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Department of Immunology, University of Washington, Seattle, WA 98195
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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(LT), LT
, TNF-, and
TNF-. With the exception of LT, these molecules are type II
membrane proteins characterized by a homologous sequence in the
extracellular C terminus. The crystallization of TNFR1, LT, and TNF
illustrates that the receptor-ligand interactions between these
families involve one homotrimeric ligand binding to three cross-linked
receptors organized in a predictable cluster (3, 4).
The TNF family members 4-1BBL, CD27L, CD30L, CD40L, FasL, OX40L, and
TNF- all exhibit the capacity to reverse signal, to transduce a
signal inward upon binding specific receptors (2, 5, 6, 7, 8).
Bipolar signaling may turn out to be the norm for this family of
molecules, thereby blurring the distinction between receptor and
ligand. Although it is known that these ligands are mainly
membrane-bound molecules that primarily interact with their receptors
by direct cell-cell contact, little is known about the mechanism of
reverse signaling. It is likely that the cytoplasmic domains of these
molecules are involved in signal transduction, as the cytoplasmic tails
of TNF family members are distinct from each other yet conserved across
species (1).
Our work has focused on bipolar signaling by FasL. Previous in vitro studies from our laboratory provide compelling evidence that FasL acts as a costimulatory receptor in CD8+ T cells. From our initial studies (7), three lines of evidence support a role for FasL in positive reverse signaling. First, multiple CD8+ CTL cell lines, independently derived from FasL- gld mice on a C57BL/6 (B6) background, exhibit depressed proliferation upon alloantigenic stimulation compared with CTL lines derived from wild-type B6 mice or Fas- lpr mice. This was not due to differences in kinetics of proliferation among the cell lines, overexpression of Fas on the gld cell lines (which could result in increased apoptosis of these cells), or a general defect in signaling in the gld cells. Our findings suggest that the presence of FasL is important for inducing maximal proliferation of CTLs. The second major line of evidence implicating FasL in positive signaling used soluble FasIgG fusion protein (a chimeric molecule of human IgG and the ligand-binding portion of murine Fas) to block cell surface FasL-Fas interactions. In a dose-dependent manner, soluble FasIgG, but not isotype-matched control human IgG, attenuated the proliferation of wild-type B6 CTLs down to the level achieved by gld responders. A third line of evidence showed that cross-linking FasL with plate-bound FasIgG, in conjunction with suboptimal amounts of anti-CD3, delivered a costimulatory signal for proliferation to FasL+ but not FasL- CTL. We extended these initial studies to examine the role of FasL costimulation in CD8+ vs CD4+ peripheral T cells (9). We found that naive CD4+ T cells can also respond to FasL costimulation when Fas-mediated death is prevented, and that the observed differences between CD4+ and CD8+ T cells stem partly from the differential control of FasL expression in these two cell types. FasL costimulation occurs very early during the course of an MLC, at a time when FasL expression is induced on both CD4+ and CD8+ T cells meeting Ag for the first time. In contrast, Fas-mediated death occurs late in an immune response when high levels of FasL expression are maintained on CD4+ T cells, rendering them sensitive to apoptosis, whereas CD8+ T cells are relatively insensitive to this signal. Taken together, these in vitro data provide evidence that FasL plays dual functions in the regulation of CD8+ and CD4+ T cells, having the capacity to both positively and negatively regulate the peripheral T cell compartment.
The in vivo consequence of FasL costimulation is the focus of the current study. In general, costimulation has multiple functions. First, costimulation may regulate the production of cytokines and their receptors that together help regulate cell proliferation. Second, costimulation may strengthen the TCR signal in both amplitude and duration, possibly by enhancing the redistribution and clustering of raft microdomains at the TCR engagement site (10, 11). Third, costimulation may enhance the long-term survival of Ag-activated T lymphocytes (12). Fourth, costimulation may play a role in the differentiation of T cells into mature effector subsets by altering the Th1/Th2 balance (13, 14) and regulating CTL effector development (15).
The following work brings us one step closer to deciphering the function of FasL costimulation by demonstrating that maximizing the Ag-driven in vivo proliferation of both primary and Ag-experienced CD8+ T cells requires FasL costimulation. Thus, proliferative signals delivered to Ag-reactive T cells during the course of an immune response can be further amplified by the engagement of a molecule that can also initiate a death signal through its receptor. With the discovery of increasing numbers of accessory molecules that possess costimulatory function, the activation of T cells is becoming more complex than previously envisioned, involving multiple ligand-receptor interactions that occur at the interface of the T cell and the APC. FasL can now be placed into that group of subtle regulators of T cell proliferation.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B6, B6.MRL-Faslpr (B6.lpr),
B6Smn.C3H-Faslgld (B6.gld),
B6.SJL-Ptprca
Pep3b/BoyJ.Ly5.1 (B6.Ly5.1), and C3H.
MRL-Faslpr (C3H.lpr) mice were
purchased from The Jackson Laboratory (Bar Harbor, ME) and used at 6–9
wk of age. OT-1 TCR-transgenic mice on a B6 background (OT-1) were
derived to express the
V2+V5+ TCR from a
CD8+ CTL clone specific for chicken OVA presented
by H-2Kb and have been previously described
(16). These mice were maintained as heterozygotes by
crossing with B6 mice and screening PBLs for V5 expression by flow
cytometry. FasL-deficient OT-1 mice (OT-1.gld) were obtained
by cross/backcross breeding of OT-1 and B6.gld mice,
screening for V5 expression by flow cytometry and for the
gld mutation by a PCR protocol provided by A.
Marshak-Rothstein (Boston University, Boston, MA). The PCR using tail
DNA was performed (50 cycles: 1 min at 94°C, 1 min at 51°C, and 1
min at 72°C) using the specific primer
5'-TCTCAACTCTCTCTGATCAATTTTGAGGAATCTAAGGCC-3' and the reverse
primer 5'-CTCTCATTCAAGACAATATTCCTG-3' on a DNA ThermalCycler 480
(Perkin-Elmer/Cetus, Emeryville, CA). The PCR product of wild-type, but
not gld, FasL contains a diagnostic StuI
restriction enzyme site. Reaction products digested with
StuI (Roche, Indianapolis, IN) were separated on a 6%
polyacrylamide gel to reveal a single band of 135 bp for gld
FasL or two bands of 96 and 39 bp for wild-type FasL. Fas-deficient
OT-1 mice (OT-1.lpr) were obtained by cross/backcross
breeding of OT-1 and B6.lpr mice, screening for V5
expression by flow cytometry and for the lpr mutation by a
PCR protocol (17).
B6.PL-Thy-1a/Cy (B6.Thy-1.1) and C3HeB/FeJ
(C3H) mice were bred and maintained in our animal facility using
founders obtained from The Jackson Laboratory. Thy-1 congenic OT-1 mice
(OT-1.Thy-1.1) were generated by cross/backcross breeding of OT-1 and
B6.Thy-1.1 mice, screening for V5 and Thy-1.1 expression by flow
cytometry.
Reagents
PE-conjugated anti-CD8 (53-6.7) mAb, biotin-conjugated
anti-Thy-1.2 (30-H12) and anti-V2 (B20.2) mAbs, and
FITC-conjugated anti-V5 (MR9-4), anti-Ly5.1 (A20), and
anti-Thy-1.1 (HIS51) mAbs were all purchased from PharMingen (San
Diego, CA). TriColor-conjugated streptavidin was purchased from Caltag
(Burlingame, CA). The H-2Kb-binding peptides,
OVA257–264 (OVAp, SIINFEKL) and the vesicular
stomatitis virus (VSV)-NP52–59 (VSVp, RGYVYQGL),
were prepared as described previously (18).
Flow cytometry
PBLs, splenocytes, and lymph node (LN) cells were stained as described previously (19) and analyzed on a FACScan using CellQuest software (Becton Dickinson, Mountain View, CA). Dead cells were excluded on the basis of forward and side scatter profiles, and at least 104 live-gated events were collected.
Generation and maintenance of Ag-specific CTLs
Alloreactive H-2k-specific CTLs were
generated by incubating naive spleen and LN cells from age-matched B6,
B6.lpr, and B6.gld mice with an equal number of
irradiated (3000 rad) C3H splenocytes, as previously described
(7). Lines were maintained by stimulation every 8–10
days. After the third stimulation, the medium was supplemented with 50
mM -methyl mannoside and 5% supernatant from rat cells stimulated
with 3 µg/ml Con A for 2 days. All CTL lines were routinely monitored
by flow cytometry and CTL assay.
Adoptive transfers
For the transfer of wild-type OT-1 cells into B6.wt,
B6.lpr, and B6.gld recipients, 20 x
106 unseparated spleen and LN cells (
7 x
106 CD8+ T cells) were
injected i.v. into unirradiated hosts, and recipients were i.p.
injected daily on days 1–3 after cellular transfer with 100 nmol of
either OVAp or VSVp in PBS. For the transfer of OT-1.lpr
cells into B6.wt and B6.lpr recipients, 20 x
106 unseparated spleen and LN cells (4 x
106 CD8+ T cells) were
injected i.v. into unirradiated hosts, and recipients were i.p.
injected daily on days 1–3 after cellular transfer with either PBS or
100 nmol of OVAp. On day 4, the accumulation of the donor population
was quantitated by flow cytometry and the cytolytic activity by CTL
assay. For the adoptive transfer of mixed wild-type OT-1 and
OT-1.gld donor cells, unseparated spleen and LN cell
populations were analyzed before transfer by flow cytometry to ensure
the coinjection of 15 x 106
CD8+V2+V5+
T cells of each origin. The Ly5 congenic adoptive hosts were primed
daily on days 1–3 of cellular transfer with 25–100 nmol of OVAp, and
splenocytes were analyzed on day 4 by flow cytometry. The donor
CD8+ T cell populations were distinguished from
the host and from each other by Ly5 and Thy-1 expression. In each case,
>98% of the recovered donor T cells were found to be
V2+V5+. For the
transfer of Ag-experienced CTL lines into unirradiated B6.Thy-1.1
recipients, 3 x 106
H-2k-specific CTLs, harvested 8–10 days after
their last stimulation, were i.v. injected along with their cell-borne
Ag (107 viable anti-Thy-1.2 plus
complement-pretreated C3H splenocytes). Before transfer, CTLs were
separated from fragments of antigenic cells over Ficoll
(7). Splenocytes and LN cells from the recipients were
analyzed for donor cell expansion on day 4 using flow cytometry to
distinguish the donor and host CD8+ T cells by
Thy-1 expression. All cellular transfers were analyzed over a short
time course of 4 days to avoid potential rejection of the donor cells
across Ly5 and Thy-1 differences.
Cytotoxicity assay
Cytolytic activity of splenocytes from adoptive transfer
recipients was quantitated directly ex vivo by a
51Cr release assay. Serial dilutions of effector
cells and 2 x 103 tumor targets labeled
with [51Cr]sodium chromate were plated in
round-bottom 96-well plates in a final volume of 200 µl. After a 4-h
incubation at 37°C, 100 µl of supernatant was collected, and the
percent specific lysis was determined as 100 x [(cpm released by
CTL - spontaneous release)/(cpm released by detergent -
spontaneous release)]. Target cells were spleen cells incubated for 2
days with 2 µg/ml Con A or EL4 cells pulsed with either OVAp or VSVp.
Where indicated, 0.5 mM EGTA was added to the responders simultaneously
with the targets to determine the level of
Ca2+-independent Fas-mediated lysis. To assay the
cytolytic activity of the progeny of individual resting CTL,
splenocytes were plated in limiting dilution from 6-wk-old B6.wt and
B6.gld mice 8 days after i.p. infection with 5 x
106 PFU of VSV. Responders were incubated for 7
days at 37°C in the presence of exogenous IL-2 and irradiated,
low-density B6 spleen cells coated with VSVp. Wells containing
effectors were identified as those with cytolytic activity at least 3
SD above the mean of spontaneous lysis. RMA-S cells (TAP-2 mutant,
H-2b) pulsed with VSVp were used as CTL targets.
The anti-VSV CTL frequency was previously determined to be 1 in
600 CD8+ T cells for B6.gld and 1 in
1200 CD8+ T cells for B6.wt (data not shown). The
dilution at which 1000 CD8+ T cells were plated
was used for the calculation of percent specific lysis by individual
CTL; the frequency of positive wells at this dilution was <5%.
Spontaneous release of target cells in the absence of CTL was <20% in
all experiments.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Cells from OT-1 TCR-transgenic mice were used to track the
fate of a homogeneous CD8+ T cell population
proliferating to Ag in the presence or the absence of FasL
costimulation. Adoptive transfer of wild-type OT-1 cells into B6.wt,
B6.lpr, or B6.gld hosts followed by OVAp
injections (schematic diagram, Fig. 1
A) showed that the
OVAp-driven accumulation of OT-1 cells in Fas+
(B6.wt and B6.gld) mice greatly exceeded that in
Fas- (B6.lpr) hosts. This difference
in Ag-mediated accumulation of the OVA-specific OT-1 cells between the
hosts was observed by both flow cytometry (Fig. 1B) and
cytotoxic activity (Fig. 1C). The Ag specificity of this
accumulation is shown by the lack of donor cell expansion upon
injection of VSVp, another Kb-binding peptide. No
OVAp-driven expansion of host cells was measurable (data not
shown).
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Because B6.lpr mice express increased levels of FasL
relative to wild-type B6 mice, it is possible that the lower level of
donor cell accumulation seen in the lpr hosts shown in Fig. 1
is due to Fas-mediated cell death of donor OT-1 cells delivered by
up-regulated FasL on the lpr host cells. To test this
possibility, cells from OT-1.lpr mice, which are incapable
of undergoing Fas-mediated lysis, were transferred into adoptive hosts
in the presence or absence of FasL costimulation. Adoptive transfer of
OT-1.lpr donor cells into B6.wt and B6.lpr hosts
followed by OVAp injection (schematic diagram, Fig. 2A) demonstrates that the
reduced accumulation of Ag-specific donor cells in
Fas- hosts (Figs. 1B and
2B) is not a reflection of donor cell elimination by the
host, but is more likely due to the lack of FasL costimulation. This
difference was observed both by flow cytometry (Fig. 2B and
data not shown), where two to four times the number of
OT-1.lpr CD8+ T cells were recovered
from wild-type B6 relative to B6.lpr hosts, and by CTL assay
(Fig. 2C). No OVAp-driven expansion of host cells was
measurable (data not shown). Taken together, these data suggest that
FasL costimulation is required to maximize the accumulation of
Ag-specific CD8+ T cells in vivo.
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To corroborate the previous results, we performed adoptive
transfer experiments in which the donor cells, rather than the host
cells, differed in expression of functional FasL. To avoid
host-to-host variations in comparing the two donor types, the
CD8+V2+V5+ T cells from
the spleens and LNs of wild-type OT-1 (Thy-1.1) and OT-1.gld
(Thy-1.2) mice were mixed in a 1:1 ratio and injected into unirradiated
B6.Ly5.1 recipients (schematic diagram, Fig. 3A). Splenocytes from the
recipients were analyzed by flow cytometry (Fig. 3B) on day
4 to quantitate wild-type OT-1 (CD8+,
Ly5.1-, Thy-1.2-) and
OT-1.gld donor cells (CD8+,
Ly5.1-, Thy-1.2+).
Expression of the
V2+V5+-transgenic TCR
by these responders was verified by flow cytometry (data not shown). As
shown in Fig. 3, B and C, approximately twice the
number of wild-type OT-1 compared with OT-1.gld
CD8+ T cells was recovered from the adoptive
hosts. These data further demonstrate the effect of FasL costimulation
on the expansion of CD8+ T cells in a primary
immune response.
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To determine whether FasL costimulation influences the expansion
of previously activated T cells, CTL lines were used as donor cells in
an adoptive transfer experiment (Fig. 4).
Resting oligoclonal anti-H-2k CTL lines
derived from B6.wt and B6.gld mice were coinjected with
their cell-borne Ag (T cell-depleted C3H or C3H.lpr
splenocytes) into unirradiated B6.Thy-1.1 hosts (schematic diagram,
Fig. 4). Recipient splenocytes and LN cells were analyzed by flow
cytometry 4 days later. As with the previous transfers of naive
CD8+ donor T cells expressing a uniform TCR
(
Figs. 1–3), the expansion of previously activated oligoclonal
wild-type donor CTL exceeded that of FasL- CTL
by a factor of 2–4 (p = 0.03, by Student’s
t test; Fig. 4 and data not shown).
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CTLs can kill target cells through two different lytic pathways:
the degranulation pathway, involving perforin and granzymes, and the
Fas lytic pathway involving FasL on the CTL and Fas on the target cell
(reviewed in Refs. 20 and 21). Our previous
studies have determined that the cytolytic activity of B6.wt and
B6.gld responders is indistinguishable on a recovered cell
basis despite the absence of functional FasL in B6.gld
effector cells (7). Furthermore, no traces of
EGTA-independent (Fas-dependent) lysis are detectable in our 4-h
51Cr release assay, and alloantigen-specific
lysis by wild-type responders is identical on both lpr and
wild-type targets (Fig. 5). The absence
of Fas-mediated target cell lysis in these assay conditions allows a
comparison of the CTL burst size generated by
FasL+ and FasL-
responders. To quantitate the influence of FasL costimulation at the
level of a single CD8+ T cell using T lymphocytes
that express diverse TCRs, we plated splenocytes from age-matched
VSV-primed B6.wt and B6.gld mice in limiting dilution to
quantitate the frequency of responding CTL (schematic diagram, Fig. 6, top). The cytolytic
activity in each well calculated initially to contain a single resting
VSV-specific T cell was used as a measure of CTL burst size. Cultures
were maintained for 7 days, and targets were added directly to the
wells to assay for specific lysis. The percent specific lysis was
significantly higher for the wells plated with
FasL+ anti-VSV CTL than for those plated with
FasL- anti-VSV CTL (Fig. 6, bottom), revealing the larger burst size for the former
compared with the latter responder cells.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-transgenic Ag-specific CD8+ T cells
were transferred into age-matched hosts that could
(Fas+ mice) and could not
(Fas- mice) provide costimulation to
FasL+ responders (Fig. 1). The limited
Ag-specific accumulation of wild-type OT-1 cells in B6.lpr
hosts underscores the important role the host plays in providing FasL
costimulation to maximize the Ag-specific response by incoming donor
cells. The second adoptive transfer protocol (Fig. 2) indicates that
this deficit in Ag-specific accumulation in lpr hosts is not
due to Fas-mediated killing of the incoming donor cells, but is more
likely a result of the absence of FasL costimulation.
In the third set of experiments, TCR-transgenic
CD8+ donor T cells with
(FasL+ wild-type OT-1) and without
(FasL- OT-1.gld) the capacity to
receive FasL costimulation were mixed at a 1:1 ratio before transfer
into a common host (Fig. 3). The absence of FasL costimulation leads to
a 2-fold decrease in the yield of OT-1.gld donor cells
following antigenic challenge. One caveat with these data is the
identification of the
Ly5.1-CD8+Thy-1.2-
population as the wild-type OT-1 donor population (Fig. 3B).
The presence of any
Thy-1-CD8+ contaminants,
such as CD8+ dendritic cells (22),
would influence the cell numbers obtained in this category. However,
the frequency of these cells is low, and the contaminants are unlikely
to be isolated and transferred by the protocol used in this
experiment.
Although the inability of donor cells to receive FasL costimulation
translates into a 2-fold difference in cell yields (Fig. 3), the
inability of lpr hosts to provide FasL costimulation results
in a greater difference in donor cell recovery (Figs. 1 and 2). One
possible explanation for this difference is that incoming wild-type
OT-1 donor cells could be more efficiently eliminated through
Fas-mediated death, initiated by the up-regulated FasL in
B6.lpr mice (23, 24). The intermediate
recoveries observed with the transfer of lpr OT-1 donor
cells into lpr hosts shown in Fig. 2 (lower recoveries than
wild-type donor cells from wild-type hosts, but higher than wild-type
donor cells from lpr hosts, both shown in Fig. 1) suggest
some involvement of Fas-mediated donor cell elimination by the host.
Therefore, the most likely explanation for the results observed in Fig. 1 is a combination of Fas-mediated cell death and deficient FasL
costimulation resulting in reduced wild-type OT-1 donor cell numbers
recovered from the B6.lpr adoptive host. The third adoptive
transfer experiment (Fig. 3) used a common host, thus allowing a direct
comparison of the two donor populations while minimizing the effects of
host-to-host variation (including any pre-existing conditions and
differences in the efficiency of priming) and potential differences in
the two donor populations (including cytokine production and APC
number). However, these data could be complicated by a potential
increase in Fas-mediated death of gld
CD8+ donor cells relative to coinjected wild-type
cells as a result of increased Fas expression by gld T
cells. This explanation is unlikely, given the relative insensitivity
to Fas-mediated death of CD8+ T cells
(25) and the fact that four times the number of
OT-1.lpr T cells accumulate in wild-type as in
lpr recipients (Fig. 2).
The adoptive transfer experiments described above used naive
TCR-transgenic CD8+ donor T cells in a primary
immune response induced by antigenic challenge. Our final set of cell
transfer experiments tested the role of FasL costimulation in an
oligoclonal response with Ag-experienced CD8+ T
cells by quantitating the recovery of adoptively transferred, resting,
H-2k-specific CTLs (Fig. 4). In line with the
transfer of primary CD8+ T cells (Fig. 3), FasL
costimulation enhanced the capacity of the Ag-experienced oligoclonal
CD8+ T cell population to expand by an average of
2-fold (Fig. 4 and data not shown). The donor cell recovery was much
lower than that obtained with the transfer of primary cells, probably
due to the unusual homing patterns of CTL lines (26) and
the competition for Ag recognition from the immune system of the
unirradiated host. It is important to note that this outcome was not
altered by immunization with C3H.lpr spleen cells (data not
shown), demonstrating that Fas-mediated differences in the rate of Ag
clearance by FasL+ and
FasL- CTLs were not a factor. In this context,
it should also be mentioned that the use of C3H.lpr APCs
does not affect the ability of FasL+ donor CTLs
to receive FasL costimulation, perhaps due to the ability of
Fas+ responders and non-APCs to costimulate
through FasL (data not shown).
In each of these adoptive transfer experiments, FasL costimulation was
measured at the population level using T cells expressing either a
uniform or a restricted TCR repertoire. Examination of the burst size
of individual resting CTLs in a polyclonal response against VSV
demonstrates that FasL costimulation operates at the level of the
individual CTL and influences the fate of cells expressing a diverse
set of TCRs (Fig. 6). The degree of proliferation, mediated in part by
FasL costimulation, ultimately influences the cytolytic activity of the
CTL culture. The comparisons made between wild type and gld
CTLs in this experiment are validated by the results obtained in Fig. 5, showing that Ca2+-independent, Fas-mediated
lysis (27) is not operative in our assay. Neither the
addition of EGTA at the time of target cell coincubation
(28) nor the use of lpr targets demonstrated
any difference in the cytolytic level between wild-type and
gld CTLs. Therefore, the difference in cytotoxicity observed
for individual wild-type and gld CTLs in Fig. 6 is not due
to the presence or the absence of Fas-mediated lysis by the CTLs, but
more likely is a result of the differential contribution of FasL
costimulation to the burst size of the two responder cell
populations.
FasL costimulation affects T cell responses, from monoclonal to
polyclonal, regardless of their antigenic history (naive vs Ag
experienced), and is measurable at the single-cell level as well as at
the population level. It is important to note that this costimulation
appears to be Ag specific, being required early after TCR ligation
(9) and only in contexts in which T cell stimulation uses
the TCR. Stimuli that are subject to FasL costimulation include
cell-bound Ag, anti-CD3, and Con A, all of which initiate signaling
directly through the TCR. However, FasL costimulation is not
demonstrable with stimuli that bypass the TCR, such as PMA plus
ionomycin or anti-Thy-1, despite their ability to up-regulate FasL
expression (I. Suzuki and P. Fink, unpublished observations). Cells
receiving FasL costimulation display an 2-fold advantage in
Ag-mediated expansion compared with cells that have not been
costimulated through FasL (Figs. 3 and 4). This finding is consistent
with the report of a 2-fold reduction in the expansion of
gld compared with wild-type donor cells in a parent 
F1 model of acute graft-vs-host disease
(29). It remains to be determined whether this difference
in expansion in either system reflects a role for FasL costimulation in
triggering the entry of Ag-responsive cells into the memory pathway. It
is likely, however, that the influence of Ag-specific, FasL-mediated
costimulation on CTL burst size will affect the efficiency with which
an individual clears an invading pathogen.
In summary, the work described in this paper demonstrates the in vivo function of FasL as a costimulatory receptor for peripheral CD8+ T cells, a function previously defined in vitro for CD8+ (7) and CD4+ T cells (9). Multiple levels of costimulation such as that provided by FasL appear to be a mechanism used by the immune system in fine-tuning T cell reactivity.
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Acknowledgments |
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Footnotes |
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2 Current address: Clinical Research Group, Department of Dermatology, University of Freiburg, Hauptstrasse 7, D-79104 Freiburg, Germany.
3 Address correspondence and reprint requests to Dr. Pamela J. Fink, Department of Immunology, University of Washington, Box 357650, Room H574A, Seattle, WA 98195.
4 Abbreviations used in this paper: TNFR, TNF receptor; B6, C57BL/6; B6.gld, B6Smn.C3H-Faslgld; B6.lpr, B6.MRL-Faslpr; B6.Ly5.1, B6.SJL-Ptprca Pep3b/BoyJ (Ly5.1); B6.Thy-1.1, B6.PL-Thy-1a/Cy; B6.wt, wild-type C57BL/6; C3H, C3HeB/FeJ; L, ligand; LN, lymph node; LT, lymphotoxin; OT-1, line of OVA-specific TCR-transgenic mice; OT-1.gld, OT-1 on a B6.gld background; OT-1.Thy-1.1, OT-1 on a B6.Thy-1.1 background; OVAp, OVA peptide; VSV, vesicular stomatitis virus; VSVp, VSV peptide.
Received for publication February 28, 2000. Accepted for publication August 21, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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  M. Paulsen, B. Mathew, J. Qian, M. Lettau, D. Kabelitz, and O. Janssen FasL cross-linking inhibits activation of human peripheral T cells Int. Immunol., May 1, 2009; 21(5): 587 - 598. [Abstract] [Full Text] [PDF]
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 L. E. Makaroff, D. W. Hendricks, R. E. Niec, and P. J. Fink Postthymic maturation influences the CD8 T cell response to antigen PNAS, March 24, 2009; 106(12): 4799 - 4804. [Abstract] [Full Text] [PDF]
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 M. R. Olson and S. M. Varga Fas Ligand Is Required for the Development of Respiratory Syncytial Virus Vaccine-Enhanced Disease J. Immunol., March 1, 2009; 182(5): 3024 - 3031. [Abstract] [Full Text] [PDF]
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I. Puliaeva, R. Puliaev, A. Shustov, M. Haas, and C. S. Via Fas Expression on Antigen-Specific T Cells Has Costimulatory, Helper, and Down-Regulatory Functions In Vivo for Cytotoxic T Cell Responses but Not for T Cell-Dependent B Cell Responses J. Immunol., November 1, 2008; 181(9): 5912 - 5929. [Abstract] [Full Text] [PDF]
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 M. M. Kavurma, N. Y. Tan, and M. R. Bennett Death Receptors and Their Ligands in Atherosclerosis Arterioscler Thromb Vasc Biol, October 1, 2008; 28(10): 1694 - 1702. [Abstract] [Full Text] [PDF]
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 M. Loeffler, G. Le'Negrate, M. Krajewska, and J. C. Reed Inhibition of Tumor Growth Using Salmonella Expressing Fas Ligand J Natl Cancer Inst, August 6, 2008; 100(15): 1113 - 1116. [Abstract] [Full Text] [PDF]
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R. Dudani, M. Russell, H. van Faassen, L. Krishnan, and S. Sad Mutation in the Fas Pathway Impairs CD8+ T Cell Memory J. Immunol., March 1, 2008; 180(5): 2933 - 2941. [Abstract] [Full Text] [PDF]
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M. Sun, S. Lee, S. Karray, M. Levi-Strauss, K. T. Ames, and P. J. Fink Cutting Edge: Two Distinct Motifs within the Fas Ligand Tail Regulate Fas Ligand-Mediated Costimulation J. Immunol., November 1, 2007; 179(9): 5639 - 5643. [Abstract] [Full Text] [PDF]
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M. Sun and P. J. Fink A New Class of Reverse Signaling Costimulators Belongs to the TNF Family J. Immunol., October 1, 2007; 179(7): 4307 - 4312. [Abstract] [Full Text] [PDF]
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M. Sun, K. T. Ames, I. Suzuki, and P. J. Fink The Cytoplasmic Domain of Fas Ligand Costimulates TCR Signals J. Immunol., August 1, 2006; 177(3): 1481 - 1491. [Abstract] [Full Text] [PDF]
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A. S. Mohamood, C. J. Trujillo, D. Zheng, C. Jie, F. M. Murillo, J. P. Schneck, and A. R. A. Hamad Gld mutation of Fas ligand increases the frequency and up-regulates cell survival genes in CD25+CD4+ TR cells Int. Immunol., August 1, 2006; 18(8): 1265 - 1277. [Abstract] [Full Text] [PDF]
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J. S. Hale, T. E. Boursalian, G. L. Turk, and P. J. Fink Thymic output in aged mice PNAS, May 30, 2006; 103(22): 8447 - 8452. [Abstract] [Full Text] [PDF]
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 N. Cahuzac, W. Baum, V. Kirkin, F. Conchonaud, L. Wawrezinieck, D. Marguet, O. Janssen, M. Zornig, and A.-O. Hueber Fas ligand is localized to membrane rafts, where it displays increased cell death-inducing activity Blood, March 15, 2006; 107(6): 2384 - 2391. [Abstract] [Full Text] [PDF]
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 C. Shi, J. Wolfe, J. Q. Russell, K. Fortner, N. Hardin, J. Anguita, and R. C. Budd Fas Ligand Deficiency Impairs Host Inflammatory Response against Infection with the Spirochete Borrelia burgdorferi Infect. Immun., February 1, 2006; 74(2): 1156 - 1160. [Abstract] [Full Text] [PDF]
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 W. Baum, V. Kirkin, S. B. M. Fernandez, R. Pick, M. Lettau, O. Janssen, and M. Zornig Binding of the Intracellular Fas Ligand (FasL) Domain to the Adaptor Protein PSTPIP Results in a Cytoplasmic Localization of FasL J. Biol. Chem., December 2, 2005; 280(48): 40012 - 40024. [Abstract] [Full Text] [PDF]
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 Valerie Pasqualetto, Florence Vasseur, Flora Zavala, Elke Schneider, and Sophie Ezine Fas receptor signaling is requisite for B cell differentiation J. Leukoc. Biol., November 1, 2005; 78(5): 1106 - 1117. [Abstract] [Full Text] [PDF]
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G. Shi, J. Mao, G. Yu, J. Zhang, and J. Wu Tumor Vaccine Based on Cell Surface Expression of DcR3/TR6 J. Immunol., April 15, 2005; 174(8): 4727 - 4735. [Abstract] [Full Text] [PDF]
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C.-R. Yang, S.-L. Hsieh, F.-M. Ho, and W.-W. Lin Decoy Receptor 3 Increases Monocyte Adhesion to Endothelial Cells via NF-{kappa}B-Dependent Up-Regulation of Intercellular Adhesion Molecule-1, VCAM-1, and IL-8 Expression J. Immunol., February 1, 2005; 174(3): 1647 - 1656. [Abstract] [Full Text] [PDF]
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M. Cippitelli, C. Fionda, D. Di Bona, M. Piccoli, L. Frati, and A. Santoni Hyperthermia Enhances CD95-Ligand Gene Expression in T Lymphocytes J. Immunol., January 1, 2005; 174(1): 223 - 232. [Abstract] [Full Text] [PDF]
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S. F. Martin, J. C. Dudda, V. Delattre, E. Bachtanian, C. Leicht, B. Burger, H. U. Weltzien, and J. C. Simon Fas-Mediated Inhibition of CD4+ T Cell Priming Results in Dominance of Type 1 CD8+ T Cells in the Immune Response to the Contact Sensitizer Trinitrophenyl J. Immunol., September 1, 2004; 173(5): 3178 - 3185. [Abstract] [Full Text] [PDF]
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J. A. Rutigliano and B. S. Graham Prolonged Production of TNF-{alpha} Exacerbates Illness during Respiratory Syncytial Virus Infection J. Immunol., September 1, 2004; 173(5): 3408 - 3417. [Abstract] [Full Text] [PDF]
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 S. A. Mikolajczak, B. Y. Ma, T. Yoshida, R. Yoshida, D. J. Kelvin, and A. Ochi The Modulation of CD40 Ligand Signaling by Transmembrane CD28 Splice Variant in Human T Cells J. Exp. Med., April 5, 2004; 199(7): 1025 - 1031. [Abstract] [Full Text] [PDF]
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Z. Guo, M. Zhang, H. An, W. Chen, S. Liu, J. Guo, Y. Yu, and X. Cao Fas ligation induces IL-1{beta}-dependent maturation and IL-1{beta}-independent survival of dendritic cells: different roles of ERK and NF-{kappa}B signaling pathways Blood, December 15, 2003; 102(13): 4441 - 4447. [Abstract] [Full Text] [PDF]
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O. V. Proussakova, N. A. Rabaya, A. B. Moshnikova, E. S. Telegina, A. Turanov, M. G. Nanazashvili, and I. P. Beletsky Oligomerization of Soluble Fas Antigen Induces Its Cytotoxicity J. Biol. Chem., September 19, 2003; 278(38): 36236 - 36241. [Abstract] [Full Text] [PDF]
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T. E. Boursalian and P. J. Fink Mutation in Fas Ligand Impairs Maturation of Thymocytes Bearing Moderate Affinity T Cell Receptors J. Exp. Med., July 21, 2003; 198(2): 349 - 360. [Abstract] [Full Text] [PDF]
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D. R. Beisner, I. H. Chu, A. F. Arechiga, S. M. Hedrick, and C. M. Walsh The Requirements for Fas-Associated Death Domain Signaling in Mature T Cell Activation and Survival J. Immunol., July 1, 2003; 171(1): 247 - 256. [Abstract] [Full Text] [PDF]
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M. Cippitelli, C. Fionda, D. Di Bona, A. Lupo, M. Piccoli, L. Frati, and A. Santoni The Cyclopentenone-Type Prostaglandin 15-Deoxy-{Delta}12,14-Prostaglandin J2 Inhibits CD95 Ligand Gene Expression in T Lymphocytes: Interference with Promoter Activation Via Peroxisome Proliferator-Activated Receptor-{gamma}-Independent Mechanisms J. Immunol., May 1, 2003; 170(9): 4578 - 4592. [Abstract] [Full Text] [PDF]
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 S. Kasahara, K. Ando, K. Saito, K. Sekikawa, H. Ito, T. Ishikawa, H. Ohnishi, M. Seishima, S. Kakumu, and H. Moriwaki Lack of Tumor Necrosis Factor Alpha Induces Impaired Proliferation of Hepatitis B Virus-Specific Cytotoxic T Lymphocytes J. Virol., February 15, 2003; 77(4): 2469 - 2476. [Abstract] [Full Text] [PDF]
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X. Wan, J. Zhang, H. Luo, G. Shi, E. Kapnik, S. Kim, P. Kanakaraj, and J. Wu A TNF Family Member LIGHT Transduces Costimulatory Signals into Human T Cells J. Immunol., December 15, 2002; 169(12): 6813 - 6821. [Abstract] [Full Text] [PDF]
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G. Shi, H. Luo, X. Wan, T. W. Salcedo, J. Zhang, and J. Wu Mouse T cells receive costimulatory signals from LIGHT, a TNF family member Blood, October 16, 2002; 100(9): 3279 - 3286. [Abstract] [Full Text] [PDF]
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M. Surquin, A. Le Moine, V. Flamand, N. Nagy, K. Rombaut, F.-X. Demoor, P. Stordeur, I. Salmon, J.-C. Guery, M. Goldman, et al. Skin Graft Rejection Elicited by {beta}2-Microglobulin as a Minor Transplantation Antigen Involves Multiple Effector Pathways: Role of Fas-Fas Ligand Interactions and Th2-Dependent Graft Eosinophil Infiltrates J. Immunol., July 1, 2002; 169(1): 500 - 506. [Abstract] [Full Text] [PDF]
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T.-L. Hsu, Y.-C. Chang, S.-J. Chen, Y.-J. Liu, A. W. Chiu, C.-C. Chio, L. Chen, and S.-L. Hsieh Modulation of Dendritic Cell Differentiation and Maturation by Decoy Receptor 3 J. Immunol., May 15, 2002; 168(10): 4846 - 4853. [Abstract] [Full Text] [PDF]
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 Y. Seko, N. Kayagaki, K.-i. Seino, H. Yagita, K. Okumura, and R. Nagai Role of Fas/FasL pathway in the activation of infiltrating cells in murine acute myocarditis caused by Coxsackievirus B3 J. Am. Coll. Cardiol., April 17, 2002; 39(8): 1399 - 1403. [Abstract] [Full Text] [PDF]
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M. Cippitelli, C. Fionda, D. Di Bona, F. Di Rosa, A. Lupo, M. Piccoli, L. Frati, and A. Santoni Negative Regulation of CD95 Ligand Gene Expression by Vitamin D3 in T Lymphocytes J. Immunol., February 1, 2002; 168(3): 1154 - 1166. [Abstract] [Full Text] [PDF]
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M. C. G. Marcondes, E. M. E. Burudi, S. Huitron-Resendiz, M. Sanchez-Alavez, D. Watry, M. Zandonatti, S. J. Henriksen, and H. S. Fox Highly Activated CD8+ T Cells in the Brain Correlate with Early Central Nervous System Dysfunction in Simian Immunodeficiency Virus Infection J. Immunol., November 1, 2001; 167(9): 5429 - 5438. [Abstract] [Full Text] [PDF]
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M. I. Kafrouni, G. R. Brown, and D. L. Thiele Virally Infected Hepatocytes Are Resistant to Perforin-Dependent CTL Effector Mechanisms J. Immunol., August 1, 2001; 167(3): 1566 - 1574. [Abstract] [Full Text] [PDF]
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