Nucleolar Protein B23.1 Binds to Retinoblastoma Protein and Synergistically Stimulates DNA Polymerase   Activity

Abstract

Phosphorylated retinoblastoma protein and nucleolar protein B23 are putative stimulatory factors for DNA polymerase alpha. We showed that these two factors interacted with each other and stimulated the activity of DNA polymerase alpha synergistically. B23 exists in two isoforms designated as B23.1 and B23.2. While B23.1 bound to a retinoblastoma protein-conjugated column, B23.2 did not. These results indicate that B23.1 can directly bind to retinoblastoma protein. It was also shown that B23 was co-immunoprecipitated with both retinoblastoma protein and DNA polymerase alpha from a HeLa cell extract by monoclonal antibodies raised against these components. These results suggest that these three proteins exist as a complex in cells, at least in part. The simultaneous addition of both B23.1 and retinoblastoma protein caused stimulation of DNA polymerase alpha activity that is much higher than the sum of the stimulation by retinoblastoma protein and B23.1 alone. The maximal stimulation was attained at the molar ratio of DNA polymerase alpha/retinoblastoma protein/B23.1 = 1:1:12. Since B23 exists as a hexamer in solution, it may act as a stimulator of DNA polymerase alpha in a form of double-hexamer, in concert with the phosphorylated retinoblastoma protein.

NSC348884, a nucleophosmin inhibitor disrupts oligomer formation and induces apoptosis in human cancer cells

“…The structural features of NPM consist of an oligomerization domain, a metal-binding motif, a bipartite nuclear localization signal, phosphorylation sites and a nucleolar localization signal (Wang et al, 1993;Wang et al, 2005). Mounting evidence supports that NPM interacts with a variety of proteins including nucleolin (Li et al, 1996), cell cycle-related protein p120 (Valdez et al, 1994), human immunodeficiency virus (HIV)-1 Rev protein (Fankhauser et al, 1991), Human MDM2 (HDM2) (Kurki et al, 2004), tumor suppressor ARF (Itahana et al, 2003;Bertwistle et al, 2004;Korgaonkar et al, 2005), p53 (Colombo et al, 2002;Li et al, 2004;Maiguel et al, 2004) and pRb (Takemura et al, 1999).…”

“…In addition, NPM is frequently found in chromosomal translocations associated with hematological malignancies (Naoe et al, 2006). The role of NPM as an oncogene is further enhanced as it binds several tumor suppressor genes, including pRb (Takemura et al, 1999), p14 ARF (Brady et al, 2004;Zhang, 2004;Lee et al, 2005;Gjerset, 2006) and p53 (Colombo et al, 2002;Li et al, 2004;Maiguel et al, 2004). The p53/p14 ARF /Mdm2 (Hdm2) stress response pathway plays a central role in mediating cellular responses to oncogene activation, genome instability and therapy-induced DNA damage.…”

Increased expression of nucleophosmin/B23 in hepatocellular carcinoma and correlation with clinicopathological parameters

“…It has also been shown that NPM is a direct target of the Myc transcription factor during ribosomal biogenesis (Zeller et al, 2001) and cyclin E/CDK2 kinase activity during centrosome duplication (Okuda et al, 2000). Nucleophosmin physically interacts with several other cellular factors such as the retinoblastoma susceptibility gene product pRB (Takemura et al, 1994(Takemura et al, , 1999, the cell cycle G2 -M checkpoint factor Gadd45a (Gao et al, 2005), and the tumour suppressor p14 ARF (Itahana et al, 2003). Mounting evidence has linked excessive NPM to cellular transformation and ontogenesis (Pulford et al, 2001).…”

Proteome analysis of proliferative response of bystander cells adjacent to cells exposed to ionizing radiation

“…This protein is mainly localized in granular regions of the nucleolus and is associated with ribosome biogenesis. It also plays a substantial role in cell-cycle progression and cell division [26][27][28]. NPM/B23 binds to retinoblastoma protein (pRb) and synergistically stimulates DNA polymerase-α [26].…”

“…It also plays a substantial role in cell-cycle progression and cell division [26][27][28]. NPM/B23 binds to retinoblastoma protein (pRb) and synergistically stimulates DNA polymerase-α [26]. Down-regulation of NPM/B23 mRNA delays the entry of cells into mitosis [27].…”