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,§
*
Virginia Mason Research Center, Seattle, WA 98101;
Department of Molecular and Cellular Biology, Roswell Park Cancer Institute, Buffalo, NY 14263; and Departments of
Immunology and
§
Biological Structure, University of Washington, Seattle, WA 98195
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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-chain; IL-7R is paired with
the common 
-chain (c) in the IL-7 receptor complex and the unique
TSLP-R chain in the TSLP receptor complex. Although TSLP and IL-7 both
induce tyrosine phosphorylation of the transcription factor Stat5, only
IL-7-mediated signal transduction could be associated with activation
of Janus family kinases (Jaks). Because Stat5 phosphorylation following
cytokine stimulation is generally mediated by Jaks, the lack of Jak
activation after TSLP treatment suggested the possibility that
tyrosine-phosphorylated Stat5 may be nonfunctional. Herein, we
demonstrate that TSLP induces a functional Stat5 transcription factor
in that TSLP stimulation results in Stat5-DNA complex formation and
transcription of the Stat5-responsive gene CIS. We also show that the
TSLP receptor complex is functionally reconstituted using TSLP-R and
IL-7R and that TSLP-mediated signal transduction requires Stat5.
Moreover, TSLP-mediated signaling is inhibited by suppressor of
cytokine signaling (SOCS)-1 and a kinase-deficient version of Tec but
not by kinase-deficient forms of Jak1 and Jak2. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-chain (IL-7R) have developmentally arrested B cells (1, 2). We have recently identified a novel cytokine designated
thymic stromal lymphopoietin
(TSLP)3. TSLP is
similar to IL-7 in that both cytokines can costimulate thymocytes and
mature T cells, support B lymphopoiesis in long-term cultures of fetal
liver cells, and sustain the factor-dependent, fetal liver-derived
NAG8/7 pre-B cell line (3, 4, 5). But, while TSLP facilitates
B cell development to the
B220+/IgM+ immature B cell
stage, IL-7 supports development only to the less mature
B220+/IgM- pre-B cell
stage (5).
The similarity between TSLP and IL-7 extends to their receptor
complexes. The IL-7 receptor complex is composed of IL-7R and the
IL-2R -chain (also known as the common -chain or c; Refs.
6 and 7). The TSLP receptor complex also
employs IL-7R, but here IL-7R associates with a novel receptor
chain designated TSLP-R
(5).4 The
sharing of cytokine receptor subunits by different cytokines is well
documented. IL-2Rß is shared by IL-2 and IL-15 (8);
IL-4R is shared by IL-4 and IL-13 (9); and common
ß-chain is shared by IL-3, IL-5, and GM-CSF (10).
Moreover, common -chain is shared by the receptors for IL-2, IL-4,
IL-7, IL-9, and IL-15 (11) but is not a component of the
TSLP receptor complex (5).
A prevalent feature of cytokine-mediated signal transduction is activation of the Janus family kinase (Jak)/Stat pathway. Seven mammalian Stat proteins (Stat1 to -4, -5a, -5b, and -6) and four Janus protein kinases (Jak1 to -3 and Tyk2) have been identified (for review see Ref. 11). Cytokine-induced receptor aggregation activates the Jak/Stat pathway by bringing the receptor-associated Janus kinases (Jaks) into mutual proximity, thus allowing for transphosphorylation and activation. The activated Jaks then phosphorylate the cytokine receptor, thereby providing a docking site for SH2 containing proteins, including members of the Stat transcription factor family. Upon binding to the phosphorylated cytokine receptor, Stats themselves become tyrosine phosphorylated, are released from the receptor complex, dimerize, and translocate to the nucleus where they bind DNA with sequence specificity to initiate transcription of target genes.
Our previous investigations of the biochemical signals induced by TSLP
and IL-7 revealed that TSLP and IL-7 both trigger tyrosine
phosphorylation of Stat5a and Stat5b (the products of two distinct
genes that are 95% identical). Although IL-7 stimulation leads to
tyrosine phosphorylation and subsequent activation of Jak1 and Jak3, we
were unable to detect activation of any of the four known Jak kinases
following TSLP stimulation (5). This lack of Jak
activation suggested the possibility that phosphorylated Stat5 in
TSLP-treated cells may be nonfunctional. Here we show that TSLP
stimulation results in a functional Stat5 transcription factor. Using
the TSLP and IL-7 dependent NAG8/7 pre-B cell line (3), we
demonstrate that TSLP induces Stat5-DNA complex formation,
transcription of the Stat5-responsive gene CIS, and regulation of the
luciferase reporter gene when fused to the CIS promoter. Furthermore,
we show that the TSLP receptor complex is functionally reconstituted
using TSLP-R and IL-7R and that TSLP-mediated signal transduction
requires Stat5, is blocked by overexpression of suppressor of cytokine
signaling (SOCS)-1 or kinase-deficient Tec, yet is unaffected by
overexpression of kinase-deficient Jak1 or Jak2.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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NAG8/7 cells (3) were maintained in RPMI 1640 complete medium (Life Technologies, Grand Island, NY) supplemented with 10% FCS, 45 U/ml penicillin, 45 µg/ml streptomycin, 34 mM HEPES, 3.8 mM L-glutamine, 1 mM sodium pyruvate, 20 µM 2-ME, and, unless otherwise specified, 1 ng/ml murine IL-7. The human IL-2-dependent T cell line CTLL-2 was maintained as above with the addition of 50 U/ml human rIL-2 (Chiron, Emeryville, CA). Murine IL-7 and TSLP were obtained by transfection of the p3XAG cell line with cDNAs to the appropriate cytokine in the BCMGS expression vector (12). Cytokine concentrations were determined by comparison with purified cytokine. Human hepatoma HepG2 cells (13) were cultured in DMEM containing 10% FCS and antibiotics.
EMSA
NAG8/7 cells were washed three times in PBS, cultured without
cytokine for 4 h, and then stimulated at 106
cells/ml with either 10 ng/ml IL-7, 24 ng/ml TSLP, or left
unstimulated. After 20- and 60-min incubations, nuclear extracts were
prepared as described (14). A probe for Stat DNA-binding
activity was generated by annealing incompletely overlapping
oligonucleotides (5'-TCGAGTATTTCCCAGAAAAGGAAC-3' and
5'-AGCTGTTCCTTTTCTGGGAAATAC-3') corresponding to the sense and
antisense strands of the Stat-responsive DNA element from the FcRI
promoter (15). The probe was
32P-labeled by an end-filling T4 polymerase
reaction and purified with Bio-gel P30 spin columns (Bio-Rad, Hercules,
CA). Nuclear extracts (
2–4 µg total protein) in 50 mM KCl, 15 mM
HEPES (pH 7.9), 15% glycerol, 0.75 mM DTT, and 87.5 mg/ml poly(dI-dC)
were incubated with the 32P-labeled probe (5000
cpm) at room temperature for 20 min. For the supershift reactions, 100
ng of Stat5 rabbit polyclonal Abs (Santa Cruz Biotechnology, Santa
Cruz, CA) or 100 ng rabbit polyclonal preimmune serum were added to the
reaction after the first 10 min. Samples were electrophoresed at 4°C
through a 4% native polyacrylamide, 0.25x TBE gel followed by
autoradiography.
Northern blots
NAG8/7 and CTLL-2 cells were washed, starved, and stimulated as was done for the EMSA experiments, except that CTLL-2 cells were stimulated with 50 U/ml IL-2. After the indicated incubation times, total RNA was isolated using Trizol reagent (Life Technologies). Twenty micrograms of total RNA from each sample was denatured for 5 min at 55°C in 50% DMSO, 0.5x TBE, and 0.05% bromphenol blue and then electrophoresed through a 1% agarose, 0.5x TBE gel. The size-separated RNA was capillary transferred to Transfer-IT plus nylon membrane (CPG, Lincoln Park, NJ) and cross-linked to the membrane by UV irradiation. Blots were prehybridized at 42°C in a heat-sealed pouch with hybridization buffer (50% formamide, 5x SSC, 1x Denhart’s solution, 20 mM mono and dibasic sodium phosphate, 100 µg/ml salmon sperm DNA, 10% dextran sulfate, and 1% SDS). DNA probes were 32P-labeled using the high-prime labeling kit (Boehringer Mannheim, Indianapolis, IN) and purified with Bio-gel P30 spin columns (Bio-Rad). After an overnight hybridization at 42°C, blots were washed several times, each time with increasing stringency. The final wash, before autoradiography, consisted of 0.1x SSC, 0.1% SDS at ambient temperature (CIS probe) or 65°C (oncostatin M (OSM) and GAPDH probes). Before the blots were reprobed, they were stripped by immersion in boiling water for 5 min.
Transient transfections and luciferase assay in NAG8/7 cells
The -404 CIS promoter-luciferase construct and -694 OSM promoter-luciferase construct have been described (16, 17). After a 5-h incubation in complete medium lacking cytokine, 10 µg of the CIS- or OSM-luciferase constructs were transiently transfected into 3 x 107 NAG8/7 or CTLL-2 cells by electroporation (250V, 960 µF) using a Genepulser (Bio-Rad). Following transfection, cells were washed in complete medium and divided into individual wells of a 12-well culture plate. Cells were cultured in duplicate in the presence of either no cytokine, 1 ng/ml IL-7, 21 ng/ml TSLP, or 100 U/ml IL-2. After 5 h of culture, cells were washed with PBS and lysed for 15 min at room temperature with 1x passive lysis buffer (Promega, Madison, WI). Cell lysates were centrifuged, and supernatants were examined for luciferase activity as determined by measuring light emitted for 30 s using a luminometer (Lumat LB9507, EG+G Berthold, Nashua, NH) and Promega’s luciferase assay reagent.
Transient transfection and chloramphenicol acetyl transferase (CAT) assay in HepG2 cells
Constructs.
p(8xHRRE)-CAT contains eight tandem copies of the 27 bp hemopoietin
receptor response element in pCAT (18). Expression vectors
for the following molecules have been described: IL-7R and c
(6), the internal transfection control mouse major urinary
protein (MUP; Ref. 19), rat Stat5b (20), the
truncated, dominant-negative mutant Stat5b
40C (21), the
murine, kinase-deficient form of Jak2 (22), and the human,
kinase-deficient form of Tec called TecKM
(23). TSLP-R4 and murine SOCS-1
(24, 25, 26, 27) were cloned into the pcDNA3.1 expression vector
(Invitrogen, Carlsbad, CA) using a PCR-based approach, and the
sequences were verified by DNA sequence analysis. A kinase-deficient
form of murine Jak1 (28, 29), in which the codon for the
conserved lysine residue in the ATP-binding site (aa 907) was changed
to an aspartic acid codon, was generated using PCR-based mutagenesis.
The mutant Jak1 construct was verified by DNA sequence analysis and
cloned into the expression vector pDC302 (30). The human
C-terminal Src kinase (Csk; Ref. 31) was expressed in
pDC302 as well (30).
Transfection and analysis. HepG2 cells were transfected by the calcium phosphate method (32) using 20 µg/ml total plasmid DNA and 2–3 x 105 cells. After an overnight recovery period, cultures were released from the plate with trypsin and divided into six-well culture plates. After an additional 24 h, subcultures were treated for 24 h with serum-free medium containing 100 ng/ml IL-7, TSLP, IL-6, or OSM. Medium was then collected and subjected to immunoelectrophoresis to quantitate expression of the cotransfected control MUP plasmid. CAT activity for each culture was determined, normalized to the amount of MUP expression, and calculated relative to the control cultures in each experimental series (defined as 1.0). Data are presented as the mean of three to five trials.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The cytokines TSLP and IL-7 both employ IL-7R as part of their
receptor complexes (5, 6).4 In addition,
both cytokines trigger tyrosine phosphorylation of the two Stat5
isoforms, Stat5a and Stat5b. Yet, in the case of TSLP, Stat5 tyrosine
phosphorylation occurs in the absence of detectable Jak activation
(5). This finding suggests not only that Stat5 activation
may be occurring by an alternate mechanism, but also that TSLP and IL-7
signal via different biochemical pathways despite sharing the IL-7R
receptor subunit. The fact that no detectable Jak activation occurs
following TSLP treatment does raise the possibility, however, that,
although TSLP induces Stat5 tyrosine phosphorylation, a functionally
active Stat5 transcription factor may not result.
To determine whether TSLP-activated Stat5 enters the nucleus and binds
a Stat-responsive DNA element, we performed EMSA. Nuclear extracts were
generated from NAG8/7 cells that were treated with TSLP or IL-7 for 60
min (or 20 min, data not shown), and these extracts were tested for
DNA-binding activity using the FcRI Stat-responsive DNA element
(15). Both TSLP- and IL-7-stimulated cell extracts showed
specific DNA-binding activity whereas extracts from unstimulated cells
did not (Fig. 1
). Furthermore, addition
of an antisera to Stat5 (which recognizes both Stat5a and Stat5b
isoforms) supershifted these complexes. The lower level of complexed
Stat5 seen in TSLP-treated cells, as compared with IL-7-treated cells,
is consistent with our previous report that TSLP induced tyrosine
phosphorylation of Stat5 less efficiently and with delayed kinetics
relative to IL-7 (5). Thus, TSLP induces changes in Stat5
that result in nuclear translocation and DNA-binding activity, despite
the lack of concomitant Jak activation.
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A functional Stat5 transcription factor must do more than
translocate to the nucleus and bind DNA. It must be able to initiate
transcription of Stat5 target genes. The promoters of both the CIS and
OSM genes contain Stat5 target sequences, and both genes are induced
upon Stat5 activation by IL-2, IL-3, and erythropoietin (17, 33, 34). Therefore, we used Northern blot analysis to test whether
CIS and OSM were transcribed following TSLP stimulation. As shown in
Fig. 2, CIS transcription was induced in
NAG8/7 cells by both TSLP and IL-7, although to a lesser extent by
TSLP. When the Northern blots were stripped and reprobed for OSM
expression, OSM transcripts were detected in neither the TSLP- nor the
IL-7-stimulated lanes. OSM transcripts were readily apparent, however,
in control lanes containing comparable amounts of RNA isolated from
IL-2-stimulated CTLL-2 cells (data not shown). These data suggest that
the OSM gene is either not active or not regulated by these cytokines
in NAG8/7 cells.
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). However, luciferase activity was
induced in IL-2-treated CTLL-2 cells after transfection with the
OSM-luciferase construct, indicating that this reporter gene is
functional (data not shown). In NAG8/7 cells there was less activity
induced from the CIS reporter construct following TSLP stimulation than
following IL-7 stimulation, and this reduction was proportional to the
levels of Stat5 phosphorylation previously seen (5). We do
not attribute the lower luciferase activity seen after TSLP stimulation
to limiting amounts of cytokine because using 10 ng/ml, instead of 21
ng/ml, of TSLP gave identical results, suggesting that both levels were
saturating (data not shown). Together, the Northern blot and luciferase
assay results reveal that TSLP- and IL-7-mediated signal transduction
include transcription of the Stat5 target gene CIS, but not OSM, and
uncover another similarity in the signaling pathways of these
cytokines. Furthermore, these results highlight the fact that, although
CIS and OSM are both regulated by Stat5, they are regulated
differently, and Stat5 activation is not sufficient for OSM
induction.
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To further address the role of Stat5 in TSLP-mediated signal transduction, we used reconstitution of defined receptor combinations in a cell culture system. This system involves cotransfection of the human hepatoma cell line HepG2 with cDNA clones encoding receptor subunits, as well as a CAT-reporter plasmid containing eight copies of the cytokine-inducible hemopoietin receptor response element (HRRE-CAT; Refs. 6 and 18). The ability of the transfected cells to signal when treated with the appropriate cytokine is measured by CAT activity.
As shown in Fig. 4A, cells
cotransfected with TSLP-R and IL-7R generated a TSLP-specific signal
only when cDNAs encoding Stat5b (or Stat5a, data not shown) were
included in the cotransfection. On the other hand, cells cotransfected
with TSLP-R and c were unable to activate the cytokine-inducible
promoter irrespective of which cytokine was used and whether or not
Stat5b was included in the cotransfection. Moreover, HepG2 cells
cotransfected with IL-7R and c responded to IL-7 in the absence
of cotransfected Stat5b (consistent with earlier findings by Ziegler et
al.; Ref. 6), but the response was enhanced if Stat5b
cDNAs were included. These results suggest several things. First, they
demonstrate that a functional TSLP receptor complex can be
reconstituted with TSLP-R and IL-7R. Second, HepG2 cells express
abundant amounts of Stat1 and Stat3, but only very low levels of Stat5.
Therefore, these results suggest that TSLP-R is unable to engage
endogenous signal transduction pathways in these cells, including those
mechanisms used by the IL-7 receptor and other hemopoietin receptors,
unless Stat5 is also provided. In contrast, IL-7-mediated signal
transduction is able to utilize the endogenous signaling pathways,
which may include Stat1 and/or Stat3, in addition to experimentally
enhanced Stat5. This finding is consistent with previous reports of
IL-7-mediated activation of Stat1, Stat3, and Stat5 (35, 36).
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A, we
tested whether cotransfection of a dominant-negative version of Stat5b
(Stat5b40C) would inhibit reconstituted TSLP signal transduction in
the HepG2 system (20). For these experiments, HepG2 cells
were cotransfected with cDNAs encoding TSLP-R, IL-7R, and Stat5b. In
addition, increasing amounts of the dominant-negative Stat5b construct,
or vector alone, were included. As shown in Fig. 4B,
expression of dominant-negative Stat5b inhibited TSLP-mediated
signaling in a dose-dependent manner. Together, these data support the
premise that the TSLP receptor complex is composed of the TSLP-R and
IL-7R subunits and that TSLP requires Stat5 for effective gene
induction. Protein tyrosine kinase involvement in TSLP-mediated signal transduction
TSLP receptor engagement results in tyrosine phosphorylation, and
subsequent activation, of Stat5 in the absence of concomitant Jak
kinase activation (5). To confirm and extend these
findings, we used the HepG2 receptor reconstitution system to
overexpress kinase-deficient versions of Jak1 or Jak2 (two Jaks that
are endogenously expressed in HepG2 cells; Ref. 22). As
shown in Fig. 5A,
overexpression of inactive Jak1 was unable to inhibit TSLP-mediated
signaling, thus providing further evidence that Jak1 is not a required
component of the TSLP signaling pathway. On the other hand,
overexpression of kinase-deficient Jak1 did reduce IL-7- and
OSM-mediated CAT activation, corroborating previous findings that Jak1
is necessary for IL-7- and OSM-mediated signaling
(37, 38, 39). Similarly, overexpression of kinase-deficient
Jak2 had no appreciable effect on TSLP, IL-7, IL-6, and OSM signaling
but did decrease thrombopoietin-induced CAT activity in
c-mpl-transfected cells (data not shown and Ref. 22).
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B, expression of
SOCS-1 abolished TSLP-mediated induction of CAT activity in HepG2 cells
cotransfected with TSLP-R, IL-7R, Stat5b, and SOCS-1. This result,
combined with our findings on the lack of Jak kinase involvement in
TSLP-mediated signaling (Ref. 5 and Fig. 5A),
implicates a kinase that is inhibited by SOCS-1, such as a Tec family
kinase and/or a novel Jak kinase, in the TSLP-signaling pathway.
Conversely, overexpression of Csk, a negative regulator of the Src
family kinases (31, 40), had no appreciable effect on TSLP
receptor signaling, suggesting that in this assay Src family kinases
are not involved (Fig. 5B). To further test for the
involvement of a Tec family kinase, we cotransfected a kinase-deficient
version (TecKM) of the founding family member Tec
(23). Overexpression of TecKM
decreased TSLP-mediated signaling, thereby further implicating Tec or a
similar protein tyrosine kinase in the TSLP signaling pathway (Fig. 5C). |
Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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. The differences
may be due to the presence of receptor subunits unique to each complex:
c for the IL-7 receptor (6, 7) and TSLP-R for the TSLP
receptor (Ref. 5 , Fig. 4A).4 The relationship between TSLP
and IL-7 is suggestive of that between IL-4 and IL-13. The functional
IL-4 receptor complex includes the IL-4R -chain and c whereas the
IL-13 receptor complex includes the unique IL-13R plus the IL-4R
-chain. IL-4 and IL-13 have overlapping, as well as distinct,
biochemical and biological activities (9). A similar story
is emerging for TSLP and IL-7.
Here we have shown that TSLP, like IL-7, functionally activates the
Stat5 transcription factor. Specifically, TSLP or IL-7 treatment of the
NAG8/7 cell line induces Stat5 tyrosine phosphorylation
(5) and DNA binding activity (Fig. 1). Moreover, treatment
with these cytokines results in transcription of CIS and activation of
a Stat5-responsive reporter gene. Although TSLP and IL-7 both induce
transcription of CIS, they do not induce transcription of OSM, another
Stat5-responsive gene (Fig. 2 and 3). Thus, although Stat5 activation
results in expression of CIS and OSM in Ba/F3 cells (41),
our results with TSLP and IL-7 demonstrate that Stat5 activation is not
sufficient for OSM expression. Exactly why TSLP and IL-7 induce CIS but
not OSM transcription, while IL-2, IL-3, and erythropoietin activate
the transcription of both genes, remains to be determined (17, 33, 34). Recently it has become apparent that a number of
factors, in addition to Stat tyrosine phosphorylation, can regulate the
degree and specificity of Stat-regulated transcription (reviewed in
Ref. 42). These include Stat serine phosphorylation; the
number, nucleotide composition, and spacing of Stat binding sequences
present in the promoters of target genes; hetero- vs homodimerization
and tetramerization of the activated Stats; and the possible necessity
of additional proteins.
We have previously shown that Stat5 tyrosine phosphorylation occurs in TSLP-treated cells without concomitant tyrosine phosphorylation of any of the four known Jaks (5). Here, we have used overexpression of kinase-deficient versions of Jak1 and Jak2 to confirm and extend those findings. Clearly, TSLP-mediated signaling activates Stat5, but it still remains to be answered how Stat5 gets phosphorylated. Although there are numerous examples of cytokine stimulation leading to Jak-mediated Stat activation (reviewed in Ref. 11), there are only a few reports suggesting Jak-independent Stat activation. For example, Stat activation following stimulation with epidermal growth factor requires intrinsic kinase activation of the epidermal growth factor receptor but does not require Jak1 phosphorylation (43). Moreover, Saharinen et al. have shown that expression in COS cells of Bmx, a member of the Tec tyrosine kinase family, induces activation of endogenous Stat1, Stat3, and Stat5 without the activation of endogenous Jak kinases (44).
The ability of Bmx to induce Stat activation without concomitant Jak activation is of particular interest considering our findings that overexpression of SOCS-1, a negative regulator of Jak and Tec protein kinases (24, 25, 26, 27), or a kinase-deficient version of the protein tyrosine kinase Tec (23) inhibited TSLP-mediated signal transduction whereas overexpression of Csk, a negative regulator of Src family kinases (31, 40), had no appreciable effect on TSLP signaling. In addition, the Src family kinase inhibitor PP1 was unable to inhibit TSLP-mediated Stat5 tyrosine phosphorylation (S. D. Levin, unpublished observations). Together, these results suggest the possibility that a Tec, and not a Src, family kinase is involved in TSLP-mediated Stat5 activation.
Using the HepG2 receptor reconstitution system, we have also shown that TSLP signaling requires Stat5 and cannot utilize Stat1 and Stat3. Although these experiments were not performed in lymphocyte or lymphocyte progenitor cells, results in HepG2 cells examining IL-2, IL-4, IL-6, IL-7, and GCSF signal transduction have been faithfully recapitulated in lymphocytes (6, 18, 45). In addition, we detected TSLP-mediated CIS induction using the HepG2 receptor reconstitution system (unpublished observations), which is consistent with the results in NAG8/7 pre-B cells presented herein. Since our results suggest that TSLP cannot utilize Stat1 or Stat3 and since there was no detectable decrease in the number of thymocytes or peripheral B cells in mutant mice lacking both Stat5a and Stat5b (although the ability of IL-7 to induce bone marrow colony formation was compromised; Ref. 46), one might predict that TSLP will be found unnecessary for B lymphopoiesis. Alternatively, TSLP may be discovered to signal through an additional, non-Stat pathway.
The functional TSLP receptor complex is composed of TSLP-R and
IL-7R. Engagement of this complex activates Stat5 and induces CIS
transcription through a novel mechanism. Our data suggest Stat5
activation may be through a Tec family kinase. Identification of the
kinase that phosphorylates Stat5 and identification of the other
components of TSLP-mediated signal transduction remain challenges for
future studies.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Steven F. Ziegler, Virginia Mason Research Center, 1201 9th Avenue, Seattle, WA 98101. E-mail address:
3 Abbreviations used in this paper: TSLP, thymic stromal lymphopoietin; c, common -chain; Jak, Janus family kinase; OSM, oncostatin M; HRRE, hemopoietin receptor response element; CAT, chloramphenicol acetyl transferase; MUP, mouse major urinary protein; Csk, C-terminal Src kinase; SOCS, suppressor of cytokine signaling.
4 L. S. Park, U. Martin, K. Garka, B. Gliniak, J. P. DiSanto, W. Muller, D. A. Largaespada, N. G. Copeland, N. A. Jenkins, A. G. Farr, S. F. Ziegler, P. J. Morrissey, R. Paxton, and J. Sims. Cloning of the murine TSLP receptor: formation of a functional heteromeric complex requires IL-7 receptor. Submitted for publication.
Received for publication May 28, 1999. Accepted for publication September 21, 1999.
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 N. Carpino, W. E. Thierfelder, M.-s. Chang, C. Saris, S. J. Turner, S. F. Ziegler, and J. N. Ihle Absence of an Essential Role for Thymic Stromal Lymphopoietin Receptor in Murine B-Cell Development Mol. Cell. Biol., March 15, 2004; 24(6): 2584 - 2592. [Abstract] [Full Text] [PDF]
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M. J. Osborn, P. L. Ryan, N. Kirchhof, A. Panoskaltsis-Mortari, F. Mortari, and K.-S. R. S. Tudor Overexpression of murine TSLP impairs lymphopoiesis and myelopoiesis Blood, February 1, 2004; 103(3): 843 - 851. [Abstract] [Full Text] [PDF]
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D. E. Isaksen, H. Baumann, B. Zhou, S. Nivollet, A. G. Farr, S. D. Levin, and S. F. Ziegler Uncoupling of Proliferation and Stat5 Activation in Thymic Stromal Lymphopoietin-Mediated Signal Transduction J. Immunol., April 1, 2002; 168(7): 3288 - 3294. [Abstract] [Full Text] [PDF]
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 C. J. Greenhalgh and D. J. Hilton Negative regulation of cytokine signaling J. Leukoc. Biol., September 1, 2001; 70(3): 348 - 356. [Abstract] [Full Text] [PDF]
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P. A. Reche, V. Soumelis, D. M. Gorman, T. Clifford, M.-r. Liu, M. Travis, S. M. Zurawski, J. Johnston, Y.-J. Liu, H. Spits, et al. Human Thymic Stromal Lymphopoietin Preferentially Stimulates Myeloid Cells J. Immunol., July 1, 2001; 167(1): 336 - 343. [Abstract] [Full Text] [PDF]
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K. J. Hare, E. J. Jenkinson, and G. Anderson An Essential Role for the IL-7 Receptor During Intrathymic Expansion of the Positively Selected Neonatal T Cell Repertoire J. Immunol., September 1, 2000; 165(5): 2410 - 2414. [Abstract] [Full Text] [PDF]
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 D. Krebs and D. Hilton SOCS: physiological suppressors of cytokine signaling J. Cell Sci., January 8, 2000; 113(16): 2813 - 2819. [Abstract] [PDF]
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