Nek1 shares structural and functional similarities with NIMA kinase

AbstractThe Aspergillus NIMA serine/threonine kinase plays a pivotal role in controlling entrance into mitosis. A major function attributed to NIMA is the induction of chromatin condensation. We show here that the founder murine NIMA-related kinase, Nek1, is larger than previously reported, and that the full-length protein conserves the structural hallmarks of NIMA. Even though Nek1 bears two classical nuclear localization signals (NLS), the endogenous protein localizes to the cytoplasm. Ectopic overexpression of various Nek1 constructs suggests that the C-terminus of Nek1 bears cytoplasmic localization signal(s). Overexpression of nuclear constructs of Nek1 resulted in abnormal chromatin condensation, with the DNA mainly confined to the periphery of the nucleus. Advanced condensation phenotype was associated with nuclear pore complex dispersal. The condensation was not accompanied by up-regulation of mitotic or apoptotic markers. A similar phenotype has been described following NIMA overexpression, strengthening the notion that the mammalian Nek1 kinase has functional similarity to NIMA

A Novel Role for Microphthalmia-Associated Transcription Factor–Regulated Pigment Epithelium-Derived Factor during Melanoma Progression

Microphthalmia-associated transcription factor (MITF) acts via pigment epithelium-derived factor (PEDF), an antiangiogenic protein, to regulate retinal pigment epithelium migration. PEDF expression and/or regulation during melanoma development have not been investigated previously. Using immunohistochemistry, we determined expression of PEDF in common and dysplastic melanocytic nevi, melanoma in situ, invasive melanoma, and metastatic melanoma (n = 102). PEDF expression was consistently decreased in invasive and metastatic melanoma, compared with nevi and melanoma in situ (P < 0.0001). PEDF was lost in thicker melanomas (P = 0.003), and correlated with depth of invasion (P = 0.003) and distant metastasis (P = 0.0331), but only marginally with mitotic index, AJCC stage, nodal metastasis, or blood vascular density (0.05 < P < 0.10). Quantitative real-time PCR and microarray analyses confirmed PEDF down-regulation at the mRNA level in several melanoma lines, compared with melanocytes. MITF positively correlated with PEDF expression in invasive melanomas (P = 0.0003). Searching for PEDF regulatory mechanisms revealed two occupied conserved E-boxes (DNA recognition elements) in the first intron of the human and mouse PEDF promoter regions, confirmed by binding assays. Dominant-negative and siRNA approaches in vivo demonstrated direct transcriptional influence of MITF on PEDF, establishing the PEDF gene (SERPINF1) as a MITF target in melanocytes and melanoma cells. These findings suggest that loss of PEDF expression promotes early invasive melanoma growth.ISSN:0002-9440ISSN:1525-219