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Mouse Immunogenetics, U462, Institut National de la Santé et de la Recherche Médicale, Institute of Hematology, Saint-Louis Hospital, Paris, France; and
Institut für Anthropologie und Humangenetik, Ludwig-Maximilians-Universität, Munich, Germany
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 Top Abstract IntroductionMaterials and MethodsResults and DiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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1 and
2 domains of the class Ia heavy chains form the peptide binding
groove (2), and most polymorphisms in these molecules are around this
binding groove. At the cell surface, the peptide and class Ia complex
is recognized by peptide-specific and allospecific CTL. The MHC class Ib molecules in humans include principally HLA-E, HLA-F, and HLA-G molecules. Like the classical MHC class I loci, the nonclassical HLA class I genes are highly transcribed in many tissues (3), but they display only little genetic variation. One such molecule, HLA-E, is currently of particular immunological interest. In vitro studies have shown that HLA-E preferentially binds a peptide derived from amino acid residues 3–11 of the signal sequences of most HLA-A, -B, -C, and -G molecules (4, 5, 6, 7). Recent studies have revealed that HLA-E molecules function as ligands for NK cell inhibitory receptors (8, 9, 10, 11), and that the recruitment of HLA-E on the surface of transfected cells by the addition of class I signal sequence-derived peptides is enough to protect target cells from lysis by NK cell clones (9, 10, 11).
Until recently (11), the lack of HLA-E-specific reagents rendered in vivo study of HLA-E cell-surface expression in humans impossible; the only data available concerning HLA-E cell-surface expression were obtained using human (class Ia-defective) transfectants (6) or mouse cells transfected with HLA-E genes (12). To facilitate the production of such reagents, and to further the study of HLA-E functional properties, we have derived transgenic mice expressing HLA-E molecules. With the help of these transgenic mice, we have produced (13) a mAb (V16) that binds specifically to the cell surface of HLA-E-expressing cells, whether this molecule is naturally expressed (in human cells) or is the product of a transfected gene (in mouse cells). We report here that HLA-E behaves as a transplantation Ag in mice, and that a significant portion of the mouse CTL response involved in the recognition of HLA-E recognizes HLA-E as an intact molecule and not as an HLA-E-derived peptide presented by a mouse MHC molecule.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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All mouse strains used in this study were bred and maintained in our own colony (Mouse Immunogenetics, U462, Institut National de la Santé et de la Recherche Médicale, Saint-Louis Hospital, Paris, France).
Cells
Con A-induced blasts (Con A blasts) used as targets in cell-mediated lympholysis (CML)4 were prepared by incubating 4 ml of spleen cell suspension (5 x 106/ml) with Con A (5 µg/ml; Sigma, St. Louis, MO) for 48 h in culture medium. The following human cell lines were also used as targets in CML: the homozygous EBV-transformed B cell line HO104 (HLA-A3, B7, Cw7; IHW 9082 (14)) and the HLA class I-negative cell line Daudi (IHW 9366 (14)).
Monoclonal Abs
The following mAbs were used: B9.12.1, HLA class I-specific (15, 16); TGD 15, human ß2m-specific (17); AF6-88.5, H-2Kb-specific (PharMingen, San Diego, CA); and SF1-1.1.1, H-2Kd-specific (ATCC HB159).
Generation of HLA-E transgenic mice
Transgenic mice were produced as described (18). Briefly, DNA
fragments were purified free of vector DNA and flanking human genomic
DNA by preparative agarose gel electrophoresis and were microinjected
into fertilized oocytes. Embryos surviving microinjection were
reimplanted into the oviducts of pseudopregnant females, and offsprings
were tested for the integration of the transgene by Southern blot
analysis of tail-derived DNA using a PCR product encoding exon 3 of the
HLA-E gene as a probe (see Fig. 1
).
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Cells (5 x 105) were successively incubated with saturating concentrations of mAb and FITC-conjugated goat F(ab')2 anti-mouse Ig. Both incubations were conducted on ice for 30 min, followed by two washing steps. Cytofluorometry was conducted on a Becton Dickinson (Mountain View, CA) FACScan and analyzed with Cell Quest software.
Skin grafting
Skin grafting was performed as described (19). Briefly, skin from tail of donor mice was grafted onto the flank of a recipient mouse. The graft was covered with gauze and plaster that were removed on day 8. Grafts were scored daily until rejection (defined as >80% of grafted tissue rejected) or until 150 days after transplantation.
In vivo generation of HLA-E-reactive CTL
Recipient mice were injected with 107 irradiated (25
Gy) spleen cells in the hind footpads. After 3 days, cell suspensions
were prepared from draining lymph nodes and cells were cultured in
vitro for 4 more days in the absence of any stimulating cells in
culture medium containing Con A-stimulated rat spleen cell supernatant
as a lymphokine source (50 U IL-2/ml). The culture medium was MEM
-medium supplemented with 100 U/ml penicillin, 100 µg/ml
streptomycin, 2 mM L-glutamine, 10% heat-inactivated FCS
(all products from Life Technologies, Gaithersburg, MD), and 5 x
10-5 M 2-ME (Sigma).
In vitro generation of HLA-E-reactive CTL
Bulk lymphocyte cultures were set up confronting spleen cells from the naive or primed mice to irradiated (25 Gy) HLA-E-expressing H-2-matched spleen cells. The HLA-E-reactive cytotoxic T cell line (CIA) was propagated by weekly restimulation with HLA-E-expressing H-2b spleen cells.
Split-well analysis
Cells (in various dilutions) harvested from the draining lymph nodes were cultured in round-bottom wells (200 µl/well) for 4 days in culture medium supplemented with Con A-stimulated rat spleen cell supernatant as a lymphokine source (50 U IL-2/ml). Cell suspensions were then divided into three aliquots of 60 µl and transferred into wells containing 5 x 103 51Cr-labeled target cells in 140 µl medium. After a 4-h incubation, the supernatants were harvested and the 51Cr release was counted. Wells in which the experimental 51Cr release exceeded the mean (six replicate wells) spontaneous release of a given target plus three times the SD value were considered positive.
CML assay
Five thousand 51Cr-labeled target cells were incubated with effector cells at various E:T ratios in round-bottom wells for 4 h. The percentage of specific 51Cr release was calculated as (experimental - spontaneous release)/(maximum - spontaneous release) x 100.
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Results and Discussion |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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An 8-kb or a 21-kb fragment (Fig. 1
)
of the genomic HLA-E cosmid clone cd3.14 (20) was microinjected into
fertilized oocytes. These fragments contain the complete
E*01033 gene. Southern blot hybridization of an HLA-E probe
(Fig. 1) with offspring tail DNA identified nine independent founder
mice, two (nos. 36 and 45) containing the 21-kb fragment, and seven
with the 8-kb fragment. Both founders carrying 21-kb fragments
transmitted transgenes to their offsprings, whereas only five (nos. 72,
81, 82, 86, and 89) of the founder animals with 8-kb fragments
transmitted the HLA-E gene to their offsprings. Northern blot analysis
of liver and spleen RNA revealed (data not shown) that six of seven
transgenic lines (nos. 36, 45, 72, 81, 82, and 86) express the HLA-E
gene.
Each of the founder mice was mated to mice of an already established C57BL/10 transgenic line expressing human ß2m (M-TGM). Both transgenes segregated in backcross matings as simple Mendelian traits. Thus, in offsprings four types of mice were obtained: single transgenic mice carrying only the transgene for the HLA-E heavy chain (E-TGM) or human ß2m (M-TGM), double transgenic mice carrying both human genes (EM-TGM), and mice without any transgene (non-TGM). At present, it is not known where in the mouse genome the transgenes are incorporated. They do not occur in linkage with H-2 (data not shown). To obtain HLA-E congenic lines expressing H-2b or H-2d haplotypes, the mice were typed for H-2, and double transgenic mice (EM-TGM) carrying H-2b or H-2d were repeatedly backcrossed to C57BL/10 background mice with H-2b (B10) or H-2d (B10.D2) haplotypes.
Expression of the HLA-E transgene products detected by flow cytometric analysis
Cell-surface expression was analyzed by flow cytometry of lymph
node cells from double (EM) and single (E) transgenic mice. As a
control, transgenic mice with human ß2m (M-TGM) were
used. For this analysis, mAbs were required that could detect the
transgenic HLA-E molecules in association with murine
ß2m. One of the most widely used serological reagents for
detecting human class I Ag expression in human or in transfected mouse
cells is the monomorphic mAb W6/32 (21). However, Lemonnier and
colleagues (15) have shown that the HLA-B7 epitope recognized by W6/32
on HLA-B7-expressing mouse cells is abolished when the molecule is
associated with murine ß2m as opposed to human
ß2m. In contrast, it was shown that the alternate HLA-B27
epitope recognized by B9.12.1 mAb was unaffected by the species origin
of the associated ß2m (15). In view of this, cell-surface
expression of HLA-E in the current study was assessed in flow cytometry
using this mAb. TGD 15 mAb was used for detection of human
ß2m expression. Flow cytometry analysis showed a
detectable level of HLA-E cell-surface expression on lymph node cells
originating from double transgenic mice of the 72, 81, 82, and 86 lines
and a very low level of expression on lymph node cells from mice of
line 45. No HLA-E expression was detected on the lymph node cells from
double transgenic mice of the 36 line and on lymph node cells from
single transgenic mice (E-TGM, line 81), carrying only the transgene
encoding the HLA-E heavy chain. The results (Fig. 2A) clearly indicate that
detectable levels of HLA-E molecules were observed only in the presence
of human ß2m on lymph node cells from double transgenic
mice (EM-TGM).
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). Cells
from transgenic mice with human ß2m (M-TGM) served as
controls. Similarly as in Fig. 2A, the B9.12.1 mAb showed
clearly the expression of HLA-E molecules on the surface of double
transgenic mouse spleen cells. Rather low but reproducible binding of
B9.12.1 mAb was obtained on peripheral blood cells. The level of
H-2Kb-specific-mAb staining on peripheral blood and spleen
cells from double transgenic mice (EM-TGM) was not significantly
different from that observed on M-TGM cells. Thus, for the 81
transgenic line, the level of endogenous class I Ag expression
(H-2Kb) in double EM-TGM mice was not altered, despite the
efficient expression of the transgenic HLA-E molecule. Similarly, no
difference was found in the level of endogenous class I expression on
the surface of lymph node cells from single (E) or double (EM)
transgenic mice (data not shown).
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). A significant percentage of the
population stained positively for HLA-E or H-2Kb at an
intensity similar to spleen cells, while the remaining part was
negative. A similar distribution of H-2Kb was obtained for
HLA-E negative thymocytes originating from M-TGM.
As mentioned, the HLA-E transgene was transferred into two H-2 congenic
strains (B10 and B10.D2). It has been reported (22) that the level of
HLA class I transgene expression on the cell surface is dependent on
the host H-2 haplotype. To determine whether the H-2b and
H-2d haplotypes are able to alter the expression of the
HLA-E transgene, we have compared the cell-surface expression of HLA-E
molecules on the lymph node cells from double HLA-E trangenic mice
carrying either of these H-2 haplotypes. The flow cytometry analysis
shown in Fig. 4 suggests that there is no
significant difference in the level of HLA-E expression on the surface
of lymph node cells from double EM-TGM expressing H-2b or
H-2d haplotypes. As shown in Fig. 4, the Con A activation
did not lead to an increase of HLA-E expression. Similarly to lymph
node cells, there was no difference in HLA-E expression between the Con
A blasts carrying H-2b or H-2d haplotypes.
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To determine whether the HLA-E transgene product in HLA-E
transgenic mice is expressed in a form that could be detected by the
immune system of mice, we conducted skin graft experiments. All mice
used for these experiments were originated from the 81 line. As already
mentioned, the founder (no. 81) transgenic for HLA-E was mated to mice
of an already established C57BL/10 transgenic line expressing human
ß2m. The double transgenic mice (EM) carrying
H-2b were repeatedly backcrossed to C57BL/10 background
mice with H-2b (B10) haplotypes. The mice used in our study
derived from the F3 generation of backcross matings.
Recipient mice expressing human ß2m were engrafted with
three tail skin grafts: an autologous graft, a graft originated from a
littermate expressing only human ß2m (control
transplants), and a graft originated from a littermate expressing both
HLA-E and human ß2m (HLA-E transplants). All tail skin
autografts were accepted. Table I shows
that all HLA-E grafts were rejected with a mean survival time of 23.6
days. Nearly 50% of control transplants (HLA-E-negative) were also
rejected, however with a very prolonged mean survival time (58.9 days).
The rejection of control transplants might be due to remaining minor
histocompatibility Ag disparities. These results reveal that HLA-E
molecules are recognized as functional transplantation Ags in mice.
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Cell-mediated cytotoxicity assays were performed to determine
whether normal mice could generate a cytotoxic T cell response against
HLA-E-expressing transgenic spleen cells. The T cell response directed
against HLA-E was first investigated by carrying out in vitro assays
for cytotoxicity. No cytotoxic T cell activity was detected when CTL
were generated in vitro in unidirectional mixed lymphocyte cultures
using spleen cells from naive B10 (H-2b) mice as responder
cells and irradiated spleen cells from H-2-matched HLA-E transgenic
mice (EM-TGM) as stimulator cells. However, when a primary CTL response
was induced in vivo, a significant CTL response was obtained (Fig. 5, A and B). This
in vivo experimental approach is based on our previous observations
(23, 24) that injection of spleen cells from allogeneic mice into the
hind footpads leads to the development of CTL within the draining lymph
nodes. In the present experiments, spleen cells from double EM-TGM
originating from line 45 (Fig. 5A) or from line 81 (Fig. 5B) carrying H-2b haplotypes were injected into
the hind footpads of nontransgenic H-2-compatible mice. After 3 days,
cell suspensions were prepared from draining lymph nodes and cells were
cultured for 4 more days in the absence of any stimulating cells. After
this period, which was required to allow full differentiation of
sensitized CTL precursors (25), lymph node cells were tested for the
presence of CTL in a 51Cr release assay using Con A blasts
as target cells. As shown in Fig. 5, A and B, CTL
generated in this way lysed (15–35% at an E:T ratio of 20:1) target
cells from HLA-E transgenic mice (EM-TGM) but did not lyse target cells
from human ß2m transgenic mice (M-TGM). Despite the fact
that the flow cytometry analysis showed a very low level of HLA-E
expression on lymph node cells from mice of line 45 (Fig. 2A), it was sufficient to elicit a primary CTL response
(Fig. 5A).
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C show that
Con A blasts from both H-2b and H-2d EM-TGM
were killed. The percent specific 51Cr release achieved in
this secondary response was about twice higher (70% at an E:T ratio of
20:1) than that obtained in the primary in vivo-induced response. These
CTL were also capable of killing human EBV-transformed cells (60% at
an E:T ratio of 20:1). No killing was observed with HLA-E-negative
mouse cells, nor with Daudi cells, a human cell line that fails to
express both cell surface class Ia and Ib molecules due to a defective
ß2m gene. Lysis of human targets and of H-2-mismatched
HLA-E-positive mouse cells indicate that the elicited CTL recognize
HLA-E as an intact molecule and not as an HLA-E-derived peptide
presented by a mouse MHC molecule (H-2-unrestricted recognition). Induction of anti-HLA-E cytotoxic T cell response in HLA class Ia transgenic recipients
The experimental protocol using i.p. immunized mice was further
applied to attempt to induce an anti-HLA-E CTL response in HLA-B27
transgenic mice (27M-TGM), in which the HLA class Ia transgenic
products are tolerated as a part of self-MHC. Spleen cells from
transgenic H-2b mice expressing HLA-E molecules were i.p.
injected in H-2-compatible HLA-B27-expressing recipients. Spleen cells
from the injected mice were restimulated in vitro with irradiated
spleen cells from EM-TGM and tested for cytotoxic activity in a
51Cr release assay against HLA-E transgenic
(H-2b and H-2d) target cells and human cell
lines (Fig. 5D). Significant killing of both
H-2b and H-2d HLA-E-positive mouse target cells
was obtained with effector cells from HLA-B27 transgenic mice (60%
specific lysis at an E:T ratio of 20:1). There was no 51Cr
released from HLA-E-negative H-2 syngeneic targets above background
levels. These CTL were also capable of killing human EBV-transformed
cells (35% at an E:T ratio of 20:1). Daudi, HLA class I-negative human
cells were not killed. These results indicate that HLA-E expressed on
transgenic spleen cells can serve as an immunogenic element in HLA
class Ia transgenic mice.
Analysis of primary in vivo induced anti-HLA-E CTL
To analyze in detail the primary anti-HLA-E cytotoxic T cells
and especially those involved in graft rejection, we have utilized the
in vivo priming approach together with a split-well analysis technique.
This test permits us to estimate how many CTL cultures could be
generated that are only specific for the H-2-matched HLA-E transgenic
targets. We examined 60 individual microcultures set up from draining
lymph nodes of nontransgenic H-2b mice that were injected
into the hind footpads with H-2-matched HLA-E-expressing spleen cells.
The results from a representative experiment (Fig. 6) show that from the total of 39
reactive wells, 28 wells contained CTL that lysed HLA-E-positive
H-2-matched (H-2b) as well as H-2-mismatched
(H-2d) targets. Thus, these primary CTL (72% of reactive
wells) displayed an H-2-unrestricted reaction pattern. Seven wells
(18% of reactive wells) contained CTL recognizing only
H-2b (syngeneic) HLA-E-positive targets and included mouse
CTL that recognized HLA-E in an H-2-restricted manner. The levels of
51Cr release from wells with CTL exhibiting H-2-restricted
or H-2-nonrestricted reactivity pattern were comparable. Several
split-well analysis experiments were performed using the same
donor-recipient combination. We observed that the recipients
originating from a given inbred strain exhibited an individual
variation in their HLA-E-induced CTL response. The percentage of
reactive wells containing CTL that recognize HLA-E Ags in an
H-2-unrestricted manner ranged from 64% to 79% and those with CTL
recognizing HLA-E in an H-2-restricted manner ranged from 4% to 21%.
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The flow cytometry analysis shown in Fig. 2A indicates
that no HLA-E molecules could be detected on the surface of the cells
from single transgenic mice (E-TGM) carrying only the transgene for the
HLA-E heavy chain. A CTL line (CIA) was derived from primary in vivo
induced HLA-E-reactive CTL and maintained by periodic stimulation with
irradiated H-2b EM-TGM spleen cells. The CIA line was
assayed (Fig. 2B) on a panel of Con A blasts originating
from double (EM, left panel) and single (E, right
panel) transgenic mice coming from various HLA-E transgenic lines
(lines 36, 45, 72, 81, 82, and 86). The HLA-E-reactivity of this line
is demonstrated by its cytotoxic activity toward target cells from
HLA-E transgenic mice from the 36, 45, 72, 81, 82 and 86 lines but not
from the human ß2m transgenic line (M-TGM). It is
noteworthy that the CIA cytotoxic T cell line was cytotoxic for cells
from both single E- and double EM-transgenic mice. As shown in Fig. 2B (left panel), the lysis of Con A blasts
originating from transgenic mice of the 36 and 45 transgenic lines was
less efficient (55% at an E:T ratio of 20:1) than that of blasts from
mice of the 72, 81, 82 and 86 transgenic lines (75–80% at an E:T
ratio of 20:1). The CIA T cell line lysed efficiently (Fig. 2B, right panel) the Con A blasts from single
E-TGM of the 72, 81, 82, and 86 transgenic lines (40–50% at an E:T
ratio of 20:1). Only marginal lysis of Con A blasts from single E-TGM
of the 45 transgenic line was observed (15% at an E:T ratio of 20:1).
There was no 51Cr released above background levels from
targets originating from single E-TGM of the 36 transgenic line.
The CIA line is able to lyse both H-2b (Con A blasts from lines 36, 45, 72, 82, and 86) and H-2d (Con A blasts from line 81) HLA-E transgenic cells. Lysis of H-2-mismatched HLA-E-positive targets indicates that this line recognizes HLA-E in an H-2-unrestricted manner.
Flow cytometry analysis (Fig. 2A) of HLA-E cell-surface
expression in double (EM) transgenic lines shows that in mice from four
(72, 81, 82, and 86) transgenic lines a substantial expression of HLA-E
was detected by the B9.12.1 mAb. Low or no cell-surface expression of
HLA-E was detected in transgenic mice from line 45 and line 36,
respectively. Thus, the observed differences in efficiency of lysis of
double (EM) transgenic targets by the CIA cytotoxic T cell line might
be due to the differences in HLA-E cell-surface expression detected by
flow cytometry (Fig. 2A). No cell-surface expression of
HLA-E molecules was detected in single (E) transgenic lines from all
transgenic lines when tested with B9.12.1 mAb (Fig. 2A for
the 81 line) or with various HLA class I-specific mAb (data not shown).
Thus, it appears that the TCR of the CIA cytotoxic cell line is able to
detect HLA-E gene products on cells with serologically undetectable
HLA-E cell-surface expression.
HLA-E cell-surface expression in transgenic mice: the role of human ß2m
Our motives for generating transgenic mice expressing HLA-E
molecules were to obtain a reliable source of cells expressing HLA-E
molecules and to provide a means to investigate the biological function
of HLA-E molecules in vivo. However, before such mice can be utilized,
expression of HLA-E molecules on the cell surface and their recognition
by T cells must be demonstrated. The results presented in this paper
indicate that both DNA fragments (8 kb and 21 kb) containing the gene
for the human HLA nonclassical class I Ag HLA-E can be functionally
expressed on the cell surface in transgenic mice (Fig. 2).
It has been reported that HLA-E heavy chain polypeptides are not
efficiently accumulated on the surface of mouse cells unless human
ß2m is also present. In mouse myeloma cells (X63)
transfected only with HLA-E heavy chain genes, no HLA-E Ag could be
detected on the cell surface even upon adding exogenous human
ß2m (26). This result suggests that no HLA-E molecules
detectable with HLA class I specific mAbs are transported to the cell
surface in the absence of endogenous human ß2m. The flow
cytometry analysis shown in Fig. 2A indicates that no HLA-E
molecules could be detected on the surface of the cells from mice
carrying only the transgene for HLA-E heavy chain. However,
HLA-E-reactive mouse CTL (Fig. 2B) that recognize HLA-E in
an H-2-unrestricted manner lysed efficiently the target cells from
single transgenic mice suggesting that the TCR of mouse CTL recognizes
HLA-E gene products on the cells with serologically undetectable HLA-E
cell-surface expression. Recently, by limiting dilution of the CIA
line, we have derived a CTL clone (TER-1) bearing an ß receptor
that specifically recognizes HLA-E gene products complexed with MHC
class I signal sequence-derived peptides (27). This clone lysed
efficiently target cells from both single E and double EM transgenic
mice. Taken together, our data show that the HLA-E protein can be
transported to the cell surface in the absence of human
ß2m, presumably by association with murine
ß2m.
Alloantigenic function of HLA-E on transgenic cells
From the experiments reported here several conclusions can be
drawn about interactions between HLA-E molecules and murine T cells.
First, despite the relatively low level of HLA-E cell-surface
expression in HLA-E transgenic mice, all EM-TGM skin grafts were
rejected (Table I), demonstrating that HLA-E functions as a strong
transplantation Ag in mice.
Second, HLA-E on the surface of transgenic spleen cells can trigger the
generation of CTL in normal mice, which are capable of killing both
HLA-E-positive mouse targets and human EBV-transformed cells. CTL
reactive with allogeneic and xenogeneic MHC class I molecules are
readily detectable in unprimed mice. While the level of HLA-E expressed
was rather low compared with that of endogenous H-2 class I (Fig. 3),
it was sufficient to elicit a CTL response in unprimed animals (Fig. 5, A and B). As shown in this figure,
HLA-E-expressing spleen cells from transgenic line 45 carrying the
21-kb fragment (Fig. 5A) as well as those from line 81 with
the 8-kb fragment (Fig. 5B) were able to induce in vivo a
significant CTL response, suggesting that there are no functional
differences in alloantigenicity of HLA-E molecules expressed in these
two transgenic lines. Moreover HLA-E expressed on transgenic spleen
cells was shown (Fig. 5D) to elicit
anti-HLA-E CTL in HLA-B27 transgenic recipients, in which the HLA
class Ia transgenic products are tolerated.
Finally, in the present report we have described the analysis of CTL
activated in vivo in response to a local graft of spleen cells
expressing HLA-E molecules (Fig. 6). The results with HLA-E-reactive
CTL induced in vivo are consistent with our previous studies (24)
showing that the anti-HLA-B27 CTL response is predominantly
H-2-unrestricted. Our experimental protocol allowed us to detect two
populations of CTL: one not restricted by H-2, and the other
H-2-restricted. Thus xenogeneic HLA-E molecules can be perceived by TCR
of the mouse as MHC molecules similar to their own, or as Ags
recognized in the context of mouse self-MHC products. However,
according to our results, the latter type of xenogeneic recognition
represents a minor part of the primary anti-HLA-E CTL response
activated in graft-draining lymph nodes.
In summary, nonclassical HLA-E and classical HLA class I molecules display identical alloantigenic behavior when expressed in transgenic mice. We are currently investigating whether HLA-E might play a role as a restriction element for viral proteins in vivo. Nonclassical MHC class I molecules are rapidly emerging as key mediators of immune recognition. HLA-E transgenic mice provide a powerful model for further rational investigation of the biological function of HLA nonclassical class I genes without need for the ethically difficult experimentation with human.
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Acknowledgments |
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Footnotes |
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2 Current address: Mouse Immunogenetics, U462 Institut National de la Santé et de la Recherche Médicale, Institute of Hematology, Saint-Louis Hospital, Paris, France
3 Address correspondence and reprint requests to Dr. Marika Pla, Mouse Immunogenetics, U462 Institut National de la Santé et de la Recherche Médicale, Institute of Hematology, Saint-Louis Hospital, 1 avenue Claude Vellefaux, 75475 Paris Cedex 10, France. E-mail address:
4 Abbreviations used in this paper: CML, cell-mediated lympholysis; ß2m, ß2-microglobulin; TGM, transgenic mouse/mice.
Received for publication October 15, 1998. Accepted for publication February 12, 1999.
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References |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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-helices residues of the MHC class I molecule. J. Immunol. 161:148.This article has been cited by other articles:
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  T. Tilburgs, D. L. Roelen, B. J. van der Mast, G. M. de Groot-Swings, C. Kleijburg, S. A. Scherjon, and F. H. Claas Evidence for a Selective Migration of Fetus-Specific CD4+CD25bright Regulatory T Cells from the Peripheral Blood to the Decidua in Human Pregnancy J. Immunol., April 15, 2008; 180(8): 5737 - 5745. [Abstract] [Full Text] [PDF]
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J. D. Miller, D. A. Weber, C. Ibegbu, J. Pohl, J. D. Altman, and P. E. Jensen Analysis of HLA-E Peptide-Binding Specificity and Contact Residues in Bound Peptide Required for Recognition by CD94/NKG2 J. Immunol., August 1, 2003; 171(3): 1369 - 1375. [Abstract] [Full Text] [PDF]
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 C. Romagnani, G. Pietra, M. Falco, E. Millo, P. Mazzarino, R. Biassoni, A. Moretta, L. Moretta, and M. C. Mingari Identification of HLA-E-specific alloreactive T lymphocytes: A cell subset that undergoes preferential expansion in mixed lymphocyte culture and displays a broad cytolytic activity against allogeneic cells PNAS, August 20, 2002; 99(17): 11328 - 11333. [Abstract] [Full Text] [PDF]
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J. Li, I. Goldstein, E. Glickman-Nir, H. Jiang, and L. Chess Induction of TCR V{beta}-Specific CD8+ CTLs by TCR V{beta}-Derived Peptides Bound to HLA-E J. Immunol., October 1, 2001; 167(7): 3800 - 3808. [Abstract] [Full Text] [PDF]
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 A. Horuzsko, F. Lenfant, D. H. Munn, and A. L. Mellor Maturation of antigen-presenting cells is compromised in HLA-G transgenic mice Int. Immunol., March 1, 2001; 13(3): 385 - 394. [Abstract] [Full Text] [PDF]
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Y. Szpak, J.-C. Vieville, T. Tabary, M.-C. Naud, M. Chopin, C. Edelson, J. H. M. Cohen, J. Dausset, Y. de Kozak, and M. Pla Spontaneous retinopathy in HLA-A29 transgenic mice PNAS, February 8, 2001; (2001) 51595998. [Abstract] [Full Text]
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Y. Szpak, J.-C. Vieville, T. Tabary, M.-C. Naud, M. Chopin, C. Edelson, J. H. M. Cohen, J. Dausset, Y. de Kozak, and M. Pla Spontaneous retinopathy in HLA-A29 transgenic mice PNAS, February 27, 2001; 98(5): 2572 - 2576. [Abstract] [Full Text] [PDF]
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), 45
(x), 72 (
), 81 (
), 82 (+), and 86 (
) as targets.
Con A blasts from human ß2m transgenic mice (
) were
used as negative controls. All target cells carried H-2b
haplotypes, with the exception of targets from line 81 which carry
H-2d haplotypes.
