title: The NF90/NF45 complex participates in DNA break repair via nonhomologous end joining.
abstract: Nuclear factor 90 (NF90), an RNA-binding protein implicated in the regulation of gene expression, exists as a heterodimeric complex with NF45. We previously reported that depletion of the NF90/NF45 complex results in a multinucleated phenotype. Time-lapse microscopy revealed that binucleated cells arise by incomplete abscission of progeny cells followed by fusion. Multinucleate cells arose through aberrant division of binucleated cells and displayed abnormal metaphase plates and anaphase chromatin bridges suggestive of DNA repair defects. NF90 and NF45 are known to interact with the DNA-dependent protein kinase (DNA-PK), which is involved in telomere maintenance and DNA repair by nonhomologous end joining (NHEJ). We hypothesized that NF90 modulates the activity of DNA-PK. In an in vitro NHEJ assay system, DNA end joining was reduced by NF90/NF45 immunodepletion or by RNA digestion to an extent similar to that for catalytic subunit DNA-PKcs immunodepletion. In vivo, NF90/NF45-depleted cells displayed increased γ-histone 2A.X foci, indicative of an accumulation of double-strand DNA breaks (DSBs), and increased sensitivity to ionizing radiation consistent with decreased DSB repair. Further, NF90/NF45 knockdown reduced end-joining activity in vivo. These results identify the NF90/NF45 complex as a regulator of DNA damage repair mediated by DNA-PK and suggest that structured RNA may modulate this process.
authors.affiliation: Departments of Biochemistry and Molecular Biology, UMDNJ-New Jersey Medical School, and Graduate School of Biomedical Sciences, UMDNJ, Newark, New Jersey 07101-1709, USA.
authors.forename: Raghavendra A
authors.initials: RA
authors.lastname: Shamanna
authors.forename: Mainul
authors.initials: M
authors.lastname: Hoque
authors.forename: Anita
authors.initials: A
authors.lastname: Lewis-Antes
authors.forename: Edouard I
authors.initials: EI
authors.lastname: Azzam
authors.forename: David
authors.initials: D
authors.lastname: Lagunoff
authors.forename: Tsafi
authors.initials: T
authors.lastname: Pe'ery
authors.forename: Michael B
authors.initials: MB
authors.lastname: Mathews
mesh_terms: D034961:Antigens, Nuclear / Q000378:metabolism; D002459:Cell Fusion; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000378:metabolism / Q000528:radiation effects; D053903:DNA Breaks, Double-Stranded; D059766:DNA End-Joining Repair; D051747:DNA-Activated Protein Kinase / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D057075:Enzyme Assays; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D000072200:Ku Autoantigen; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D046912:Multiprotein Complexes / Q000378:metabolism*; D051903:Nuclear Factor 45 Protein / Q000235:genetics / Q000378:metabolism*; D050779:Nuclear Factor 90 Proteins / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D034622:RNA Interference; D059008:Time-Lapse Imaging
chemical_list: D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; C494013:ILF2 protein, human; D046912:Multiprotein Complexes; D051903:Nuclear Factor 45 Protein; D050779:Nuclear Factor 90 Proteins; D009687:Nuclear Proteins; D004247:DNA; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen