Protocols

2742:

Western Blotting

Background

BLM, a member of the RecQ family of DNA helicases, is part of the BRCA1-associated genome surveillance complex (BASC) that responds to DNA damage, stalled replication forks and S phase arrest (1-4). Phosphorylation of BLM helicase at Thr99 and Thr122 occurs in response to genotoxic stress (4), and phosphorylation of Ser144 appears to be important in regulating chromosome stability during mitosis (5). Typical BLM protein resides in the nucleus and forms part of a dynamic protein complex that acts in response to DNA damage during specific periods of the cell cycle (6). Although RecQ helicases are rarely considered as essential enzymes, they function at the interface between DNA recombination and repair and are required for global genome stability maintenance. Mutations in BLM helicase are responsible for development of Bloom Syndrome, a recessive genetic disorder clinically characterized by short stature, immunodeficiency and elevated risk of malignancy (7). Similar alterations to genes encoding the related RecQ helicases RecQ4 and WRN also result in recessive genetic disorders associated with genomic instability (8,9). Cells from Bloom Syndrome patients exhibit genomic instability and increased frequency of sister chromatid exchange (10).

1. Wang, Y. et al. (2000) Genes Dev. 14, 927-939.
2. Langland, G. et al. (2002) Cancer Res. 62, 2766-2770.
3. Sengupta, S. et al. (2003) EMBO J. 22, 1210-1222.
4. Davies, S.L. et al. (2004) Mol. Cell. Biol. 24, 1279-1291.
5. Leng, M. et al. (2006) Proc. Natl. Acad. Sci. USA 103, 11485-11490.
6. Bischof, O. et al. (2001) J. Cell Biol. 153, 367-380.
7. van Brabant, A.J. et al. (2000) Annu. Rev. Genomics Hum. Genet. 1, 409-459.
8. Kitao, S. et al. (1999) Nat. Genet. 22, 82-84.
9. Yu, C.E. et al. (1996) Science 272, 258-262.
10. Chaganti, R.S. et al. (1974) Proc. Natl. Acad. Sci. USA 71, 4508-4512.

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