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1.
Anticancer Res ; 39(5): 2251-2258, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31092416

RESUMO

Cancer is characterized by uncontrolled cell proliferation due to the aberrant activity of various proteins. Cell cycle-related proteins are thought to be important in several functions, such as proliferation, invasion and drug resistance in human malignancies. Never in mitosis gene A-related kinase 2 (NEK2) is a cell cycle-related protein. NEK2 is highly expressed in various tumor types and cancer cell lines. NEK2 expression is correlated with rapid relapse and poor outcome in multiple cancer types. Several researchers have demonstrated that NEK2 inhibition results in anticancer effects against many types of cancers, both in vitro and in vivo. Recent research strongly indicates the advantages of NEK2-targeted therapy for cancer. This review focuses on the current understanding of NEK2 in cancer and the rationale of a xenograft cancer model for cancer treatment. A possible therapeutic strategy, such as inhibitor and nucleic acid medicine targeting of NEK2, is also discussed.


Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Quinases Relacionadas a NIMA/genética , Neoplasias/genética , Animais , Ciclo Celular/genética , Proliferação de Células/genética , Humanos , Camundongos , Mitose/genética , Quinases Relacionadas a NIMA/antagonistas & inibidores , Quinases Relacionadas a NIMA/uso terapêutico , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
2.
Int J Oncol ; 54(4): 1295-1305, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30968157

RESUMO

The mechanisms through which cancer­upregulated gene 2 (CUG2), a novel oncogene, affects Wnt/ß­catenin signaling, essential for tumorigenesis, are unclear. In this study, we aimed to elucidate some of these mechanisms in A549 lung cancer cells. Under the overexpression of CUG2, the protein levels and activity of ß­catenin were evaluated by western blot analysis and luciferase assay. To examine a biological consequence of ß­catenin under CUG2 overexpression, cell migration, invasion and sphere formation assay were performed. The upregulation of ß­catenin induced by CUG2 overexpression was also accessed by xenotransplantation in mice. We first found that CUG2 overexpression increased ß­catenin expression and activity. The suppression of ß­catenin decreased cancer stem cell (CSC)­like phenotypes, indicating that ß­catenin is involved in CUG2­mediated CSC­like phenotypes. Notably, CUG2 overexpression increased the phosphorylation of ß­catenin at Ser33/Ser37, which is known to recruit E3 ligase for ß­catenin degradation. Moreover, CUG2 interacted with and enhanced the expression and kinase activity of never in mitosis gene A­related kinase 2 (NEK2). Recombinant NEK2 phosphorylated ß­catenin at Ser33/Ser37, while NEK2 knockdown decreased the phosphorylation of ß­catenin, suggesting that NEK2 is involved in the phosphorylation of ß­catenin at Ser33/Ser37. Treatment with CGK062, a small chemical molecule, which promotes the phosphorylation of ß­catenin at Ser33/Ser37 through protein kinase C (PKC)α to induce its degradation, reduced ß­catenin levels and inhibited the CUG2­induced features of malignant tumors, including increased cell migration, invasion and sphere formation. Furthermore, CGK062 treatment suppressed CUG2­mediated tumor formation in nude mice. Taken together, the findings of this study suggest that CUG2 enhances the phosphorylation of ß­catenin at Ser33/Ser37 by activating NEK2, thus stabilizing ß­catenin. CGK062 may thus have potential for use as a therapeutic drug against CUG2­overexpressing lung cancer cells.


Assuntos
Carcinogênese/efeitos dos fármacos , Proteínas Cromossômicas não Histona/metabolismo , Quinases Relacionadas a NIMA/metabolismo , Neoplasias/tratamento farmacológico , beta Catenina/metabolismo , Células A549 , Acrilatos/farmacologia , Acrilatos/uso terapêutico , Animais , Carcinogênese/patologia , Cromanos/farmacologia , Cromanos/uso terapêutico , Feminino , Humanos , Masculino , Camundongos , Camundongos Nus , Quinases Relacionadas a NIMA/genética , Neoplasias/patologia , Fosforilação/efeitos dos fármacos , Estabilidade Proteica/efeitos dos fármacos , RNA Interferente Pequeno/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , beta Catenina/genética
3.
Biosci Rep ; 39(1)2019 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-30578380

RESUMO

A consensus about the prognostic role of NIMA-related kinase 2 (NEK2) expression in various solid tumors has not been made yet. Thus, this meta-analysis aimed to systematically assess the prognostic role of NEK2 expression in patients with solid tumors. The eligible studies were identified through searching PubMed, Web of Science, and EMBASE. The hazard ratios (HRs) with their corresponding 95% confidence intervals (CIs) were used to evaluate the link between NEK2 overexpression and overall survival (OS) and disease-free survival/recurrence-free survival (DFS/RFS) of patients with solid tumors. A total of 17 studies with 4897 patients were included in this meta-analysis. Among these studies, all of them explored the association between NEK2 expression and OS of patients with solid tumors. Our pooled analysis indicated that NEK2 overexpression was significantly related to adverse OS (HR = 1.66; 95% CI: 1.38-2.00; P = 0.001). Additionally, there were six studies with 854 patients that investigated the association between NEK2 expression and DFS/RFS. Our pooled result indicated that there was a substantial relationship between NEK2 overexpression and poorer DFS/RFS (HR = 2.00; 95% CI: 1.61-2.48; P = 0.003). In conclusion, our meta-analysis indicated that NEK2 may be a useful predictor of prognosis and an effective therapeutic target in solid tumors. Nevertheless, more high-quality studies are warranted to further support our conclusions because of several limitations in our meta-analysis.


Assuntos
Biomarcadores Tumorais/genética , Quinases Relacionadas a NIMA/genética , Neoplasias/genética , Prognóstico , Intervalo Livre de Doença , Humanos , Neoplasias/patologia
4.
EBioMedicine ; 40: 77-91, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30594554

RESUMO

BACKGROUND: Meningioma is the most frequent primary intracranial tumour. Surgical resection remains the main therapeutic option as pharmacological intervention is hampered by poor knowledge of their proteomic signature. There is an urgent need to identify new therapeutic targets and biomarkers of meningioma. METHODS: We performed proteomic profiling of grade I, II and III frozen meningioma specimens and three normal healthy human meninges using LC-MS/MS to analyse global proteins, enriched phosphoproteins and phosphopeptides. Differential expression and functional annotation of proteins was completed using Perseus, IPA® and DAVID. We validated differential expression of proteins and phosphoproteins by Western blot on a meningioma validation set and by immunohistochemistry. FINDINGS: We quantified 3888 proteins and 3074 phosphoproteins across all meningioma grades and normal meninges. Bioinformatics analysis revealed commonly upregulated proteins and phosphoproteins to be enriched in Gene Ontology terms associated with RNA metabolism. Validation studies confirmed significant overexpression of proteins such as EGFR and CKAP4 across all grades, as well as the aberrant activation of the downstream PI3K/AKT pathway, which seems differential between grades. Further, we validated upregulation of the total and activated phosphorylated form of the NIMA-related kinase, NEK9, involved in mitotic progression. Novel proteins identified and validated in meningioma included the nuclear proto-oncogene SET, the splicing factor SF2/ASF and the higher-grade specific protein, HK2, involved in cellular metabolism. INTERPRETATION: Overall, we generated a proteomic thesaurus of meningiomas for the identification of potential biomarkers and therapeutic targets. FUND: This study was supported by Brain Tumour Research.


Assuntos
Meningioma/metabolismo , Fosfoproteínas/metabolismo , Proteoma , Proteômica , Linhagem Celular Tumoral , Cromatografia Líquida , Biologia Computacional/métodos , Regulação Neoplásica da Expressão Gênica , Chaperonas de Histonas/genética , Chaperonas de Histonas/metabolismo , Humanos , Cinesina/genética , Cinesina/metabolismo , Meningioma/genética , Meningioma/patologia , Mutação , Quinases Relacionadas a NIMA/genética , Quinases Relacionadas a NIMA/metabolismo , Fosfopeptídeos/metabolismo , Proteômica/métodos , Estabilidade de RNA , Reprodutibilidade dos Testes , Espectrometria de Massas em Tandem , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Microambiente Tumoral/genética
5.
Proc Natl Acad Sci U S A ; 115(40): E9371-E9380, 2018 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-30232264

RESUMO

The NLRP3 inflammasome is an important regulator of inflammation and immunity. It is a multimolecular platform formed within cells that facilitates the activation of proinflammatory caspases to drive secretion of cytokines such as interleukin-1ß (IL-1ß). Knowledge of the mechanisms regulating formation of the NLRP3 inflammasome is incomplete. Here we report Cl- channel-dependent formation of dynamic ASC oligomers and inflammasome specks that remain inactive in the absence of K+ efflux. Formed after Cl- efflux exclusively, ASC specks are NLRP3 dependent, reversible, and inactive, although they further prime inflammatory responses, accelerating and enhancing release of IL-1ß in response to a K+ efflux-inducing stimulus. NEK7 is a specific K+ sensor and does not associate with NLRP3 under conditions stimulating exclusively Cl- efflux, but does after K+ efflux, activating the complex driving inflammation. Our investigation delivers mechanistic understanding into inflammasome activation and the regulation of inflammatory responses.


Assuntos
Proteínas Adaptadoras de Sinalização CARD/metabolismo , Cloretos/metabolismo , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Multimerização Proteica , Animais , Proteínas Adaptadoras de Sinalização CARD/genética , Feminino , Inflamassomos/genética , Inflamação/genética , Inflamação/metabolismo , Inflamação/patologia , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Transporte de Íons/genética , Masculino , Camundongos , Camundongos Knockout , Quinases Relacionadas a NIMA/genética , Quinases Relacionadas a NIMA/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Potássio/metabolismo
6.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 34(3): 247-252, 2018 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-29773107

RESUMO

Objective To observe the effect of microRNA-151a-3p (miR-151a-3p) up-regulation on the proliferation and migration of prostate cancer cells and explore the possible molecular mechanism. Methods The expression of miR-151a-3p in PC-3M, C4-2B, 22RV1, DU-145, PC-3, LNCap human prostate cancer cells and RWPE-1 human normal prostate epithelial cells was detected by real-time fluorescence quantitative PCR. PC-3 cells with the lowest expression of miR-151a-3p were used for subsequent experiments. Bioinformatics and dual-luciferase reporter assay were performed to predict and test potential target genes of miR-151a-3p. The miR-151a-3p mimics or negative control microRNAs (miR-NCs) were transfected into PC-3 cells. Real-time fluorescence quantitative PCR was used to detect the expression of miR-151a-3p and potential target gene mRNA. The protein expressions of target genes and downstream signaling pathway proteins were analyzed by Western blotting. The proliferation of PC-3 cells was examined by MTT assay, and the migration of PC-3 cells was detected by TranswellTM assay. Results The expression level of miR-151a-3p in the prostate cancer cells was significantly lower than that in RWPE-1 normal human prostate epithelial cells. PC-3 cells had the lowest expression level of miR-151a-3p. The bioinformatics and dual-luciferase reporter assay showed that NEK2 was the potential target gene for miR-151a-3p. After transfection with miR-151a-3p mimics, the expression of miR-151a-3p in PC-3 cells significantly increased and the expression of NEK2 mRNA significantly decreased. The protein expressions of PI3K-AKT-mTOR signaling pathway were also reduced. Up-regulation of miR-151a-3p significantly inhibited the proliferation and migration of PC-3 cells. Conclusion The expression of miR-151a-3p is reduced in prostate cancer cells. Up-regulation of miR-151a-3p can inhibit the proliferation and migration of P-3 in prostate cancer by decreasing the expression of NEK2 and PI3K-AKT-mTOR signaling pathway proteins.


Assuntos
Movimento Celular , Proliferação de Células , MicroRNAs/genética , Neoplasias da Próstata/fisiopatologia , Apoptose , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , MicroRNAs/metabolismo , Quinases Relacionadas a NIMA/genética , Quinases Relacionadas a NIMA/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo
7.
PLoS Genet ; 14(4): e1007313, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29608564

RESUMO

Molting is an essential process in the nematode Caenorhabditis elegans during which the epidermal apical extracellular matrix, termed the cuticle, is detached and replaced at each larval stage. The conserved NIMA-related kinases NEKL-2/NEK8/NEK9 and NEKL-3/NEK6/NEK7, together with their ankyrin repeat partners, MLT-2/ANKS6, MLT-3/ANKS3, and MLT-4/INVS, are essential for normal molting. In nekl and mlt mutants, the old larval cuticle fails to be completely shed, leading to entrapment and growth arrest. To better understand the molecular and cellular functions of NEKLs during molting, we isolated genetic suppressors of nekl molting-defective mutants. Using two independent approaches, we identified CDC-42, a conserved Rho-family GTPase, and its effector protein kinase, SID-3/ACK1. Notably, CDC42 and ACK1 regulate actin dynamics in mammals, and actin reorganization within the worm epidermis has been proposed to be important for the molting process. Inhibition of NEKL-MLT activities led to strong defects in the distribution of actin and failure to form molting-specific apical actin bundles. Importantly, this phenotype was reverted following cdc-42 or sid-3 inhibition. In addition, repression of CDC-42 or SID-3 also suppressed nekl-associated defects in trafficking, a process that requires actin assembly and disassembly. Expression analyses indicated that components of the NEKL-MLT network colocalize with both actin and CDC-42 in specific regions of the epidermis. Moreover, NEKL-MLT components were required for the normal subcellular localization of CDC-42 in the epidermis as well as wild-type levels of CDC-42 activation. Taken together, our findings indicate that the NEKL-MLT network regulates actin through CDC-42 and its effector SID-3. Interestingly, we also observed that downregulation of CDC-42 in a wild-type background leads to molting defects, suggesting that there is a fine balance between NEKL-MLT and CDC-42-SID-3 activities in the epidermis.


Assuntos
Actinas/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Quinases Relacionadas a NIMA/metabolismo , Proteínas Tirosina Quinases/metabolismo , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans/genética , Caenorhabditis elegans/crescimento & desenvolvimento , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ciclo Celular/genética , Endocitose , Epiderme/metabolismo , Proteínas de Ligação ao GTP/genética , Microscopia Confocal , Muda/genética , Mutação , Quinases Relacionadas a NIMA/genética , Transporte Proteico , Proteínas Tirosina Quinases/genética , Interferência de RNA , Transdução de Sinais/genética
8.
Medicine (Baltimore) ; 97(3): e9630, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29504992

RESUMO

The NIMA-related kinase 3 (NEK3) plays an important role in cell migration, cell proliferation, and cell viability. Recently, NEK3 was reported to enhance the malignancy of breast cancer. However, its role in gastric cancer has not been completely characterized. In this study, we explored the prognostic significance of NEK3 in human gastric cancer. Reverse transcription-polymerase chain reaction and western blot were performed to detect the NEK3 mRNA and protein expression in 6 paired fresh human gastric cancer tissues and surrounding normal tissues. NEK3 levels in gastric cancer and its adjacent normal samples of 168 cases were detected by immunohistochemistry, and the relationships between the NEK3 level and various clinicopathological features were analyzed. NEK3 mRNA and protein were significantly overexpressed in gastric cancer tissues, compared with adjacent normal tissues. Immunohistochemistry staining assay showed the percentage of high NEK3 expression in gastric cancer samples was higher than that in adjacent normal samples. NEK3 overexpression was significantly correlated with pT stage, pathologic TNM stage, lymph node metastasis, and poor prognosis of gastric cancer. Cox multivariate regression analyses suggested that NEK3 was an independent prognostic factor for survival of patients with gastric cancer. The data demonstrate that NEK3 is overexpressed in gastric cancer, which promotes the malignancy of gastric cancer. NEK3 may be as a prognostic biomarker and a potential therapeutic target for gastric cancer.


Assuntos
Quinases Relacionadas a NIMA/genética , Neoplasias Gástricas/genética , Neoplasias Gástricas/mortalidade , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Quinases Relacionadas a NIMA/biossíntese , Prognóstico , Neoplasias Gástricas/metabolismo , Taxa de Sobrevida
9.
Development ; 145(5)2018 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-29440300

RESUMO

Tip growth is driven by turgor pressure and mediated by the polarized accumulation of cellular materials. How a single polarized growth site is established and maintained is unclear. Here, we analyzed the function of NIMA-related protein kinase 1 (MpNEK1) in the liverwort Marchantia polymorpha In the wild type, rhizoid cells differentiate from the ventral epidermis and elongate through tip growth to form hair-like protrusions. In Mpnek1 knockout mutants, rhizoids underwent frequent changes in growth direction, resulting in a twisted and/or spiral morphology. The functional MpNEK1-Citrine protein fusion localized to microtubule foci in the apical growing region of rhizoids. Mpnek1 knockouts exhibited increases in both microtubule density and bundling in the apical dome of rhizoids. Treatment with the microtubule-stabilizing drug taxol phenocopied the Mpnek1 knockout. These results suggest that MpNEK1 directs tip growth in rhizoids through microtubule organization. Furthermore, MpNEK1 expression rescued ectopic outgrowth of epidermal cells in the Arabidopsis thaliana nek6 mutant, strongly supporting an evolutionarily conserved NEK-dependent mechanism of directional growth. It is possible that such a mechanism contributed to the evolution of the early rooting system in land plants.


Assuntos
Marchantia , Quinases Relacionadas a NIMA/fisiologia , Rizoma/crescimento & desenvolvimento , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Sequência Conservada , Embriófitas , Evolução Molecular , Regulação da Expressão Gênica de Plantas , Marchantia/genética , Marchantia/crescimento & desenvolvimento , Quinase 1 Relacionada a NIMA/genética , Quinases Relacionadas a NIMA/genética , Desenvolvimento Vegetal/genética , Plantas Geneticamente Modificadas , Rizoma/genética
10.
Exp Cell Res ; 362(2): 412-423, 2018 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-29225051

RESUMO

Never in mitosis A-related kinase 2A (Nek2A), a centrosomal serine/threonine kinase, is involved in mitotic progression by regulating the centrosome cycle. Particularly, Nek2A is necessary for dissolution of the intercentriole linkage between the duplicated centrosomes prior to mitosis. Nek2A activity roughly parallels its cell cycle-dependent expression levels, but the precise mechanism regulating its activity remains unclear. In this study, we found that γ-taxilin co-localized with Nek2A at the centrosome during interphase and interacted with Nek2A in yeast two-hybrid and pull-down assays and that γ-taxilin regulated centrosome disjunction in a Nek2A-dependent manner. γ-Taxilin depletion increased the number of cells with striking splitting of centrosomes. The precocious splitting of centrosomes induced by γ-taxilin depletion was attenuated by Nek2A depletion, suggesting that γ-taxilin depletion induces the Nek2A-mediated dissolution of the intercentriole linkage between the duplicated centrosomes nevertheless mitosis does not yet begin. Taken together with the result that γ-taxilin protein expression levels were decreased at the onset of mitosis, we propose that γ-taxilin participates in Nek2A-mediated centrosome disjunction as a negative regulator through its interaction with Nek2A.


Assuntos
Centríolos/genética , Centrossomo , Quinases Relacionadas a NIMA/genética , Proteínas de Transporte Vesicular/genética , Proteínas de Ciclo Celular/genética , Regulação da Expressão Gênica/genética , Células HeLa , Humanos , Mitose/genética , Proteínas Serina-Treonina Quinases/genética , Técnicas do Sistema de Duplo-Híbrido
11.
BMC Cell Biol ; 18(1): 33, 2017 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-29141582

RESUMO

BACKGROUND: The primary cilium is an extension of the cell membrane that encloses a microtubule-based axoneme. Primary cilia are essential for transmission of environmental cues that determine cell fate. Disruption of primary cilia function is the molecular basis of numerous developmental disorders. Despite their biological importance, the mechanisms governing their assembly and disassembly are just beginning to be understood. Cilia growth and disassembly are essential events when cells exit and reenter into the cell cycle. The kinases never in mitosis-kinase 2 (Nek2) and Aurora A (AurA) act to depolymerize cilia when cells reenter the cell cycle from G0. RESULTS: Coexpression of either kinase with its kinase dead companion [AurA with kinase dead Nek2 (Nek2 KD) or Nek2 with kinase dead AurA (AurA KD)] had different effects on cilia depending on whether cilia are growing or shortening. AurA and Nek2 are individually able to shorten cilia when cilia are growing but both are required when cilia are being absorbed. The depolymerizing activity of each kinase is increased when coexpressed with the kinase dead version of the other kinase but only when cilia are assembling. Additionally, the two kinases act additively when cilia are assembling but not disassembling. Inhibition of AurA increases cilia number while inhibition of Nek2 significantly stimulates cilia length. The complex functional relationship between the two kinases reflects their physical interaction. Further, we identify a role for a PP1 binding protein, PPP1R42, in inhibiting Nek2 and increasing ciliation of ARPE-19 cells. CONCLUSION: We have uncovered a novel functional interaction between Nek2 and AurA that is dependent on the growth state of cilia. This differential interdependence reflects opposing regulation when cilia are growing or shortening. In addition to interaction between the kinases to regulate ciliation, the PP1 binding protein PPP1R42 directly inhibits Nek2 independent of PP1 indicating another level of regulation of this kinase. In summary, we demonstrate a complex interplay between Nek2 and AurA kinases in regulation of ciliation in ARPE-19 cells.


Assuntos
Aurora Quinase A/metabolismo , Cílios/enzimologia , Proteínas dos Microtúbulos/metabolismo , Quinases Relacionadas a NIMA/metabolismo , Receptores de Neuropeptídeo Y/agonistas , Aurora Quinase A/antagonistas & inibidores , Aurora Quinase A/genética , Azepinas/farmacologia , Linhagem Celular , Cílios/efeitos dos fármacos , Células Epiteliais/citologia , Células Epiteliais/enzimologia , Células Epiteliais/metabolismo , Humanos , Quinases Relacionadas a NIMA/genética , Ligação Proteica/fisiologia , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/farmacologia
12.
Cancer Res ; 77(18): 4785-4796, 2017 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-28720575

RESUMO

Cancer cells frequently possess extra amplified centrosomes clustered into two poles whose pseudo-bipolar spindles exhibit reduced fidelity of chromosome segregation and promote genetic instability. Inhibition of centrosome clustering triggers multipolar spindle formation and mitotic catastrophe, offering an attractive therapeutic approach to selectively kill cells with amplified centrosomes. However, mechanisms of centrosome clustering remain poorly understood. Here, we identify a new pathway that acts through NIMA-related kinase 6 (Nek6) and Hsp72 to promote centrosome clustering. Nek6, as well as its upstream activators polo-like kinase 1 and Aurora-A, targeted Hsp72 to the poles of cells with amplified centrosomes. Unlike some centrosome declustering agents, blocking Hsp72 or Nek6 function did not induce formation of acentrosomal poles, meaning that multipolar spindles were observable only in cells with amplified centrosomes. Inhibition of Hsp72 in acute lymphoblastic leukemia cells resulted in increased multipolar spindle frequency that correlated with centrosome amplification, while loss of Hsp72 or Nek6 function in noncancer-derived cells disturbs neither spindle formation nor mitotic progression. Hence, the Nek6-Hsp72 module represents a novel actionable pathway for selective targeting of cancer cells with amplified centrosomes. Cancer Res; 77(18); 4785-96. ©2017 AACR.


Assuntos
Neoplasias da Mama/patologia , Centrossomo/patologia , Proteínas de Choque Térmico HSP72/metabolismo , Neuroblastoma/patologia , Animais , Apoptose , Aurora Quinase A/genética , Aurora Quinase A/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proliferação de Células , Centrossomo/metabolismo , Feminino , Proteínas de Choque Térmico HSP72/genética , Humanos , Camundongos , Mitose/fisiologia , Quinases Relacionadas a NIMA/genética , Quinases Relacionadas a NIMA/metabolismo , Neuroblastoma/genética , Neuroblastoma/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Fuso Acromático/metabolismo , Fuso Acromático/patologia , Células Tumorais Cultivadas
13.
Kidney Int ; 92(6): 1544-1554, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28754558

RESUMO

Polycystic kidney disease (PKD) is among the leading causes of end-stage renal disease. Increasing evidence exists that molecular therapeutic strategies targeted to cyst formation and growth might be more efficacious in early disease stages, highlighting the growing need for sensitive biomarkers. Here we apply quantitative magnetic resonance imaging techniques of T2 mapping and diffusion-weighted imaging in the jck mouse model for PKD using a clinical 3.0 T scanner. We tested whether kidney T2 values and the apparent diffusion coefficient (ADC) are superior to anatomical imaging parameters in the detection of early cystogenesis, as shown on macro- and histopathology. We also tested whether kidney T2 values and ADC have the potential to monitor early treatment effects of therapy with the V2 receptor antagonist Mozavaptane. Kidney T2 values and to a lesser degree ADC were found to be highly sensitive markers of early cystogenesis and superior to anatomical-based imaging parameters. Furthermore, kidney T2 values exhibited a nearly perfect correlation to the histological cystic index, allowing a clear separation of the two mouse genotypes. Additionally, kidney T2 values and ADC were able to monitor early treatment effects in the jck mouse model in a proof-of-principle experiment. Thus, given the superiority of kidney T2 values and ADC over anatomical-based imaging in mice, further studies are needed to evaluate the translational impact of these techniques in patients with PKD.


Assuntos
Antagonistas dos Receptores de Hormônios Antidiuréticos/uso terapêutico , Benzazepinas/uso terapêutico , Cistos/diagnóstico por imagem , Rim/diagnóstico por imagem , Doenças Renais Policísticas/diagnóstico por imagem , Adulto , Animais , Cistos/tratamento farmacológico , Cistos/genética , Cistos/patologia , Imagem de Difusão por Ressonância Magnética/métodos , Modelos Animais de Doenças , Diagnóstico Precoce , Feminino , Humanos , Processamento de Imagem Assistida por Computador , Rim/patologia , Estudos Longitudinais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Terapia de Alvo Molecular/métodos , Mutação , Quinases Relacionadas a NIMA/genética , Doenças Renais Policísticas/tratamento farmacológico , Doenças Renais Policísticas/genética , Doenças Renais Policísticas/patologia , Estudo de Prova de Conceito , Fatores de Tempo , Resultado do Tratamento , Adulto Jovem
14.
J Cell Biol ; 216(8): 2339-2354, 2017 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-28630147

RESUMO

In mitosis, cells undergo a precisely orchestrated series of spatiotemporal changes in cytoskeletal structure to divide their genetic material. These changes are coordinated by a sophisticated network of protein-protein interactions and posttranslational modifications. In this study, we report a bifurcation in a signaling cascade of the NIMA-related kinases (Neks) Nek6, Nek7, and Nek9 that is required for the localization and function of two kinesins essential for cytokinesis, Mklp2 and Kif14. We demonstrate that a Nek9, Nek6, and Mklp2 signaling module controls the timely localization and bundling activity of Mklp2 at the anaphase central spindle. We further show that a separate Nek9, Nek7, and Kif14 signaling module is required for the recruitment of the Rho-interacting kinase citron to the anaphase midzone. Our findings uncover an anaphase-specific function for these effector kinesins that is controlled by specific Nek kinase signaling modules to properly coordinate cytokinesis.


Assuntos
Anáfase , Citocinese , Cinesina/metabolismo , Quinases Relacionadas a NIMA/metabolismo , Fuso Acromático/enzimologia , Trifosfato de Adenosina/metabolismo , Cromatografia de Afinidade , Células HeLa , Humanos , Hidrólise , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Cinesina/genética , Quinases Relacionadas a NIMA/genética , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/metabolismo , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Interferência de RNA , Transdução de Sinais , Espectrometria de Massas em Tandem , Fatores de Tempo , Transfecção
15.
Tumour Biol ; 39(5): 1010428317699754, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28475000

RESUMO

NIMA-related kinase 2B has been known to be an important centrosome regulatory factor. The aim of this study was to investigate the effect of NIMA-related kinase 2B on the sensitivity of breast cancer to paclitaxel. We detected the expression of NIMA-related kinase 2B messenger RNA in MCF-10 cells, including MCF-10A, MCF-10AT, MCF-10DCIS.com , and MCF-10CA1a. The influence of NIMA-related kinase 2B in nude mouse was also detected. The association between NIMA-related kinase 2B and clinicopathological factors was explored in invasive ductal carcinoma tissues. NIMA-related kinase 2B was lowly expressed in the precancerous cells, MCF-10A and MCF-10AT, and it was highly expressed in carcinomatous cells, MCF-10DCIS.com and MCF-10CA1a. The upregulation of NIMA-related kinase 2B can introduce the growth of MCF-10AT cells, knockdown of NIMA-related kinase 2B could remarkably inhibit cell proliferation in MCF-10DCIS.com and MCF-10 CA1a cells. Comparing the volume of the xenografts in nude mouse, we found that the tumors treated by NIMA-related kinase 2B small interfering RNA associated with paclitaxel were the smallest among all the groups. Expression of NIMA-related kinase 2B messenger RNA was associated with higher histological grades, positive lymph node, and high Ki67 index (>20%). The partial response rates were 75.0% in NIMA-related kinase 2B negative (NIMA-related kinase 2B-) patients and 15.8% in NIMA-related kinase 2B++ patients. The progressive disease rates were 10.0% in NIMA-related kinase 2B- patients and 52.6% in NIMA-related kinase 2B++ patients ( p = 0.002). Our findings suggested that NIMA-related kinase 2B could play a role in the development and progression of breast cancer. Combination treatment using NIMA-related kinase 2B small interfering RNA and paclitaxel might be a novel potential therapy method for breast cancer.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Quinases Relacionadas a NIMA/genética , Paclitaxel/administração & dosagem , Idoso , Animais , Apoptose/efeitos dos fármacos , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos , Pessoa de Meia-Idade , Quinases Relacionadas a NIMA/biossíntese , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Thorac Cancer ; 8(4): 304-311, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28514100

RESUMO

BACKGROUND: MicroRNA-128 (miR-128) serves as a regulator by inducing cancer cell apoptosis, differentiation, the epithelial-to-mesenchymal transition process, and tumor growth by mediating different targets. NIMA-related kinase 2 (NEK2) is aberrantly expressed in lung cancer. The miR-128/NEK2 pathway has been reported to predict prognosis in colorectal cancer; however, the determination of a relationship between miR-128 and NEK2 in lung cancer has remained elusive. We explored the association between miR-128 and NEK2 in lung cancer. METHODS: MiR-128 and NEK2 expression were examined in 15 lung cancer tissues by real time-PCR. Lung cancer SK-MES-1 cells were transfected with miR-128 mimic, an inhibitor or a negative control. MiR-128 and NEK2 expression levels were detected using quantitative real time-PCR and Western blot. SK-MES-1 cell apoptosis was performed by flow cytometry. RESULTS: Compared to adjacent non-tumor tissues, miR-128 was downregulated and NEK2 was upregulated in 15 lung cancer tissues. Lung cancer SK-MES-1 cells transfected with miR-128 mimic induced a higher apoptotic rate than those transfected with the negative control. Dual luciferase assay further confirmed that NEK2 was a direct target of miR-128 in lung cancer, and transfection with miR-128 mimic could decrease the NEK2 protein level while the miR-128 inhibitor increased NEK2 expression. Finally, the apoptotic effect of lung cancer cells induced by miR-128 mimic could be reversed by NEK2 overexpression. CONCLUSIONS: NEK2 was regulated by miR-128 in lung cancer and miR-128 induced lung cancer cell apoptosis by mediating NEK2 expression.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , MicroRNAs/genética , Quinases Relacionadas a NIMA/genética , Regiões 3' não Traduzidas , Apoptose , Regulação Neoplásica da Expressão Gênica , Humanos , Prognóstico , Análise de Sobrevida
17.
Gene ; 625: 72-77, 2017 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-28479381

RESUMO

Pancreatic neuroendocrine tumors are relatively rare pancreatic neoplasms over the world. Investigations about molecular biology of PNETs are insufficient for nowadays. We aimed to explore the expression of messenger RNA and regulatory processes underlying pancreatic neuroendocrine tumors from different views. The expression profile of GSE73338 were downloaded, including samples with pancreatic neuroendocrine tumors. First, the Limma package was utilized to distinguish the differentially expressed messenger RNA. Gene Ontology classification and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis were performed to explore the functions and pathways of target genes. In addition, we constructed a protein-protein interaction network. NEK2, UBE2C, TOP2A and PPP1R1A were revealed with continuous genomic alterations in higher tumor stage. 91 up-regulated and 36 down-regulated genes were identified to be differentially expressed in malignant PNETs. Locomotory behavior was significantly enriched for biological processes of metastasis PNETs. GCGR and GNAS were identified as the hub of proteins in the protein-protein interaction sub-network of malignant PNETs. We showed the gene expression differences in PNETs according to different clinicopathological aspects. NEK2, UBE2C, TOP2A are positively associated with high tumor grade, and PPP1R1A negatively. GCGR and GNAS are regarded as the hub of the PPI sub-network. CXCR4 may affect the progression of PNETs via the CXCR4-CXCL12-CXCR7 chemokine receptor axis. However, more studies are required.


Assuntos
Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes , Tumores Neuroendócrinos/genética , Neoplasias Pancreáticas/genética , Mapas de Interação de Proteínas , Antígenos de Neoplasias/genética , Quimiocinas/genética , Cromograninas/genética , DNA Topoisomerases Tipo II/genética , Proteínas de Ligação a DNA/genética , Subunidades alfa Gs de Proteínas de Ligação ao GTP/genética , Humanos , Quinases Relacionadas a NIMA/genética , Metástase Neoplásica , Tumores Neuroendócrinos/patologia , Neoplasias Pancreáticas/patologia , Proteínas de Ligação a Poli-ADP-Ribose , Proteína Fosfatase 1/genética , Enzimas de Conjugação de Ubiquitina/genética
18.
Cell Rep ; 18(12): 2918-2931, 2017 03 21.
Artigo em Inglês | MEDLINE | ID: mdl-28329684

RESUMO

Alterations in distal regulatory elements that control gene expression underlie many diseases, including cancer. Epigenomic analyses of normal and diseased cells have produced correlative predictions for connections between dysregulated enhancers and target genes involved in pathogenesis. However, with few exceptions, these predicted cis-regulatory circuits remain untested. Here, we dissect cis-regulatory circuits that lead to overexpression of NEK6, a mitosis-associated kinase, in human B cell lymphoma. We find that only a minor subset of predicted enhancers is required for NEK6 expression. Indeed, an annotated super-enhancer is dispensable for NEK6 overexpression and for maintaining the architecture of a B cell-specific regulatory hub. A CTCF cluster serves as a chromatin and architectural boundary to block communication of the NEK6 regulatory hub with neighboring genes. Our findings emphasize that validation of predicted cis-regulatory circuits and super-enhancers is needed to prioritize transcriptional control elements as therapeutic targets.


Assuntos
Linfócitos B/metabolismo , Linfócitos B/patologia , Transformação Celular Neoplásica/genética , Elementos Facilitadores Genéticos , Fator de Ligação a CCCTC/metabolismo , Transformação Celular Neoplásica/patologia , Cromatina/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Células Jurkat , Linfoma Folicular/genética , Linfoma Folicular/patologia , Quinases Relacionadas a NIMA/genética , Quinases Relacionadas a NIMA/metabolismo
19.
Cancer Discov ; 7(3): 288-301, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28213356

RESUMO

Medulloblastoma is the most frequent malignant pediatric brain tumor and is divided into at least four subgroups known as WNT, SHH, Group 3, and Group 4. Here, we characterized gene regulation mechanisms in the most aggressive subtype, Group 3 tumors, through genome-wide chromatin and expression profiling. Our results show that most active distal sites in these tumors are occupied by the transcription factor OTX2. Highly active OTX2-bound enhancers are often arranged as clusters of adjacent peaks and are also bound by the transcription factor NEUROD1. These sites are responsive to OTX2 and NEUROD1 knockdown and could also be generated de novo upon ectopic OTX2 expression in primary cells, showing that OTX2 cooperates with NEUROD1 and plays a major role in maintaining and possibly establishing regulatory elements as a pioneer factor. Among OTX2 target genes, we identified the kinase NEK2, whose knockdown and pharmacologic inhibition decreased cell viability. Our studies thus show that OTX2 controls the regulatory landscape of Group 3 medulloblastoma through cooperative activity at enhancer elements and contributes to the expression of critical target genes.Significance: The gene regulation mechanisms that drive medulloblastoma are not well understood. Using chromatin profiling, we find that the transcription factor OTX2 acts as a pioneer factor and, in cooperation with NEUROD1, controls the Group 3 medulloblastoma active enhancer landscape. OTX2 itself or its target genes, including the mitotic kinase NEK2, represent attractive targets for future therapies. Cancer Discov; 7(3); 288-301. ©2017 AACR.This article is highlighted in the In This Issue feature, p. 235.


Assuntos
Neoplasias Cerebelares/genética , Cromatina/metabolismo , Meduloblastoma/genética , Fatores de Transcrição Otx/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Neoplasias Cerebelares/patologia , Cromatina/genética , Elementos Facilitadores Genéticos , Regulação Neoplásica da Expressão Gênica , Humanos , Meduloblastoma/patologia , Células-Tronco Mesenquimais/fisiologia , Quinases Relacionadas a NIMA/genética , Quinases Relacionadas a NIMA/metabolismo , Fatores de Transcrição Otx/metabolismo
20.
Mol Cell ; 65(5): 818-831.e5, 2017 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-28216227

RESUMO

Telomeric repeat binding factor 1 (TRF1) is essential to the maintenance of telomere chromatin structure and integrity. However, how telomere integrity is maintained, especially in response to damage, remains poorly understood. Here, we identify Nek7, a member of the Never in Mitosis Gene A (NIMA) kinase family, as a regulator of telomere integrity. Nek7 is recruited to telomeres and stabilizes TRF1 at telomeres after damage in an ATM activation-dependent manner. Nek7 deficiency leads to telomere aberrations, long-lasting γH2AX and 53BP1 foci, and augmented cell death upon oxidative telomeric DNA damage. Mechanistically, Nek7 interacts with and phosphorylates TRF1 on Ser114, which prevents TRF1 from binding to Fbx4, an Skp1-Cul1-F box E3 ligase subunit, thereby alleviating proteasomal degradation of TRF1, leading to a stable association of TRF1 with Tin2 to form a shelterin complex. Our data reveal a mechanism of efficient protection of telomeres from damage through Nek7-dependent stabilization of TRF1.


Assuntos
Dano ao DNA , Quinases Relacionadas a NIMA/metabolismo , Estresse Oxidativo , Proteínas de Ligação a Telômeros/metabolismo , Telômero/enzimologia , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Sítios de Ligação , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Células HEK293 , Células HeLa , Histonas/metabolismo , Humanos , Quinases Relacionadas a NIMA/genética , Fosforilação , Complexo de Endopeptidases do Proteassoma/metabolismo , Ligação Proteica , Estabilidade Proteica , Interferência de RNA , Telômero/genética , Telômero/efeitos da radiação , Proteínas de Ligação a Telômeros/genética , Fatores de Tempo , Transfecção , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/genética , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/metabolismo , Ubiquitinação
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