Background Genome-wide association studies identified a PTGER4 expression-modulating region on chromosome 5p13.1 as Crohn's disease (CD) susceptibility region. The study aim was to test this association in a large cohort of patients with inflammatory bowel disease (IBD) and to elucidate genotypic and phenotypic interactions with other IBD genes. Methodology/Principal Findings A total of 7073 patients and controls were genotyped: 844 CD and 471 patients with ulcerative colitis and 1488 controls were analyzed for the single nucleotide polymorphisms (SNPs) rs4495224 and rs7720838 on chromosome 5p13.1. The study included two replication cohorts of North American (CD: n = 684; controls: n = 1440) and of German origin (CD: n = 1098; controls: n = 1048). Genotype-phenotype, epistasis and transcription factor binding analyses were performed. In the discovery cohort, an association of rs4495224 (p = 4.10×10？5; 0.76 [0.67–0.87]) and of rs7720838 (p = 6.91×10？4; 0.81 [0.71–0.91]) with susceptibility to CD was demonstrated. These associations were confirmed in both replication cohorts. In silico analysis predicted rs4495224 and rs7720838 as essential parts of binding sites for the transcription factors NF-κB and XBP1 with higher binding scores for carriers of the CD risk alleles, providing an explanation of how these SNPs might contribute to increased PTGER4 expression. There was no association of the PTGER4 SNPs with IBD phenotypes. Epistasis detected between 5p13.1 and ATG16L1 for CD susceptibility in the discovery cohort (p = 5.99×10？7 for rs7720838 and rs2241880) could not be replicated in both replication cohorts arguing against a major role of this gene-gene interaction in the susceptibility to CD. Conclusions/Significance We confirmed 5p13.1 as a major CD susceptibility locus and demonstrate by in silico analysis rs4495224 and rs7720838 as part of binding sites for NF-κB and XBP1. Further functional studies are necessary to confirm the results of our in silico analysis and to analyze if changes in PTGER4 expression modulate CD susceptibility.
References
[1]
Podolsky DK (2002) Inflammatory bowel disease. N Engl J Med 347: 417–429.
[2]
Xavier RJ, Podolsky DK (2007) Unravelling the pathogenesis of inflammatory bowel disease. Nature 448: 427–434.
[3]
Sartor RB (2006) Mechanisms of disease pathogenesis of Crohn's disease and ulcerative colitis. Nat Clin Pract Gastroenterol Hepatol 3: 390–407.
[4]
Cho JH, Weaver CT (2007) The genetics of inflammatory bowel disease. Gastroenterology 133: 1327–1339.
[5]
Ogura Y, Bonen DK, Inohara N, Nicolae DL, Chen FF, et al. (2001) A frameshift mutation in NOD2 associated with susceptibility to Crohn's disease. Nature 411: 603–606.
[6]
Hugot JP, Chamaillard M, Zouali H, Lesage S, Cezard JP, et al. (2001) Association of NOD2 leucine-rich repeat variants with susceptibility to Crohn's disease. Nature 411: 599–603.
[7]
Duerr RH, Taylor KD, Brant SR, Rioux JD, Silverberg MS, et al. (2006) A genome-wide association study identifies IL23R as an inflammatory bowel disease gene. Science 314: 1461–1463.
[8]
Glas J, Seiderer J, Wetzke M, Konrad A, T？r？k HP, et al. (2007) rs1004819 is the main disease-associated IL23R variant in German Crohn's disease patients: combined analysis of IL23R, CARD15, and OCTN1/2 variants. PLoS ONE 2: e819.
[9]
Hampe J, Franke A, Rosenstiel P, Till A, Teuber M, et al. (2007) A genome-wide association scan of nonsynonymous SNPs identifies a susceptibility variant for Crohn disease in ATG16L1. Nat Genet 39: 207–211.
[10]
Rioux JD, Xavier RJ, Taylor KD, Silverberg MS, Goyette P, et al. (2007) Genome-wide association study identifies new susceptibility loci for Crohn disease and implicates autophagy in disease pathogenesis. Nat Genet 39: 596–604.
[11]
Glas J, Konrad A, Schmechel S, Dambacher J, Seiderer J, et al. (2007) The ATG16L1 gene variants rs2241879 and rs2241880 (T300A) are strongly associated with susceptibility to Crohn's disease in the German population. Am J Gastroenterol 103: 682–691.
[12]
Parkes M, Barrett JC, Prescott NJ, Tremelling M, Anderson CA, et al. (2007) Sequence variants in the autophagy gene IRGM and multiple other replicating loci contribute to Crohn's disease susceptibility. Nat Genet 39: 830–832.
[13]
Brand S, Hofbauer K, Dambacher J, Schnitzler F, Staudinger T, et al. (2006) Increased expression of the chemokine fractalkine in Crohn's disease and association of the fractalkine receptor T280M polymorphism with a fibrostenosing disease phenotype. Am J Gastroenterol 101: 99–106.
[14]
Brand S, Staudinger T, Schnitzler F, Pfennig S, Hofbauer K, et al. (2005) The role of Toll-like receptor 4 Asp299Gly and Thr399Ile polymorphisms and CARD15/NOD2 mutations in the susceptibility and phenotype of Crohn's disease. Inflamm Bowel Dis 11: 645–652.
[15]
Schnyder-Candrian S, Togbe D, Couillin I, Mercier I, Brombacher F, et al. (2006) Interleukin-17 is a negative regulator of established allergic asthma. J Exp Med 203: 2715–2725.
[16]
Seiderer J, Schnitzler F, Brand S, Staudinger T, Pfennig S, et al. (2006) Homozygosity for the CARD15 frameshift mutation 1007fs is predictive of early onset of Crohn's disease with ileal stenosis, entero-enteral fistulas, and frequent need for surgical intervention with high risk of re-stenosis. Scand J Gastroenterol 41: 1421–1432.
[17]
Seiderer J, Brand S, Herrmann KA, Schnitzler F, Hatz R, et al. (2006) Predictive value of the CARD15 variant 1007fs for the diagnosis of intestinal stenoses and the need for surgery in Crohn's disease in clinical practice: results of a prospective study. Inflamm Bowel Dis 12: 1114–1121.
[18]
Thalmaier D, Dambacher J, Seiderer J, Konrad A, Schachinger V, et al. (2006) The +1059G/C polymorphism in the C-reactive protein (CRP) gene is associated with involvement of the terminal ileum and decreased serum CRP levels in patients with Crohn's disease. Aliment Pharmacol Ther 24: 1105–1115.
[19]
Schnitzler F, Brand S, Staudinger T, Pfennig S, Hofbauer K, et al. (2006) Eight novel CARD15 variants detected by DNA sequence analysis of the CARD15 gene in 111 patients with inflammatory bowel disease. Immunogenetics 58: 99–106.
[20]
Glas J, Seiderer J, Nagy M, Fries C, Beigel F, et al. (2010) Evidence for STAT4 as a common autoimmune gene: rs7574865 is associated with colonic Crohn's disease and early disease onset. PLoS One 5: e10373.
[21]
Libioulle C, Louis E, Hansoul S, Sandor C, Farnir F, et al. (2007) Novel Crohn disease locus identified by genome-wide association maps to a gene desert on 5p13.1 and modulates expression of PTGER4. PLoS Genet 3: e58.
[22]
Kabashima K, Saji T, Murata T, Nagamachi M, Matsuoka T, et al. (2002) The prostaglandin receptor EP4 suppresses colitis, mucosal damage and CD4 cell activation in the gut. J Clin Invest 109: 883–893.
[23]
Nitta M, Hirata I, Toshina K, Murano M, Maemura K, et al. (2002) Expression of the EP4 prostaglandin E2 receptor subtype with rat dextran sodium sulphate colitis: colitis suppression by a selective agonist, ONO-AE1-329. Scand J Immunol 56: 66–75.
[24]
Jiang GL, Nieves A, Im WB, Old DW, Dinh DT, et al. (2007) The prevention of colitis by E Prostanoid receptor 4 agonist through enhancement of epithelium survival and regeneration. J Pharmacol Exp Ther 320: 22–28.
[25]
Cox DG, Crusius JB, Peeters PH, Bueno-de-Mesquita HB, Pena AS, et al. (2005) Haplotype of prostaglandin synthase 2/cyclooxygenase 2 is involved in the susceptibility to inflammatory bowel disease. World J Gastroenterol 11: 6003–6008.
[26]
Subbaramaiah K, Yoshimatsu K, Scherl E, Das KM, Glazier KD, et al. (2004) Microsomal prostaglandin E synthase-1 is overexpressed in inflammatory bowel disease. Evidence for involvement of the transcription factor Egr-1. J Biol Chem 279: 12647–12658.
[27]
Cuzzocrea S, Ianaro A, Wayman NS, Mazzon E, Pisano B, et al. (2003) The cyclopentenone prostaglandin 15-deoxy-delta(12,14)- PGJ2 attenuates the development of colon injury caused by dinitrobenzene sulphonic acid in the rat. Br J Pharmacol 138: 678–688.
[28]
Franke A, Hampe J, Rosenstiel P, Becker C, Wagner F, et al. (2007) Systematic association mapping identifies NELL1 as a novel IBD disease gene. PLoS ONE 2: e691.
[29]
McGovern DP, Gardet A, T？rkvist L, Goyette P, Essers J, et al. (2010) Genome-wide association identifies multiple ulcerative colitis susceptibility loci. Nat Genet 42: 332–337.
[30]
Lennard-Jones JE (1989) Classification of inflammatory bowel disease. Scand J Gastroenterol Suppl 170: 2–6 discussion 16–19.
[31]
Silverberg MS, Satsangi J, Ahmad T, Arnott ID, Bernstein CN, et al. (2005) Toward an integrated clinical, molecular and serological classification of inflammatory bowel disease: Report of a Working Party of the 2005 Montreal World Congress of Gastroenterology. Can J Gastroenterol 19 Suppl A: 5–36.
[32]
T？r？k HP, Glas J, Tonenchi L, Lohse P, Müller-Myhsok B, et al. (2005) Polymorphisms in the DLG5 and OCTN cation transporter genes in Crohn's disease. Gut 54: 1421–1427.
[33]
Heinemeyer T, Wingender E, Reuter I, Hermjakob H, Kel AE, et al. (1998) Databases on transcriptional regulation: TRANSFAC, TRRD and COMPEL. Nucleic Acids Res 26: 362–7.
[34]
Kaser A, Lee AH, Franke A, Glickman JN, Zeissig S, et al. (2008) XBP1 links ER stress to intestinal inflammation and confers genetic risk for human inflammatory bowel disease. Cell 134: 743–56.
[35]
Franke A, McGovern DP, Barrett JC, Wang K, Radford-Smith GL, et al. (2010) Genome-wide meta-analysis increases to 71 the number of confirmed Crohn's disease susceptibility loci. Nat Genet 42: 1118–1125.
[36]
Anderson CA, Boucher G, Lees CW, Franke A, D'Amato M, et al. (2011) Meta-analysis identifies 31 additional ulcerative colitis risk loci, increasing the number of confirmed associations to 49. Nat Genet 43: 246–252.
[37]
Jostins L, Ripke L, Weersma R, Duerr RH, McGovern DP, et al. (2012) Host-microbe interactions have shaped the genetic architecture of inflammatory bowel disease. Nature 491: 119–24.
[38]
Okamoto T, Uemoto S, Tabata Y (2012) Prevention of trinitrobenzene sulfonic acid-induced experimental colitis by oral administration of a poly(lactic-coglycolic acid microsphere containing prostaglandin E2 receptor subtype 4 agonist. J Pharmacol Exp Ther 341: 340–349.
[39]
Clark P, Rowland SE, Denis D, Mathieu MC, Stocco R, et al. (2008) MF498 [N-{[4-(5,9-Diethoxy-6-oxo-6,8-dihydro-7？H-pyrrolo[3,4-g]quinolin-7-yl)-3-methylb？enzyl]sulfonyl}-2-(2-methoxyphenyl)aceta？mide],a selective E prostanoid receptor 4 antagonist, relieves joint inflammation and pain in rodent models of rheumatoid and osteoarthritis. J Pharmacol Exp Ther 325: 425–434.
[40]
Chen Q, Muramoto K, Masaaki N, Ding Y, Yang H, et al. (2010) A novel antagonist of the prostaglandin E(2) EP(4) receptor inhibits Th1 differentiation and Th17 expansion and is orally active in arthritis models. Br J Pharmacol 160: 292–310.
[41]
Yao C, Sakata D, Esaki Y, Li Y, Matsuoka T, et al. (2009) Prostaglandin E2-EP4 signaling promotes immune inflammation through Th1 cell differentiation and Th17 cell expansion. Nat Med 15: 633–640.
[42]
Boniface K, Bak-Jensen KS, Li Y, Blumenschein WM, McGeachy MJ, et al. (2009) Prostaglandin E2 regulates Th17 cell differentiation and function through cyclic AMP and EP2/EP4 receptor signaling. J Exp Med 206: 535–548.
[43]
Brand S (2009) Crohn's disease: Th1, Th17 or both? The change of a paradigm: new immunological and genetic insights implicate Th17 cells in the pathogenesis of Crohn's disease. Gut 2009 58: 1152–1167.
[44]
Atreya I, Atreya R, Neurath MF (2008) NF-kappaB in inflammatory bowel disease. J Intern Med 263: 591–596.
[45]
Morteau O (2000) Prostaglandins and inflammation: the cyclooxygenase controversy. Arch Immunol Ther Exp (Warsz) 48: 473–480.
[46]
Dey I, Lejeune M, Chadee K (2006) Prostaglandin E2 receptor distribution and function in the gastrointestinal tract. Br J Pharmacol 149: 611–623.
[47]
Gookin JL, Galanko JA, Blikslager AT, Argenzio RA (2003) PG-mediated closure of paracellular pathway and not restitution is the primary determinant of barrier recovery in acutely injured porcine ileum. Am J Physiol Gastrointest Liver Physiol 285: G967–G979.
[48]
Morteau O, Morham SG, Sellon R, Dieleman LA, Langenbach R, et al. (2000) Impaired mucosal defense to acute colonic injury in mice lacking cyclooxygenase-1 or cyclooxygenase-2. J Clin Invest 105: 469–478.
[49]
Kurz T, Hoffjan S, Hayes MG, Schneider D, Nicolae R, et al. (2006) Fine mapping and positional candidate studies on chromosome 5p13 identify multiple asthma susceptibility loci. J Allergy Clin Immunol 118: 396–402.
[50]
Kim SH, Kim YK, Park HW, Jee YK, Bahn JW, et al. (2007) Association between polymorphisms in prostanoid receptor genes and aspirin-intolerant asthma. Pharmacogenet Genomics 17: 295–304.
[51]
Kabashima K, Sakata D, Nagamachi M, Miyachi Y, Inaba K (2003) Narumiya S. Prostaglandin E2-EP4 signaling initiates skin immune responses by promoting migration and maturation of Langerhans cells. Nat Med 9: 744–749.
[52]
Niess JH, Brand S, Gu X, Landsman L, Jung S, et al. (2005) CX3CR1-mediated dendritic cell access to the intestinal lumen and bacterial clearance. Science 307: 254–258.
[53]
Barrett JC, Hansoul S, Nicolae DL, Cho JH, Duerr RH, et al. (2008) Genome-wide association defines more than 30 distinct susceptibility loci for Crohn's disease. Nat Genet 40: 955–962.
[54]
Stoll M, Corneliussen B, Costello CM, Waetzig GH, Mellgard B, et al. (2004) Genetic variation in DLG5 is associated with inflammatory bowel disease. Nat Genet 36: 476–480.
[55]
Glas J, Seiderer J, Pasciuto G, Tillack C, Diegelmann J, et al. (2009) rs224136 on chromosome 10q21.1 and variants in PHOX2B, NCF4, and FAM92B are not major genetic risk factors for susceptibility to Crohn's disease in the German population. Am J Gastroenterol 104: 665–672.
[56]
Glas J, Seiderer J, Tillack C, Pfennig S, Beigel F, et al. (2010) The NOD2 single nucleotide polymorphisms rs2066843 and rs2076756 are novel and common Crohn's disease susceptibility gene variants. PLoS One 5: e14466.
[57]
Jürgens M, Brand S, Laubender RP, Seiderer J, Glas J, et al. (2010) The presence of fistulas and NOD2 homozygosity strongly predict intestinal stenosis in Crohn's disease independent of the IL23R genotype. J Gastroenterol 45: 721–31.
