Nonerythroid αII spectrin is required for recruitment of FANCA and XPF to nuclear foci induced by DNA interstrand cross-links

Abstract

The events responsible for repair of DNA interstrand cross-links in mammalian cells, the proteins involved and their interactions with each other are poorly understood. The present study demonstrates that the structural protein nonerythroid α spectrin (αSpIIΣ*), present in normal human cell nuclei, plays an important role in repair of DNA interstrand cross-links. These results show that αSpIIΣ* relocalizes to nuclear foci after damage of normal human cells with the DNA interstrand cross-linking agent 8-methoxypsoralen plus ultraviolet A (UVA) light and that FANCA and the known DNA repair protein XPF localize to the same nuclear foci. That αSpIIΣ* is essential for this re-localization is demonstrated by the finding that in cells from patients with Fanconi anemia complementation group A (FA-A), which have decreased ability to repair DNA interstrand cross-links and decreased levels of αSpIIΣ*, there is a significant reduction in formation of damage-induced XPF as well asαSpIIΣ* nuclear foci, even though levels of XPF are normal in these cells. In corrected FA-A cells, in which levels of αSpIIΣ*are restored to normal, numbers of damage-induced nuclear foci are also returned to normal. Co-immunoprecipitation studies show thatαSpIIΣ*, FANCA and XPF co-immunoprecipitate with each other from normal human nuclear proteins. These results demonstrate thatαSpIIΣ*, FANCA and XPF interact with each other in the nucleus and indicate that there is a close functional relationship between these proteins. These studies suggest that an important role for αSpIIΣ* in the nucleus is to act as a scaffold, aiding in recruitment and alignment of repair proteins at sites of damage.