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The Jackson Laboratory, Bar Harbor, ME 04609
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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88% of their genome with NOD mice including
the H2g7 haplotype, NOR mice remain free of T
cell-mediated autoimmune diabetes (IDDM), due to non-MHC genes of
C57BLKS/J (BKS) origin. NOR IDDM resistance was previously found to be
largely controlled by the Idd13 locus within an 24 cM
segment on Chromosome 2 encompassing BKS-derived alleles for
H3a, B2m, Il1, and
Pcna. NOD stocks carrying subcongenic intervals of NOR
Chromosome 2 were utilized to more finely map and determine possible
functions of Idd13. NOR- derived H3a-Il1
(6.0 cM) and Il1-Pcna (1.2 cM) intervals both
contribute components of IDDM resistance. Hence, the Idd13
locus is more complex than originally thought, since it consists of at
least two genes. B2m variants within the
H3a-Il1 interval may represent one of these. Monoclonal Ab
binding demonstrated that dimerizing with the
ß2ma (NOD type) vs
ß2mb isoform (NOR type) alters the structural
conformation, but not total expression levels of H2g7 class
I molecules (e.g. Kd, Db).
ß2m-induced alterations in H2g7 class I
conformation may partially explain findings from bone marrow chimera
analyses that Idd13 modulates IDDM development at the level
of non-hematopoietically derived cell types controlling selection of
diabetogenic T cells and/or pancreatic ß cells targeted by these
effectors. Since trans-interactions between relatively
common and functionally normal allelic variants may contribute to IDDM
in NOD mice, the search for Idd genes in humans should not
be limited to functionally defective variants. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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That common allelic variants can acquire a diabetogenic function is best illustrated by the fact that while the Kd and/or Db class I gene products encoded by the H2g7 MHC haplotype of NOD mice are also expressed by many strains without obvious autoimmune proclivity, they play an essential role in the development of IDDM. This was demonstrated by the finding that NOD mice made deficient in MHC class I expression by congenic transfer of a functionally inactivated ß2-microglobulin (B2m) allele (designated NOD.B2mnull mice) remain completely free of IDDM (9, 10, 11, 12). We subsequently found this is due to the fact that the earliest initiative phases of autoimmune ß cell destruction in NOD mice are mediated by CD8+ T cells that recognize Ags presented by the common Kd and/or Db class I gene products of the H2g7 MHC haplotype (13). IDDM is also inhibited in a stock of NOD mice congenic for the MHC haplotype of the CTS strain (H2ct) that shares the class II, but not the class I alleles, of the H2g7 haplotype (14). Thus, it appears that rather than developing at a high frequency in the presence of any MHC class I molecules, autoimmune IDDM in NOD mice is most readily promoted by expression of the particular class I alleles that constitute the H2g7 haplotype. Similarly, recent evidence indicates that some relatively common MHC class I variants, such as HLA-A2, may also contribute to enhanced IDDM susceptibility in humans (15, 16). The mechanism by which relatively common MHC class I gene products can acquire diabetogenic activity in both humans and NOD mice remains unknown. However, it seems likely that they do so through interactions with non-MHC associated Idd susceptibility alleles, many of which may also represent physiologically normal variants found in humans and mouse strains characterized by IDDM resistance.
Insights to diabetogenic interactions between MHC and non-MHC
genes have been provided by analysis of IDDM-resistant
NOR mice. The NOR/Lt strain is a
recombinant congenic stock that derives 88% of its genome from NOD
including the H2g7 MHC haplotype, but contains
genetic material of C57BLKS/J (BKS) origin on regions of chromosomes
(Chr.) 1, 2, 4, 5, 7, 11, 12, and 18 (17, 18). The BKS strain itself
represents a recombinant congenic strain, carrying defined genomic
contributions primarily from a C57BL/6 (B6) donor, but also from a
"DBA/2-like" donor (19). Constitutive levels of MHC class I
expression in NOD and NOR mice are equivalent to that of the BKS
control strain (20). However, the ability of macrophages from NOD and
NOR mice to futher up-regulate expression of H2g7 MHC class
I molecules in response to stimulation with IFN-
is differentially
controlled by genes within chromosomal regions distinguishing these two
strains. IFN- fails to up-regulate H2g7 MHC class I
expression in macrophages from NOD mice, but does so normally in NOR
macrophages (20). The failure of NOD macrophages to up-regulate MHC
class I expression in response to IFN- could diminish the capacity
of these APC to activate tolerogenic mechanisms that normally delete or
inactivate diabetogenic CD8+ T cells. Autoreactive
CD8+ T cells that are generated as a result of such
tolerogenic defects in NOD mice could then be efficiently targeted to
the pancreatic ß cells, since these cells regulate H2g7
MHC class I expression in a normal fashion.
(NODxNOR)F2 segregation analysis demonstrated that the
major genetic component contributing to IDDM resistance in NOR is the
Idd13 locus on Chromosome 2 in linkage with BKS-derived
genes for ß2m, both isoforms of IL-1
(Il1a and Il1b), and proliferating
cell nuclear Ag (Pcna)(18). Existence of the
Idd13 locus was confirmed by the induction of IDDM
resistance in a stock of NOD mice congenic for an 24 cM segment of
NOR Chromosome 2 that contains these linkage markers flanked by the
H3a minor histocompatability and adenosine deaminase
(Ada) genes (18, 21). However, it is unknown
which gene(s) within this segment of Chromosome 2 contributes to IDDM
susceptibility in NOD and resistance in NOR mice, and if this effect is
mediated through a modulation of H2g7 MHC class
I expression or function. The present study was conducted to gain
insight to these questions through an analysis of IDDM development and
H2g7 MHC class I expression and function in NOD stocks
carrying variable truncations of the originally defined congenic
segment of NOR Chromosome 2 found to confer Idd13-mediated
resistance.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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NOD/Lt and NOR/Lt mice (both H2g7 = Kd, Ag7, Enull, Db) have been maintained at The Jackson Laboratory by brother-sister mating. Currently, IDDM develops in 90% of female and 63% of male NOD/Lt mice by one year of age, whereas both sexes of NOR/Lt mice are IDDM resistant. A stock of NOD mice carrying a previously defined congenic segment of NOR Chromosome 2 conferring Idd13-mediated IDDM resistance was initially utilized at the N12 backcross generation (18, 21). To further refine the localization of genes contributing to IDDM resistance within the NOR-derived Idd13 locus, the Chromosome 2 congenic segment present in this N12 stock was variably truncated by further backcrossing to the NOD parental strain. Backcross mice were screened for further recombination events on Chromosome 2 using the genotyping methodologies described below. At the 20th backcross (N21) generation, the indicated NOR-derived Chromosome 2 congenic segments were fixed to homozygosity on the NOD background by brother-sister matings. C57BLKS/J (H2d) mice were supplied by the Animal Resources Unit of The Jackson Laboratory. All mice were maintained under specific pathogen-free (SPF) conditions and allowed free access to autoclaved chow diet (diet 96WA, Emory Morse, Guilford, CT.) and acidified drinking water.
Assessment of diabetes and insulitis development
Mice from each of the indicated strains were simultaneously
monitored at weekly intervals for the development of glycosuria with
Ames Diastix (kindly supplied by Miles Diagnostics, Elkhart, IN).
Glycosuric values of 
3 were considered diagnostic of diabetes onset.
Development of stable glycosuria was confirmed by weekly urinalysis for
2 weeks after the initial diagnosis. A subset of non-diabetic mice were
necropsied for pancreatic histology at 1 year of age. Pancreata were
stained with aldehyde fuchsin to detected granulated ß cells and then
counterstained with hematoxylin and eosin.
Genotyping methodologies
DNA samples used for genotypic analyzes were extracted from tail clips. Most polymorphic markers were typed either by PCR or Southern blotting as previously described (18, 22). Allelic variations at the H3a minor histocompatibility gene were typed as previously described (21) by assessing the sensitivity of splenic leukocytes from the various congenic stocks to lysis by cloned lines of gene product-specific cytotoxic T-lymphocytes (CTL).
Effect of B2m polymorphisms on the conformation of MHC class I molecules expressed by splenic leukocytes
Splenic leukocytes from the indicated strains were prepared as previously described (23), and resuspended at 2 x 107/ml in FACS buffer (PBS containing 0.1% sodium azide with 2% FBS). Aliquots of 1 x 106 cells (50 µl) were incubated for 30 min at 4°C with FITC-conjugated mAbs specific for the allelically variable regions of the H2Kd (SF1–1.1) or H2Db (28–14–4) MHC class I molecules, or an epitope within the MHC class I constant region that undergoes dimerization with ß2m (M1/42) and is shared by all alleles. The cells were washed in FACS buffer after staining. Data for the extent of MHC class I Ab binding are presented as mean channel of log fluorescence (MFI) ± SEM, as determined by FACScan (Becton Dickinson, San Jose, CA.) using the Cell Quest 3.0 data reduction program.
Regulation of MHC class I expression by IFN- in peritoneal
macrophage cultures
Thioglycollate-elicted peritoneal macrophages were isolated from
the indicated male mice using our previously described protocols (20).
Macrophages were suspended at 2.0 x 106/ml in
the previously described culture medium (23) in the presence and
absence of 50 U/ml rat recombinant IFN- (kindly supplied by P. van
de Meide, Rijswijk, Netherlands) and then incubated for 6 days at
37°C. At this time macrophages were harvested by washing with
calcium- and magnesium-free HBSS and subsequent treatment with
enzyme-free cell dissociation buffer (Life Technologies, Gaithersburg,
MD). To assess IFN--regulated levels of cell surface MHC class I
expression, the macrophages were stained with the FITC-conjugated mAb
31–3–4S that recognizes the Kd MHC class I molecule
shared by all of the strains used for these experiments. Levels of
total Kd MHC class I expression in IFN--treated and
untreated macrophages are presented as MFI as determined by FACScan
analysis.
Production of bone marrow chimeras
Females from the indicated strains were lethally irradiated (1200R from a 137Cs source) at 4 wk of age, and then reconstituted as previously described (24) with 5 x 106 bone marrow cells isolated from the indicated 8-wk-old female donors. Bone marrow chimeras were then monitored through a 21-wk postreconstitution for the development of IDDM as described above. Upon the onset of IDDM or at 21 wk of postreconstitution, chimerization was assessed by genotyping splenic DNA for donor or recipient type B2m polymorphisms by Southern blot analysis of a BglI restriction fragment length variant (B2ma = 801 bp fragment; B2mb = 575 and 226 bp fragments). A B2m-specific probe was generated by PCR amplification of C57BL/10J genomic DNA with the primer set 5'-CACGCCACCCACCGGAGAATG-3' and 5'-GATGCTGATCACATGTCTCG-3'.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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As shown in Table I
, three congenic
intervals derived from NOR Chromosome 2 were fixed to homozygosity on
the NOD inbred background at the 20th backcross generation. The first
of these congenic stocks carries the largest segment of NOR Chromosome
2 spanning a 31.5 cM interval encompassing the linkage markers
D2Mit490 through D2Mit144 (designated
NOD.D2Mit490-Mit144NOR). This large congenic
segment contains all markers previously found to be linked to
Idd13-mediated IDDM resistance in NOR mice (18). This
segment of NOR-derived Chromosome 2 is originally derived from the B6
strain contribution to the BKS genome (18) such that the
H3aa and B2mb alleles, as
well as the microsatellite-based markers are the same as found in the
B6 genome. The present analysis extends the proximal boundary of this
NOR-derived Chromosome 2 congenic segment by 4 cM to the
D2Mit490 rather than the H3a marker, and the
distal boundary by 3 cM to D2Mit144 rather than
Ada. Smaller intervals derived from this large segment of
NOR Chromosome 2 through selection of recombinants have been fixed to
homozygosity in two other NOD congenic stocks. One of these carries an
6.0 cM congenic interval of NOR Chromosome 2 spanning the linkage
markers H3a through Il1a (designated
NOD.H3a-Il1NOR). The other carries an 1.2 cM
segment of NOR Chromosome 2 spanning the interval delineated by
Il1a and Pcna (designated
NOD.Il1-PcnaNOR).
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, by 30 wk of age the
female incidence of IDDM in the
NOD.D2Mit490-Mit144NOR congenic stock (16%) was
significantly less than in standard NOD mice (100%). By 52 wk, IDDM
incidence in these congenic females had increased to only 26% (5 of
19), and no diabetes was observed in a group of 18
NOD.D2Mit490-Mit144NOR congenic males (compared
with 50% incidence in a group of 18 standard NOD/Lt males). Histologic
analysis of insulitis in pancreata from of three of these non-diabetic
congenic mice of each sex necropsied at a year of age showed widespread
perivascular/periductular leukocytic infiltrates. When associated with
islets, these were primarily peri-insular, with granulated ß cell
mass largely preserved in >80% of the islets. This histopathology was
similar to that observed in year-old NOR/Lt mice of both sexes. This
confirmed the results originally observed at an earlier backcross
generation (18) that Idd13-mediated IDDM resistance in NOR
mice did not prevent the pancreatic T-lymphoaccumulation characteristic
of NOD mice, but rather retarded activation of autoimmune effector
functions. In this regard, the pancreatic histopathology in
non-diabetic NOD.D2Mit490-Mit144NOR congenic
mice was very reminiscent of that observed in a NOD stock made IDDM
resistant through the elimination of B-lymphocyte development by
congenic transfer of a null mutation in the Igµ heavy chain
locus (25).
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31.5 cM region of NOR Chromosome 2
spanning D2Mit490 through D2Mit144 was partially
attenuated upon differential truncation of this interval in two
subcongenic stocks of NOD mice (Fig. 1
). By 30 wk of age, the female
incidence of IDDM in both the NOD.H3a-Il1NOR
(56%) and NOD.Il1-PcnaNOR (62%) congenic
stocks was significantly lower than in standard NOD mice, but
significantly higher than in the
NOD.D2Mit490-Mit144NOR congenic stock.
Similarly, the male incidence of IDDM in these two subcongenic stocks
was significantly less than that observed in standard NOD mice. By a
year of age, the cumulative incidence of IDDM in the
NOD.H3a-Il1NOR stock reached 62% (10 of 16) in
females and 25% (4 of 16) in males. In the
NOD.Il1-PcnaNOR stock, IDDM developed in 75% (9
of 12) of females and 12% (2 of 17) of males by 46 wk of age. Thus,
the H3a-Il1 and/or Il1-Pcna intervals on
Chromosome 2 carry at least one, but not the complete set of
polymorphic genes contributing to Idd13 locus-mediated IDDM
susceptibility in NOD and resistance in NOR mice.
Contributions of the Idd13 locus to differential
regulation of H2g7 MHC class I expression in
IFN--stimulated macrophages from NOD and NOR mice
While no defects in constitutive expression were observed,
our previous studies demonstrated that IFN- stimulation fails to
further up-regulate H2g7 MHC class I levels in NOD
macrophages (20). Such a defect could conceivably reduce the ability of
these APC in NOD mice to activate tolerogenic mechanisms that normally
delete or inactivate diabetogenic CD8+ T cells. We
hypothesized that this possible APC tolerogenic defect might be
controlled by a gene(s) within the Idd13 locus since
H2g7 MHC class I expression is up-regulated normally in
IFN--stimulated macrophages from IDDM-resistant NOR mice. To test
this hypothesis, we compared the pattern of IFN--regulated MHC class
I expression in macrophages from the
NOD.D2Mit490-Mit144NOR congenic stock at the N12
backcross generation to similarly treated macrophages from NOD, NOR,
and BKS control mice. The BKS strain that has provided the genetic
source of IDDM resistance to the NOR strain is characterized by the
standard H2d haplotype, and thus expresses a
Kd MHC class I gene product also encoded within
H2g7. As expected, FACS analysis utilizing the
Kd specific mAb 31–3–4S confirmed that expression of this
MHC class I molecule was up-regulated normally in IFN--treated
macrophages from both NOR and BKS control mice (Fig. 2). However, as previously observed,
IFN- stimulation failed to up-regulate Kd expression in
NOD macrophages. Stimulation with IFN- also failed to up-regulate
Kd expression in macrophages from the
NOD.D2Mit490-Mit144NOR congenic stock. While not
excluding the possibility that aberrant regulation of MHC class I
expression in IFN--stimulated macrophages contributes to autoimmune
IDDM susceptibility in NOD mice, these results conclusively demonstrate
that this defect is not controlled by the Idd13 locus on
Chromosome 2.
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We have previously found that the relatively common MHC class I gene products of the H2g7 haplotype (e.g., Kd and/or Db) mediate T cell responses essential to the initiation of autoimmune IDDM in NOD mice (11, 13). It is likely that these same H2g7 MHC class I molecules do not mediate diabetogenic T cell responses in NOR mice, due to functions exerted by resistance variants of non-MHC Idd genes. It has been reported that interacting with different isoforms of ß2m may lead to alterations in the structural conformation of MHC class I molecules (26, 27, 28), which can skew the array of Ags they bind and present to CD8+ T cells (29). The B2m gene maps within the H3a-Il1 interval on Chromosome2, which we have shown to contain at least one component of Idd13. Thus, the ß2m isoforms distinguishing NOD (ß2ma) from the B6-derived allele present in NOR mice (ß2mb) may respectively provide one component of Idd13-mediated IDDM susceptibility or resistance through an ability to differentially alter the structural conformation, and hence the function of H2g7 MHC class I molecules shared by these two strains.
To test this possibility, splenic leukocytes from NOD mice plus the
NOD.H3a-Il1NOR, and
NOD.Il1-PcnaNOR congenic stocks were stained
with FITC-conjugated mAbs specific for allelically variable regions of
the Kd (SF1–1.1) and Db (28–14–8)
H2g7 MHC class I gene products, or an epitope within the
MHC class I constant region shared by all allelic variants that
undergoes dimerization with ß2m (M1/42). As shown in
Table II, the Kd and
Db allele-specific Abs bound equivalently to splenocytes
from NOD mice and both NOD stocks congenic for intervals of NOR
Chromosome 2 carrying partial sets of Idd13 resistance
alleles. However, the level of M1/42 binding to the constant region of
these MHC class I molecules was significantly different on
ß2mb positive splenocytes from the
NOD.H3a-Il1NOR congenic stock than on
ß2ma positive splenocytes from NOD mice or
the NOD.Il1-PcnaNOR congenic stock. Thus, while
the total expression levels of H2g7 MHC class I molecules
are equivalent on splenocytes from NOD mice and both NOD stocks
congenic for intervals of NOR Chromosome 2 carrying partial sets of
Idd13 resistance alleles (based on Kd and
Db allele-specific staining), the structural conformation
of these molecules (assessed by M1/42 staining) appears to differ
depending on the isoform of ß2m with which they have
dimerized.
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The studies described above demonstrated that dimerization with
different isoforms of ß2m may alter the structural
conformation of H2g7 MHC class I molecules. Such
B2m-controlled alterations in the structural conformation of
H2g7 MHC class I molecules expressed on cell types
regulating the original selection of diabetogenic T cells (thymic
epithelium and hematopoietically derived APC) and/or on the pancreatic
ß cells targeted by these autoreactive effectors may represent one
mechanism by which the Idd13 locus contributes to IDDM
susceptibility or resistance. This possibility was tested by assessing
IDDM development in a series of bone marrow chimeras in which
Idd13 susceptibility or resistance variants could be
differentially expressed in cell types controlling T cell selection and
in pancreatic ß cells. As expected, IDDM developed in 75% of NOD
females over a 21-wk period following reconstitution with syngeneic NOD
marrow (Table III). However,
reconstitution with NOD bone marrow elicited a significantly lower
incidence of IDDM (15.4%) in NOR females. Conversely, reconstitution
with NOR marrow elicited a significantly higher incidence of IDDM in
NOD (55.5%) than in syngeneic NOR female recipients (0%). Thus, some
portion of the Idd susceptibility or resistance variants
distinguishing NOD from NOR mice control functions in
non-hematopoietically derived cell types that regulate the development
and/or functional activation of the diabetogenic T cells that normally
differentiate from NOD bone marrow. Providing support that
Idd13 variants are at least partial contributors to this
process was the finding that reconstitution with NOD bone marrow
resulted in a significantly lower incidence of IDDM in
NOD.D2Mit490-Mit144NOR (37.5%) than in
syngeneic NOD female recipients (75%). Furthermore, IDDM developed in
77.8% of standard NOD females reconstituted with
NOD.D2Mit490-Mit144NOR marrow. This was
significantly greater than the IDDM incidence in
NOD.D2Mit490-Mit144NOR recipients of syngeneic
marrow (26.7%), but did not differ from that of NOD recipients
reconstituted with syngeneic marrow.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-stimulated
H2g7 MHC class I expression in hematopoietically
derived macrophages from NOD and NOR mice (20). Indeed, the present
studies indicate that at least some portion of the multiple genes
contributing to Idd13-mediated IDDM susceptibility or
resistance exert their effects in non-hematopoietically derived cell
types. These could include thymic epithelial cells contributing to the
original selection of the T cell repertoire, and/or the pancreatic ß
cells targeted by autoreactive T cells in IDDM. Along these lines it
should be noted that Idd13 represents the first locus we
have found that does not exert a pathogenic or protective effect at the
level of hematopoietically derived cell types (30). The H3a-Il1 and Il1-Pcna intervals on Chromosome 2 carry at least one, but not the complete set, of polymorphic genes contributing to Idd13-mediated IDDM susceptibility in NOD and resistance in NOR mice. Since the H3a-Il1 and Il1-Pcna intervals overlap, it is possible that they share a gene contributing to IDDM susceptibility or resistance. One seemingly good candidate would be the structural genes for both isoforms of IL-1 that are contained within both of these intervals. Their candidacy is based on the findings that NOD macrophages are poor producers of IL-1 (23, 31), and that IDDM is inhibited in NOD mice treated with IL-1 in vivo (31). However, arguing against the candidacy of NOD-derived Il1 alleles as contributors to IDDM is that despite the presence of BKS-derived Il1 variants, macrophages from NOR mice are also defective in IL-1 production (17). The unlikely contribution of Il1 variants to IDDM susceptibility or resistance supports the possibility that H3a-Il1 and Il1-Pcna intervals carry separate components of Idd13. However, regardless of whether the H3a-Il1 and Il1-Pcna intervals are characterized by shared or different Idd genes, it remains possible that additional polymorphic genes outside of these regions, but within the larger segment flanked by D2Mit490 and D2Mit144, also contribute to Idd13-mediated IDDM susceptibility or resistance. Subcongenic analysis of the Idd10 region on Chromosome 3 (32) and the Idd2 region on Chromosome 9 (this laboratory, unpublished) are similarly providing evidence that Idd loci initially inferred to be composed of single genes in fact represent contributions from multiple genes.
Within the H3a-Il1 interval, allelic variants of B2m represent an excellent candidate for representing one component of Idd13-mediated IDDM susceptibility in NOD and resistance in NOR mice. The ß2ma (NOD type) and ß2mb (NOR type) isoforms exert trans-acting effects that alter the structural conformation, but not the total expression levels, of H2g7 MHC class I molecules shared by NOD and NOR mice. Such alterations in the structural conformation of H2g7 MHC class I molecules elicited by dimerization with different isoforms of ß2m may skew their ability to bind and present certain Ags, and hence contribute to IDDM susceptibility or resistance by effecting the selection and/or targeting of ß cell autoreactive CD8+ T cells. Support for this possibility is provided by previous reports that dimerization with different ß2m isoforms can alter the structural conformation of MHC class I molecules (26, 27, 28), which may in turn skew the array of antigenic peptides they bind and present to CD8+ T cells (29). Our bone marrow chimera studies indicate that any such effects are manifest at the level of non-hematopoietically derived thymic epithelial cells contributing to the original selection of the T cell repertoire, or at the level of pancreatic ß cells targeted by autoreactive T cells in IDDM. Supporting this hypothesis is that in retrospect, all nominally resistant mouse stocks that developed IDDM following reconstitution with NOD marrow have been at least heterozygous for the B2ma allele (24, 33, 34, 35, 36, 37). Further evidence that heterozygous expression of the B2ma allele may be sufficient to support IDDM development is provided by our previous finding that the NOR-derived Idd13 locus (containing B2mb) can only inhibit disease when in the homozygous state (18). It should also be noted that neither the B2ma or B2mb variant can be considered to represent a deleterious mutant allele since the gene products encoded by both function normally in terms of mediating the transport to and stable expression of MHC class I molecules on cell surfaces. Thus, if future studies ultimately demonstrate that B2m variants represent actual Idd susceptibility and resistance alleles, it would strongly support the previously proposed hypothesis (8) that autoimmune IDDM is not produced by rare mutations, but rather a set of common genetic variants that have coalesced in a dysfunctional array.
The present findings in no way support previous assertions by a single laboratory (38, 39, 40) that NOD splenic leukocytes are characterized by aberrantly low constitutive levels of MHC class I molecules as a consequence of defects in expression of the intra-MHC Tap1 gene. None of the findings from this aforementioned laboratory have been replicated by other investigators (10, 11, 20, 41, 42, 43, 44, 45). The erroneous conclusion that constitutive MHC class I expression was decreased can be explained by the fact that T lymphocyte numbers, which normally express lower levels of class I than other other leukocyte populations, are proportionally increased in NOD lymphoid organs relative to that observed in diabetes-resistant strains (42, 44). A polymorphism in an intron of the NOD Tap1 gene was shown by others not to affect loading of Ag to MHC class I molecules or their presentation to CTLs (21, 46). The conclusion that mice lacking a functional B2m gene developed spontaneous autoimmune diabetes (38) has also not been replicated (9, 10, 11, 12). As emphasized above, NOD and NOR mice are H2g7-identical, and thus share the same Tap genes. Neither strain is ß2m deficient; the allelic polymorphism distinguishing NOD from NOR entails a single amino acid difference at position 85 (47). Dimerization with these subtly different isoforms of ß2m may affect the structural conformation of the H2g7 MHC class molecules, but not their overall (normal) level of expression.
In conclusion, our studies have demonstrated that multiple polymorphic genes within the originally defined Idd13 locus on Chromosome 2 contribute to IDDM susceptibility in NOD mice and resistance in the H2g7 identical NOR strain. It is possible that normal allelic variants of B2m represent one component of Idd13 through an ability to differentially alter the structural conformation of the relatively common H2g7 MHC class I molecules and hence promote or inhibit their ability to select and/or target diabetogenic T cells. If correct, this would indicate that some of the processes that underlie the development of autoimmune IDDM in NOD mice are controlled by trans-interactions between relatively common and functionally normal allelic variants. That trans-interactions between relatively common and functionally normal allelic variants may contribute to autoimmune IDDM in NOD mice indicates that the search for Idd genes in humans should not be limited to functionally defective variants.
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Footnotes |
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2 Address correspondence and reprint requests to Dr. David V. Serreze, The Jackson Laboratory, Bar Harbor, Maine 04609.
Received for publication July 29, 1997. Accepted for publication October 9, 1997.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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 M. A. Wallet, R. R. Flores, Y. Wang, Z. Yi, C. J. Kroger, C. E. Mathews, H. S. Earp, G. Matsushima, B. Wang, and R. Tisch MerTK regulates thymic selection of autoreactive T cells PNAS, March 24, 2009; 106(12): 4810 - 4815. [Abstract] [Full Text] [PDF]
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 J. M. Fletcher, M. A. Jordan, S. L. Snelgrove, R. M. Slattery, F. D. Dufour, K. Kyparissoudis, G. S. Besra, D. I. Godfrey, and A. G. Baxter Congenic Analysis of the NKT Cell Control Gene Nkt2 Implicates the Peroxisomal Protein Pxmp4 J. Immunol., September 1, 2008; 181(5): 3400 - 3412. [Abstract] [Full Text] [PDF]
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L. A. Benoit and R. Tan Xenogeneic beta2-Microglobulin Substitution Affects Functional Binding of MHC Class I Molecules by CD8+ T Cells J. Immunol., September 15, 2007; 179(6): 3588 - 3595. [Abstract] [Full Text] [PDF]
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E. A. Ivakine, S. M. Mortin-Toth, O. M. Gulban, A. Valova, A. Canty, C. Scott, and J. S. Danska The Idd4 Locus Displays Sex-Specific Epistatic Effects on Type 1 Diabetes Susceptibility in Nonobese Diabetic Mice Diabetes, December 1, 2006; 55(12): 3611 - 3619. [Abstract] [Full Text] [PDF]
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P. A. Silveira, H. D. Chapman, J. Stolp, E. Johnson, S. L. Cox, K. Hunter, L. S. Wicker, and D. V. Serreze Genes within the Idd5 and Idd9/11 Diabetes Susceptibility Loci Affect the Pathogenic Activity of B Cells in Nonobese Diabetic Mice J. Immunol., November 15, 2006; 177(10): 7033 - 7041. [Abstract] [Full Text] [PDF]
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E. A. Ivakine, O. M. Gulban, S. M. Mortin-Toth, E. Wankiewicz, C. Scott, D. Spurrell, A. Canty, and J. S. Danska Molecular genetic analysis of the idd4 locus implicates the IFN response in type 1 diabetes susceptibility in nonobese diabetic mice. J. Immunol., March 1, 2006; 176(5): 2976 - 2990. [Abstract] [Full Text] [PDF]
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A. Ueno, S. Cho, L. Cheng, Z. Wang, B. Wang, and Y. Yang Diabetes Resistance/Susceptibility in T Cells of Nonobese Diabetic Mice Conferred by MHC and MHC-Linked Genes J. Immunol., October 15, 2005; 175(8): 5240 - 5247. [Abstract] [Full Text] [PDF]
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G. F. Beilhack, R. R. Landa, M. A. Masek, and J. A. Shizuru Prevention of Type 1 Diabetes with Major Histocompatibility Complex-Compatible and Nonmarrow Ablative Hematopoietic Stem Cell Transplants Diabetes, June 1, 2005; 54(6): 1770 - 1779. [Abstract] [Full Text] [PDF]
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E. A. Ivakine, C. J. Fox, A. D. Paterson, S. M. Mortin-Toth, A. Canty, D. S. Walton, K. Aleksa, S. Ito, and J. S. Danska Sex-Specific Effect of Insulin-Dependent Diabetes 4 on Regulation of Diabetes Pathogenesis in the Nonobese Diabetic Mouse J. Immunol., June 1, 2005; 174(11): 7129 - 7140. [Abstract] [Full Text] [PDF]
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E. J. Gordon, L. S. Wicker, L. B. Peterson, D. V. Serreze, T. G. Markees, L. D. Shultz, A. A. Rossini, D. L. Greiner, and J. P. Mordes Autoimmune Diabetes and Resistance to Xenograft Transplantation Tolerance in NOD Mice Diabetes, January 1, 2005; 54(1): 107 - 115. [Abstract] [Full Text] [PDF]
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C. L. Welch, S. Bretschger, P.-Z. Wen, M. Mehrabian, N. Latib, J. Fruchart-Najib, J. C. Fruchart, C. Myrick, and A. J. Lusis Novel QTLs for HDL levels identified in mice by controlling for Apoa2 allelic effects: confirmation of a chromosome 6 locus in a congenic strain Physiol Genomics, March 12, 2004; 17(1): 48 - 59. [Abstract] [Full Text] [PDF]
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S. E. Eckenrode, Q. Ruan, P. Yang, W. Zheng, R. A. McIndoe, and J.-X. She Gene Expression Profiles Define a Key Checkpoint for Type 1 Diabetes in NOD Mice Diabetes, February 1, 2004; 53(2): 366 - 375. [Abstract] [Full Text]
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L. M. Esteban, T. Tsoutsman, M. A. Jordan, D. Roach, L. D. Poulton, A. Brooks, O. V. Naidenko, S. Sidobre, D. I. Godfrey, and A. G. Baxter Genetic Control of NKT Cell Numbers Maps to Major Diabetes and Lupus Loci J. Immunol., September 15, 2003; 171(6): 2873 - 2878. [Abstract] [Full Text] [PDF]
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T. Pearson, T. G. Markees, D. V. Serreze, M. A. Pierce, M. P. Marron, L. S. Wicker, L. B. Peterson, L. D. Shultz, J. P. Mordes, A. A. Rossini, et al. Genetic Disassociation of Autoimmunity and Resistance to Costimulation Blockade-Induced Transplantation Tolerance in Nonobese Diabetic Mice J. Immunol., July 1, 2003; 171(1): 185 - 195. [Abstract] [Full Text] [PDF]
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T. Pearson, T. G. Markees, L. S. Wicker, D. V. Serreze, L. B. Peterson, J. P. Mordes, A. A. Rossini, and D. L. Greiner NOD Congenic Mice Genetically Protected From Autoimmune Diabetes Remain Resistant to Transplantation Tolerance Induction Diabetes, February 1, 2003; 52(2): 321 - 326. [Abstract] [Full Text] [PDF]
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T. C. Brodnicki, F. Quirk, and G. Morahan A Susceptibility Allele From a Non-Diabetes-Prone Mouse Strain Accelerates Diabetes in NOD Congenic Mice Diabetes, January 1, 2003; 52(1): 218 - 222. [Abstract] [Full Text] [PDF]
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S. Lesage, S. B. Hartley, S. Akkaraju, J. Wilson, M. Townsend, and C. C. Goodnow Failure to Censor Forbidden Clones of CD4 T Cells in Autoimmune Diabetes J. Exp. Med., November 4, 2002; 196(9): 1175 - 1188. [Abstract] [Full Text] [PDF]
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Y. Yang, M. Bao, and J.-W. Yoon Intrinsic Defects in the T-Cell Lineage Results in Natural Killer T-Cell Deficiency and the Development of Diabetes in the Nonobese Diabetic Mouse Diabetes, December 1, 2001; 50(12): 2691 - 2699. [Abstract] [Full Text] [PDF]
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P. A. Lyons, N. Armitage, C.J. Lord, M. S. Phillips, J. A. Todd, L. B. Peterson, and L. S. Wicker Mapping by Genetic Interaction: High-Resolution Congenic Mapping of the Type 1 Diabetes Loci Idd10 and Idd18 in the NOD Mouse Diabetes, November 1, 2001; 50(11): 2633 - 2637. [Abstract] [Full Text] [PDF]
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E. E. Hamilton-Williams, D. V. Serreze, B. Charlton, E. A. Johnson, M. P. Marron, A. Mullbacher, and R. M. Slattery Transgenic rescue implicates beta 2-microglobulin as a diabetes susceptibility gene in nonobese diabetic (NOD) mice PNAS, September 25, 2001; 98(20): 11533 - 11538. [Abstract] [Full Text] [PDF]
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 P. C. Reifsnyder, G. Churchill, and E. H. Leiter Maternal Environment and Genotype Interact to Establish Diabesity in Mice Genome Res., October 1, 2000; 10(10): 1568 - 1578. [Abstract] [Full Text]
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J. Verdaguer2, A. Amrani2, B. Anderson, D. Schmidt, and P. Santamaria Two Mechanisms for the Non-MHC-Linked Resistance to Spontaneous Autoimmunity J. Immunol., April 15, 1999; 162(8): 4614 - 4626. [Abstract] [Full Text] [PDF]
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,
n = 18), NOD.D2Mit490-Mit144NOR
(
, n = 19), NOD.H3a-Il1NOR
(•, n = 17), NOD.Il1-PcnaNOR
(
, n = 21). * IDDM incidence significantly less
(p < 0.0001, Kaplan Meier life table analysis)
than in standard NOD mice. 
IDDM incidence significantly
greater (p < 0.003, Kaplan Meier life table
analysis) than in the congenic
NOD.D2Mit490-Mit144NOR stock.