Research Article The Role of Telomere Length Modulation in Delayed Chromosome Instability Induced by Ionizing Radiation in Human Primary Fibroblasts Francesco Berardinelli, 1,2 Antonio Antoccia, 1,2 Rossella Buonsante, 1 Silvia Gerardi, 3 Roberto Cherubini, 3 Viviana De Nadal, 3 Caterina Tanzarella, 1,2 and Antonella Sgura 1,2 * 1 Dipartimento Di Scienze, Universit  a “Roma Tre”, Rome, Italy 2 INFN-“Roma Tre”, Rome, Italy 3 INFN-Laboratori Nazionali di Legnaro (LNL), Legnaro (Padova), Italy Telomere integrity is important for chromosome sta- bility. The main objective of our study was to investi- gate the relationship between telomere length modulation and mitotic chromosome segregation induced by ionizing radiation in human primary fibroblasts. We used X-rays and low-energy protons because of their ability to induce different telomeric responses. Samples irradiated with 4 Gy were fixed at different times up to 6 days from exposure and telomere length, anaphase abnormalities, and chro- mosome aberrations were analyzed. We observed that X-rays induced telomere shortening in cells har- vested at 96 hrs, whereas protons induced a signifi- cant increase in telomere length at short as well as at long harvesting times (24 and 96 hrs). Consistent with this, the analysis of anaphase bridges at 96 hrs showed a fourfold increase in X-ray- compared with proton-irradiated samples, suggesting a correlation between telomere length/dysfunction and chromo- some missegregation. In line with these findings, the frequency of dicentrics and rings decreased with time for protons whereas it remained stable after X- rays irradiation. Telomeric FISH staining on ana- phases revealed a higher percentage of bridges with telomere signals in X-ray-treated samples than that observed after proton irradiation, thus suggest- ing that the aberrations observed after X-ray irradia- tion originated from telomere attrition and consequent chromosome end-to-end fusion. This study shows that, beside an expected “early” chro- mosome instability induced shortly after irradiation, a delayed one occurs as a result of alterations in telomere metabolism and that this mechanism may play an important role in genomic stability. Environ. Mol. Mutagen. 54:172–179, 2013. V C 2013 Wiley Periodicals, Inc. Key words: DNA damage; telomeres; chromosome instability; anaphases bridge INTRODUCTION Telomeres are physically located at the end of eukaryotic chromosome arms and play a “protective” role by prevent- ing chromosome end attrition. Telomeric chromatin is char- acterized by the association of telomeric DNA with specialized proteins that organize the linear chromosome end into a stable structure that is not recognized by the cell as a chromosome break [Bailey and Cornforth, 2007]. Ionizing radiation (IR) has been associated with both telomere dysfunction and shortening. It is well known that X-rays produce reactive oxygen species (ROS) and that ROS play a central role in cellular senescence [Pas- sos and Von Zglinicki, 2005] because they increase telo- mere shortening [Richter and Von Zglinicki, 2007] and providing a link between X-ray exposure and telomere shortening [Sabatino et al., 2012]. Other authors have demonstrated that X-rays induce telomere dysfunction, persistent telomeric induced dysfunctional foci (TIF) for- mation [Urushibara et al., 2004], and deficiency in DSBs repair near telomeres [Miller et al., 2011; Fumagalli et al., 2012; Hewitt et al., 2012] Degradation of telomeres in the absence of proteins involved in telomere mainte- nance may represent a source of DNA damage that can lead to chromosomal rearrangements. Dysfunctional telo- meres that are due to alterations in the telomere-associ- ated proteins required for end-capping or to alterations Grant sponsor: INFN-CSN5 (Experiments SHEILA and EXCALIBUR). Grant sponsor: ASI MoMa Project. *Correspondence to: Antonella Sgura, Department of Biology, V.le G.Marconi 446, 00146-Rome, Italy. E-mail: antonella.sgura@uniroma3.it Received 6 July 2012; and in final form 19 December 2012 DOI 10.1002/em.21761 Published online 9 February 2013 in Wiley Online Library (wileyonlinelibrary.com). V C 2013 Wiley Periodicals, Inc. Environmental and Molecular Mutagenesis 54:172^179 (2013)