Inhibition of Progesterone-induced Xenopus Oocyte Maturation by Nm23 1 Sun Young Kim, James E. Ferrell, Jr., Suhn-Kee Chae, and Kong-Joo Lee 2 Center for Cell Signaling Research, Division of Molecular Life Sciences and College of Pharmacy, Ewha Womans University, Seoul 120-750, Korea [S. Y. K., K-J. L.]; Department of Molecular Pharmacology, Stanford University School of Medicine, Stanford, California 94305- 5332 [J. E. F.]; and Division of Life Sciences, Paichai University, Taejon 302-735, Korea [S-K. C.] Abstract The Nm23 protein has been implicated in a wide variety of biological processes, including suppression of metastasis, phytochrome responses in plants, and regulation of differentiation. Here we examine whether Nm23 is involved in Xenopus laevis oocyte maturation. We found that Nm23 is present in oocytes, indicating that it has the potential to be a regulator of maturation. Furthermore, modest overexpression of Nm23 inhibited progesterone-induced oocyte maturation. This maturation-inhibitory activity was shared by both the acidic Nm23-H1 isoform and the basic Nm23-H2 isoform and by Nm23 mutants that lack nucleoside diphosphate kinase activity (Nm23-H1 H118F and Nm23-H2 H118F). Expression of Nm23 proteins delayed the accumulation of Mos and the activation of p42 mitogen-activated protein kinase (MAPK) in progesterone-treated oocytes but had no discernible effect on Mos-induced p42 MAPK activation. Therefore, Nm23 appears to act upstream of the Mos/mitogen- activated protein/extracellular signal-regulated kinase kinase/p42 MAPK cascade. These findings suggest a novel biological role for Nm23. Introduction Nm23 was originally identified as a possible metastasis sup- pressor gene through the discovery that its expression is associated with low metastatic potential murine melanoma cell lines (1, 2). Nm23 is a member of a gene family that is highly conserved in both prokaryotes and in eukaryotes. Six human Nm23 family members have been identified: (a) Nm23-H1 (3); (b) Nm23-H2 (4); (c) DR-nm23 (5); (d) Nm23-H4 (6); (e) Nm23-H5 (7); and (f) Nm23-H6 (8). Several lines of evidence suggest that Nm23 has a role in regulating differentiation and development. Nm23 is a ho- mologue of the Drosophila awd (abnormal wing disc) gene (9), and mutations in awd are associated with a variety of developmental defects. During mouse embryonic develop- ment, the accumulation of Nm23 protein coincides with dif- ferentiation of the embryonic nervous system, heart, and various epithelial tissues during mouse organogenesis (10), again pointing to a possible role in development. The iden- tification of a differentiation-inhibiting factor (denoted “I fac- tor”) in cell lysates and conditioned media from mouse my- eloid leukemia cells as a member of the Nm23 family suggests a role for Nm23 in suppressing differentiation (11). Nm23 expression decreases during the differentiation of he- matopoietic cells (12), again suggesting a role as an antidif- ferentiation factor. In other contexts, Nm23 may promote differentiation. For example, overexpression of Nm23 pro- motes neurite outgrowth in PC12 cells (13, 14). The best-characterized Nm23 proteins exist as homo- or hetero-hexamers of two 152-amino acid isoforms, the acidic Nm23-H1 protein and the basic Nm23-H2 protein (15). These and other Nm23 proteins possess NDP 3 kinase activity, cat- alyzing the transfer of the terminal phosphate of nucleoside triphosphates to NDPs. In some contexts, the NDP kinase activity of Nm23 appears to be critical for its biological ef- fects. For example, whereas wild-type Nm23 promotes neu- rite outgrowth, a point mutant of Nm23 (H118A) that lacks NDP kinase activity does not promote neurite outgrowth and, in fact, suppresses nerve growth factor-induced neurite out- growth (13, 14). However, in other contexts, the NDP kinase activity of Nm23 appears to be unimportant. Mutated Nm23 proteins lacking NDP kinase activity still score as the differentiation- inhibiting “I factor” mentioned above (16). In addition, Nm23-H2 can bind to a nuclease hypersensitive element on the human c-myc promoter and transactivate gene expression in vitro and in vivo (17, 18), and this transactivation is independent of NDP kinase activity (19). Thus, some Nm23 functions appear to be independent of its NDP kinase activity. Nm23 proteins also function as histidine/aspartic acid-specific protein ki- nases (20) and as serine/threonine-specific protein kinases (21) and have been proposed to link receptors to G proteins (22). However, it is not yet clear how relevant any of these activities are for the biological effects of Nm23, nor is it clear where Nm23 exerts its effects. It may act in the nucleus (as Received 5/10/00; revised 7/7/00; accepted 7/10/00. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indi- cate this fact. 1 Supported by MOE Grant 1998-019-F00070, by KOSEF through the Center for Cell Signaling Research at Ewha Womans University, and by NIH Grant GM46383. S. Y. K. is supported financially by the Brain Korea 21 Project. 2 To whom requests for reprints should be addressed, at Center for Cell Signaling Research, Division of Molecular Life Sciences and College of Pharmacy, Ewha Womans University, Seoul 120-750, Korea. Phone: 82- 2-3277-3038; Fax: 82-2-3277-3760; E-mail: kjl@mm.ewha.ac.kr. 3 The abbreviations used are: NDP, nucleoside diphosphate; GVBD, ger- minal vesicle breakdown; MAPK, mitogen-activated protein kinase; MEK, mitogen-activated protein/extracellular signal-regulated kinase kinase; cAMP, cyclic AMP. 485 Vol. 11, 485– 490, September 2000 Cell Growth & Differentiation