Identification of nucleophosmin/B23, an acidic nucleolar protein, as a stimulatory factor for in vitro replication of adenovirus DNA complexed with viral basic core proteins

The processes governing chromatin remodeling and assembly, which occur prior to and/or after transcription and replication, are not completely understood. To understand the mechanisms of transcription and replication from chromatin templates, we have established in vitro replication and transcription systems using adenovirus (Ad) DNA complexed with viral basic core proteins, called Ad core, as a template. Using this system, we have previously identified, from HeLa cells, template activating factor-I as a stimulatory factor for the Ad core DNA replication. Here, using this system as a tool, we identified and purified a novel template activating factor activity that consists of two acidic polypeptides whose apparent molecular masses are 38 kDa and 37 kDa. These two polypeptides correspond to two splicing variants of nucleolar phosphoprotein, nucleophosmin/B23. Recombinant B23 proteins stimulate the Ad core DNA replication, and the acidic regions of B23 proteins are important for its activity. In addition, B23 proteins directly bind to core histones and transfer them to naked DNA. Furthermore, chromatin components such as histories and topoisomerase II are co-immunoprecipitated with B23 from cell extracts. These observations lead to a hypothesis that nucleophosmin/B23 is involved in structural changes of chromatin, thereby regulating transcription and replication within the ribosomal DNA region or maintaining the nucleolar structure. (C) 2001 Academic Press.