The Bloom's and Werner's syndrome proteins are DNA structure-specific helicases

Abstract

BLM and WRN, the products of the Bloom's and Werner's syndrome genes, are members of the RecQ family of DNA helicases. Although both have been shown previously to unwind simple, partial duplex DNA substrates with 3'-->5' polarity, little is known about the structural features of DNA that determine the substrate specificities of these enzymes. We have compared the substrate specificities of the BLM and WRN proteins using a variety of partial duplex DNA molecules, which are based upon a common core nucleotide sequence. We show that neither BLM nor WRN is capable of unwinding duplex DNA from a blunt-ended terminus or from an internal nick. However, both enzymes efficiently unwind the same blunt-ended duplex containing a centrally located 12 nt single-stranded 'bubble', as well as a synthetic X-structure (a model for the Holliday junction recombination intermediate) in which each 'arm' of the 4-way junction is blunt-ended. Surprisingly, a 3'-tailed duplex, a standard substrate for 3'-->5' helicases, is unwound much less efficiently by BLM and WRN than are the bubble and X-structure substrates. These data show conclusively that a single-stranded 3'-tail is not a structural requirement for unwinding of standard B-form DNA by these helicases. BLM and WRN also both unwind a variety of different forms of G-quadruplex DNA, a structure that can form at guanine-rich sequences present at several genomic loci. Our data indicate that BLM and WRN are atypical helicases that are highly DNA structure specific and have similar substrate specificities. We interpret these data in the light of the genomic instability and hyper-recombination characteristics of cells from individuals with Bloom's or Werner's syndrome.

Pyrimidine pool imbalance induced by BLM helicase deficiency contributes to genetic instability in Bloom syndrome

“…We demonstrate here that the helicase activity of BLM regulates CDA promoter function either directly or indirectly. The preferred substrates for the recombinant BLM protein are G-quadruplex structures 18,19 . However, we identified no potential intramolecular G-quadruplex-forming sequences or particular sequences likely to form secondary structures in the CDA promoter (using Quad Parser, website http://www.quadruplex.com), suggesting that CDA expression is not repressed by secondary structures formed in the absence of BLM.…”

The Bloom syndrome protein interacts and cooperates with p53 in regulation of transcription and cell growth control

“…Keywords: Bloom syndrome; BLM protein; p53; interaction; two-hydrid system; transactivation Bloom syndrome results from mutations inactivating BLM gene that encodes a RecQ family DNA helicase (Mohaghegh et al, 2001). Patients with Bloom syndrome are hypersensitive to UV radiation, su ering from sun-sensitive facial erythema, and show predisposition to tumor development (German, 1993).…”

Inhibition of Werner Syndrome Helicase Activity by Benzo[c]phenanthrene Diol Epoxide dA Adducts in DNA Is Both Strand-and Stereoisomer-dependent

“…It was subsequently shown that WRN facilitates polymerase ␦ synthesis through hairpin and tetraplex structures that impede the polymerase, suggesting that WRN may remove secondary structure at a stalled replication fork to allow polymerase ␦ synthesis to proceed (22). The results to be reported here suggest that WRN may not function properly to ensure replication fork progression if the fork is blocked by a covalent DNA adduct, even though WRN may disrupt other DNA structures such as tetraplexes (13,23) or triplexes (24) that might impede a replication fork.…”

“…However, unidirectional translocation of WRN on a DNA lattice remains to be demonstrated. Although the DNA substrate specificity of WRN (11) and other RecQ helicases (12,13) has been studied, the effects of DNA damage on helicases of this family have not been characterized prior to this study.…”