Abstract
Double-strand breaks (DSBs) are generated within genome DNA after exposure to Ionizing Radiation (IR) or DNA damaging agents. Cells recognize DSBs immediately then activate cell cycle checkpoints and initiate repair of the damaged DNA. H2AX, histone H2A variant, is a candidate of DNA damage sensor protein, because H2AX is phosphorylated in an earlier step by PI3KK (phosphatidylinositol 3-kinase like kinase) and this phosphorylation is indispensable for foci formation of NBS1 protein at DSB sites. Moreover, H2AX forms a complex with NBS1 and ATM, which play an important role for DNA damage response. To clarify the role of H2AX in DNA damage response, we attempted to identify H2AX-binding proteins by pull-down assay using GST-tagged H2AX and GST-tagged H2AX (S139E). Thereby, we identified nucleolin as a candidate of H2AX-interacting protein by mass spectrometry analysis. Nucleolin is a major nucleolar protein in eukaryotic cells and directly involved in the regulation of ribosome biogenesis and maturation. Moreover, nucleolin re-localized from nucleolus to nucleoplasm in response to DNA damage and interacted with RPA, which is crucial for DNA damage response after DNA replication stress. These results suggest the functional interaction of nucleolin with H2AX.
