title: Association of Ribonuclease T2 Gene Polymorphisms With Decreased Expression and Clinical Characteristics of Severity in Crohn's Disease.
abstract: Variants in the tumor necrosis factor superfamily member 15 gene (TNFSF15, also called TL1A) have been associated with risk for inflammatory bowel disease (IBD). TL1A affects expression of multiple cytokines to promote mucosal inflammation. Little is known about the TL1A-response pathways that regulate cytokine expression. We investigated T-cell gene expression patterns to determine the mechanisms by which TL1A regulates cytokine production, and whether these associate with outcomes of patients with Crohn's disease (CD).
  
  
  
  Peripheral T cells isolated from normal donors were cultured with TL1A. We performed gene expression profile analysis by RNA sequencing of subsets of interferon gamma (IFNG)-producing and non-producing cells purified by flow cytometry. Unsupervised hierarchical clustering analysis was used to identify gene expression differences between these subsets. Ribonuclease T2 gene (RNASET2) expression and methylation were assessed by quantitative trait loci analyses. Clinical characteristics of patients (complications, resistance to therapy, and recurrence time) were associated with single nucleotide polymorphisms in RNASET2. We performed motif screening to identify polymorphisms that disrupt transcription factor binding sites. Levels of RNASET2 were knocked down with small interfering RNA in CD4+T cells and the effect on protein expression was determined by proteomic analysis and cytokine production. Cell aggregation was measured by flow cytometry.
  
  
  
  We identified 764 genes with at least a 2-fold difference in TL1A-mediated expression between IFNG-secreting and non-secreting T cells (P < 1 × 10-5). Many of these genes were located near IBD susceptibility variants. RNASET2 was the only IBD risk-associated gene with >5-fold down-regulation in the IFNG-secreting subset. RNASET2 disease risk variants were associated with decreased expression in peripheral and mucosal tissues and DNA hypermethylation in CD patients requiring surgical intervention. RNASET2 disease risk variants were associated in CD patients with more complicated disease or resistance to therapy, defined in part by failed response to treatment, increased length of intestinal resection, shorter time to repeat surgery, and high Rutgeerts score (>2) in postoperative endoscopy. The RNASET2 variant rs2149092 was predicted to disrupt a consensus binding site for the transcription factor ETS within an enhancer region. Expression of RNASET2 correlated with expression of ETS. RNASET2 knockdown in T cells increased expression of IFNG and intercellular adhesion molecule 1 (ICAM1) and induced T-cell aggregation. A blocking antibody against (ILFA1), disrupting the lymphocyte function-associated antigen 1-intercellular adhesion molecule 1 interaction, reduced T-cell production of IFNG.
  
  
  
  We identified decreased expression of RNASET2 as a component of TL1A-mediated increase in production of IFNG and as a potential biomarker for patients with severe CD. Further study of the role of RNASET2 in regulating mucosal inflammation may lead to development of novel therapeutic targets.
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California.
authors.forename: Rivkah
authors.initials: R
authors.lastname: Gonsky
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California.
authors.forename: Phillip
authors.initials: P
authors.lastname: Fleshner
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California.
authors.forename: Richard L
authors.initials: RL
authors.lastname: Deem
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California.
authors.forename: Eva
authors.initials: E
authors.lastname: Biener-Ramanujan
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California.
authors.forename: Dalin
authors.initials: D
authors.lastname: Li
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California.
authors.forename: Alka A
authors.initials: AA
authors.lastname: Potdar
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California.
authors.forename: Janine
authors.initials: J
authors.lastname: Bilsborough
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California.
authors.forename: Shaohong
authors.initials: S
authors.lastname: Yang
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California.
authors.forename: Dermot P B
authors.initials: DPB
authors.lastname: McGovern
authors.affiliation: F. Widjaja Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California. Electronic address: Stephan.Targan@cshs.org.
authors.forename: Stephan R
authors.initials: SR
authors.lastname: Targan
mesh_terms: D000483:Alleles; D002449:Cell Aggregation; D002478:Cells, Cultured; D003424:Crohn Disease / Q000235:genetics* / Q000601:surgery; D019175:DNA Methylation; D015536:Down-Regulation; D005786:Gene Expression Regulation / Q000187:drug effects*; D020868:Gene Silencing; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics; D007371:Interferon-gamma / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000378:metabolism; D016169:Lymphocyte Function-Associated Antigen-1 / Q000276:immunology; D020641:Polymorphism, Single Nucleotide; D050783:Proto-Oncogene Proteins c-ets / Q000235:genetics; D012260:Ribonucleases / Q000235:genetics*; D012720:Severity of Illness Index; D013601:T-Lymphocytes / Q000187:drug effects* / Q000378:metabolism; D059467:Transcriptome; D053380:Tumor Necrosis Factor Ligand Superfamily Member 15 / Q000494:pharmacology*; D025521:Tumor Suppressor Proteins / Q000235:genetics*
chemical_list: C573420:ICAM1 protein, human; D016169:Lymphocyte Function-Associated Antigen-1; D050783:Proto-Oncogene Proteins c-ets; C117026:TNFSF15 protein, human; D053380:Tumor Necrosis Factor Ligand Superfamily Member 15; D025521:Tumor Suppressor Proteins; D018799:Intercellular Adhesion Molecule-1; D007371:Interferon-gamma; D012260:Ribonucleases; C425232:RNASET2 protein, human
keywords: Genetics; Prognosis; Risk Factor; SNP