ADPribosylation of nuclear proteins labeled with [3H]adenosine: changes during the HeLa cycle

Abstract

Cell cycle variations in the modification of histones and nonhistones by ADPribosylation were investigated. Proteins of HeLa interphase nuclei and metaphase chromosomes were radioactively labeled in vivo with [3H]adenosine. Histones of metaphase chromosomes were extensively modified by ADPribosylation, with H2B, H2A and H4 being predominant acceptors of [3H]adenosine label. For histones of interphase nuclei from synchronized cells, the highest level of 3H labeling was observed by two-dimensional gel electrophoresis to occur in S phase. The minimum level was noted in G1 phase. ADPribosylation of histones is, however, significant during all phases of the cell cycle. These conclusions were confirmed by experiments using [32P]NAD. The results with the specific inhibitor of ADPribosylation, 3-aminobenzamide, and with snake venom phosphodiesterase indicated that the radioactive isotopes were incorporated as ADPribose. Two-dimensional gels of HeLa nonhistones labeled with [3H]adenosine showed strikingly different patterns for interphase and metaphase samples. Over 100 ADPribosylated species were found for interphase nuclei, but poly(ADPribose) polymerase was the only major acceptor for metaphase chromosomes. A simple pattern was also revealed for nuclear scaffolds, with the 'lamins' and poly(ADPribose) polymerase being identifiable as modified species.