Congenic Mapping of the Type 1 Diabetes Locus,Idd3, to a 780-kb Region of Mouse Chromosome 3: Identification of a Candidate Segment of Ancestral DNA by Haplotype Mapping

Type 1 diabetes in the nonobese diabetic (NOD) mouse arises as a consequence of T cell-mediated destruction of the insulin-producing β cells of the pancreas. Although little is known of the events that initiate and subsequently drive β-cell destruction it is clear that the entire process is under complex genetic control. At present 19 loci have been mapped that influence the development of diabetes either at the level of initiation of insulitis or at the level of progression from insulitis to overt diabetes, or both. Previously, we have mapped one of these loci,Idd3, to a 0.35-cM interval on proximal mouse chromosome 3. In the present study we have narrowed the map position of this locus to an interval of 0.15 cM by a combination of novel congenic strains and an ancestral haplotype analysis approach. We have constructed a physical contig in bacterial artificial chromosome (BAC) clones across the minimal interval. Restriction mapping of the BAC contig placed the maximum size of theIdd3interval at 780 kb between the markersD3Nds36andD3Nds76. To refine further theIdd3interval we developed a series of novel single nucleotide polymorphisms (SNPs) and carried out haplotype analysis on DNA from mouse strains known to carry eitherIdd3susceptibility or protective alleles. This haplotype analysis identified a 145-kb segment of ancestral DNA between the microsatellite markerD3Nds6and the SNP81.3. One haplotype of this ancestral segment of DNA is found in mouse strains carrying anIdd3susceptibility allele and another is found in mouse strains carrying anIdd3protective allelle. Within the 780-kb congenically defined interval this 145-kb segment represents the most likely location forIdd3. TheIl2gene, which encodes the cytokine interleukin 2 (IL2), maps to this interval and is a strong candidate forIdd3. To investigate whether sequence variation exists in the promoter region of theIl2gene, which might alter its expression, we sequenced the promoter region of theIl2gene from mouse strains carrying either anIdd3susceptibility or resistance allele. Two sequence variants were identified, neither of which fell in known regulatory elements within theIl2promoter. In agreement with this observation steady-stateIl2mRNA levels showed no variation between susceptible and resistant mouse strains. These data suggest that the profound protection from diabetes seen in congenic mice carrying anIdd3protective allele is unlikely to be due to differences in the level of expression of theIl2gene. Instead, all of the current data support our hypothesis thatIdd3corresponds to amino acid variation at the amino terminus ofIl2.[Sequence data reported in this paper have been deposited in GenBank and assigned the following accession numbers: AF19594, AF19595, and AF19596.]