Study of Biological Roles of Nucleophosmin/B23 in Cancer

Nucleophosmin/B23 was first identified as a nucleolar protein expressed at higher levels in the cancer cells as compared to normal cells. Nucleophosmin/B23 has long been thought to have a role in tumor formation. With our previous efforts and others, nucleophosmin/B23 proves to be a crucial gene in control of cancer. In this study, we continue to elucidate the biological role of nucleophosmin/B23 and determine how nucleophosmin/B23 can contribute to tumorigenesis. Malignant progression is a complex and poorly understood process that appears to involve genetic and epigenetic factors. Considerable effort devoted to elucidating the intracellular signaling pathways that control cellular proliferation has identified the Ras, c-Myc and E2F1 as critical components that are key for the control of cellular growth and differentiation. The questions of whether nucleophosmin/B23 is functionally associated with oncogenes and how nucleophosmin/B23 is being elevated in tumor growth stimulation are presently unanswered. Attempts will be made to explore the signaling pathway in tumor growth promotion with the linkages among oncogenes, transcriptional factors, kinases activation and nucleophosmin/B23. Furthermore, our recent results have indicated nucleophosmin/B23 is involved in regulating E2F1 activity, possibly through the interplay of NF-κB and pRB. It is therefore important for us to determine whether phosphorylation and dephosphorylation of nucleophosmin/B23 plays a role in such interplay and the mechanisim of how nucleophosmin/B23 activates E2F1. Phosphorylation mutants (point mutations at various phosphorylation sites) will be constructed for the transfection functional experiments. Attempts will also be made to identify the specific kinases and phosphatases that are involved in modifying nucleophosmin/B23 to activate E2F1. Furthermore, our recent results have demonstrated that the expression of nucleophosmin/B23 correlates with that of PCNA. Increased nucleophosmin/B23 expression results in elevated PCNA transcription level but evidence for the direct regulation of PCNA by nucleophosmin/B23 is lacking. Attempt has been made to determine whether nucleophosmin/B23 regulates PCNA through interaction with transcription factor YY1. Our preliminary results have shown by CHIP assay the recruitment of nucleophosmin/B23 to PCNA promoter during its activation. Therefore, nucleophosmin/B23 appears to be an important player in transcription of PCNA and nucleophosmin/B23 could be critically involved in regulating gene transcriptions. Moreover, our recent transcription factor-transcription factor binding array analysis of nuclear lysate have indicated that nucleophosmin/B23 could be a binding partner with AP-2α, YY1, NF-κB, and EGR-1. Nucleophosmin/B23 being possibly associated with transcriptional factors such as YY1, NFκB, AP-2α, EGR-1 in gene regulations, attempts will therefore be made to uncover the target genes of nucleophosmin/B23 with those partners in cancer cellular growth and differentiation and in response to UV. Our studies overall will focus on elucidating the transcriptional regulations of nucleophosmin/B23 and its downstream target genes in cancer cells. The experimental strategy in this proposal will open a novel avenue to understanding the nuclear functions of nucleophosmin/B23 in cancer cell growth. The specific aims will thus be the following: I. To elucidate whether there are important linkages among oncogenes (such as Ras, c-Myc, etc) and nucleophosmin/B23 in cancer cells during growth stimulation or in response to UV and DNA-damaging agents such as doxorubicin. II. To study the transcriptional regulation of E2F1 by nucleophosmin/B23; The effects of over-expression of nucleophosmin/B23 and its mutants (point mutations at phosphorylation, acetylation sites) on E2F1 promoter activation and recruitment of NFκB, E2F1 and pRB to the promoter will be determined. III. To elucidate the Interplay of nucleophosmin/B23 and YY1 in regulation of PCNA gene expression ; In order to extend our understanding of the physiological role of nucleophosmin/B23.YY1 binding to the PCNA promoter, we will address specifically the impact of cell crowding, UV and doxorubicin (DNA damage) treatments on the promoter activation and recruitment of YY1 and nucleophosmin/B23 to the promoter. IV. To study the transcriptional factors involved in regulation of nucleophosmin/B23 during induction of differentiation. V. To uncover the target genes of nucleophosmin/B23 in cancer cellular growth and differentiation and their biological roles in cancer ; To demonstrate the possible interaction of nucleophosmin/B23 with AP-2αand to address the transcriptional function of the nucleophosmin/B23-AP2α complex.

Project ID:PC9706-0783
External Project ID:NSC96-2320-B182-027-MY3