A naturally occurring allele of BRCA1 coding for a temperature-sensitive mutant protein

Cancer Biol Ther. Sep-Oct 2002;1(5):497-501. doi: 10.4161/cbt.1.5.164.

Abstract

Recent evidence suggests that the breast and ovarian cancer susceptibility gene product BRCA1 is involved in at least two fundamental cellular processes: transcriptional regulation and DNA repair. However, the mechanism of action of BRCA1 in either of these processes is still unknown. Here, we report the characterization of a disease-predisposing allele of BRCA1, identified in a family with several cases of ovarian cancer, coding for a protein that displays temperature-sensitive activity in transcriptional activation. The mutant protein differs from the wild type protein at a single amino acid, R1699W that occurs in a region at the N-terminal BRCT domain that is highly conserved among BRCA1 homologs. When the C-terminus of the mutant protein (aa 1560-1863) was fused to a heterologous GAL4 DNA-binding domain and expressed in yeast or mammalian cells, it was able to activate transcription of a reporter gene to levels observed for wild type BRCA1 at the permissive temperature (30 degrees C) but exhibited significantly less transcription activity at the restrictive temperature (37 degrees C or 39 degrees C). Our results indicate that the transcriptional activity of the R1699W mutant can be modulated as a function of temperature and provide a novel experimental approach which can be utilized to dissect the molecular mechanism(s) of BRCA1 in processes related to transcription.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Alleles
  • Amino Acid Sequence
  • Amino Acid Substitution
  • BRCA1 Protein / chemistry
  • BRCA1 Protein / genetics*
  • BRCA1 Protein / physiology*
  • Cell Line
  • Conserved Sequence
  • DNA-Binding Proteins
  • Gene Expression Regulation
  • Genes, BRCA1
  • Genes, Reporter
  • Humans
  • Models, Molecular
  • Molecular Sequence Data
  • Point Mutation
  • Protein Conformation
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae / growth & development
  • Saccharomyces cerevisiae Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Temperature
  • Transcription Factors / metabolism
  • Transcriptional Activation

Substances

  • BRCA1 Protein
  • DNA-Binding Proteins
  • GAL4 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • Transcription Factors