title: Upregulated expression of hITF in Crohn's disease and screening of hITF Interactant by a yeast two-hybrid system.
abstract: OBJECTIVE
  To study the expression of human intestinal trefoil factor (hITF) mRNA in Crohn's disease and to screen the cellular proteins that can interact with the hITF protein by a yeast two-hybrid system in order to explore the mechanism of hITF in protecting intestinal mucosa from injury.
  
  
  METHODS
  Seventy-eight patients underwent double-balloon enteroscopy (DBE). Expression of hITF mRNA was detected by quantitative real-time polymerase chain reaction analysis (qRT-PCR). The hITF gene was amplified by PCR and cloned into vector pDEST32. The yeast cells cotransformed with pDEST32-hITF and the human jejunal cDNA library were plated in a selective SC-Leu-Trp-His-Ura medium. The subsequent screen was performed with χ-gal detection, and true-positive clones were sequenced and analyzed with bioinformatics. Co-immunoprecipitation (Co-IP) was performed to confirm the binding of putative proteins to the hITF protein.
  
  
  RESULTS
  Thirty-nine patients were diagnosed with Crohn's disease. We found that the expression of hITF mRNA is significantly increased in Crohn's disease compared to normal controls. A total of ten colonies were selected and sequenced. Among these, six colonies were Homo sapiens zinc finger protein 193 (ZNF193), three colonies were Homo sapiens Aldo-keto reductase family 1C 1 (AKR1C1), and one colony was of an unknown gene. A reverse two-hybrid experiment and Co-IP indicated that ZNF193 and AKR1C1 might interact with hITF.
  
  
  CONCLUSIONS
  The expression of hITF mRNA is increased in Crohn's disease. ZNF193 and AKR1C1 are proteins that can interact with the hITF protein by a yeast two-hybrid system and Co-IP, hITF may contribute to the mucosal repair through this interaction.
authors.affiliation: Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan, Hubei Province, China.
authors.forename: Huaxiu
authors.initials: H
authors.lastname: Shi
authors.forename: Jianlin
authors.initials: J
authors.lastname: Ren
authors.forename: Hongzhi
authors.initials: H
authors.lastname: Xu
authors.forename: Jinshui
authors.initials: J
authors.lastname: Pan
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Hao
authors.forename: Lydia Leyland
authors.initials: LL
authors.lastname: Barlow
authors.forename: Weiguo
authors.initials: W
authors.lastname: Dong
mesh_terms: D002460:Cell Line; D003424:Crohn Disease / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D015870:Gene Expression / Q000502:physiology; D015723:Gene Library; D005820:Genetic Testing / Q000379:methods*; D006706:Homeostasis / Q000502:physiology; D006801:Humans; D007413:Intestinal Mucosa / Q000473:pathology / Q000502:physiology; D007583:Jejunum / Q000473:pathology / Q000502:physiology; D007668:Kidney / Q000166:cytology; D010455:Peptides / Q000235:genetics*; D010957:Plasmids / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000071164:Trefoil Factor-2; D020798:Two-Hybrid System Techniques; D015854:Up-Regulation / Q000502:physiology
chemical_list: D010455:Peptides; D012333:RNA, Messenger; D000071164:Trefoil Factor-2