title: Functional and computational assessment of missense variants in the ataxia-telangiectasia mutated (ATM) gene: mutations with increased cancer risk.
abstract: The functional consequences of missense variants are often difficult to predict. This becomes especially relevant when DNA sequence changes are used to determine a diagnosis or prognosis. To analyze the consequences of 12 missense variants in patients with mild forms of ataxia-telangiectasia (A-T), we employed site-directed mutagenesis of ataxia-telangiectasia mutated (ATM) cDNA followed by stable transfections into a single A-T cell line to isolate the effects of each allele on the cellular phenotype. After induction of the transfected cells with CdCl2, we monitored for successful ATM transcription and subsequently assessed: 1) intracellular ATM protein levels; 2) ionizing radiation (IR)-induced ATM kinase activity; and 3) cellular radiosensitivity. We then calculated SIFT and PolyPhen scores for the missense changes. Nine variants produced little or no correction of the A-T cellular phenotype and were interpreted to be ATM mutations; SIFT/PolyPhen scores supported this. Three variants corrected the cellular phenotype, suggesting that they represented benign variants or polymorphisms. SIFT and PolyPhen scores supported the functional analyses for one of these variants (c.1709T>C); the other two were predicted to be "not tolerated" (c.6188G>A and c.6325T>G) and were classified as "operationally neutral." Genotype/phenotype relationships were compared: three deleterious missense variants were associated with an increased risk of cancer (c.6679C>T, c.7271T>G, and c.8494C>T). In situ mutagenesis represents an effective experimental approach for distinguishing deleterious missense mutations from benign or operationally neutral missense variants.
authors.affiliation: Department of Pathology and Laboratory Medicine, The David Geffen School of Medicine at the University of California, Los Angeles (UCLA), Los Angeles, California 90095-1732, USA.
authors.forename: M
authors.initials: M
authors.lastname: Mitui
authors.forename: S A
authors.initials: SA
authors.lastname: Nahas
authors.forename: L T
authors.initials: LT
authors.lastname: Du
authors.forename: Z
authors.initials: Z
authors.lastname: Yang
authors.forename: C H
authors.initials: CH
authors.lastname: Lai
authors.forename: K
authors.initials: K
authors.lastname: Nakamura
authors.forename: S
authors.initials: S
authors.lastname: Arroyo
authors.forename: S
authors.initials: S
authors.lastname: Scott
authors.forename: A
authors.initials: A
authors.lastname: Purayidom
authors.forename: P
authors.initials: P
authors.lastname: Concannon
authors.forename: M
authors.initials: M
authors.lastname: Lavin
authors.forename: R A
authors.initials: RA
authors.lastname: Gatti
mesh_terms: D001260:Ataxia Telangiectasia / Q000235:genetics; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000235:genetics* / Q000378:metabolism; D019295:Computational Biology; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D020022:Genetic Predisposition to Disease; D014644:Genetic Variation; D005838:Genotype; D006801:Humans; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense; D009369:Neoplasms / Q000235:genetics*; D010641:Phenotype; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000378:metabolism; D012307:Risk Factors; D014162:Transfection; D025521:Tumor Suppressor Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D025521:Tumor Suppressor Proteins; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases