Nonerythroid alphaII spectrin is required for recruitment of FANCA and XPF to nuclear foci induced by DNA interstrand cross-links.
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 alpha spectrin (alphaSpIISigma*), present in normal human cell nuclei, plays an important role in repair of DNA interstrand cross-links. These results show that alphaSpIISigma* 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 alphaSpIISigma* 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 alphaSpIISigma*, there is a significant reduction in formation of damage-induced XPF as well as alphaSpIISigma* nuclear foci, even though levels of XPF are normal in these cells. In corrected FA-A cells, in which levels of alphaSpIISigma* are restored to normal, numbers of damage-induced nuclear foci are also returned to normal. Co-immunoprecipitation studies show that alphaSpIISigma*, FANCA and XPF co-immunoprecipitate with each other from normal human nuclear proteins. These results demonstrate that alphaSpIISigma*, 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 alphaSpIISigma* in the nucleus is to act as a scaffold, aiding in recruitment and alignment of repair proteins at sites of damage.
Sridharan
Deepa
D
Department of Pathology and Laboratory Medicine, UMDNJ - New Jersey Medical School and the Graduate School of Biomedical Sciences, Newark, NJ 07103, USA.
Brown
Monique
M
Lambert
W Clark
WC
McMahon
Laura W
LW
Lambert
Muriel W
MW
D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D018736:DNA Adducts / Q000378:metabolism; D004249:DNA Damage; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000378:metabolism; D004307:Dose-Response Relationship, Radiation; D052217:Fanconi Anemia Complementation Group A Protein; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D008730:Methoxsalen / Q000494:pharmacology; D009687:Nuclear Proteins / Q000378:metabolism; D011233:Precipitin Tests / Q000379:methods; D011506:Proteins / Q000378:metabolism*; D013049:Spectrin / Q000378:metabolism; D013997:Time Factors; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D018736:DNA Adducts; D004268:DNA-Binding Proteins; C498471:FANCA protein, human; D052217:Fanconi Anemia Complementation Group A Protein; D009687:Nuclear Proteins; D011506:Proteins; C095888:xeroderma pigmentosum group F protein; D013049:Spectrin; D008730:Methoxsalen