Limits...
Intestinal DMBT1 expression is modulated by Crohn's disease-associated IL23R variants and by a DMBT1 variant which influences binding of the transcription factors CREB1 and ATF-2.

Diegelmann J, Czamara D, Le Bras E, Zimmermann E, Olszak T, Bedynek A, Göke B, Franke A, Glas J, Brand S - PLoS ONE (2013)

Bottom Line: Several DMBT1 SNPs were associated with CD susceptibility.All haplotype groups tested showed highly significant associations with CD (including omnibus P-values as low as 6.1 × 10(-18)).Intestinal DMBT1 expression is decreased in CD patients carrying the rs2981804 CD risk allele.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine II - Grosshadern, Ludwig-Maximilians-University (LMU), Munich, Germany ; Department of Preventive Dentistry and Periodontology, Ludwig-Maximilians-University (LMU), Munich, Germany.

ABSTRACT

Objectives: DMBT is an antibacterial pattern recognition and scavenger receptor. In this study, we analyzed the role of DMBT1 single nucleotide polymorphisms (SNPs) regarding inflammatory bowel disease (IBD) susceptibility and examined their functional impact on transcription factor binding and downstream gene expression.

Methods: Seven SNPs in the DMBT1 gene region were analyzed in 2073 individuals including 818 Crohn's disease (CD) patients and 972 healthy controls in two independent case-control panels. Comprehensive epistasis analyses for the known CD susceptibility genes NOD2, IL23R and IL27 were performed. The influence of IL23R variants on DMBT1 expression was analyzed. Functional analysis included siRNA transfection, quantitative PCR, western blot, electrophoretic mobility shift and luciferase assays.

Results: IL-22 induces DMBT1 protein expression in intestinal epithelial cells dependent on STAT3, ATF-2 and CREB1. IL-22 expression-modulating, CD risk-associated IL23R variants influence DMBT1 expression in CD patients and DMBT1 levels are increased in the inflamed intestinal mucosa of CD patients. Several DMBT1 SNPs were associated with CD susceptibility. SNP rs2981804 was most strongly associated with CD in the combined panel (p = 3.0 × 10(-7), OR 1.42; 95% CI 1.24-1.63). All haplotype groups tested showed highly significant associations with CD (including omnibus P-values as low as 6.1 × 10(-18)). The most strongly CD risk-associated, non-coding DMBT1 SNP rs2981804 modifies the DNA binding sites for the transcription factors CREB1 and ATF-2 and the respective genomic region comprising rs2981804 is able to act as a transcriptional regulator in vitro. Intestinal DMBT1 expression is decreased in CD patients carrying the rs2981804 CD risk allele.

Conclusion: We identified novel associations of DMBT1 variants with CD susceptibility and discovered a novel functional role of rs2981804 in regulating DMBT1 expression. Our data suggest an important role of DMBT1 in CD pathogenesis.

Show MeSH

Related in: MedlinePlus

IL23R CD risk-increasing variants are associated with higher DMBT1 expression in minor allele carriers and intestinal inflammation increases DMBT expression levels in CD patients.(A) Intestinal DMBT1 expression and IL23R genotypes were determined in 75 biopsies from 27 CD patients. DMBT1 expression is presented as a quotient derived from dividing expression in minor allele carriers of the respective IL23R SNP by the expression in WT carriers. ORs for the respective SNPs were available from a previous study [8]. (B) Intestinal DMBT1 expression is significantly higher in inflamed colonic biopsies (n = 30) compared to not inflamed regions (n = 45) from 27 CD patients. Expression was normalized to β-actin in the respective samples. Each dot represents one biopsy.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3818382&req=5

pone-0077773-g003: IL23R CD risk-increasing variants are associated with higher DMBT1 expression in minor allele carriers and intestinal inflammation increases DMBT expression levels in CD patients.(A) Intestinal DMBT1 expression and IL23R genotypes were determined in 75 biopsies from 27 CD patients. DMBT1 expression is presented as a quotient derived from dividing expression in minor allele carriers of the respective IL23R SNP by the expression in WT carriers. ORs for the respective SNPs were available from a previous study [8]. (B) Intestinal DMBT1 expression is significantly higher in inflamed colonic biopsies (n = 30) compared to not inflamed regions (n = 45) from 27 CD patients. Expression was normalized to β-actin in the respective samples. Each dot represents one biopsy.

Mentions: We have recently demonstrated that CD-associated variants in IL23R modulate IL-22 expression. Higher IL-22 serum levels were found in carriers of CD risk-associated IL23R SNPs and lower IL-22 serum levels in carriers of CD-protective IL23R SNPs [34] (summarized in table 3). Having shown that IL-22 induces DMBT1 expression, we further analyzed these IL23R variants regarding their association with DMBT1 expression. DMBT1 mRNA expression was determined by quantitative PCR in a total of n = 75 inflamed and not inflamed intestinal biopsies collected from 27 CD patients (for patient characteristics, see table 2). DMBT1 expression in carriers of the minor allele (homozygous and heterozygous) of the respective IL23R SNPs was divided by the expression in homozygous carriers of the wild-type (WT) allele (table 3, Fig. 3A). The odds ratios (ORs) for the analyzed IL23R SNPs were available from a previous study [8]. In 5 out of 10 IL23R SNPs, there were significant differences (p<0.05) in DMBT1 expression between carriers of the minor allele and WT carriers (table 3). Similar to our previous results obtained for IL-22 [34], there was a high correlation of OR regarding CD susceptibility and DMBT expression ratio minor allele vs. WT (r (OR/DMBT) = 0.766) and between IL-22 and DMBT1 ratios minor allele vs WT (r(DMBT/IL-22) = 0.754). For all SNPs except rs7517847, patients with risk-increasing IL23R variants had higher DMBT1 expression when carrying the minor allele (OR>1 and relative DMBT1 expression minor allele vs WT>1), while in patients with IBD risk-decreasing IL23R variants, DMBT1 expression was lower in minor allele carriers compared to WT carriers (OR<1 and relative expression minor vs. WT<1; table 3, Fig. 3A). The correlation of OR and DMBT1 expression was independent of the inflammation status of the analyzed biopsies (figure S3).


Intestinal DMBT1 expression is modulated by Crohn's disease-associated IL23R variants and by a DMBT1 variant which influences binding of the transcription factors CREB1 and ATF-2.

Diegelmann J, Czamara D, Le Bras E, Zimmermann E, Olszak T, Bedynek A, Göke B, Franke A, Glas J, Brand S - PLoS ONE (2013)

IL23R CD risk-increasing variants are associated with higher DMBT1 expression in minor allele carriers and intestinal inflammation increases DMBT expression levels in CD patients.(A) Intestinal DMBT1 expression and IL23R genotypes were determined in 75 biopsies from 27 CD patients. DMBT1 expression is presented as a quotient derived from dividing expression in minor allele carriers of the respective IL23R SNP by the expression in WT carriers. ORs for the respective SNPs were available from a previous study [8]. (B) Intestinal DMBT1 expression is significantly higher in inflamed colonic biopsies (n = 30) compared to not inflamed regions (n = 45) from 27 CD patients. Expression was normalized to β-actin in the respective samples. Each dot represents one biopsy.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3818382&req=5

pone-0077773-g003: IL23R CD risk-increasing variants are associated with higher DMBT1 expression in minor allele carriers and intestinal inflammation increases DMBT expression levels in CD patients.(A) Intestinal DMBT1 expression and IL23R genotypes were determined in 75 biopsies from 27 CD patients. DMBT1 expression is presented as a quotient derived from dividing expression in minor allele carriers of the respective IL23R SNP by the expression in WT carriers. ORs for the respective SNPs were available from a previous study [8]. (B) Intestinal DMBT1 expression is significantly higher in inflamed colonic biopsies (n = 30) compared to not inflamed regions (n = 45) from 27 CD patients. Expression was normalized to β-actin in the respective samples. Each dot represents one biopsy.
Mentions: We have recently demonstrated that CD-associated variants in IL23R modulate IL-22 expression. Higher IL-22 serum levels were found in carriers of CD risk-associated IL23R SNPs and lower IL-22 serum levels in carriers of CD-protective IL23R SNPs [34] (summarized in table 3). Having shown that IL-22 induces DMBT1 expression, we further analyzed these IL23R variants regarding their association with DMBT1 expression. DMBT1 mRNA expression was determined by quantitative PCR in a total of n = 75 inflamed and not inflamed intestinal biopsies collected from 27 CD patients (for patient characteristics, see table 2). DMBT1 expression in carriers of the minor allele (homozygous and heterozygous) of the respective IL23R SNPs was divided by the expression in homozygous carriers of the wild-type (WT) allele (table 3, Fig. 3A). The odds ratios (ORs) for the analyzed IL23R SNPs were available from a previous study [8]. In 5 out of 10 IL23R SNPs, there were significant differences (p<0.05) in DMBT1 expression between carriers of the minor allele and WT carriers (table 3). Similar to our previous results obtained for IL-22 [34], there was a high correlation of OR regarding CD susceptibility and DMBT expression ratio minor allele vs. WT (r (OR/DMBT) = 0.766) and between IL-22 and DMBT1 ratios minor allele vs WT (r(DMBT/IL-22) = 0.754). For all SNPs except rs7517847, patients with risk-increasing IL23R variants had higher DMBT1 expression when carrying the minor allele (OR>1 and relative DMBT1 expression minor allele vs WT>1), while in patients with IBD risk-decreasing IL23R variants, DMBT1 expression was lower in minor allele carriers compared to WT carriers (OR<1 and relative expression minor vs. WT<1; table 3, Fig. 3A). The correlation of OR and DMBT1 expression was independent of the inflammation status of the analyzed biopsies (figure S3).

Bottom Line: Several DMBT1 SNPs were associated with CD susceptibility.All haplotype groups tested showed highly significant associations with CD (including omnibus P-values as low as 6.1 × 10(-18)).Intestinal DMBT1 expression is decreased in CD patients carrying the rs2981804 CD risk allele.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine II - Grosshadern, Ludwig-Maximilians-University (LMU), Munich, Germany ; Department of Preventive Dentistry and Periodontology, Ludwig-Maximilians-University (LMU), Munich, Germany.

ABSTRACT

Objectives: DMBT is an antibacterial pattern recognition and scavenger receptor. In this study, we analyzed the role of DMBT1 single nucleotide polymorphisms (SNPs) regarding inflammatory bowel disease (IBD) susceptibility and examined their functional impact on transcription factor binding and downstream gene expression.

Methods: Seven SNPs in the DMBT1 gene region were analyzed in 2073 individuals including 818 Crohn's disease (CD) patients and 972 healthy controls in two independent case-control panels. Comprehensive epistasis analyses for the known CD susceptibility genes NOD2, IL23R and IL27 were performed. The influence of IL23R variants on DMBT1 expression was analyzed. Functional analysis included siRNA transfection, quantitative PCR, western blot, electrophoretic mobility shift and luciferase assays.

Results: IL-22 induces DMBT1 protein expression in intestinal epithelial cells dependent on STAT3, ATF-2 and CREB1. IL-22 expression-modulating, CD risk-associated IL23R variants influence DMBT1 expression in CD patients and DMBT1 levels are increased in the inflamed intestinal mucosa of CD patients. Several DMBT1 SNPs were associated with CD susceptibility. SNP rs2981804 was most strongly associated with CD in the combined panel (p = 3.0 × 10(-7), OR 1.42; 95% CI 1.24-1.63). All haplotype groups tested showed highly significant associations with CD (including omnibus P-values as low as 6.1 × 10(-18)). The most strongly CD risk-associated, non-coding DMBT1 SNP rs2981804 modifies the DNA binding sites for the transcription factors CREB1 and ATF-2 and the respective genomic region comprising rs2981804 is able to act as a transcriptional regulator in vitro. Intestinal DMBT1 expression is decreased in CD patients carrying the rs2981804 CD risk allele.

Conclusion: We identified novel associations of DMBT1 variants with CD susceptibility and discovered a novel functional role of rs2981804 in regulating DMBT1 expression. Our data suggest an important role of DMBT1 in CD pathogenesis.

Show MeSH
Related in: MedlinePlus