INDUCTION OF RNA-BINDING PROTEINS IN MAMMALIAN-CELLS BY DNA-DAMAGING AGENTS

A technique to detect RNA-binding proteins (RBP) involving hybridizati on of RNA probe to proteins transferred to a membrane was used to stud y RBP in different mammalian cells and in cells after genotoxic stress . With this approach, up to 13 proteins of different sizes were detect ed in crude nuclear extracts by using a viral RNA probe consisting of the trans-activation-responsive (TAR) element of human immunodeficienc y virus type 1 (HIV-1). The TAR RNA probe contains a stem-loop structu re found in nascent HIV-1 transcripts. A G+C-rich probe with similar s tructure also bound to many of these RBP. Only a 102-kDa protein Stron gly bound to other RNA probes lacking this structure, while a probe wi th an A+U-rich stem-loop structure fail to bind most RBP, thus indicat ing a RNA secondary structure preference. The expression of these RBP varied substantially in nine different human and hamster cell lines, w ith no detectable RBP in two human myeloid lines. Evidence for inducti on ion of these RBP was found in six of seven lines after treatment wi th DNA-damaging agents; UV radiation was the most effective agent. In Chinese hamster ovary cells, which showed the strongest response, all five RBP present in untreated cells rapidly increased in activity afte r UV irradiation, and eight additional RBP were detected. The inductio n of these RBP by DNA-damaging agents indicates one or more possible r oles for these proteins in the cellular response to genotoxic stress a nd in viral activation after such stress.