Phosphorylation of a 72-kDa nucleoprotein (NP-72) in HL-60 cells is mediated by the double-stranded DNA-dependent protein kinase (DNA-PK).

Nuclear extracts prepared from human leukemic cells grown in the presence of cell differentiation-inducing agents showed a significant increase in the phosphorylation of a nuclear protein referred to as NP-72. The NP-72 phosphorylation was greatly increased by the addition of purified HeLa DNA-PK and was reduced by the inclusion of a DNA-PK-specific monoclonal antibody to the assay mixture. Phosphoamino acid analysis showed serine (60%) and threonine (40%) residues to be the phosphate acceptors. Western blot analysis of control and TPA-treated nuclear extracts detected multiple immunoreactive protein bands. The two most prominent species migrated as 300- and 210-kD proteins on SDS-PAGE, possibly representing an intact and a processed form of DNA-PK. The 300-kD form of DNA-PK was only detected in TPA-treated nuclear extracts, raising the possibility that cell differentiation is associated with the down-regulation and/or the inhibition of a protease(s) capable of regulating DNA-PK turnover.