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BRCA1 Protein Structure
Image courtesy of Protein Data Bank http://www.rcsb.org/pdb/
In 1994, the BReast-CAncer susceptibility gene, BRCA1, was identified by positional cloning; subsequently, this tumor supressor gene has been the subject of intensive research effort (Miki et al., 1994). BRCA1 is composed of 22 coding exons distributed over 100 kb of genomic DNA, this gene encodes for an 1863-residue protein (NCBI amino acid sequence) involved in gene regulation and repair processes following DNA damage (Tavtigan et al., 1998). More than 200 different germline mutations associated with cancer susceptibility have been identified. Many disease-predisposing alleles of BRCA1 have loss-of function mutations, the majority of which result in premature truncation of the protein (Tavtigan et al.,1998). Because only 45% of familial breast cancers showed evidence of linkage to BRCA1, the search for a second breast cancer susceptibility gene continued, and in 1995, the BRCA2 gene was identified at chromosome 13q12.36, (Fig.1). Mutations in BRCA1 and BRCA2 are also associated with an increased susceptibility to ovarian, pancreatic, and prostate cancers (Venkitaraman, 2002).
Figure 1. Features of the Human BRCA ProteinsBRCA1 contains an N-terminal RING domain, nuclear localization signals (NLSs), and two C-terminal BRCT domains of ~110 residues (also found in several proteins with functions in DNA repair or cell cycle control). Interacting proteins discussed in the text are shown below approximate regions of binding. BRCA2 contains eight repeats of the ~40 residue BRC motifs. Six of the eight motifs in human BRCA2 can bind directly to RAD51 when expressed in vitro (Image Permit Pending Venkitaranam, 2002 )
Functions of the BRCA1 protein
BRCA1 and BRCA2 exhibit similar patterns of expression and sub-cellular localization. They are both expressed in many tissues in a cell-cycle-dependent manner, their levels are highest during S phase, which is suggestive of functions during DNA replication. Both are localized to the nucleus in somatic cells, where they co-exist in characteristic sub-nuclear foci that redistribute following DNA damage (Venkitaraman, 2001). Current analyses of the protein encoded by BRCA1 seem to be consistent with 2 functions: a role in DNA recombination and/or repair and a role in transcriptional regulation.
Role in DNA repair
Initial evidence suggesting a role of BRCA1 in the repair of damaged DNA was derived from the observation that BRCA1 is hyperphosphorylated in response to DNA damage and relocated to sites of replication forks marked by proliferating cell nuclear antigen (PCNA) . It is likely, therefore, that BRCA1 is phosphorylated at multiple residues by different kinases after DNA damage (Table 1.). However, how each type of phosphorylation affects the functions of BRCA1 remains o bscure (Scully et. al., 1997) .
Phosphorylating Enzyme Phosphorylation target Ataxia-telangiectasia mutated (ATM Ser1387 ATM-related kinase (ATR). Ser1457 G2/M control kinase, CHK2, Ser988
Table 1. The table above shows in colum 1 three different kinases and their amino acid targets in the BERCA1 protein sequence. NOTE: The information presented in the table above comes from (Scully et. al., 1997).
Studies demonstrated that BRCA1 and BRCA2 were involved in protein complexes that activate the repair of DNA doublestranded breaks (DSBs) and initiate homologous recombination (HR), maintaining tumor suppression roles (Venkitaraman, 2001).
- BRCA1 and BRCA2 co-localize with Rad51 to form a protein complex. Rad51 proteins are required for recombination during mitosis and meiosis, as well as for HR repair of DSBs. Co-localization strongly suggests that BRCAs have a role in both the detection and the repair of DSBs (Tavtigan et. al., 1998 ). However, accumulating evidence suggests that BRCA1 might not directly regulate Rad51, since interactions between BRCA1 and Rad51 are indirect (Venkitaraman, 2002).
- Other studies have shown that BRCA1 co-localizes and coimmunoprecipitates with Rad50, an apparently functions as a regulator of the Rad50-Mre11-NBS1 complex. Mre11 encodes nuclease activity, BRCA1 binds DNA directly and inhibits Mre11, regulating the length and the persistence of ss-DNA generation at sites of DNA damage. Since ss-DNA is a substrate for DNA repair by HR, BRCA1 might play an essential role in HR-mediated repair of DSBs by inactivating Mre11 (Venkitaraman, 2001).
Role in transcriptional response to DNA damage
Based in several studies, it is believed that BRCA1 is involved in the transcriptional regulation of several genes activated in response to DNA damage (Fig. 2) . Such genes include those encoding the p21 CIP1 cyclin-dependent kinase inhibitor, and the GADD45 tumour suppressor (Venkitaraman, 2001 and 2002 ). Evidence of this is that the C-terminus of BRCA1 (amino acids 1528– 1863) binds and activates the basal transcription machinery. Studies demonstrated that the C-terminus of human BRCA1 complexes with RNA polymerase II through RNA helicase A. This interaction appears to involve several proteins associated with the core polymerase complex ( Scully et. al., 1997) .
- BRCA1 was also shown to regulate transcription in a purified in vitro system. It is now clear that BRCA1 is present as a heterodimer with BARD1 (BRCA1-associated RING domain protein 1), which has been implicated in the control of RNA processing following DNA damage (Venkitaraman, 2001).
- The internal portion of BRCA1 binds to a large number of transcriptional factors, which may mediate signals to RNA polymerase II. Indeed, transcriptional activation by BRCA1 is supported by its ability to interact directly or indirectly with several transcriptional factors. Finding target genes regulated by BRCA1 would shed considerable light on the transcriptional role of BRCA1 (Scully et. al., 1997).
- BRCA1 also binds to ZBRK1, a transcriptional factor binding specifically to the DNA sequence GGGXXXCAGXXXTTT. 50) This binding motif is present in the promoter region of many transcriptional targets for BRCA1, such as p21, GADD45, and EGR1. Therefore, co-expression of BRCA1 and ZBRK1 was found to repress GADD45 promoter (Venkitaraman, 2002).
Fig 2. Protein partners of BRCA1 in DNA damage responses. There is accumulating evidence that BRCA1 performs multiple functions in the cellular response to DNA damage through its interactions with different protein partners. The list of BRCA1-interacting proteins indicated here is not exhaustive but illustrates points made in the text (Image Permit Pending Venkitaraman,2001 ).
- Miki Y, Swensen J, Shattuck-Eidens D, Futreal PA, Harshman K, Tavtigian S, Liu Q, Cochran C, Bennett LM, Ding W et al. 1994. A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1. Science 266: 66–71.
- Scully R, Chen J, Ochs RL, Keegan K, Hoekstra M, Feunteun J, and Livingston DM. 1997. Dynamic changes of BRCA1 subnuclear location and phosphorylation state are initiated by DNA damage. Cell 90: 425–35.
- Tavtigan SV, Thomas A, Thomas FS, and Sckolnick MH. 1998. The BRCA1 gene and its protein products characterization, therapeutic and diagnostic implications. Gynecologic Oncology Associates. < http://www.gynoncology.com/common_files/BRCA.htm > Accessed 2005 Feb 10.
- Venkitaraman AR. 2001 Functions of BRCA1 and BRCA2 in the biological response to DNA damage. Journal of Cell Science 114: 3591-3598 <http://jcs.biologists.org/cgi/content/full/114/20/3591>.
- Venikataram AR. 2002. Cancer susceptibility and the functions of BRCA1 and BRCA2. Cell 108: 171-182 <doi:10.1016/S0092-8674(02)00615-3>
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