BRCA1 proteins are transported to the nucleus in the absence of serum and splice variants BRCA1a, BRCA1b are tyrosine phosphoproteins that associate with E2F, cyclins and cyclin dependent kinases

Oncogene. 1997 Jul 10;15(2):143-57. doi: 10.1038/sj.onc.1201252.

Abstract

BRCA1, a familial breast and ovarian cancer susceptibility gene encodes nuclear phosphoproteins that function as tumor suppressors in human breast cancer cells. Previously, we have shown that overexpression of a BRCA1 splice variant BRCA1a accelerates apoptosis in human breast cancer cells. In an attempt to determine whether the subcellular localization of BRCA1 is cell cycle regulated, we have studied the subcellular distribution of BRCA1 in asynchronous and growth arrested normal, breast and ovarian cancer cells using different BRCA1 antibodies by immunofluorescence and immunohistochemical staining. Upon serum starvation of NIH3T3, some breast and ovarian cancer cells, most of the BRCA1 protein redistributed to the nucleus revealing a new type of regulation that may modulate the activity of BRCA1 gene. We have also characterized two new variant BRCA1 proteins (BRCA1a/p110 and BRCA1b/ p100) which are phosphoproteins containing phosphotyrosine. Immunofluorescence and Western blotting analysis indicate cytoplasmic and nuclear localization of BRCA1a and BRCA1b proteins. To elucidate the biological function of BRCA1, we created a bacterial fusion protein of glutathione-transferase (GST) and BRCA1 zinc finger domain and detected two cellular proteins with molecular weights of approximately 32 and 65 kD, one of which contains phosphotyrosine designated p32 and p65 BRCA1 interacting proteins (BIP) that specifically interact with BRCA1. Western blot analysis of BIP with cyclins/CDKs and E2F antisera indicated association with cdc2, cdk2, cdk4, cyclin B, cyclin D, cyclin A and E2F-4 but not with cdk3, cdk5, cdk6, E2F-1, E2F-2, E2F-3, E2F-5 and cyclin E. Furthermore, we have also demonstrated a direct interaction of in vitro translated BRCA1a and BRCA1b proteins with recombinant cyclin A, cyclin B1, cyclin D1, cdc2, cdk2 and E2F fusion proteins in vitro. Taken together these results seem to suggest that BRCA1 could be an important negative regulator of cell cycle that functions through interaction with E2F transcriptional factors and phosphorylation by cyclins/cdk complexes with the zinc ring finger functioning as a major protein-protein interaction domain. If the interactions we observe in vitro is also seen in vivo then it may be possible that lack or impaired binding of the disrupted BRCA1 proteins to E2F, cyclins/CDKs in patients with mutations in the zinc finger domain could deprive the cell of an important mechanism for braking cell proliferation leading to the development of breast and ovarian cancers.

Publication types

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

MeSH terms

  • 3T3 Cells
  • Animals
  • BRCA1 Protein / analysis
  • BRCA1 Protein / metabolism*
  • Biological Transport
  • Blood Physiological Phenomena
  • Carrier Proteins*
  • Cell Cycle Proteins*
  • Cell Nucleus / metabolism*
  • Cyclin-Dependent Kinases / metabolism*
  • Cyclins / metabolism*
  • DNA-Binding Proteins*
  • E2F Transcription Factors
  • E2F1 Transcription Factor
  • E2F2 Transcription Factor
  • E2F3 Transcription Factor
  • E2F4 Transcription Factor
  • E2F5 Transcription Factor
  • Humans
  • Mice
  • Retinoblastoma-Binding Protein 1
  • Transcription Factor DP1
  • Transcription Factors / metabolism*
  • Tyrosine / metabolism

Substances

  • Arid4a protein, mouse
  • BRCA1 Protein
  • Carrier Proteins
  • Cell Cycle Proteins
  • Cyclins
  • DNA-Binding Proteins
  • E2F Transcription Factors
  • E2F1 Transcription Factor
  • E2F1 protein, human
  • E2F2 Transcription Factor
  • E2F2 protein, human
  • E2F3 Transcription Factor
  • E2F4 Transcription Factor
  • E2F4 protein, human
  • E2F5 Transcription Factor
  • E2f1 protein, mouse
  • E2f4 protein, mouse
  • Retinoblastoma-Binding Protein 1
  • Transcription Factor DP1
  • Transcription Factors
  • Tyrosine
  • Cyclin-Dependent Kinases