RESUMEN
Fanconi anemia (FA) is a heritable human cancer-susceptibility disorder, delineating a genetically heterogenous pathway for the repair of replication-blocking lesions such as interstrand DNA cross-links. Here we demonstrate that one component of this pathway, FANCJ, is a structure-specific DNA helicase that dissociates guanine quadruplex DNA (G4 DNA) in vitro. Moreover, in contrast with previously identified G4 DNA helicases, such as the Bloom's helicase (BLM), FANCJ unwinds G4 substrates with 5'-3' polarity. In the FA-J human patient cell line EUFA0030 the loss of FANCJ G4 unwinding function correlates with the accumulation of large genomic deletions in the vicinity of sequences, which match the G4 DNA signature. Together these findings support a role for FANCJ in the maintenance of potentially unstable genomic G/C tracts during replication.
Asunto(s)
Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , ADN Helicasas/metabolismo , Proteínas del Grupo de Complementación de la Anemia de Fanconi/metabolismo , G-Cuádruplex , RecQ Helicasas/metabolismo , Unión Competitiva , Línea Celular , Línea Celular Tumoral , Reactivos de Enlaces Cruzados/farmacología , Replicación del ADN , Eliminación de Gen , Predisposición Genética a la Enfermedad , Genoma , Humanos , Conformación de Ácido Nucleico , Hibridación de Ácido NucleicoRESUMEN
Dishevelled (Dvl) is the essential signal transduction component of both canonical and non-canonical Wnt signaling pathways. The cysteine-rich protein Idax acts as a negative regulator of Wnt signaling in mammals by interaction with Dvl in the region of the PDZ domain. In an effort to clarify the structural basis of this interaction, we used nuclear magnetic resonance spectroscopy to study the interaction of the Dvl PDZ domain with Idax. We first confirmed that the C-terminal region of Idax consisting of residues 109-198 binds to the PDZ domain of mouse Dvl-1 at the conventional C-terminal peptide-binding groove. However, instead of the C-terminus of Idax, we showed that a peptide of an internal sequence of Idax containing a KTXXXI motif is important in the interaction with a binding affinity estimated at 56 microM. Such internal motif identified in this study suggests a new type of sequence motif recognition for Dvl PDZ domain.