Connecting defective replication initiation to inherited human diseases: Biochemical and genetic analysis of the RecQL4 helicase. (360G-Wellcome-081437_Z_06_Z)

We have found that the RecQL4 protein, mutated in human diseases characterizedby chromosome fragility, developmental malformations and premature aging, plays an unexpected role in the initiation of DNA replication (Sangrithi et al. (2005) Cell 121, 887). RecQL4 contains an N-terminal domain homologous to yeast Sld2, and a RecQ-domain, but lacks other features conserved in RecQ helicases. Our first key goal is to determine how RecQL4 promotes assembly of the replication machinery at origins. We find that the Sld2-like domain is phosphorylated by cyclin/CDK activity, and interacts with Cut5/TopBP1, and will investigate the functional effects of phosphorylation on RecQL4 activity and function. In an ongoing collaboration with Shamoo's lab in Houston, a structural analysis of RecQL4 domains is underway, with encouraging preliminary progress. We will perform structure-based analyses of RecQL4 function using this platform. Our second key goal is to develop a vertebrate genetic system to study the functional effects of disease-associated RecQL4 mutations on replication-associated processes like chromatid cohesion that maintain chromosomal integrity. The system will also facilitate genetic analysis of thebiological pathways dependent on RecQL4 activity. Our work should provide insight into the fundamental mechanisms controlling replication initiation, and into how their dysregulation triggers human genetic diseases.