DNA‐dependent protein kinase (DNA‐PK) phosphorylates nuclear DNA helicase II/RNA helicase A and hnRNP proteins in an RNA‐dependent manner

An RNA‐dependent association of Ku antigen with nuclear DNA helicase II (NDH II), alternatively named RNA helicase A (RHA), was found in nuclear extracts of HeLa cells by immunoprecipitation and by gel filtration chromatography. Both Ku antigen and NDH II were associated with hnRNP complexes. Two‐dimensional gel electrophoresis showed that Ku antigen was most abundantly associated with hnRNP C, K, J, H and F, but apparently not with others, such as hnRNP A1. Unexpectedly, DNA‐dependent protein kinase (DNA‐PK), which comprises Ku antigen as the DNA binding subunit, phosphorylated hnRNP proteins in an RNA‐dependent manner. DNA‐PK also phosphorylated recombinant NDH II in the presence of RNA. RNA binding assays displayed a preference of DNA‐PK for poly(rG), but not for poly(rA), poly(rC) or poly(rU). This RNA binding affinity of DNA‐PK can be ascribed to its Ku86 subunit. Consistently, poly(rG) most strongly stimulated the DNA‐PK‐catalyzed phosphorylation of NDH II. RNA interference studies revealed that a suppressed expression of NDH II altered the nuclear distribution of hnRNP C, while silencing DNA‐PK changed the subnuclear distribution of NDH II and hnRNP C. These results support the view that DNA‐PK can also function as an RNA‐dependent protein kinase to regulate some aspects of RNA metabolism, such as RNA processing and transport.