Negroni A, Venturelli D, Tanno B, Amendola R, Ransac S, Cesi V, Calabretta B, Raschellà G
Section of Toxicology and Biomedical Sciences, Ente Nuove Tecnologie e Ambiente (ENEA) Via Anguillarese 301, 00060 Rome, Italy.
Cell Death Differ. 2000 Sep;7(9):843-50. doi: 10.1038/sj.cdd.4400720.
DR-nm23 belongs to a gene family which includes nm23-H1, originally identified as a candidate metastasis suppressor gene. Nm23 genes are expressed in different tumor types where their levels have been alternatively associated with reduced or increased metastatic potential. Nm23-H1, -H2, DR-nm23 and nm23-H4 all possess NDP kinase activity. Overexpression of DR-nm23 inhibits differentiation and promotes apoptosis in hematopoietic cells. By contrast, it induces morphological and biochemical changes associated with neural differentiation in neuroblastoma cells. In this study, we show that mutations in the catalytic domain and in the serine 61 phosphorylation site, possibly required for protein-protein interactions, impair the ability of DR-nm23 to induce neural differentiation. Moreover, neuroblastoma cells overexpressing wild-type or mutant DR-nm23 are less sensitive to apoptosis triggered by serum withdrawal. By subcellular fractionation, wild-type and mutant DR-nm23 localize in the cytoplasm and prevalently in the mitochondrial fraction. In co-immunoprecipitation experiments, wild-type DR-nm23 binds other members of nm23 family, but mutations in the catalytic and in the RGD domains and in serine 61 inhibit the formation of hetero-multimers. Thus, the integrity of the NDP kinase activity and the presence of a serine residue in position 61 seem essential for the ability of DR-nm23 to trigger differentiation and to bind other Nm23 proteins, but not for the anti-apoptotic effect in neuroblastoma cells. These studies underline the tissue specificity of the biological effects induced by DR-nm23 expression.
DR-nm23属于一个基因家族，该家族包括最初被鉴定为候选转移抑制基因的nm23-H1。Nm23基因在不同肿瘤类型中表达，其水平与转移潜能的降低或增加呈交替相关。Nm23-H1、-H2、DR-nm23和nm23-H4均具有NDP激酶活性。DR-nm23的过表达抑制造血细胞的分化并促进其凋亡。相比之下，它在神经母细胞瘤细胞中诱导与神经分化相关的形态和生化变化。在本研究中，我们表明催化结构域和丝氨酸61磷酸化位点的突变（可能是蛋白质-蛋白质相互作用所必需的）损害了DR-nm23诱导神经分化的能力。此外，过表达野生型或突变型DR-nm23的神经母细胞瘤细胞对血清剥夺触发的凋亡不太敏感。通过亚细胞分级分离，野生型和突变型DR-nm23定位于细胞质中，且主要定位于线粒体部分。在免疫共沉淀实验中，野生型DR-nm23与nm23家族的其他成员结合，但催化结构域、RGD结构域和丝氨酸61的突变抑制了异源多聚体的形成。因此，NDP激酶活性的完整性和61位丝氨酸残基的存在似乎是DR-nm23触发分化和结合其他Nm23蛋白能力所必需的，但对于神经母细胞瘤细胞的抗凋亡作用并非必需。这些研究强调了DR-nm23表达所诱导的生物学效应的组织特异性。
