By
From the * Basel Institute for Immunology, CH-4005 Basel, Switzerland; and the Unité d'Immunité
Cellulaire Antivirale, Institut Pasteur, 75724 Paris-Cedex 15, France
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Abstract |
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The potential involvement of early growth response (Egr)-1, a zinc-finger transcription factor
belonging to the immediate-early genes, in positive/negative selection of thymocytes has been
implicated by its expression in the population of CD4+CD8+ double positive (DP) cells undergoing selection. To further investigate this possibility, transgenic mice overexpressing Egr-1 in
thymocytes were bred with a transgenic mouse line expressing a T cell receptor (TCR) recognizing the H-Y male antigen in the context of H-2b class I major histocompatibility complex
(MHC) molecules. In Egr-1/TCR H-Y double-transgenic mice, efficient positive selection of
H-Y CD8+ T cells occurred, even in mice on either a nonselecting H-2d background or a 2-microglobulin (
2m)-deficient background in which the expression of class I MHC heavy
chains is extremely low; no positive selection was observed on a Kb
/
Db
/
2m
/
background where class I MHC expression is entirely absent. Similarly, when the Egr-1 transgene
was introduced into a class II MHC-restricted TCR transgenic mouse line, Egr-1/TCR double-transgenic mice revealed increased numbers of CD4+ T cells selected by class II MHC, as
well as significant numbers of CD8+ T cells selected by class I MHC (for which the transgenic
TCR might have weak affinity). Thus, Egr-1 overexpression allows positive selection of thymocytes via TCR-MHC interactions of unusually low avidity, possibly by lowering the
threshold of avidity required for positive selection. Supporting this possibility, increased numbers of alloreactive T cells were positively selected in Egr-1 transgenic mice, resulting in a strikingly enhanced response against allo-MHC. These results suggest that expression of Egr-1 and/or its target gene(s) may directly influence the thresholds required for thymocyte selection.
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Introduction |
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Positive/negative selection of thymocytes at the
CD4+CD8+ double positive (DP)1 stage is the key
checkpoint for thymocyte maturation, at which their
fatewhether they develop further to CD4+ or CD8+ single positive (SP) cells or die by clonal deletion
is decided (1, 2). Accumulating evidence indicates that the avidity between T cell receptors (TCRs) expressed on thymocytes
and MHC/antigen-peptide complexes displayed on the
surface of APCs appears to define the type of intracellular
signals generated, which promote thymocytes to be either
positively selected (further developing to the SP stage) or
negatively selected (clonally deleted); avidity is dependent on TCR-MHC affinity and the expression levels of both
complexes (3, 4). It is believed that there are thresholds in
the strength of TCR-MHC avidity that determine the nature and consequences of subsequent TCR signaling (3, 4).
When thymocytes and APCs interact with high avidity,
cells are negatively selected through the apoptotic pathway,
whereas thymocytes are promoted to the SP cell stage
when the avidity is moderate but sufficient. If the avidity is
too low, cells cannot undergo either type of selection, resulting in their death as "neglected cells." However, the
molecular mechanism that defines these thresholds for selection events is unclear.
Signals generated during positive/negative selection must be differentially controlled; hence, rapidly responding transcriptional regulators able to elicit a cascade of changes in gene expression should be important. Immediate-early genes, expression of which is rapidly induced after cell-surface receptor ligation without de novo protein synthesis, are strong candidates for such a rapid response mediator (5). Shao et al. reported recently that the expression of one of these immediate-early genes, early growth response (Egr)-1, a zinc-finger transcription factor rapidly induced by TCR ligation, appears to correlate with selection events, as it is expressed at much lower levels in DP cells from mutant mice deficient for both MHC class I and class II molecules than in DP cells from wild-type mice. In addition, Egr-1 expression in the mutant DP cells was able to be upregulated by anti-CD3 mAb ligation (6).
We recently generated transgenic mouse lines overexpressing Egr-1 in thymocytes under the control of the
lck-proximal promoter (7), and showed that in Egr-1 transgenic mice on a recombination-activating gene (RAG)-
deficient background, thymocytes bypassed the block at the
CD25+CD44 DN stage and matured to the immature
single positive (ISP) cell stage. Here, the effect of Egr-1 expression on positive/negative selection is extensively analyzed by breeding Egr-1 transgenic mice with various
TCR transgenic mice, as well as by evaluating positive selection of alloreactive T cells in Egr-1 transgenic mice.
This report provides interesting insight into what is actually required for positive selection and what transcriptional
pathways may be involved in the selection events.
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Materials and Methods |
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Mice.
All mice used here were maintained in the specific pathogen-free facility of the Basel Institute for Immunology. Matched sets of littermates with transgene-positive and -negative genotypes were used in all experiments.Serological Reagents and Flow Cytometry.
Reagents used for staining T cells and subsets thereof were as described (7-9, and references therein). Thymus, lymph node, and spleen cells were stained with saturating levels of mAbs and analyzed using a FACSCalibur® cytometer (Becton Dickinson, San Jose, CA).Cell Survival Assay.
H-Y CD8+ SP cells were sorted after staining thymocytes for CD4, CD8, T3.70, and peanut agglutinin receptor (PNAr). 5 × 105 cells from each type of mice were cultured in triplicate in 96-well flat-bottomed microculture plates in complete DMEM culture medium including 10% FCS. 24, 48, 72, 96, and 120 h after starting the culture, the number of living cells was evaluated by trypan blue exclusion and plotted as a survival curve. H-Y CD8+ SP cells from all types of mice showed a similar number of living cells at each time point.Purification of CD8+ T Cells.
CD8+ T cells were purified by incubating spleen cells for 30 min on ice with 10% (vol/vol) culture supernatant of hybridomas RL172 (IgM antibody to CD4) and M5/114 (antibody to class II MHC), washed, and then incubated for an additional 1 h at 37°C in 10% Lo-Tox complement (Cedarlane Labs Ltd., Hornby, Ontario, Canada). Dead cells were excluded by Ficoll gradation. Living cells were washed twice and resuspended in complete DMEM.T Cell Proliferation Assay.
Purified CD8+ T cells (5 × 105/ well) were cultured in the presence of various concentrations of either immobilized anti-CD3 mAb (KT-3; provided by Dr. C. Benoist, Strasbourg, France) or Con A (Sigma Chemical Co., St. Louis, MO). The KT-3 mAb was immobilized by preincubation of wells at 4°C overnight. All cultures were performed for 3 d, and proliferation was assessed by [3H]thymidine incorporation in the last 16 h of culture.In Vivo and In Vitro Recognition of H-Y Antigen.
This is a modification of a similar experiment described elsewhere (10). In brief, purified CD8+ T cells including 2 × 106 H-Y CD8+ T cells calculated by the percentage of T3.70+ population in the CD8+ cells after fluorocytometric analysis, were suspended in 200 µl of PBS, and injected intravenously into 6-8-wk-old male and female CMLRs.
Responders (B cell-depleted lymph node cells) were cultured with stimulators (3,000-rad irradiated spleen cells) at various ratios for 5 d in complete DMEM in flat-bottomed 96-well microculture plates. Proliferation was assessed by [3H]thymidine incorporation in the last 16 h of culture.Bone Marrow Chimeras.
To construct bone marrow chimeras, recipient mice were 10-Gy (from a cobalt source) irradiated. 24 h later, they were injected intravenously with 107 bone marrow cells from which mature T and B cells had been depleted by treatment with anti-CD4 (RL172) and anti-CD8 (31M) mAbs plus Lo-Tox complement (Cedarlane Labs Ltd.). After grafting, mice were rested for 5 wk to allow reconstitution before analysis. ![]() |
Results |
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As described in our previous report, Egr-1 transgenic mice (Egr-1-TG) exhibit transgene expression at levels far exceeding endogenous Egr-1, when analyzed by Northern blotting using thymic RNA from transgenic and negative littermate mice (7). As observed for many other transgenes driven by the lck-proximal promoter (11), Egr-1 transgene expression was detected in a wide range of developmental stages of thymocytes of Egr-1-TG, including DP cells, when analyzed by the reverse transcription PCR method (12) using RNA from sorted cells from each population (data not shown). No obvious transgene expression was detected either in spleen or lymph node by Northern blot analysis, indicating thymocyte-specific transgene expression (7).
To assess the possible effect of Egr-1 expression on thymocyte selection, Egr-1-TG were cross-bred with transgenic mice expressing a TCR recognizing the H-Y male
antigen in the context of H-2b class I, Db molecules. It is
well-established that thymocytes expressing H-Y-specific
TCRs are positively selected in female H-2b/b mice and
negatively selected in male H-2b/b mice, but do not undergo either positive or negative selection in female or
male mice on an H-2d/d background (8, 13). Fig. 1 a shows
CD4/CD8 profiles of mature thymocytes bearing the H-Y
TCR defined as T3.70 (mAb against H-Y TCR
chain)high,
PNArlow. As shown in the upper panels, in H-2b/b females,
both Egr-1-TG with an H-Y TCR transgene (Egr/H-Y)
and Egr-1-TG-negative littermate mice with an H-Y
TCR transgene (NL/H-Y) harbored mature H-Y CD8+
SP thymocytes. The CD8+ SP cells in these profiles do not
contain immature CD8 SPs (ISPs), which exhibit high levels of PNAr (7). However, in H-Y mice, since expression
levels of transgenic H-Y TCR in DN cells (which are
PNArlow) are as high as those in mature SP cells (8), a significant proportion of DN cells as well as mature SP cells
are plotted in these profiles. The numbers of total thymocytes
were slightly larger in Egr/H-Y mice than in NL/H-Y
mice (Table 1). However, the absolute numbers of mature
H-Y CD8+ SP cells were ~1.7 times higher in Egr/H-Y
than in NL/H-Y mice (Table 1). In line with this, the percentages of T cells bearing the transgenic
chain (T3.70+)
in the DP and the total mature CD8+ populations were
larger in Egr/H-Y than in NL/H-Y mice (Table 1). Thus,
H-Y CD8+ T cells appear to be more efficiently positively
selected in the presence of Egr-1 overexpression. More
surprisingly, significant numbers of mature H-Y CD8+ SP
thymocytes were also observed in female Egr/H-Y mice
on either a nonselecting H-2d/d background or a
2m-deficient (
2m0) H-2b/b background (14, 15), in which class I
MHC heavy chains are expressed at extremely low levels
(Fig. 1 a, middle and bottom; absolute numbers of both total
thymocytes and mature H-Y CD8+ SP cells are shown in
Table 1). Very few (<5 × 103) mature H-Y CD8+ SP cells
were detected in NL/H-Y mice on both H-2d/d and
2m0
backgrounds.
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H-Y CD8+ T cells were observed in peripheral lymphoid tissues in Egr/H-Y mice on H-2b/b, H-2d/d, and
2m0 backgrounds as well as in H-2b/b NL/H-Y female
mice (numbers shown in Table 1). As observed in the thymus, the percentage of T cells bearing the transgenic
chain (T3.70+) in splenic CD8+ cells was larger in H-2b/b
Egr/H-Y mice than in H-2b/b NL/H-Y mice (Table 1).
Peripheral H-Y CD8+ T cells revealed similar expression
patterns of several activation and memory markers such as
CD25, CD44, and Mel-14 in all types of mice (data not
shown).
Thus, in the presence of Egr-1 overexpression, H-Y
thymocytes appear to be efficiently positively selected even
by H-2d class I molecules and by the extremely low levels
of class I expressed in the absence of 2m, and mature to
become peripheral H-Y CD8+ T cells.
Whether positive selection of H-Y CD8+ T cells can
occur in the complete absence of class I MHC molecules
was assessed by grafting bone marrow cells from female
2m0 Egr/H-Y (H-2b/b) mice into female Kb
/
Db
/
2m
/
(I0
2m0; H-2b/b) recipient mice, which completely lack class I MHC (16, 17). As shown in Fig. 1 b, no
mature H-Y CD8+ SP thymocytes could be observed in
the thymi from I0
2m0 recipients into which either
2m0
Egr/H-Y or
2m0 NL/H-Y bone marrow cells were
transplanted. In contrast, female
2m0 (H-2b/b) recipients
into which
2m0 Egr/H-Y bone marrow cells had been
transferred harbored a significant number (6.2 ± 0.6 × 106;
n = 2) of mature H-Y CD8+ SP thymocytes. This result
strongly indicates that H-Y CD8+ T cells in H-2d/d and
2m0 Egr/H-Y mice were positively selected by H-2d class
I molecules or by the extremely low levels of class I expressed in the absence of
2m, but not by other molecules
like class II MHC. This was supported by the analysis of
2m, class II MHC doubly deficient (
2m0II0) Egr/H-Y
mice, which harbored grossly the same numbers of mature H-Y CD8+ SP thymocytes as
2m0 Egr/H-Y mice (Fig. 1 c).
Whether H-Y CD8+
T cells from H-2b/b, H-2d/d, and 2m0 Egr/H-Y mice
were functional was determined using matched cells from
H-2b/b female NL/H-Y mice as controls. First, purified
CD8+ T cells from each type of mouse were stimulated by
anti-CD3 mAb or Con A. As shown in Fig. 2, a and b,
CD8+ T cells from Egr/H-Y mice on all three backgrounds responded to both anti-CD3 mAb and Con A to a
comparable degree as those from H-2b/b female NL/H-Y
mice.
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Next, to determine whether the H-Y CD8+ T cells specifically recognize the H-Y male antigen, equal numbers of
H-Y CD8+ T cells from each type of mouse were injected
intravenously into either male or female H-2b/b C-deficient mice (which lack T cells; reference 18), and the number of H-Y CD8+ T cells in recipient spleens was evaluated
4 d after injection. As shown in Fig. 2 c, significant numbers of H-Y CD8+ T cells were detected in male recipients
into which either Egr/H-Y or NL/H-Y T cells were transplanted. In contrast, no obvious H-Y CD8+ T cells were
detected in female recipients. These results suggest that
H-Y CD8+ T cells from all types of mice recognized and
specifically responded to the male antigen; expansion of the
H-Y CD8+ T cells resulted in accumulation of a significant
population of those T cells in recipient spleens. Interestingly, the number of H-Y CD8+ T cells in male recipients
into which cells from
2m0 Egr/H-Y mice were injected
was slightly lower (~30% less) than those in recipients into
which cells from other types of mice were injected. This
may be due to rejection by NK cells, since NK cells recognize and kill
2m0 T cell blasts because of their extremely
low class I MHC expression (19, 20). Supporting this possibility, the in vitro response of H-Y CD8+ T cells from
2m0 Egr/H-Y mice to irradiated spleen cells from male
C57BL/6 (B6: H-2b/b) mice (containing NK cells that are
no longer able to kill target cells due to irradiation) was
comparable to the response of H-Y CD8+ cells from other
types of mice (Fig. 2 d). As observed in in vivo experiments, cells from all types of mice also failed to respond to
female spleen cells in vitro.
Thus, the H-Y CD8+ T cells positively selected by low
avidity ligands in H-2d/d or 2m0 Egr/H-Y mice were
functionally indistinguishable from cells positively selected
in NL/H-Y female H-2b/b mice in the absence of the Egr-1
transgene.
In male H-2b/b thymi, H-Y thymocytes are negatively selected and die through the apoptotic pathway (8). As
shown in Fig. 3, both Egr/H-Y and NL/H-Y thymocytes
have undergone clonal deletion. Interestingly, even more
efficient negative selection appears to occur in Egr/H-Y
mice, judging from the lower number of thymocytes in
Egr/H-Y mice. However, male H-2d/d and 2m0 Egr/H-Y
mice exhibited positive selection of H-Y CD8+ SP cells,
and the numbers of mature H-Y CD8+ SP cells were comparable to those of females (data not shown).
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Egr-1 transgenic mice were also
bred with another type of transgenic mouse expressing a
TCR recognizing the moth cytochrome C peptide in the
context of class II MHC I-Ek (called AND mice; reference
9). The AND TCR has a weak affinity for I-Ab class II
MHC molecules, and therefore, CD4+ SP thymocytes are
positively selected in an H-2b/b background (9, 21). The
upper panels of Fig. 4 show the CD4/CD8 profiles of mature thymocytes bearing the AND TCR (defined as transgenic TCR [V3]high, PNArlow) from Egr-1 transgene-positive AND mice (Egr/AND) and Egr-1 transgene-negative AND mice (NL/AND), both of which are on an H-2b/b
background. Egr/AND mice contained increased numbers
of both total thymocytes (1.1 ± 0.1 × 108 in Egr/AND
versus 0.8 ± 0.2 × 108 in NL/AND; n = 4 each) and
AND CD4+ SP thymocytes (3.5 ± 0.3 × 107 in Egr/
AND versus 2.2 ± 0.3 × 107 in NL/AND; n = 4 each).
This is reminiscent of the increased numbers of H-Y CD8+
SP cells in H-2b/b Egr/H-Y mice and, again, suggests more
efficient positive selection in the presence of the Egr-1
transgene. Interestingly, a significant number (8.0 ± 0.6 × 105; n = 4) of mature AND CD8+ SP cells were observed
in Egr/AND mice. It is possible that the AND TCR also
has a weak affinity for class I MHC (22) which is not normally sufficient to induce signal(s) for positive selection. Egr-1 overexpression might lower the threshold for positive selection, resulting in positive selection of AND cells
by class I MHC, creating a population of AND CD8+ SP
cells. This hypothesis is well-supported by analysis of Egr/ AND mice on a
2m0 background, where the number of
AND CD8+ SP cells strikingly decreased (Fig. 4, middle).
However, the difference between H-Y CD8+ SP cells and
AND CD8+ SP cells in
2m0 Egr-1 transgenic mice is intriguing; very efficient positive selection of H-Y CD8+ SP
cells was observed, while almost no positive selection of AND CD8+ SP cells was achieved, in the
2m0 background. This may be explained by a possible difference in the original affinity of H-Y TCR and AND TCR for class I
MHC molecules. The former may have a relatively high
affinity, which creates sufficient avidity for positive selection
even with extremely low expression levels of class I MHC,
when Egr-1 overexpression lowers the threshold. In contrast, the latter may have a low affinity, which contributes to
sufficient avidity with normal expression levels of class I
MHC in the presence of Egr-1 overexpression, but not with
the extremely low levels of class I MHC in
2m0 mice.
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In a class II MHC-deficient (II0) background, in which
"leaky" expression of class II molecules is not detected
(23), AND CD4+ cells virtually disappeared in both Egr/
AND and NL/AND mice (Fig. 4, bottom). This is consistent with failed positive selection of H-Y CD8+ T cells on
an I02m0 background, which entirely lacks class I MHC
expression. Deficiency of positive selection in the complete
absence of MHC molecules indicates a requirement of
TCR-mediated signaling to promote DP cells to the SP
stage, even in the presence of Egr-1 overexpression.
Alloreactive T cells, which recognize allo-MHC
complexes, are believed to be positively selected by auto-MHC/antigen-peptide complexes for which the alloreactive TCRs have sufficient, but not too much, avidity (24).
Therefore, whether Egr-1 expression might affect the selection of alloreactive T cells was determined using "normal" (no TCR transgenes introduced) Egr-1 transgenic mice. Fig. 5 a demonstrates allo-MLR responses of H-2b/b
Egr-1-TG and NL mice to m12 (H-2b with a mutated
class II) and BALB/c (H-2d/d) spleen cells. Transgenic T
cells exhibited enhanced responses to both
m12 and
BALB/c spleen cells. Similar enhanced allo-responses of
H-2d/d transgenic T cells were elicited towards B6 (H-2b/b)
spleen cells (Fig. 5 b). These enhanced allo-responses of
transgenic T cells appear to be due to an increased frequency of alloreactive T cells, but not to hyperproliferation
of each T cell, because fewer transgenic T cells responded
to the stimulators, as shown by the limiting dilution of responding cells in Fig. 5, a and b; and transgenic and negative littermate T cells exhibited comparable proliferative
responses to Con A (Fig. 5 c). Overall, Egr-1 overexpression appears to allow positive selection of a larger number
of alloreactive T cells, some of which originally might not
have had sufficient affinity with the auto-MHC to be positively selected in normal mice.
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Discussion |
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This is the first description of a transcription factor behaving as a modulator of positive selection; Egr-1 overexpression allows positive selection of thymocytes mediated by low avidity interactions with ligand(s) that are usually not sufficient, perhaps by lowering the threshold of avidity required for positive selection.
One might argue that Egr-1 overexpression induces survival effectors, such as antiapoptotic factors (25) in thymocytes. This may result in accumulation of the few leaky
mature SP cells, such as the very small number of H-Y
CD8+ SP cells that are detectable in both H-2d/d and 2m0
NL/H-Y mice, thus forming a seemingly positively selected cell population. However, this is unlikely for at least
three reasons. First, purified H-Y CD8+ SP cells from
H-2b/b, H-2d/d, or
2m0 female Egr/H-Y mice and H-2b/b
female NL/H-Y mice showed comparable survival curves
when cells were simply cultured in vitro (data not shown;
see Materials and Methods). Second, I0
2m0 recipient mice
into which
2m0 Egr/H-Y bone marrow cells were transplanted did not harbor the significant H-Y CD8+ SP cell
population that was seen in H-2d/d or
2m0 female Egr/
H-Y mice, although their thymi contained detectable leaky
H-Y CD8+ SP cells. Third, both male H-2b/b Egr/H-Y
and NL/H-Y thymocytes exhibited clonal deletion of H-Y cells, indicating no rescue from apoptosis of Egr/H-Y cells
undergoing negative selection. On the contrary, even more
efficient negative selection appears to occur in Egr/H-Y
mice. By these criteria, Egr-1 overexpression does not appear to induce survival effectors for thymocytes. It seems
more likely that Egr-1 overexpression allows an unusually
low avidity interaction between TCR and MHC/antigen-peptide complexes to initiate positive selection and further
development of thymocytes.
An alternative argument is that Egr-1 overexpression itself
might be sufficient to promote the DP to SP transition, with
no interaction between thymocytes and ligands required.
When the Egr-1 transgene was introduced into a RAG-deficient background, thymocytes did overcome the " selection" beyond the maturation block at the CD25+CD44
DN stage and developed into immature CD8 single positive (ISP) cells without any signaling through the TCR (7).
However, this was not the case for positive selection at the
DP stage. As we demonstrated in a previous report, although irradiated Egr-1 transgenic mice on a RAG-deficient background developed DP cells expressing the Egr-1
transgene but lacking TCR expression, neither CD4+ nor
CD8+ SP cells were observed (7). This suggests that DP
cells do require some interaction with ligand(s) to proceed
to the SP stage even in the presence of Egr-1 overexpression. This is supported convincingly by the analysis of
I0
2m0 recipient mice into which
2m0 Egr/H-Y bone
marrow cells were transplanted, in which no positive selection of H-Y CD8+ T cells occurred, as well as of Egr/
AND mice on a class II MHC-deficient background,
which harbored no AND CD4+ T cells. Hence, Egr-1 expression appears not to directly initiate transition of DP
cells to the SP stage, but may influence the threshold of
avidity required for positive selection.
Functions of T cells selected by unusually low avidity
TCR-MHC interactions in the presence of Egr-1 overexpression appear normal both in responses to various stimuli
and in specificity of antigen-peptide recognition, as demonstrated by the functional analysis of mature H-Y CD8+
T cells in H-2d/d and 2m0 Egr/H-Y mice. However,
long-term survival of a naive population of peripheral H-Y
CD8+ T cells in H-2d/d Egr/H-Y mice could be affected,
as a recent report by Tanchot et al. (16) implicated a requirement of the right MHC for survival of peripheral naive T cells. Therefore, a large proportion of peripheral H-Y
CD8+ T cells in H-2d/d Egr/H-Y mice might be newly
generated T cells. Tanchot et al. also demonstrated a requirement of only a nonspecific class I MHC for survival of
peripheral memory T cells (16). This is consistent with the
comparable expression patterns of memory markers such as
Mel-14 in the peripheral H-Y CD8+ T cells in H-2d/d
Egr/H-Y mice and H-2b/b NL/H-Y mice.
Phenotype in Egr-1 transgenic mice indicates a physiological role of Egr-1 in determining the threshold required for positive selection. Perhaps related to this, Egr-1 overexpression may also affect negative selection. As described above, more efficient negative selection appeared to be achieved by Egr-1 overexpression in male H-2b/b H-Y mice. However, thymocytes express other members of Egr family genes, such as Egr-2, -3, and -4, all of which share a highly conserved DNA-binding domain (5, 6). These members might functionally compensate for each other, and hence no alteration of thymic selection is observed in Egr-1-deficient mice as discussed previously (7, 31). Therefore, a set of genes controlled by a common binding site for Egr genes, including Egr-1, might be critical in defining the overall thresholds required for thymic selection. Further characterization of Egr target genes should shed light on the precise molecular mechanism underlying the positive/negative selection of thymocytes.
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Footnotes |
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Address correspondence to T. Miyazaki, Basel Institute for Immunology, Grenzacherstrasse 487, postfach CH-4005, Basel, Switzerland. Phone: 41-61-605-1323; Fax: 41-61-605-1364; E-mail: miyazaki{at}bii.ch
Received for publication 21 April 1998 and in revised form 3 June 1998.
F.A. Lemonnier is supported by the Association pour la Recherche sur le Cancer. Basel Institute for Immunology was founded and is supported by F. Hoffman-La Roche Ltd. (Basel, Switzerland).We thank Drs. S. Gilfillan and K. Campbell for critical reading of the manuscript; E. Wagner, W. Metzger,
and R. Zedi for help with the mice; Dr. P. Kisielow for T3.70 mAb; and Dr. C. Benoist for 2m0II0 mice
and KT-3 mAb.
Abbreviations used in this paper DP, double positive; Egr-1, early growth response 1; Egr/H-Y, Egr-1 transgene-positive H-Y transgenic; I0, class I MHC-deficient; II0, class II MHC-deficient; ISP, immature SP; NL/H-Y, Egr-1 transgene-negative H-Y transgenic; PNAr, peanut agglutinin receptor; RAG, recombination-activating gene; SP, single positive; TG, transgenic.
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References |
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