p53 centrosomal localization diagnoses ataxia-telangiectasia homozygotes and heterozygotes
Andrea Prodosmo, … , Luciana Chessa, Silvia Soddu
Andrea Prodosmo, … , Luciana Chessa, Silvia Soddu
Published February 1, 2013
Citation Information: J Clin Invest. 2013;123(3):1335-1342. https://doi.org/10.1172/JCI67289.
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Technical Advance Genetics

p53 centrosomal localization diagnoses ataxia-telangiectasia homozygotes and heterozygotes

Abstract

Ataxia-telangiectasia (A-T) is an autosomal recessive neurodegenerative disorder characterized by radiosensitivity, genomic instability, and predisposition to cancer. A-T is caused by biallelic mutations in the ataxia-telangiectasia mutated (ATM) gene, but heterozygous carriers, though apparently healthy, are believed to be at increased risk for cancer and more sensitive to ionizing radiation than the general population. Despite progress in functional and sequencing-based assays, no straightforward, rapid, and inexpensive test is available for the identification of A-T homozygotes and heterozygotes, which is essential for diagnosis, genetic counseling, and carrier prediction. The oncosuppressor p53 prevents genomic instability and centrosomal amplification. During mitosis, p53 localizes at the centrosome in an ATM-dependent manner. We capitalized on the latter finding and established a simple, fast, minimally invasive, reliable, and inexpensive test to determine mutant ATM zygosity. The percentage of mitotic lymphoblasts or PBMCs bearing p53 centrosomal localization clearly discriminated among healthy donors (>75%), A-T heterozygotes (40%–56%), and A-T homozygotes (<30%). The test is specific for A-T, independent of the type of ATM mutations, and recognized tumor-associated ATM polymorphisms. In a preliminary study, our test confirmed that ATM is a breast cancer susceptibility gene. These data open the possibility of cost-effective, early diagnosis of A-T homozygotes and large-scale screenings for heterozygotes.

Authors

Andrea Prodosmo, Andrea De Amicis, Cecilia Nisticò, Mario Gabriele, Giuliana Di Rocco, Laura Monteonofrio, Maria Piane, Enrico Cundari, Luciana Chessa, Silvia Soddu

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Figure 1

p53 centrosomal localization in mitotic LCLs.

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p53 centrosomal localization in mitotic LCLs. (A) p53 centrosomal locali...
(A) p53 centrosomal localization detected by confocal IF microscopy with anti-p53 (green) and anti–γ-tubulin (red) Abs. DNA was stained with DAPI (blue); representative image and its software analysis are shown in A. (B) Representative IF images of the indicated proteins in mitotic LCLs derived from the indicated donors. p53, centrosomes, and DNA are marked as in A. Scale bar = 5 μm. (C) Summary data of percentages of p53-MCL measured by 2 different operators who counted 100 metaphases per sample in quadruplicate. Data are expressed as mean ± SD. Four different families, each including 1 A-T patient (A-T), 1 parent (Htz), and 1 wtATM-carrying relative (WT) were evaluated. (D) Comparison of p53-MCL percentages, evaluated as in C, between LCLs derived from wtATM donors (n = 11), A-T carriers (n = 20), and A-T patients (n = 10). The 3 groups are significantly different. Box-and-whisker plot: horizontal lines within the box represent median values, and the bottom and top of the box show the lower and upper quartiles. ***P < 0.0001; 2-tailed Student’s t test.