Abstract 3924: Failure of localization of αII spectrin to telomeres after DNA interstrand cross-link damage in Fanconi anemia cells may be an important factor in their chromosome instability

Abstract

Abstract A hallmark of the cancer prone genetic disorder, Fanconi anemia (FA), is genomic instability characterized by both chromosome instability and defects in DNA interstrand crosslink (ICL) repair. The chromosome instability, in particular after exposure to ICLs, suggests telomere dysfunction. We have demonstrated that the structural protein, nonerythroid α spectrin (αIISp), is present in the nucleus of normal human cells where it is critical in repair of DNA ICLs and maintenance of chromosome stability and that in FA cells levels of αIISp are reduced due to its increased cleavage by μ-calpain, which correlate with reduced ICL repair and increased chromosomal instability. The goal of this study was to determine whether αIISp is important for telomere function after DNA ICL damage and whether telomere dysfunction in FA cells after damage can be corrected when levels of αIISp are restored to normal. Normal and FA complementation group A (FA-A) lymphoblastoid cells were treated with a DNA ICL agent, 8-methoxypsoralen plus UVA light or mitomycin C and studied using immunofluorescence and in situ hybridization (IF-FISH). Cells were probed with anti-α-spectrin and either a telomeric probe (PNA) or antibodies against two telomeric proteins, TRF1 or TRF2. The results showed that 47-50% of αIISp foci colocalized with the PNA probe, TRF1 or TRF2 in telomeres in ICL damaged but not undamaged normal cells and that this colocalization was specific for the S phase. This indicated that αIISp may play an important role in telomere function after ICLs and this view was strengthened by data showing that αIISp has markedly enhanced binding for TRF1 and TFR2 after ICL damage. IF-FISH showed that the ICL repair protein, XPF, also had enhanced association with telomeres after ICL formation. In FA-A cells, no localization of XPF to telomeres was observed; however, after αIISp levels were returned to normal by siRNA knockdown of μ-calpain, αIISp as well as XPF associated with telomeres after ICLs. The chromosomal aberrations (i.e., breaks and end fusions) observed in FA-A cells after ICL damage were also corrected. The present results suggest that, after ICL damage, αIISp associates with telomeres in S phase and this association may be important in repair of ICLs in replicating telomeres. In FA-A cells, failure of localization of αIISp and XPF to telomeres after ICL damage could lead to unrepaired ICLs, telomere dysfunction and production of the chromosomal aberrations observed in these cells. These studies show that αIISp may play an important role in telomere function after DNA damage and during telomere replication. Correction of the deficiency in αIISp in FA cells by knocking down μ-calpain may be significant in reversing telomere dysfunction and chromosomal instability after ICL damage and could have important therapeutic relevance. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3924. doi:10.1158/1538-7445.AM2011-3924