the activation
process.31,32 It has been shown that SDF-1/CXCR4
interaction results in activation of NF-B.33 We
therefore transfected C3H 10T1/2 (a mouse embryonic fibroblast cell
line) with CCR9 and human immunodeficiency virus (NF-B)3 luciferase reporter gene along with
vector control. The transfected cells were stimulated with TECK (1 µg/mL) or tumor necrosis factor (TNF)- (100 ng/mL) as a control
for 6 hours. Luciferase assay was performed. It was found that TECK was
unable to induce activation of NF-B, but TNF- dramatically
induced the activity, suggesting that activation of NF-B does not
account for the CCR9-mediated cell survival against CHX (data
not shown).
Recently, it was demonstrated that bone marrow neutrophils from
caspase-3-deficient mice were resistant to CHX-induced apoptosis, indicating that caspase-3 expression is essential for this type of cell
death.34 Expression levels of activated caspase 3 were measured by intracellular staining of MOLT4 cells with the use of
cleavage-specific caspase-3 antibody. As shown in Figure
8, both TECK and
SDF-1 significantly down-regulated cleaved caspase 3 in CHX-treated
MOLT4 cells, but did not affect the levels of cleaved caspase 3 in
MOLT4 cells stimulated by combined treatment with CHX and CH-11. CH-11
comparably augmented the levels of cleaved caspase 3 to CHX.
Interestingly, TECK, but not SDF-1, significantly down-regulated
cleaved caspase 3 in Fas-mediated apoptosis, suggesting that CCR9 and
CXCR4 exert differential effects on Fas signaling. This is consistent
with the previous annexin-V data.
Here, we report that PI-3 kinase plays a more important role than MAPK in CCR9-mediated chemotaxis and is important for survival signaling against Fas-mediated apoptosis. In the case of growth factor receptors, MAPK has been strongly implicated in both cell migration and cell survival.18 However, although evidence suggests that activation of chemokine receptors leads to activation of MAPK, the role for MAPK in chemotaxis is controversial.35 It has been shown that the chemotaxis of CXCR2-expressing 293 cells in response to interleukin-8 (IL-8) is not inhibited by PD98059, whereas the chemotaxis of monocytes and eosinophils in response to MCP-1 and eotaxin-1, respectively, is inhibited by PD98059.8,9,19,35 This suggests that cell type is an important factor for determining the role of MAPK in chemokine-mediated chemotaxis. PI-3 kinase represents a multi-enzyme complex that plays a central role
in relaying proximal signals upon activation of growth factor receptors
or G protein-coupled receptors (GPCRs) to downstream signaling
components. Class I PI-3 kinases (PI-3Ks) have been implicated in many
cellular responses downstream of tyrosine kinases. Such cellular
responses involve proliferation, antiapoptosis, and cytoskeletal
rearrangements and cell migration.36 Class IA PI-3K is a
heterodimeric lipid kinase consisting of a p85 regulatory subunit and a
p110 catalytic subunit ( Akt is a serine/threonine kinase that is activated upon ligation of
several cell surface receptors, including the receptors for insulin and
PDGF.39 The biological importance of Akt activation has
been demonstrated by its ability to protect cells from
apoptosis.20 In order for Akt to be activated, Akt needs
to be membrane localized and phosphorylated on a specific serine (S473)
and a specific threonine (T308). Akt contains an amino-terminal PH
domain, which binds PIP3. Binding of PIP3 to the PH domain has been
proposed to facilitate targeting of Akt to membrane compartments.
Because PIP3 is an end product of PI-3K, Akt activation necessarily
depends on PI-3K. Several lines of evidence have suggested that
chemokine receptor signaling activates Akt. These include IL-8, SDF-1,
MIP-1
The authors thank Audrey Carson for typing the manuscript and Dr Byoung H. Kim for instruction on how to use the Excel program.
Submitted October 24, 2000; accepted April 5, 2001.
Supported by US Public Health Service grants R01 HL56416 and RO1 DK53674 from the National Institutes of Health (H.E.B), and a Young Investigator Award of the Core Centers of Excellence in Molecular Hematology (P30 DK49218) to Indiana University School of Medicine (B.-S.Y.).
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: Byung-S. Youn, KOMED Institute for Life Science, Graduate School of Biotechnology, Korea University, Rm 640, 1,5-ka, Anam-dong, Sungbuk-ku, Seoul, Korea.
1. Ward SG, Bacon KB, Westwick J. Chemokine and T lymphocytes: more than attraction. Immunity. 1998;9:1-11[CrossRef][Medline] [Order article via Infotrieve]. 2. Rossi D, Zlotnik A. The biology of chemokines and their receptors. Ann Rev Immunol. 2000;18:217-242[CrossRef][Medline] [Order article via Infotrieve].
3.
Vila-Coro AJ, Rodriguez-Frade JM, Martin De Ana A, Moreno-Ortiz MC, Martinez-AC, Mellado M.
The chemokine SDF-1alpha triggers CXCR4 receptor dimerization and activates the JAK/STAT pathway.
FASEB J.
1999;13:1699-1710
4.
Majka M, Janowska-Wieczorek A, Ratajczak J, Kowalska MA, Vilaire G, Pan ZK, Honczarenko M, Marquez LA, Poncz M, Ratajczak MZ.
Stromal-derived factor 1 and thrombopoietin regulate distinct aspects of human megakaryopoiesis.
Blood.
2000;96:4142-4151
5.
Ganju RK, Dutt P, Wu L, et al.
Beta-chemokine receptor CCR5 signals via the novel tyrosine kinase RAFTK.
Blood.
1998;91:791-797 6. Turner L, Ward SG, Westwick J. RANTES-activated human T lymphocytes: a role for phosphoinositide 3-kinase. J Immunol. 1995;155:2437-2444[Abstract].
7.
Turner SJ, Domin J, Waterfield MD, Ward SG, Westwick J.
The CC chemokine monocyte chemotactic peptide-1 activates both the class I p85/p110 phosphatidylinositol 3-kinase and the class II P13K-C2alpha.
J Biol Chem.
1998;273:25987-25995 8. Kampen GT, Stafford S, Adachi T, et al. Eotaxin induces degranulation and chemotaxis of eosinophils through the activation of ERK2 and p38 mitogen-activated protein kinases. Blood. 2000;15:1911-1917.
9.
Boehme SA, Sullivan SK, Crowe PD, et al.
Activation of mitogen-activated protein kinase regulates eotaxin-induced eosinophil migration.
J Immunol.
1999;163:1611-1618
10.
Wang JF, Park IW, Groopman JE.
Stromal cell-derived factor-1alpha stimulates tyrosine phosphorylation of multiple focal adhesion proteins and induces migration of hematopoietic progenitor cells: roles of phosphoinositide-3 kinase and protein kinase C.
Blood.
2000;95:2505-2513 11. Youn BS, Mantel C, Broxmeyer HE. Chemokines, chemokine receptors and hematopoiesis. Immunol Rev. 2000;177:150-174[CrossRef][Medline] [Order article via Infotrieve].
12.
Youn BS, Kim CH, Smith FO, Broxmeyer HE.
TECK, an efficacious chemoattractant for human thymocytes, uses GPR-9-6/CCR9 as a specific receptor.
Blood.
1999;94:2533-2536
13.
Zaballos A, Gutierrez J, Varona R, Ardavin C, Marquez G.
Cutting edge: identification of the orphan chemokine receptor GPR-9-6 as CCR9, the receptor for the chemokine TECK.
J Immunol.
1999;162:5671-5675
14.
Zabel BA, Agace WW, Campbell JJ, et al.
Human G protein-coupled receptor GPR-9-6/CC chemokine receptor 9 is selectively expressed on intestinal homing T lymphocytes, mucosal lymphocytes, and thymocytes and is required for thymus-expressed chemokine-mediated chemotaxis.
J Exp Med.
1999;190:1241-1256
15.
Rommel C, Clarke BA, Zimmermann S, et al.
Differentiation stage-specific inhibition of the Raf-MEK-ERK pathway by Akt.
Science.
1999;286:1738-1741 16. Vanhaesebroeck B, Jones GE, Allen WE, et al. Distinct PI(3)Ks mediate mitogenic signalling and cell migration in macrophages. Nat Cell Biol. 1999;1:69-71[CrossRef][Medline] [Order article via Infotrieve].
17.
Yujiri T, Ware M, Widmann C, et al.
MEK kinase 1 gene disruption alters cell migration and c-Jun NH2-terminal kinase regulation but does not cause a measurable defect in NF-kappa B activation.
Proc Natl Acad Sci U S A.
2000;97:7272-7277
18.
Vicente-Manzanares M, Rey M, Jones DR, et al.
Involvement of phosphatidylinositol 3-kinase in stromal cell-derived factor-1 alpha-induced lymphocyte polarization and chemotaxis.
J Immunol.
1999;163:4001-4012 19. Yen H, Zhang Y, Penfold S, Rollins BJ. MCP-1-mediated chemotaxis requires activation of non-overlapping signal transduction pathways. J Leukoc Biol. 1997;61:529-532[Abstract].
20.
Datta SR, Brunet A, Greenberg ME.
Cellular survival: a play in three Akts.
Genes Dev.
1999;13:2905-2927 21. Klippel A, Kavanaugh WM, Pot D, Williams LT. A specific product of phosphatidylinositol 3-kinase directly activates the protein kinase Akt through its pleckstrin homology domain. Mol Cell Biol. 1997;17:338-344[Abstract].
22.
Gosling J, Dairaghi DJ, Wang Y, et al.
Cutting edge: identification of a novel chemokine receptor that binds dendritic cell- and T cell-active chemokines including ELC, SLC, and TECK.
J Immunol.
2000;164:2851-2856
23.
Lopez-Llasaca M, Crespo P, Giuseppe Pellici P, Gutkind JS, Wetzker R.
Linkage of G protein-coupled receptors to the MAPK signaling pathway through PI 3-kinase gamma.
Science.
1997;275:394-397
24.
Sasaki T, Irie-Sasaki J, Jones RG, et al.
Function of P13K 25. Hausler P, Papoff G, Eramo A, Reif K, Cantrell DA, Ruberti G. Protection of CD95-mediated apoptosis by activation of phosphatidylinositide 3-kinase and protein kinase B. Eur J Immunol. 19 |
Top
Abstract
Introduction
) in the presence of TECK (
)
or Z-VAD-FMK (
). Data are expressed as the mean with SD obtained
from at least 3 different experiments. (C and D) Dose-dependent
effect of TECK or SDF-1 on CHX-induced apoptosis. Data presented are
from at least 3 different experiments.
). PI-3K is activated by the
interaction of the p85 regulatory subunit with phosphorylated tyrosine
residues on activated growth factor receptors. The binding of the p85
regulatory subunit to upstream signaling molecules causes the p110
catalytic subunit to be membrane localized and activated. Class IB
PI-3K (PI-3K