Necdin is expressed predominantly in postmitotic neurons, and ectopic expression of this protein strongly suppresses cell growth. Necdin has been implicated in the pathogenesis of Prader-Willi syndrome, a human neurodevelopmental disorder associated with genomic imprinting. Here we demonstrate that ectopic expression of necdin induces a neuronal phenotype in neuroblastoma cells. Necdin was undetectable in mouse neuroblastoma N1E-115 cells under undifferentiated and differentiated conditions. N1E-115 cells transfected with necdin cDNA showed morphological differentiation such as neurite outgrowth and expression of the synaptic marker proteins synaptotagmin and synaptophysin. In addition, Western blot analysis of the retinoblastoma protein (Rb) family members Rb, p130, and p107 revealed that necdin cDNA transfectants contained an increased level of p130 and a reduced level of p107, a pattern seen in differentiated G 0 cells. The transcription factors E2F1 and E2F4 physically interacted with necdin via their carboxyl-terminal transactivation domains, but only E2F1 abrogated necdin-induced growth arrest and neurite outgrowth of neuroblastoma cells. Overexpression of E2F1 in differentiated N1E-115 cells induced apoptosis, which was antagonized by co-expression of necdin. These results suggest that necdin promotes the differentiation and survival of neurons through its antagonistic interactions with E2F1.Necdin is a 325-amino acid protein encoded in a cDNA sequence that has been isolated from a subtraction library of neurally differentiated mouse embryonal carcinoma P19 cells (1). The necdin gene is expressed in most of the terminally differentiated neurons, although its expression levels vary among neuronal cell types, being the highest in the hypothalamus and brain stem (2, 3). The human necdin gene is mapped to chromosome 15q11.2-q12, a region deleted in Prader-Willi syndrome (PWS) 1 (4 -6). PWS is a neurodevelopmental disorder associated with genomic imprinting, and its major symptoms such as feeding problems, gross obesity, and hypogonadism are consistent with a hypothalamic defect. Human and mouse necdin genes are maternally imprinted and transcribed only from the paternal allele (4, 5, 7). Necdin is not expressed in the cells prepared from PWS patients whose chromosome 15q11.2-q13 region in the paternal allele is deleted. Disruption of the paternal allele of mouse necdin gene results in early postnatal lethality (8), reduction in specific hypothalamic neurons such as luteinizing hormone-releasing hormone-and oxytocin-containing neurons, and behavioral alternations, which are reminiscent of human PWS (9). Therefore, it is likely that a defect in necdin expression is responsible, at least in part, for various clinical symptoms of PWS. However, little is known about the functional roles of necdin in neuronal differentiation and development.Accumulating evidence has suggested that necdin is a growth suppressor expressed in terminally differentiated cells. Ectopic expression of necdin suppresses the proliferation of...

Necdin is a growth suppressor expressed predominantly in postmitotic neurons. The necdin gene is involved in the etiology of the genomic imprinting-associated neurodevelopmental disorder Prader-Willi syndrome and belongs to the MAGE gene family. All the MAGE family proteins contain a large homology domain termed the MAGE homology domain (MHD). We here characterize the regions of necdin required for the protein-protein interaction, nuclear matrix targeting, and cell growth suppression. The region including entire MHD (amino acids 116-280) of necdin was required for its interaction with p53, while the regions amino acids 144-184 and 191-222 within the MHD were required for both the nuclear matrix targeting and the cell growth suppression of osteosarcoma SAOS-2 cells. The amino-terminal proline-rich acidic region (amino acids 60-100) was also necessary for cell growth suppression. Tetracycline-regulatable overexpression of necdin induced growth arrest of SAOS-2 cells in a reversible manner, and the necdin-overexpressing cells showed a large, flattened morphology with double nuclei. In contrast, a necdin mutant lacking amino acids 191-222 did not induce such changes. These findings suggest that different functions of necdin are mediated via its distinct domains.