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Departments of
*
Molecular Immunology and
Cell Biology, Institute of Development, Aging, and Cancer, Tohoku University, Sendai, Japan;
Central Institute for Experimental Animals, Kawasaki, Japan; and
Department of Immunology, Tokai University School of Medicine, Isehara, Japan
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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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Numerous studies have tried to unravel the molecular mechanisms involved when DP thymocytes choose to follow either the CD8 SP or the CD4 SP lineage. A number of signaling and apoptosis-related molecules have been reported to be involved in this decision and include a transmembrane receptor, Notch I (1, 2, 3); a coreceptor-coupled kinase, Lck (4, 5); a pathway of mitogen-activated protein kinases (6, 7); and an anti-apoptotic bcl-2 gene (8). With regard to transcription factors, however, there is only one reported example, namely, IFN-regulatory factor 1. In IRF1-/- mice, the generation of CD8 SP, but not CD4 SP, thymocytes is severely impaired (9), and the impairment appears to be intrinsic to the thymocytes themselves rather than due to the lack of MHC class I expression on thymic epithelial cells (10). Analysis of the CD4 silencer that functions in CD8 SP cells has also led to the identification of DNA-binding proteins, such as helix-loop-helix type hairy/enhancer of split homologue-1 (11) and homeodomain type silencer-associated factor (12).
AML1/Runx1 is one member of the mammalian runt gene family and encodes a DNA-binding subunit of a heteromeric transcription factor called polyomavirus enhancer binding protein 2/core binding protein (PEBP2/CBF) (13, 14). The physiological and pathological significance of AML1 is well established in the fields of hemopoietic development and human leukemogenesis (see the reviews in Refs. 15, 16, 17). AML1 is abundantly expressed not only in hemopoietic progenitors and myeloid cells, but also in T lymphocytes present in the thymus and spleen (18). Based on this expression profile, we have been studying the roles, if any, that the AML1 transcription factor may play in the differentiation and/or function of T lymphocytes. For example, we found that overexpression of AML1 renders a T cell hybridoma resistant to TCR-mediated apoptosis (19). This resistance was acquired by the down-regulation of Fas ligand expression. In addition, we generated mice in which AML1 activity is diminished compared with that in wild-type mice. These mice include AML1+/- mice (20) and mice in which a dominant interfering form of AML1 is expressed in T lymphocytes (21). In both mouse strains, positive selection is moderately to severely impaired, resulting in a reduction in the numbers of both CD8 SP and CD4 SP thymocytes.
We noted in our previous publication that in mice with diminished AML1 function the CD8 SP cell numbers were more severely reduced than the CD4 SP thymocyte numbers (21). This observation suggests that AML1 may also play a role in regulating the choice DP cells make to follow one or the other SP lineages. Thus, we assessed in this study the effect of overexpressing AML1 on the fate of DP cells. This was done by expressing AML1 as a transgene. Our data show that overexpression of AML1 promotes the development of CD8 SP cells to the extent that even MHC class II-restricted thymocytes are skewed into the CD8 SP lineage.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The expression plasmid pCX2neoHA/
B1 harbors hemagglutinin
(HA)-tagged, murine AML1b and was described previously
(19). The 1.4-kbp EcoRI fragment containing the
AML1 coding region was excised from pCX2neoHA/B1 and inserted into
the EcoRI site of human CD2 minigene (22). The
resulting plasmid was designated pCD2-HA/AML1b.
Animals
Transgenic mouse lines expressing the AML1 protein were
generated as follows. The DNA of pCD2-HA/AML1b was digested by
KpnI and XbaI, and the purified fragment
containing the AML1 coding region was microinjected into fertilized
eggs of (DBA2xC57BL/6) F1 mice. Transgenic
founders were identified by PCR on tail genomic DNA using primers
specific for Runt domain of murine AML1 as described previously
(21). Transgenic mice were maintained by back-crossing
founders to C57BL/6. 
2-Microglobulin
(2m)-deficient mice were purchased from The
Jackson Laboratory (Bar Harbor, ME). I-A-deficient mice
(23) and DO11.10 TCR transgenic mice (24)
were provided by D. Mathis and D. Loh, respectively. Four- to 12-wk-old
mice were used in this study.
Immunoblot analysis
All the procedures necessary for immunoblot analysis, including
protein extraction, electrophoresis, transfer to the filter, and
immunoreaction, were performed as described previously (25, 26). The CD4 SP and CD8 SP fractions were prepared from
thymocytes as described previously (21). The monoclonal
anti-HA Ab, 3F10, was obtained from Roche Diagnostics
(Indianapolis, IN). Raising and characterization of anti-AML1
peptide Ab were described previously (27). Briefly, the
antiserum was raised against the carboxyl-terminal, 15 aa residues of
murine AML1b/PEBP2B protein. The antiserum cross-reacts with the
products of mammalian runt gene family, since they all share
the common VWRPY sequence at their extreme carboxyl-terminal
ends.
EMSA
The procedures for preparing whole cell extract and for EMSA were described previously (28). The PEBP2/CBF binding sequence from Polyomavirus enhancer was used as a probe to detect PEBP2/CBF DNA binding activities.
Flow cytometry
Single-cell suspensions from thymus and spleen were stained with
the appropriate Abs as described previously (21).
Fluorescein-conjugated Abs with following specificities were used:
PE-CD4 (H129.19), FITC-CD4 (RM4-5), CyChrome-CD4 (RM4-5), FITC-CD8a
(53-6.7), PE-CD8a (53-6.7), FITC-TCR (H57-597), FITC-HSA (M1/69; BD
PharMingen, San Diego, CA). Biotinylated KJ1–26 and
streptavidin-RED670 (Life Technologies, Gaithersburg, MD) were used for
detection of DO11.10 TCR. The analytical flow cytometer used was
EPICS-XL (Coulter, Miami, FL), and the data were analyzed by Mac LAS
software (Management Science Associates, Pittsburgh, PA).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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We generated a transgenic mouse line in which the HA-tagged
AML1 gene is designed to be expressed under the regulation
of CD2 promoter. Immunoblot analysis using the HA-Ab revealed that the
transgene-derived AML1 protein is abundant in the thymus, but is scarce
in the spleen (Fig. 1
A, see
lanes 1 and 3).
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B). In lanes 1 and 4,
lanes 2 and 5, and lanes 3 and
6, equal amounts of protein extracts were loaded,
respectively. PEBP2/CBF DNA-binding activity detected in the transgenic
thymocytes was found to be roughly 2 times higher than that detected in
the wild-type thymocytes. Furthermore, inclusion of appropriate Abs to
the incubation mixture before electrophoresis supershifted the
protein/DNA complexes to the positions indicated by the asterisks (Fig. 1C). In the case of the extract prepared from the transgenic
thymocytes, the anti-HA Ab supershifted half of the total
PEBP2/CBF/DNA complex (lane 3), whereas the
anti-AML1 Ab supershifted the entire complex (lane
5). The results indicate that the PEBP2/CBF DNA-binding activity
in the transgenic thymocytes is composed of both HA-AML1 and endogenous
AML1. Thus, the exogenously introduced, HA-tagged AML1 protein is not
only overexpressed, but also contributes to the PEBP2/CBF DNA-binding
activity. Overexpression of AML1 increases the number of CD8 SP cells in the thymus
Flow cytometric analysis was performed to compare expression of
CD4 and CD8 in the AML1-transgenic and wild-type thymocytes
(Fig. 2A, note that hereafter
only representative profiles of flow cytometry are presented for each
genotype of mice, but that in all cases essentially similar results
were obtained for several individual mice with the identical genotype).
We found that the frequency of CD8 SP cells in the
AML1-transgenic thymus (8.2%) was 3 times higher than in
the wild-type thymus (2.8%), whereas the frequencies of the CD4 SP,
DP, and double-negative subsets did not differ significantly between
the two mouse strains. As the total number of thymocytes in the two
strains did not vary significantly from each other, this means that the
CD8 SP cell number in the AML1-transgenic thymus was
increased 3- to 4-fold compared with the wild-type thymus (Table I). Thus, high levels of AML1 protein
appear to drive the production of more CD8 SP cells in the thymus.
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B). The frequencies of TCRhigh
and TCRlow CD8 SP cells were both increased in
the AML1-transgenic thymus (8.2 x 54% and 8.2 x
46%, respectively) compared with the wild-type thymus (2.8 x
69% and 2.8 x 31%, respectively). Thus, overexpression of AML1
increases the numbers of both positively selected CD8 SP cells
(TCRhigh) and ISP cells
(TCRlow). The transgenic, thymic CD4 SP cells do
not differ markedly from wild-type cells in their expression of TCR
(data not shown). Overexpression of AML1 promotes CD8 SP cell production even when MHC class I molecules are absent
Recognition of MHC class I and class II molecules by the TCR and coreceptors leads to the development of CD8 SP and CD4 SP cells, respectively. Given that the AML1-transgenic thymocytes favor the CD8 SP lineage, we speculated that perhaps AML1 overexpression could even override the requirement of class I molecules for the CD8 SP cells to develop.
To test this possibility, we introduced the AML1 transgene
into 2m-deficient mice (Fig. 3). In the thymus of nontransgenic,
2m-/- mice,
CD4 SP cells (7.6%) could be detected, but only a few CD8 SP cells
(0.7%) were present. As the few CD8 SP cells found in these mice are
TCRlow, they are most likely to be ISP cells.
Thus, positively selected CD8 SP cells are not generated in the
2m-/- thymus,
as has been reported previously (29). In the
AML1-transgenic,
2m-/- thymus,
however, a significant population of CD8 SP cells (4.2%) was detected,
and at least some of these cells expressed TCR at higher levels
(TCRhigh) than the DP cells. Thus, overexpression
of AML1 promotes the generation of positively selected CD8 SP cells
even when MHC class I molecules are absent.
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The above observations led us to inquire whether the CD8 SP cells
found in the AML1-transgenic,
2m-/- thymus could also be generated
when there was a complete lack of MHC molecules. The AML1
transgene was thus introduced into mice that lack both
2m and class II I-A molecules (Fig. 4). TCRhigh CD8 SP
cells were extremely scarce in the thymi of both the
2m-/-:I-A-/-
and the AML1-transgenic,
2m-/-:I-A-/-
mice. Thus, the complete absence of MHC molecules abrogates the
promotional effect of AML1 overexpression on the generation of selected
CD8 SP cells.
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CD8 SP cells generated in the AML1-transgenic thymus do not complete maturation
After being positively selected, SP cells pass through a step of maturation in the thymic medulla, which is accompanied by the down-regulation of heat stable Ag (HSA) (30). We next examined whether CD8 SP cells generated in the AML1-transgenic thymus are capable of completing the maturation as the non-transgenic cells are.
When the CD8 SP subset of cells in the wild-type thymi is tested
for HSA expression, two peaks are observed, with the positively
selected CD8 SP cells being HSAlow, and the ISP
cells being HSAhigh (Fig. 5A). Wild-type DP cells are
HSAmed, with a fluorescence intensity below that
of ISP and above that of selected CD8 SP cells. However, when the CD8
SP subset prepared from the AML1-transgenic thymus was
tested for HSA expression, a major peak was observed, the intensity of
which matched the HSAmed DP cells in the
wild-type thymi. Thus, the maturation of transgenic CD8 SP cells
appears to be impeded at the HSAmed stage. In
this sense, we note that a putative TCRmed
subfraction with the intermediate fluorescence intensity could be
recognized in the AML1-transgenic, but not the wild-type,
CD8 SP thymocytes (see Fig. 2B). This
TCRmed is also considered to reflect the CD8 SP
cells, which are positively selected but arrested in the
maturation.
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2m-/- and
2m-/-:I-A-/-
mice, the HSA expression on the CD8 SP subset that represents the ISP
cells is higher than that of DP cells (Fig. 5, B and
C). The HSA expression remained high in the CD8 SP subset
derived from the AML1-transgenic,
2m-/-:I-A-/-
thymus. Notably, overexpression of AML1 in the
2m-/- mice
could down-regulate HSA expression on the CD8 SP subset to the medium
level, but not further to the lower level. These results confirmed the
above-mentioned idea that AML1 overexpression, though promoting the
generation of selected CD8 SP cells, cannot advance the maturation of
such cells beyond the HSAmed stage.
An observation relevant to the maturation arrest shown in Fig. 5, A and B, is that the frequencies of CD8 SP cells
in the spleen did not differ significantly between the wild-type and
AML1-transgenic mice (Table II) and between the
2m-/- and the
AML1-transgenic,
2m-/- mice
(data not shown). This is in contrast to the marked increase in CD8 SP
subset in the AML1-transgenic thymus. The
AML1-overexpressing, CD8 SP cells, although positively selected, do not
appear to emigrate from the thymus efficiently due to their incomplete
maturation.
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DP thymocytes recognizing MHC class II molecules normally
differentiate into the CD4 SP lineage. That thymic CD8 SP cells are
increased in AML1-transgenic,
2m-/-, but
not AML1-transgenic,
2m-/-:I-A-/-
mice may be because overexpression of AML1 protein causes the
thymocytes that bear class II-restricted TCRs to differentiate into the
CD8 SP lineage. To test this possibility, we generated
double-transgenic mice that overexpress AML1 and bear the DO11.10 TCR.
The DO11.10 TCR recognizes a peptide derived from OVA in the context of
I-A and thus most DP thymocytes in mice that are transgenic for this
TCR differentiate into CD4 SP cells (24). We assessed the
effect of overexpression of AML1 on this profile and performed this
experiment with mice on a SCID background to avoid TCR rearrangement
and expression of endogenous TCRs (Fig. 6).
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The class II-restricted, CD8 SP cells were infrequent in the spleens of double-transgenic mice (data not shown).
AML1 does not affect the choice of thymocytes to follow the CD4 SP lineage
The flow cytometric profiles presented above indicate that AML1
overexpression does not promote the generation of CD4 SP thymocytes. We
used MHC class II-deficient mice to confirm this observation. As shown
in Fig. 7, CD4 SP thymocytes were as
infrequent in mice that expressed AML1 but not I-A as in
I-A-/- mice. Thus, overexpression
of AML1 protein cannot rescue the generation of CD4 SP cells when class
II molecules are absent. In contrast, CD8 SP cells were frequent in the
thymi of both mouse strains.
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It is noteworthy that the CD8 SP thymocytes in
AML1-transgenic, I-A-/-
mice contained both the HSAmed and
HSAlow subfractions. That the
HSAmed subfraction is present in the
AML1-transgenic, I-A-/-
thymus, but not in the I-A-/- thymus
indicates that AML1 overexpression suppresses the maturation of some of
positively selected (TCRhigh) CD8 SP cells, even
if their TCR/coreceptors correctly match the MHC class that is
present.
AML1 protein is expressed both in CD8 SP and CD4 SP thymocytes
As described above in Fig. 1, B and C, the
HA-AML1 protein is considered to exert its effect by being recruited
into the endogenous PEBP2/CBF DNA-binding activity. Finally, we
examined whether the expression profile of endogenous AML1 protein
could be more or less correlated with T cell differentiation. The
extracts from the wild-type thymocytes were processed for immunoblot
analysis using the anti-AML1 antiserum (Fig. 8). The unfractionated thymocytes in
which DP cells occupy 80% of the population gave rise to the 56/58-kDa
band (lane 1). This band represents the AML1/Runx1 gene
product as described previously (21, 27). The extracts
prepared from the purified CD4 SP and CD8 SP thymocytes likewise
contained the 56/58-kDa component to a similar degree
(lanes 2 and 3).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Two mechanisms can be considered to explain how the overexpressed AML1 promotes the generation of class II-restricted, CD8 SP thymocytes. One model is that overexpression of AML1 might have redirected the fate of otherwise CD4 SP-oriented DP thymocytes into the CD8 SP lineage. According to the instruction model, the affinities of the TCR for class II and class I molecules guide the differentiation of DP cells into the CD4 SP and CD8 SP lineages, respectively (31, 32). It is hypothesized that DP cells receive signals from their CD4/TCR or CD8/TCR molecules and distinguish each signal based on its strength and duration (33, 34, 35). The engagement of CD4 and TCR molecules may confer a strong and/or long term signal that induces the generation of CD4 SP cells, whereas the weak and/or short term signal delivered from the CD8/TCR molecules may direct the generation of CD8 SP cells. In any case, these signals from the TCR would be translated into activation or silencing of transcription factors that would then regulate genes that cause DP thymocytes to adopt one of their two possible cell fates. Therefore, it is possible that AML1 might be one such transcription factor involved in SP lineage determination.
It must be noted, however, that the numbers of CD4 SP thymocytes were comparable between the AML1 DO11.10 TCR double-transgenic and DO11.10 TCR single-transgenic mice, an observation that is incompatible with the instruction model. An alternative model is, therefore, that the overexpressed AML1 prolongs the survival of thymocytes that are mis-selected into the CD8 lineage. A candidate for such thymocytes is, for example, transitional CD4low CD8high cells that were observed in the MHC class I-/- thymus, albeit in low numbers (36). In addition, overexpression of the anti-apoptotic bcl-2 gene is reported to promote the generation of CD8 SP thymocytes in MHC class I-deficient mice (8). Thus, it is also possible that the overexpressed AML1 in the class I-/- mice might have provided the above-mentioned, transitional population with a survival signal, thereby rescuing these cells from apoptosis and promoting their differentiation into the CD8 SP lineage.
It should be noted that for AML1 to promote thymocytes into the CD8 SP lineage, an interaction between TCR and either class of MHC is obligatory. This is evident from the observation that the selected CD8 SP cells were not generated, even when AML1 was overexpressed, if both MHC classes were completely lacking. In positive selection, DP cells whose TCRs possess sufficient affinity for MHC develop resistance to apoptosis. Our observation therefore implies that the increased activity of AML1 does not per se rescue cells from death by neglect, a fate marked for most DP cells with lesser affinities for MHC.
Despite the marked numerical increase in CD8 SP thymocytes in
AML1-transgenic,
2m-/- mice
and AML1 DO11.10 TCR-double transgenic mice, class
II-restricted CD8 SP cells were infrequent in their spleens. This
appeared to correlate with the absence of mature
(HSAlow) CD8 SP thymocytes in these mice, since
only mature SP thymocytes are allowed to be released into the
periphery. It is known that a signal that is elicited continuously due
to the correct combination of TCR/CD8 and MHC is necessary for the
continued survival of previously positively selected CD8 SP cells. This
has been shown by transfer experiments of DO11.10 TCR-transgenic CD8 SP
cells or class I-restricted HY TCR-transgenic CD8 SP cells into mice
bearing mismatched class I molecules (37, 38). Thus,
although overexpression of AML1 can cause the (mis) selection of class
II-restricted CD8 SP thymocytes, it probably does not provide the cells
with the signal necessary for their continued survival and/or
maturation.
Interestingly, the mismatch between TCR/CD8 and MHC could be overcome by supplying a putative survival/maturation signal through other means. For example, in helper-deficient mice (39) and mice expressing a dominant interfering form of the Lck molecule (4), MHC class II-restricted CD8 SP cells develop in thymus and, in contrast to the AML1-transgenic cells, are released in abundant numbers into peripheral lymphoid tissues.
It must be noted that overexpression of AML1 causes an additional
abnormality in the maturation of selected CD8 SP thymocytes that is
distinct from the abnormality discussed above. We found that the
maturation of CD8 SP thymocytes was also impeded in the
AML1-transgenic, I-A-/-
mice whose combination of TCR/CD8 and MHC should be adequate for CD8 SP
cell maturation. In addition, we previously reported that the
maturation of CD8 SP thymocytes is enhanced in
AML1-diminished mice, as judged by their HSA expression and
sensitivity to TCR stimulation (21). Thus, it is also
likely that AML1, whether expressed endogenously or introduced
exogenously, acts to suppress the maturation of CD8 SP thymocytes even
when the correct combination of TCR/coreceptors and MHC molecules is
present.
The mammalian runt gene family includes other genes as well
than AML1. According to Vaillant et al. (40),
overexpression of Runx2/Cbfa1 in the thymus results in a phenotype that
is similar to that noted in the present study, namely there is a
pronounced skewing toward the generation of CD8 SP thymocytes. In mice
overexpressing Runx2/Cbfa1, most of the CD8 SP thymocytes generated are
immature ISP cells, although the frequency of
CD3high CD8 SP cells also appears to increase
somewhat. Thus, one possibility is that overexpression of either AML1
or Runx2/Cbfa1 more or less promotes double-negative thymocytes to the
ISP stage and DP thymocytes to the CD8 SP lineage. However, there is
one notable difference between the case of AML1 and that of
Runx2/Cbfa1. In immunoblot analysis, we could detect the
endogenous AML1 gene product of 56/58 kDa in the fractions
of thymocytes, whereas a band of 62 kDa, which is estimated to be the
size of Runx2/Cbfa1 (27), was missing in the lanes of Fig. 8. It is not clear, therefore, whether the phenotypic effect caused by
the overexpressed Runx2/Cbfa1 may have correspondingly physiological
relevance. It must be noted as well that the 56/58-kDa band
representing AML1 was detected in a similar amount in both CD8 SP and
CD4 SP thymocytes. In addition, a new band of 52 kDa was observed,
which is considered to be the product of Runx3 gene based on
the Mr (see AF155880 submitted by D. Levanon, V.
Negreanu, and Y. Groner, GenBank). This 52-kDa band was detected
specifically in the CD8 SP, but not CD4 SP, thymocytes. It is possible
therefore that the superimposition of Runx3 to the AML1, like the
overexpressed AML1, skews thymocytes to the CD8 SP lineage, but, unlike
the overexpressed AML1, completes the maturation of generated CD8 SP
cells. An exact role the endogenous Runx3 plays in the differentiation
of CD8 SP thymocytes remains to be elucidated in a future study.
In conclusion, the AML1/Runx1 transcription factor participates in two distinct points of thymocyte development, namely at the transition of DP cells to the SP lineage and at SP maturation.
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Acknowledgments |
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-deficient mice and the DO11.10 TCR transgenic mice,
respectively, and D. Kioussis and P. Marrack for supplying us
with the pCD2 minigene and the KJ1-26 Ab, respectively. We also thank
M. Kuji for secretarial assistance. |
Footnotes |
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2 Address correspondence and reprint requests to Dr. Masanobu Satake, Department of Molecular Immunology, Institute of Development, Aging, and Cancer, Tohoku University, Seiryo-machi 4-1, Aoba-ku, Sendai 980-8575, Japan. E-mail address: satake{at}idac.tohoku.ac.jp
3 Abbreviations used in this paper: DP, double-positive; 2m, 2-microglobulin; HA, hemagglutinin; HSA, heat-stable Ag; ISP, immature single-positive; SP, single-positive; PEP2/CBF, polyomavirus enhancer binding protein 2/core binding protein.
Received for publication June 18, 2001. Accepted for publication August 29, 2001.
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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M. Ehlers, K. Laule-Kilian, M. Petter, C. J. Aldrian, B. Grueter, A. Wurch, N. Yoshida, T. Watanabe, M. Satake, and V. Steimle Morpholino Antisense Oligonucleotide-Mediated Gene Knockdown During Thymocyte Development Reveals Role for Runx3 Transcription Factor in CD4 Silencing During Development of CD4-/CD8+ Thymocytes J. Immunol., October 1, 2003; 171(7): 3594 - 3604. [Abstract] [Full Text] [PDF]
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O. Komine, K. Hayashi, W. Natsume, T. Watanabe, Y. Seki, N. Seki, R. Yagi, W. Sukzuki, H. Tamauchi, K. Hozumi, et al. The Runx1 Transcription Factor Inhibits the Differentiation of Naive CD4+ T Cells into the Th2 Lineage by Repressing GATA3 Expression J. Exp. Med., July 7, 2003; 198(1): 51 - 61. [Abstract] [Full Text] [PDF]
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 T. Nosaka, S. Morita, H. Kitamura, H. Nakajima, F. Shibata, Y. Morikawa, Y. Kataoka, Y. Ebihara, T. Kawashima, T. Itoh, et al. Mammalian Twisted Gastrulation Is Essential for Skeleto-Lymphogenesis Mol. Cell. Biol., April 15, 2003; 23(8): 2969 - 2980. [Abstract] [Full Text] [PDF]
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M. Schwieger, J. Lohler, J. Friel, M. Scheller, I. Horak, and C. Stocking AML1-ETO Inhibits Maturation of Multiple Lymphohematopoietic Lineages and Induces Myeloblast Transformation in Synergy with ICSBP Deficiency J. Exp. Med., November 4, 2002; 196(9): 1227 - 1240. [Abstract] [Full Text] [PDF]
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F. Vaillant, K. Blyth, L. Andrew, J. C. Neil, and E. R. Cameron Enforced Expression of Runx2 Perturbs T Cell Development at a Stage Coincident with {beta}-Selection J. Immunol., September 15, 2002; 169(6): 2866 - 2874. [Abstract] [Full Text] [PDF]
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