Sumoylation of nucleophosmin/B23 regulates its subcellular localization, mediating cell proliferation and survival

Abstract

Nucleophosmin/B23 is a major multifunctional nucleolar phosphoprotein that plays a critical role in ribosome biogenesis and cell proliferation. Arf tumor suppressor binds B23 and enhances its sumoylation. However, the biological effects of this event remain unknown. Here we show that B23 is sumoylated on both Lysine 230 and 263 residues, but the latter is the major one. Mutation of K263, but not K230, into R abolishes its centrosomal and nucleolar residency. Moreover, Rb binds to wild-type B23, but fails to interact with K263R. Sumoylation enhances B23 binding to Rb. Consequently, B23 potently stimulates E2F1-mediated transcriptional activity, which is abolished in B23 K263R. Further, K263R mutation makes B23 vulnerable to caspase-3 cleavage and sensitizes cells to apoptosis. Surprisingly, K230R mutant strongly binds to phosphatidylinositol-3,4,5-trisphosphate and suppresses DNA fragmentation. Thus, B23 sumoylation regulates its subcellular localization, cell proliferation, and survival activities.centrosome ͉ nucleolus ͉ sumoylation ͉ K263 N ucleophosmin (NPM)/B23 is a multifunctional protein involved in many cellular activities, and it has been attributed with both oncogenic and tumor-suppressive functions. B23 is more abundant in tumor and growing cells than in normal resting cells (1, 2). In addition, B23 is frequently found in the chromosomal translocation associated with several hematopoietic malignancies, such as acute promyelocytic leukemia (3), anaplastic large-cell lymphomas (4), and myelodysplasia/acute myeloid leukemia (5). Furthermore, certain mutations in the NPM gene incur the cytoplasmic localization of mutated B23, which has been proposed as a biomarker for certain acute myelogenous leukemia (6, 7). Overexpression of B23 induces cell cycle arrest in normal fibroblasts, whereas it promotes S phase entry in cells lacking p53. Conversely, knocking down B23 inhibits the processing of preribosomal RNA and induces cell death (8). In agreement with these observations, overexpression of B23 decreases the susceptibility of human leukemia HL-60 cells to retinoic acid-induced differentiation and apoptosis as well as UV-induced apoptosis in NIH 3T3 cells (9-11). Nonetheless, B23 can be cleaved by active caspase-3, which may influence its antiapoptotic action (12). It has been shown that B23 suppresses hypoxia or UV-induced cell death by repressing p53 phosphorylation and transcription activity (13,14). Recently, we presented biochemical evidence revealing that B23 is a nuclear phosphatidylinositol-3,4,5-trisphosphate [PI(3,4,5)P 3 ] receptor, and this complex directly interacts with caspase-activated DNase and inhibits its DNA fragmentation activity (15). Thus, these findings demonstrate that B23 acts as a physiologically important antiapoptotic protein. However, emerging evidence demonstrates that B23 can also act as a tumor suppressor. Npm germ-line deletion reveals that it is essential for embryonic development and the maintenance of genomic stability. Npm1 heterozygosity accelerates oncogenesis both in ...

Prognostic Role of Nucleophosmin in Colorectal Carcinomas

“…It is involved in a complex network of interactions and has multiple functions involved in the ribosomal biosynthesis (Herrera et al, 1995;Savkur et al, 1998), control of centrosome reproduction (Okuda, 2002), cell apoptosis (Liu et al, 1998), molecular chaperones (Okuwaki et al, 2001), environmental stress responses (Kurki et al, 2004), the regulation of the tumor suppressor proteins p53 (Colombo et al, 2002) and p14ARF (Bertwistle et al, 2004), and post-translational modification by acetylation (Swaminathan et al, 2005), sumoylation (Liu et al, 2007), ubiquitinylation (Sato et al, 2004), and phosphorylation (Okuda et al, 2000).…”

Nucleolin and nucleophosmin: nucleolar proteins with multiple functions in DNA repair

“…NPM1 undergoes rapid and reversible translocation from the nucleolus to the nucleoplasm (nucleoplasmic translocation) upon exposure to UV irradiation, oxidative stress, heat shock or numerous DNA-damaging agents including mitomycin C, methyl methanesulfonate (MMS) and cisplatin; however, NPM1 does not alter its localization in response to ionizing radiation (IR) or exposure to double strand break (DSB)-inducing topoisomerase inhibitors such as etoposide or camptothecin (Kurki et al 2004;Vanderwaal et al 2009;Yogev et al 2008). Importantly, these localization changes do not coincide with relocalization of NCL or a generalized dissociation of the nucleolus as seen in response to RNA Pol I inhibition, and evidence is emerging for their control by particular, DNA-damage-responsive signaling pathways and/or post-translational modifications (Kurki et al 2004;Liu et al 2007;Sato et al 2004;Vanderwaal et al 2009;Yogev et al 2008). Given such evidence, it is likely that the nucleoplasmic translocation of NPM1 is an active and controlled response to particular genotoxic stress conditions and not the result of generalized nucleolar disruption.…”

“…Separate groups have also used a biochemical assay for repair of a UV-irradiated reporter plasmid to demonstrate decreased NER activity following either overexpression of NCL (Yang et al 2009) or depletion of NPM1 (Wu et al 2002b), suggesting that these proteins may have opposing activities in NER regulation. It is important to note, however, that neither of these experimental approaches are able to differentiate between direct activities of NPM1/NCL at sites of NER and global NER deficiencies arising indirectly, such as from NPM1/NCL-dependent transcriptional modulation of the NER machinery (Lin et al 2010;Liu et al 2007;Wu et al 2002a;Wu et al 2002b). Indeed, Wu et al (2002aWu et al ( , 2002b observed that expression levels of PCNA were closely correlated with those of NPM1 in their system, likely explaining the observed effects on NER efficiency.…”

An Extensive Tumor Array Analysis Supports Tumor Suppressive Role for Nucleophosmin in Breast Cancer

“…8,9 NPM also regulates the tumor suppressor proteins p53 10 -12 and p14 ARF . [13][14][15] Moreover, NPM protein is post-translationally modified by acetylation, 16 sumoylation, 17,18 ubiquitinylation, 19 and phosphorylation. 20 -23 NPM has been heavily implicated in cancer pathogenesis, but its actual role in oncogenesis is controversial.…”