A TEF-1 binding motif that interacts with a placental protein is important for the transcriptional activity of the hCS-B enhancer

DNA Cell Biol. 1994 Oct;13(10):1037-45. doi: 10.1089/dna.1994.13.1037.

Abstract

The transcriptional activity of the human placental lactogen genes (choriosomatomammotropic hormone, hCS) is controlled by tissue-specific enhancers located 4 kb downstream from their respective origins of transcription. The hCS-B enhancer is the strongest; its activity is mediated by synergism between two protein-binding sites (DF-3 and DF-4). The DF-4 site possesses a potential binding sequence for TEF-1, a known transcription factor. In this paper, we show by electrophoretic mobility-shift assays and antibody supershift experiments that TEF-1 does not bind to site DF-4. Mutations in the TEF-1-like binding motif of site DF-4 prevent formation of the DNA-protein complex, called complex f, in the presence of placental JEG-3 cell extracts. When HeLa cell extracts are used, another complex (complex c) is also affected. In transient expression experiments, TKCAT constructs linked to this mutated DF-4 site exhibit greatly reduced transcriptional activity when introduced into JEG-3 cells. Some cell lines contain both protein c and protein f (the proteins forming complexes c and f); when transfected, these lines display reduced DF-4-driven activity, suggesting that the two proteins could compete for the same DF-4 sequence. We conclude that protein f is important for the placenta-specific activity of the hCS-B enhancer. By UV cross-linking, we show that protein f is actually three polypeptides ranging in size from about 12 to 21 kD.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Cell Line
  • DNA-Binding Proteins / metabolism*
  • Enhancer Elements, Genetic*
  • HeLa Cells
  • Humans
  • Molecular Sequence Data
  • Molecular Weight
  • Nuclear Proteins / metabolism*
  • Oligodeoxyribonucleotides
  • Peptides / metabolism
  • Placental Lactogen / genetics*
  • Placental Lactogen / metabolism
  • Point Mutation
  • Protein Binding
  • Transcription Factors / metabolism*

Substances

  • DNA-Binding Proteins
  • Nuclear Proteins
  • Oligodeoxyribonucleotides
  • Peptides
  • TEAD1 protein, human
  • Transcription Factors
  • Placental Lactogen