Gli regulates MUC5AC transcription in human gastrointestinal cells

PLoS One. 2014 Aug 28;9(8):e106106. doi: 10.1371/journal.pone.0106106. eCollection 2014.

Abstract

MUC5AC is a well-known gastric differentiation marker, which has been frequently used for the classification of stomach cancer. Immunohistochemistry revealed that expression of MUC5AC decreases accompanied with increased malignant property of gastric mucosa, which further suggests the importance of MUC5AC gene regulation. Alignment of the 5'-flanking regions of MUC5AC gene of 13 mammal species denoted high homology within 200 bp upstream of the coding region. Luciferase activities of the deletion constructs containing upstream 451 bp or shorter fragments demonstrated that 15 bp region between -111 and -125 bp plays a critical role on MUC5AC promoter activity in gastrointestinal cells. We found a putative Gli-binding site in this 15 bp sequence, and named this region a highly conserved region containing a Gli-binding site (HCR-Gli). Overexpression of Gli homologs (Gli1, Gli2, and Gli3) clearly enhanced MUC5AC promoter activity. Exogenous modulation of Gli1 and Gli2 also affected the endogenous MUC5AC gene expression in gastrointestinal cells. Chromatin immunoprecipitation assays demonstrated that Gli1 directly binds to HCR-Gli: Gli regulates MUC5AC transcription via direct protein-DNA interaction. Conversely, in the 30 human cancer cell lines and various normal tissues, expression patterns of MUC5AC and Gli did not coincide wholly: MUC5AC showed cell line-specific or tissue-specific expression whereas Gli mostly revealed ubiquitous expression. Luciferase promoter assays suggested that the far distal MUC5AC promoter region containing upstream 4010 bp seems to have several enhancer elements for gene transcription. In addition, treatments with DNA demethylation reagent and/or histone deacetylase inhibitor induced MUC5AC expression in several cell lines that were deficient in MUC5AC expression. These results indicated that Gli is necessary but not sufficient for MUC5AC expression: namely, the multiple regulatory mechanisms should work in the distal promoter region of MUC5AC gene.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Binding Sites
  • Cell Line, Tumor
  • Conserved Sequence
  • Gastrointestinal Neoplasms / genetics
  • Gastrointestinal Neoplasms / metabolism
  • Gastrointestinal Neoplasms / pathology*
  • Gastrointestinal Tract / metabolism*
  • Gastrointestinal Tract / pathology
  • Gene Expression Regulation, Neoplastic
  • Humans
  • Mucin 5AC / chemistry
  • Mucin 5AC / genetics*
  • Mucin 5AC / metabolism*
  • Organ Specificity
  • Promoter Regions, Genetic*
  • Sequence Alignment
  • Transcription Factors / genetics
  • Transcription Factors / metabolism*
  • Zinc Finger Protein GLI1

Substances

  • GLI1 protein, human
  • MUC5AC protein, human
  • Mucin 5AC
  • Transcription Factors
  • Zinc Finger Protein GLI1

Grants and funding

This work was supported in part by a grant from Takeda Science Foundation, in part by a research grant from the Tokyo Society of Medical Sciences, in part by Grant-in-Aid for Scientific Research (C) from the Japan Society for the Promotion of Science, in part by Sato Memorial Foundation for Cancer Research, in part by a research grant from Nakayama Cancer Research Institute, in part by a research grant from Foundation for promotion of Cancer Research, and also in part by Grant-in-Aid for Young Scientists (B) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT). All the funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The URLs and grant numbers are as follows: A grant from Takeda Science Foundation, http://www.takeda-sci.or.jp/assist/, Grant number: N/A (Grant in 2012); a research grant from the Tokyo Society of Medical Sciences, http://square.umin.ac.jp/igakukai/02toppage/toppage.html, Grant number: N/A (Grant in 2012); Grant-in-Aid for Scientific Research (C) from the Japan Society for the Promotion of Science, http://www.jsps.go.jp/english/e-grants/index.html, Grant number: 25460381 (Grant in 2012–2014); Sato Memorial Foundation for Cancer Research, http://www.jca.gr.jp/researcher/reward/2010/sato.html, Grant number: N/A (Grant in 2011); a research grant from Nakayama Cancer Research Institute, http://ncri.or.jp/project/project.html, Grant number: N/A (Grant in 2010); a research grant from Foundation for promotion of Cancer Research, http://www.fpcr.or.jp/about/index.html, Grant number: N/A (Grant in 2010); Grant-in-Aid for Young Scientists (B) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), http://www.jsps.go.jp/english/e-grants/index.html, Grant number: 23790355 (Grant in 2010–2011).