Regulation of human myocilin/TIGR gene transcription in trabecular meshwork cells and astrocytes: role of upstream stimulatory factor

Genes Cells. 2000 Aug;5(8):661-76. doi: 10.1046/j.1365-2443.2000.00355.x.


Background: Mutations in the myocilin (MYOC)/TIGR gene are responsible for autosomal-dominant juvenile primary open-angle glaucoma (POAG). In patients with non-autosomal-dominant POAG, such mutations are rare, but the expression of MYOC/TIGR in the trabecular meshwork (TM) of the eye is considerably higher than in normals. We performed transfection, DNAse I footprinting, mutagenesis and electrophoretic mobility shift assays (EMSA) to identify elements responsible for the basal transcription of MYOC/TIGR in TM cells and astrocytes.

Results: DNAse I footprinting experiments of the human MYOC/TIGR promoter showed a major protected area between nt -106 to -77, which was not conserved in the homologous region of the mouse myoc/tigr promoter. In addition, the TATA-box was protected, as well as at least three downstream sites, including an AP-1-like sequence. Deletion of the -106 to -77 region caused a substantial loss of functional promotor activity in all cell types. Site-directed mutagenesis and EMSA experiments revealed the presence of two regulatory elements in the -106 to -77 region. Each of these cis-elements is essential for minimal promoter activity. The 5'-half of the region contains a sequence with similarities to NF-kappaB-related sites, however, binding of NF-kappaB could not be confirmed by EMSA. The 3'-half contains a canonical E-box sequence. EMSA experiments showed that the upstream regulatory factor (USF) was binding to the E-box sequence and that the binding can be supershifted by specific antibodies.

Conclusions: Several DNA-protein binding elements contribute to a transcription of MYOC/TIGR, and USF is critically required for its basal transcription in trabecular meshwork cells and astrocytes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Astrocytes / metabolism*
  • Base Sequence
  • Cytoskeletal Proteins
  • DNA Footprinting
  • DNA-Binding Proteins*
  • Deoxyribonuclease I / metabolism
  • Eye Proteins / biosynthesis
  • Eye Proteins / genetics*
  • Gene Expression Regulation
  • Glaucoma, Open-Angle / genetics*
  • Glycoproteins / biosynthesis
  • Glycoproteins / genetics*
  • Humans
  • Mice
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • NF-kappa B / metabolism
  • Promoter Regions, Genetic*
  • Protein Binding
  • Sequence Homology, Nucleic Acid
  • Trabecular Meshwork / metabolism*
  • Transcription Factor AP-1 / metabolism
  • Transcription Factors / metabolism*
  • Transcription, Genetic
  • Upstream Stimulatory Factors


  • Cytoskeletal Proteins
  • DNA-Binding Proteins
  • Eye Proteins
  • Glycoproteins
  • NF-kappa B
  • Transcription Factor AP-1
  • Transcription Factors
  • USF1 protein, human
  • Upstream Stimulatory Factors
  • Usf1 protein, mouse
  • trabecular meshwork-induced glucocorticoid response protein
  • Deoxyribonuclease I