Dual roles for Pax-6: a transcriptional repressor of lens fiber cell-specific beta-crystallin genes

Mol Cell Biol. 1998 Sep;18(9):5579-86. doi: 10.1128/MCB.18.9.5579.


It has been demonstrated previously that Pax-6, a paired domain (PD)/homeodomain (HD) transcription factor critical for eye development, contributes to the activation of the alphaB-, alphaA-, delta1-, and zeta-crystallin genes in the lens. Here we have examined the possibility that the inverse relationship between the expression of Pax-6 and beta-crystallin genes within the developing chicken lens reflects a negative regulatory role of Pax-6. Cotransfection of a plasmid containing the betaB1-crystallin promoter fused to the chloramphenicol acetyltransferase reporter gene and a plasmid containing the full-length mouse Pax-6 coding sequences into primary embryonic chicken lens epithelial cells or fibroblasts repressed the activity of this promoter by as much as 90%. Pax-6 constructs lacking the C-terminal activation domain repressed betaB1-crystallin promoter activity as effectively as the full-length protein, but the PD alone or Pax-6 (5a), a splice variant with an altered PD affecting its DNA binding specificity, did not. DNase footprinting analysis revealed that truncated Pax-6 (PD+HD) binds to three regions (-183 to -152, -120 to -48, and -30 to +1) of the betaB1-crystallin promoter. Earlier experiments showed that the betaB1-crystallin promoter sequence from -120 to -48 contains a cis element (PL2 at -90 to -76) that stimulates the activity of a heterologous promoter in lens cells but not in fibroblasts. In the present study, we show by electrophoretic mobility shift assay and cotransfection that Pax-6 binds to PL2 and represses its ability to activate promoter activity; moreover, mutation of PL2 eliminated binding by Pax-6. Taken together, our data indicate that Pax-6 (via its PD and HD) represses the betaB1-crystallin promoter by direct interaction with the PL2 element. We thus suggest that the relatively high concentration of Pax-6 contributes to the absence of betaB1-crystallin gene expression in lens epithelial cells and that diminishing amounts of Pax-6 in lens fiber cells during development allow activation of this gene.

MeSH terms

  • Animals
  • Base Sequence
  • Binding Sites
  • Cell Line
  • Chick Embryo
  • Chickens
  • Chloramphenicol O-Acetyltransferase / biosynthesis
  • Cloning, Molecular
  • Crystallins / biosynthesis*
  • Crystallins / genetics
  • DNA-Binding Proteins / biosynthesis
  • DNA-Binding Proteins / metabolism*
  • Epithelial Cells / cytology
  • Epithelial Cells / metabolism
  • Eye Proteins / metabolism
  • Gene Expression Regulation, Developmental
  • Genes, Reporter
  • Homeodomain Proteins*
  • Lens, Crystalline / cytology*
  • Lens, Crystalline / growth & development
  • Lens, Crystalline / metabolism*
  • Mice
  • Molecular Sequence Data
  • PAX6 Transcription Factor
  • Paired Box Transcription Factors
  • Promoter Regions, Genetic
  • Recombinant Fusion Proteins / biosynthesis
  • Recombinant Fusion Proteins / metabolism
  • Repressor Proteins / metabolism*
  • Transcription Factors / metabolism
  • Transfection


  • Crystallins
  • DNA-Binding Proteins
  • Eye Proteins
  • Homeodomain Proteins
  • PAX6 Transcription Factor
  • Paired Box Transcription Factors
  • Pax6 protein, mouse
  • Recombinant Fusion Proteins
  • Repressor Proteins
  • Transcription Factors
  • Chloramphenicol O-Acetyltransferase