Gastroenterology

Gastroenterology

Volume 145, Issue 2, August 2013, Pages 339-347
Gastroenterology

Original Research
Full Report: Clinical—Alimentary Tract
Association Between Variants of PRDM1 and NDP52 and Crohn's Disease, Based on Exome Sequencing and Functional Studies

https://doi.org/10.1053/j.gastro.2013.04.040Get rights and content

Background & Aims

Genome-wide association studies (GWAS) have identified 140 Crohn’s disease (CD) susceptibility loci. For most loci, the variants that cause disease are not known and the genes affected by these variants have not been identified. We aimed to identify variants that cause CD through detailed sequencing, genetic association, expression, and functional studies.

Methods

We sequenced whole exomes of 42 unrelated subjects with CD and 5 healthy subjects (controls) and then filtered single nucleotide variants by incorporating association results from meta-analyses of CD GWAS and in silico mutation effect prediction algorithms. We then genotyped 9348 subjects with CD, 2868 subjects with ulcerative colitis, and 14,567 control subjects and associated variants analyzed in functional studies using materials from subjects and controls and in vitro model systems.

Results

We identified rare missense mutations in PR domain-containing 1 (PRDM1) and associated these with CD. These mutations increased proliferation of T cells and secretion of cytokines on activation and increased expression of the adhesion molecule L-selectin. A common CD risk allele, identified in GWAS, correlated with reduced expression of PRDM1 in ileal biopsy specimens and peripheral blood mononuclear cells (combined P = 1.6 × 10−8). We identified an association between CD and a common missense variant, Val248Ala, in nuclear domain 10 protein 52 (NDP52) (P = 4.83 × 10−9). We found that this variant impairs the regulatory functions of NDP52 to inhibit nuclear factor κB activation of genes that regulate inflammation and affect the stability of proteins in Toll-like receptor pathways.

Conclusions

We have extended the results of GWAS and provide evidence that variants in PRDM1 and NDP52 determine susceptibility to CD. PRDM1 maps adjacent to a CD interval identified in GWAS and encodes a transcription factor expressed by T and B cells. NDP52 is an adaptor protein that functions in selective autophagy of intracellular bacteria and signaling molecules, supporting the role of autophagy in the pathogenesis of CD.

Section snippets

Study Subjects

We captured the exomes of 42 German patients with CD, one HapMap trio,6 and 3 unrelated German healthy control subjects by means of the NimbleGen 2.1M Human Exome Array (Madison, WI), followed by Illumina Genome Analyzer (San Diego, CA) resequencing (Supplementary Figures 1 and 2 and Supplementary Table 1). For detailed information on the whole-exome enrichment and sequencing process and sequence read alignment, variant calling, and annotation, see Supplementary Notes. Follow-up genotyping of

Discovery of Low-Frequency Variants Using Whole-Exome Sequencing

In total, we identified 117,957 SNVs in our 48 subjects with CD, including 59,076 coding and splice site SNVs (Supplementary Tables 2 and 3). The overall SNV analysis workflow of our study is shown in Supplementary Figure 3. To pinpoint a small subset of functionally important SNVs from the large amount of genetic variation, we developed a software tool for SNV annotation, categorization, and filtering named snpActs (Supplementary Table 4 and Supplementary Notes).

Results of Association Testing

Two missense SNPs,

Discussion

The outcomes of the present study highlight the importance of detailed multifaceted analyses to extend genetic insights beyond those derived from GWAS alone. By conducting whole-exome sequencing followed by rare variant analysis, as well as detailed expression and functional studies, we have substantially advanced understanding of a previously defined GWAS locus and have identified a new CD susceptibility gene that further emphasizes the contribution of autophagy and NF-κB pathways to the

Acknowledgments

The authors thank all subjects with CD or UC and their families, control subjects, and clinicians for their participation; Tanja Wesse, Tanja Henke, Sanaz Sedghpour Sabet, Sandra Greve, and Ilona Urbach for expert technical help; and the International IBD Genetics Consortium for providing summary statistics of the CD and UC meta-analyses through their website (http://www.ibdgenetics.org).

References (32)

  • J. Hampe et al.

    A genome-wide association scan of nonsynonymous SNPs identifies a susceptibility variant for Crohn disease in ATG16L1

    Nat Genet

    (2007)
  • J.D. Rioux et al.

    Genome-wide association study identifies new susceptibility loci for Crohn disease and implicates autophagy in disease pathogenesis

    Nat Genet

    (2007)
  • M. Parkes et al.

    Sequence variants in the autophagy gene IRGM and multiple other replicating loci contribute to Crohn's disease susceptibility

    Nat Genet

    (2007)
  • L. Henckaerts et al.

    Genetic variation in the autophagy gene ULK1 and risk of Crohn's disease

    Inflamm Bowel Dis

    (2011)
  • D.A. Steeber et al.

    Efficient lymphocyte migration across high endothelial venules of mouse Peyer's patches requires overlapping expression of L-selectin and beta7 integrin

    J Immunol

    (1998)
  • N. Wagner et al.

    L-selectin and beta7 integrin synergistically mediate lymphocyte migration to mesenteric lymph nodes

    Eur J Immunol

    (1998)
  • Cited by (125)

    • Molecular functions of autophagy adaptors upon ubiquitin-driven mitophagy

      2021, Biochimica et Biophysica Acta - General Subjects
      Citation Excerpt :

      The findings will provide a more nuanced understanding of the role autophagy adaptors play, which is critical given their relationship to human diseases. Mutations to OPTN have been linked to ALS and POAG, whereas missense mutations in p62 have been identified in Paget's disease of bone, frontotemporal lobar degradation, and ALS [133], and associations between NDP52 and Crohn's disease have been reported [134]. Although it remains to be determined if the onset of these disease states is linked to the ubiquitin and/or autophagy machinery, research into the molecular functions of autophagy adaptors and their recently-identified novel binding partners will provide new insights for therapeutic and pharmacological approaches.

    • A catalog of GWAS fine-mapping efforts in autoimmune disease

      2021, American Journal of Human Genetics
    View all citing articles on Scopus

    Conflicts of interest The authors disclose no conflicts.

    Funding Supported by Deutsche Forschungsgemeinschaft (DFG) grant FR 2821/2-1 and research fellowship Ti 640 1-1, the SFB 877 subproject B9, the German Ministry of Education and Research through the National Genome Research Network, the PopGen biobank (http://www.popgen.de), and the National Institutes of Health (grant GM069373). The project received infrastructure support through the DFG Clusters of Excellence “Inflammation at Interfaces” and “Multimodal Computing and Interaction.” S. Brand is supported by DFG grant BR1912/6-1 and by the Else-Kröner-Fresenius-Stiftung (stipend 2010_EKES.32). The authors acknowledge use of material from the British 1958 Birth Cohort DNA collection, funded by Medical Research Council grant G0000934 and Wellcome Trust grant 068545/Z/02 (http://www.b58cgene.sgul.ac.uk/). The authors also acknowledge the participation of all Cambridge BioResource subjects and staff (http://www.cambridgebioresource.org.uk/); the National Institute for Health Research Biomedical Research Centre awards to Guy’s & St Thomas’ National Health Service Trust/King’s College London and to Addenbrooke’s Hospital/University of Cambridge School of Clinical Medicine; the Wellcome Trust Case Control Consortium, a seed grant from the San Diego Center for Systems Biology (to A. Till and S. Subramani), National Institutes of Health grant 1R01CA141743-01 (principal investigator: R. H. Duerr), the University of Pittsburgh Genomics and Proteomics Core Laboratories for the ImmunoChip genotyping services, and DFG grants ZE 814/4-1 and ZE 814/5-1 (to S. Zeissig). M. D’Amato received support from the Swedish Research Council (VR).

    Authors share co-first authorship.

    View full text