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Phenotypic characterization of missense polymerase-δ mutations using an inducible protein-replacement system. | LitMetric

Phenotypic characterization of missense polymerase-δ mutations using an inducible protein-replacement system.

Nat Commun

Institute of Molecular Cancer Research, University of Zurich, ETH Zurich, Winterthurerstrasse 190, Zurich 8057, Switzerland.

Published: September 2014

AI Article Synopsis

Article Abstract

Next-generation sequencing has revolutionized the search for disease-causing genetic alterations. Unfortunately, the task of distinguishing the handful of causative mutations from rare variants remains daunting. We now describe an assay that permits the analysis of all types of mutations in any gene of choice through the generation of stable human cell lines, in which the endogenous protein has been inducibly replaced with its genetic variant. Here we studied the phenotype of variants of the essential replicative polymerase-δ carrying missense mutations in its active site, similar to those recently identified in familial colon cancer patients. We show that expression of the mutants but not the wild-type protein endows the engineered cells with a mutator phenotype and that the mutations affect the fidelity and/or the exonuclease activity of the isolated enzyme in vitro. This proof-of-principle study demonstrates the general applicability of this experimental approach in the study of genotype-phenotype correlations.

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Source
http://dx.doi.org/10.1038/ncomms5990DOI Listing

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