Session Type: Abstract Submissions (ACR)
Ankylosing spondylitis (AS) is the prototypic spondyloarthritis and affects approximately 0.2-0.5% of the human population. It is estimated that genetic risk factors contribute >90% of disease susceptibility, and the major genetic risk factor is HLA-B27. Approximately the other half of the genetic risk is composed by non-MHC traits. By generating interval-specific congenic (IVSC) strains utilizing the genetic combination of proteoglycan-induced SpA (PGISpA) in susceptible BALB/c mice and SpA-resistant DBA/2 mice (both carry the same MHC) we can identify non-MHC causative genes by excluding the effect of the MHC. These IVSC strains with overlapping chromosome (Chr) regions were used to narrow the (Pgis2) risk locus on mouse Chr2 (mChr2) to a small region syntenic with AS risk loci on human chromosomes 4 and 9.
PGISpA-resistant DBA/2 and –susceptible BALB/c strains were intercrossed, and then backcrossed with BALB/c mice until the entire Pgis2 locus (mChr2) was from DBA/2 on a full BALB/c background. Congenic mice were tested for PGISpA, and then backcrossed to BALB/c strain. Recombination events were genotyped until partially overlapping DBA/2 intervals (n=12) completely covered the Pgis2 locus. Heterozygous females and males with the same genomic intervals were intercrossed, homozygous offspring tested for PGISpA. Based on the overlapping Chr2 intervals, we narrowed a resistant (DBA/2 origin) to ~3.5 Mbp in size (mChr2:30.5-34.0 Mbp; syntenic with hChr9:129.-132.8 Mbp). All IVSC mice were tested with near infrared OsteoSenseTM 750 (Visen Medical) probe, spine histology, serum cytokines and cytokines in antigen-stimulated in vitro spleen cell cultures. The corresponding 3 Mbp genomic regions of PGISpA-resistant and –susceptible IVSC and parent strains (n=10) were sequenced by high-throughput methods and analyzed using the GeneSpring NGS program.
We identified a total of 4,416 mutations within the 3.5 Mbp-long genomic region . The 3.5 Mbp region contains 70 protein-coding genes, 12 antisense transcripts (RNA) and 25 different forms (small nucloelar, miRNA) of non-coding RNAs. Mutations/indels were detected (93.38%) in three relatively small genomic regions affecting 3 (Gpr107-Nsc1-Hmcn2) genes and their intergenic regions, and two other genes (St6galnac6 and Lmx1b). PGISpA was significantly reduced (p<0.001) in each IVSC strain carrying any of these three DBA/2 alleles. The Gpr107-Nsc1-Hmcn2 triplet contained mutations affecting multiple transcription binding sites, the Lmx1b transcription factor has an in-frame 18-nucleotide deletion in exon 1, and the promoter region of the St6galnac6 was hypermethylated in genomic regions of DBA/2 origin.
Although all three alleles affected PGISpA individually, it appears that an “epistatic triangle” exists among these alleles that protect against PGISpA. Overall, all mutations with predicted functional consequence are located in non-coding sequences underlining the critical role of epigenetic alterations in PGISpA (and perhaps in AS).
T. A. Rauch,
H. L. Rosenzweig,
M. A. Brown,
G. P. Thomas,
T. T. Glant,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/identification-of-genetic-and-epigenetic-alterations-in-spondyloarthritis/