Sequence‐specific DNA binding activity of rat <i>cdc</i>37‐related gene product

The BRCT (BRCA1 C-terminus) superfamily includes a large number of nuclear proteins closely involved in DNA repair, recombination, and cell-cycle control. The human cDNA clone NFBD1 (previously designated KIAA0170) encodes a novel protein (2089 amino acids in length; calculated molecular mass 226,440 D) with possible BRCT domains at its carboxy terminus (amino acid residues 1894-2089). This gene product has been described as one of the BRCT superfamily proteins. However, its biological significance has been unclarified. Expression of green fluorescent protein (GFP)-tagged full-length NFBD1 or a series of deletion mutants indicated that NFBD1 was localized to the nucleus in various mammalian cells, and a 197-amino acid segment near the amino terminus (amino acid residues 142-338) contained a nuclear targeting signal. In vitro DNA-binding experiments showed that the highly basic region of NFBD1 (amino acid residues 1841-1893) possessed DNA-binding activity. The region encoding amino acids 508-995 of NFBD1 fused inframe with GAL4 DNA-binding domain activated transcription in both yeast and mammalian cells, while the possible BRCT domains of NFBD1 failed to induce transcription in mammalian cells. Overexpression of antisense NFBD1 RNA in a p53-deficient human osteogenic sarcoma cell line (SAOS-2) resulted in remarkable suppression of SAOS-2 colony formation. These results suggest that NFBD1 is a nuclear transcriptional transactivator with possible BRCT domains and may contribute to cell growth control.

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Interaction of Rat Cdc37-Related Protein with Retinoblastoma Gene Product

Ozaki

Sato²

1996

DNA and Cell Biology

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By using in vitro binding assays and the yeast two-hybrid system, we have found that a full-length rat Cdc37-related protein (RCdc37) could associate specifically with the retinoblastoma susceptibility gene product (pRB). A series of GST-RCdc37 deletion mutants was constructed to define the amino acid sequence required for the interaction with pRB. A GST-RCdc37 (-20 approximately 229) possessed an activity to associate with pRB, whereas GST-RCdc37 (-20 approximately 165) lost its activity, indicating that the amino acid sequence between 166 and 229 of RCdc37 was essential for the association with pRB. Interestingly, there exists a highly conserved pRB-binding motif (LXCXE; X = any amino acid) that is essential for pRB binding of SV40 large T antigen, E1A, and E7 proteins. A similar experiment using a pRB deletion mutant revealed that the carboxy-terminal portion of pRB was required for binding to RCdc37.

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Sequence‐specific DNA binding activity of rat cdc37‐related gene product

Cited by 5 publications

References 15 publications

Genomic organization and the promoter region of the round-spotted pufferfish (Tetraodon fluviatilis) CDC37 gene

Genomic organization and the promoter region of the round-spotted pufferfish (Tetraodon fluviatilis) CDC37 gene

NFBD1/KIAA0170 Is a Novel Nuclear Transcriptional Transactivator with BRCT Domain

Interaction of Rat Cdc37-Related Protein with Retinoblastoma Gene Product

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