The Werner Syndrome Gene Product Co-purifies with the DNA Replication Complex and Interacts with PCNA and Topoisomerase I

Michel Lebel,Elisa A Spillare,Curtis C Harris,Philip Leder

doi:10.1074/jbc.274.53.37795

Michel Lebel, Elisa A Spillare + Show 2 more

Open Access

https://doi.org/10.1074/jbc.274.53.37795

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Abstract

Werner syndrome (WS) is a recessive disorder characterized by genomic instability and by the premature onset of a number of age-related diseases. To understand the molecular basis of this disease, we deleted a segment of the murine Wrn gene and created Wrn-deficient embryonic stem (ES) cells. At the molecular level, wild type-but not mutant-WS protein co-purifies through a series of centrifugation, chromatography, and sucrose gradient steps with the well characterized 17 S multiprotein DNA replication complex. Furthermore, wild type WS protein co-immunoprecipitates with a prominent component of the multiprotein replication complex, proliferating cell nuclear antigen (PCNA). In vitro studies also indicate that PCNA binds to a region in the N terminus portion of the WS protein containing a potential 3'-5' exonuclease domain. Finally, human WS protein also co-immunoprecipitates with both PCNA and topoisomerase I. These results suggest that the WS protein interacts with several components of the DNA replication fork.

Highlights

Werner syndrome (WS) is a recessive disorder characterized by genomic instability and by the premature onset of a number of age-related diseases
A Deletion of a Portion of the WS Helicase Domain Affects Its Interaction with the DNA Replication Complex—The human WRN gene encodes a central motif composed of seven domains constituting the helicase signature
Because both of these topoisomerases are involved in DNA replication, it is possible that the WS protein acts in concert with these enzymes as part of a replication structure

Summary

Introduction

Werner syndrome (WS) is a recessive disorder characterized by genomic instability and by the premature onset of a number of age-related diseases. Wild type WS protein co-immunoprecipitates with a prominent component of the multiprotein replication complex, proliferating cell nuclear antigen (PCNA). Human WS protein co-immunoprecipitates with both PCNA and topoisomerase I. These results suggest that the WS protein interacts with several components of the DNA replication fork. Werner syndrome (WS) is a rare disorder characterized by the premature onset of a number of processes associated with aging [1, 2]. The homologue of the human Werner syndrome gene product in Xenopus laevis is required for the formation of replication foci in egg extracts [16]

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