MODIFICATION OF NUCLEOLAR PROTEIN B23 AFTER EXPOSURE TO IONIZING-RADIATION

The responses of cells to ionizing radiation include induction and/or suppression of the expression of genes and proteins, In our investigat ions of alterations in cellular protein expression in response to ioni zing radiation, we have used the techniques of two-dimensional polyacr ylamide gel electrophoresis and silver staining. We compared the nucle ar protein profiles of control and irradiated (6 Gy, 4 h postirradiati on) radioresistant squamous carcinoma cells (SQ-20B) and observed an a lteration in the expression of a 40 kDa protein: control nuclei expres s a protein isoform with pi values between 5.4-5.8, while irradiated n uclei express a more acidic variant with pi values between 5.2-5.5. Us ing the cyanogen bromide/O-phthalaldehyde method followed by microsequ encing analysis, we obtained an internal amino acid sequence and ident ified the 40 kDa protein as nucleolar protein B23. Western blotting ex periments confirmed the internal amino acid sequencing results and sho wed both species (control, 5.4-5.8, irradiated, 5.2-5.5) to be recogni zed by an anti-B23 monoclonal antibody. Radiolabeling of control and i rradiated samples with [P-32]NAD or [P-32]orthophosphoric acid reveale d the acidic species of B23 to be both ADP-ribosylated and phosphoryla ted. Therefore, exposure of SQ-20B cells to radiation results in the i ncrease in expression of an ADP-ribosylated and phosphorylated species of B23. (C) 1995 by Radiation Research Society