Centrosome Research Articles

The function and molecular mechanism of CEP55 in anaplastic thyroid cancer.

The purpose of this study was to determine the role of centrosomal protein of 55 kDa (CEP55) in anaplastic thyroid cancer (ATC) and to further explore the mechanism, which might provide a new molecular marker for treatment of ATC. The expression level of CEP55 in clinical cases was tested by fluorescence quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Also, qRT-PCR assay was performed in different TC cell lines. The relationship between CEP55 expression and clinicopathological characteristics was statistically analyzed. Kaplan-Meier curve and Cox's proportional hazards regression model were performed in survival analysis. Further, Western blot assay was used to analyze the protein expression changes in PI3K/Akt pathway. The expression level of CEP55 in TC tissues showed a noticeable upgrade, especially in ATC. In vitro, CEP55 expression was also increased in four kinds of TC cells, in which, the highest expression was found in ATC (TA-K) cells. The clinicopathological features, including lymph node metastasis, distant metastasis, and prognostic index were found to be correlated with the expression level of CEP55. Besides, the ATC patients with higher expression of CEP55 had a statistically worse overall survival (OS) time. In univariate analyses and multivariate analyses, the CEP55 level was an independent prognosis index of patients with ATC. In vitro study, CEP55 protein expression level was significantly reduced in si-CEP55-transfected TA-K cells. Notably, the downregulation of CEP55 could suppress the phosphorylation of PI3K and AKT. This study found that CEP55 could promote ATC progression, and PI3K/AKT pathway might be the downstream target of its action. These results provided a new therapeutic direction for the treatment of ATC.

C-Cbl Acts as an E3 Ligase Against DDA3 for Spindle Dynamics and Centriole Duplication during Mitosis.

The spatiotemporal mitotic processes are controlled qualitatively by phosphorylation and qualitatively by ubiquitination. Although the SKP1-CUL1-F-box protein (SCF) complex and the anaphase-promoting complex/cyclosome (APC/C) mainly mediate ubiquitin-dependent proteolysis of mitotic regulators, the E3 ligase for a large portion of mitotic proteins has yet to be identified. Here, we report c-Cbl as an E3 ligase that degrades DDA3, a protein involved in spindle dynamics. Depletion of c-Cbl led to increased DDA3 protein levels, resulting in increased recruitment of Kif2a to the mitotic spindle, a concomitant reduction in spindle formation, and chromosome alignment defects. Furthermore, c-Cbl depletion induced centrosome over-duplication and centriole amplification. Therefore, we concluded that c-Cbl controls spindle dynamics and centriole duplication through its E3 ligase activity against DDA3.

Advances in research of monopolar spindle protein kinase 1 in tumor therapy

The Spindle assembly checkpoint (SAC) is an extremely important monitoring step in the mitosis process, ensuring that all sister chromatids are placed on the medium-term plate before the mid-mitotic transformation. Recent studies have shown that this checkpoint abnormality has a higher correlation with tumorigenesis, development and metastasis mechanisms. Monopolar spindle 1 (Mps1) is a conserved bispecific protein kinase whose primary function is centrosome replication and activation of SAC, both of which help prevent mitotic errors and prevent tumorigenesis. The latest study found that the expression level of Mps1 is related to the degree of malignancy of breast cancer. More notably, inhibition of Mps1 kinase activity or knockdown of Mps1 by siRNA results in a barrier to tumor cell division, leading to tumor cell apoptosis without affecting normal cell activity. The dysregulation of the cell cycle is also closely related to the invasive biological behavior of malignant tumors, and becomes a basic marker of tumor progression. The cell cycle is mainly regulated by a number of checkpoint regulatory pathways, which are specifically dysregulated in tumor cells and Has the potential to become a target for the design of selective treatment of tumor cells. This article reviews how Mps1 regulates SAC and its recent advances in cancer therapy Key words: Spindle assembly checkpoint; Monopolar spindle 1; Gene targeting therapy; Osteosarcoma

The Replication Protein Cdc6 Suppresses Centrosome Over-Duplication in a Manner Independent of Its ATPase Activity.

The Cdc6 protein is essential for the initiation of chromosomal replication and functions as a licensing factor to maintain chromosome integrity. During the S and G2 phases of the cell cycle, Cdc6 has been found to inhibit the recruitment of pericentriolar material (PCM) proteins to the centrosome and to suppress centrosome over-duplication. In this report, we analyzed the correlation between these two functions of Cdc6 at the centrosome. Cdc6 depletion increased the population of cells showing centrosome over-duplication and premature centrosome separation; Cdc6 expression reversed these changes. Deletion and fusion experiments revealed that the 18 amino acid residues (197–214) of Cdc6, which were fused to the Cdc6-centrosomal localization signal, suppressed centrosome over-duplication and premature centrosome separation. Cdc6 mutant proteins that showed defective ATP binding or hydrolysis did not exhibit a significant difference in suppressing centrosome over-duplication, compared to the wild type protein. In contrast to the Cdc6-mediated inhibition of PCM protein recruitment to the centrosome, the independence of Cdc6 on its ATPase activity for suppressing centrosome over-duplication, along with the difference between the Cdc6 protein regions participating in the two functions, suggested that Cdc6 controls centrosome duplication in a manner independent of its recruitment of PCM proteins to the centrosome.

Cep70 drives taxol drug resistance through regulating the acetylation α-tubulin to reduce microtubule stability

Objective To explore the Cep70 by adjusting the stability of acetylated alpha tubulin, participate in breast cancer drug resistance mechanisms. Methods ⑴ In order to induce taxol drug resistance cell line Michigan cancer foundation-7 (MCF-7)/pac, high-dose shock treatments taxol MCF-7 was used for 6 months, until the cells can grow in 3.5 μmol/L of paclitaxel. ⑵ The 3-(4, 5-dimenthylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) method was used to detect inhibition rate by taxol to MCF-7 and MCF-7/pac cell. ⑶ Immunofluorescence and Western blot were used to test acetylated alpha-tubulin and Cep70 expression levels in MCF-7 and MCF-7/pac cells. ⑷ Chemical intervention was used to acetylate apha-tubulin expression, Western blot and polymerase chain reaction (PCR) were used to detect the change of acetylated alpha-tubulin and Cep70 in MCF-7 and MCF-7/pac groups. Flow cytometry and Western blot were used to detect the change of cell cycle. Results ⑴ IC50 of MCF-7 and MCF-7/pac was 22.47 μmol/L and 31.38 μmol/L, respectively. ⑵ Immunofluorescence and Western blot results showed that the expression of acetylation of alpha-tubulin in resistant MCF-7 cell/pac was obviously decreased. ⑶ Real time polymerase chain reaction (RT-PCR) and Western blot showed Cep70 expression was consistent of acetylation of alpha-tubulin. ⑷ After incubation with paclitaxel for 24 hours, the expressions of acetylation of alpha-tubulin and Cep70 in MCF-7 and MCF-7/pac were increased, but the extent of MCF-7 cell was much higher. Instead, incubation with nocodazole after 24 hours, the acetylation of alpha-tubulin and Cep70 in MCF-7 and MCF-7/pac cells were obviously lowered. ⑸ After paclitaxel intervention, compared to the same group MCF-7 cells, the G2 phase ratio in MCF-7/pac cells was lower. In addition, given nocodazole after the intervention, compared to the same group MCF-7 cells, the ratio of G2 phase in MCF-7 cell/pac was significantly decreased. Conclusions Cep70 decreased the expression of the acetylated alpha-tubulin, reduced the stability of microtubules, which could be an important mechanism of taxol drug resistance. Key words: Centrosome; Tubulin; Breast neoplasms/DT/ME; Taxoids; Drug resistance, neoplasm

Tumor specific theranostic streptavidin-coupled superparamagnetic iron oxide nanoparticles for targeting therapeutic moieties in pancreatic cancer

Pancreatic cancer is one of the most aggressive malignancies and burdened with a dismal prognosis (1). Given that the regulation of cell division is executed with high fidelity to maintain organ homeostasis, it is not surprising that alterations of the cell cycle are a hallmark of cancer (2). Many of the genes encoding key regulators of the cell cycle are mutated in both, sporadic and hereditary forms of cancer including pancreatic cancers implicating a role in the pathogenesis of PDAC (3). Polo-like kinases (PLKs) play an important role in the centrosome cycle which suggests that their deregulation would not be unexpected in malignant tours and their oncogenesis. PLK1 overexpression has, indeed, been observed in wide range of tumor types and was often associated with a poor prognosis (4).

Function and mechanism of PAR6 gene in the occurrence and development of tumor

Cell polarity is a common feature of many different types of cells, and it is essential to the normal differentiation and function of cells. Partitioning defective 6 (PAR6) gene encodes PAR6 protein, which is crucial to asymmetric cell division and polarized growth. PAR6 protein as a member of the PAR6 polarity complex, affects the synthetic of centrosome and protein recruitment to the centrosome. The abnormal number of centrosomal and the loss of cell polarity may eventually lead to the occurrence of tumor. Key words: Gene expression regulation, neoplastic; Multiprotein complexes; Centrosome; Cell polarity; Partitioning defective

Novel role of PKC epsilon in mitotic spindle stability

Mitosis is a highly regulated process characterized by dramatic and coordinated morphological changes to ensure the fidelity of chromosome segregation. Missegregation of mitotic chromosomes leads to a condition that underlies chromosomal instability(1), which is a hallmark of cancer. In order to assure symmetry and bipolarity of the cell division process, mitotic spindle microtubules properly segregate mitotic chromosomes (2). Among the several isoforms of serine/threonine kinases, PKCe is one of the best understood for its role as a transforming oncogene, and it has been found overexpressed in different types of tumors. In 2008, Saurin and colleagues demonstrated the involvement of PKCe in the regulation of the late stage of mitosis (3). Through its association with 14-3-3 at the midbody, PKCe is essential for the successful completion of cytokinesis, and the inhibition of functional PKCe-14-3- 3 complex leads to abscission failure and multinucleated phenotype in cells. In this study, we found that PKCe is involved in mitotic spindle stability. Using fluorescence microscopy, we found that the active form of PKCe (phosphorylated at Ser-729), colocalizes to the centrosome in cells in metaphase, where the mitotic spindle nucleation occurs. Furthermore, experiments of co-immunoprecipitation revealed that, when cells are synchronized in metaphase, PKCɛ is associated to ɣ-tubulin, a member of the tubulin superfamily localized to the microtubule organizing centers and is essential for microtubule nucleation from centrosomes. Consequently modulation of PKCɛ expression affects spindle stability: PKCɛ downregulation by specific shRNA results in mitotic spindle disorganization with a reduction of the amount of centrosomal and mitotic ɣ-Tubulin and αβ-tubulin fluorescence. Mitotic spindle formation assays using Nocodazole, known to interfere with the polymerization of microtubules, revealed that cells lacking PKCɛ were unable to regrow microtubules after depolymerization. These results reveal a novel role of PKCɛ in mitotic spindle stability, which likely determinant for genome stability.

Novel role of PKC ɛ in mitotic spindle stability

Mitosis is a highly regulated process characterized by dramatic and coordinated morphological changes to ensure the fidelity of chromosome segregation. Misseg- regation of mitotic chromosomes leads to a condition that underlies chromosomal instability(1), which is a hallmark of cancer. In order to assure symmetry and bipo- larity of the cell division process, mitotic spindle microtubules properly segregate mitotic chromosomes (2). Among the several isoforms of serine/threonine kinases, PKCe is one of the best understood for its role as a transforming oncogene, and it has been found overexpressed in different types of tumors. In 2008, Saurin and col- leagues demonstrated the involvement of PKCe in the regulation of the late stage of mitosis (3). Through its association with 14-3-3 at the midbody, PKCe is essential for the successful completion of cytokinesis, and the inhibition of functional PKCe-14-3- 3 complex leads to abscission failure and multinucleated phenotype in cells. In this study, we found that PKCe is involved in mitotic spindle stability. Using fluorescence microscopy, we found that the active form of PKCe (phosphorylated at Ser-729), co- localizes to the centrosome in cells in metaphase, where the mitotic spindle nuclea- tion occurs. Furthermore, experiments of co-immunoprecipitation revealed that, when cells are synchronized in metaphase, PKCɛ is associated to ɣ-tubulin, a member of the tubulin superfamily localized to the microtubule organizing centers and is essen- tial for microtubule nucleation from centrosomes. Consequently modulation of PKCɛ expression affects spindle stability: PKCɛ downregulation by specific shRNA results in mitotic spindle disorganization with a reduction of the amount of centrosomal and mitotic ɣ-Tubulin and αβ-tubulin fluorescence. Mitotic spindle formation assays using Nocodazole, known to interfere with the polymerization of microtubules, revealed that cells lacking PKCɛ were unable to regrow microtubules after depolymerization. These results reveal a novel role of PKCɛ in mitotic spindle stability, which likely determinant for genome stability.

Changes of centrosome and related protein in malignant transformation of BEAS-2B cell induced by coal tar pitch smoke extracts

To analyze the centrosome abnormalities in the malignant transformation of human bronchial epithelial cells (BEAS-2B) induced by coal tar pitch smoke extracts and to investigate the role and action mechanism of centrosome in the lung cancer induced by coal tar pitch. Medium-temperature coal tar pitch smoke extracts were used to treat immortalized human bronchial epithelial cells (BEAS-2B) and establish a malignant transformation model. The treated BEAS-2B cells were used as exposure group, and solvent control group and normal control group were also set for passage culture. The changes of centrosome in BEAS-2B cells seeded on coverslips were evaluated by indirect immunofluorescence assay. The mRNA expression of p53, p21, and cyclin E in BEAS-2B cells was measured by real-time quantitative RT-PCR, and their protein levels in BEAS-2B cells seeded on coverslips were measured by semiquantitative immunohistochemical analysis. The overall rate of centrosome abnormalities in BEAS-2B cells at passage 20 was 6.56±1.01% in the exposure group, significantly higher than those in the normal control group (3.40±0.86%) and solvent control group (3.14±0.59%) (P < 0.05). In addition, the exposure group had a significantly higher overall rate of centrosome abnormalities in BEAS-2B cells at passage 30 compared with the normal control group and solvent control group (22.39±9.5% vs 4.34±1.04%, P < 0.05; 22.39±9.5% vs 4.33±1.20%, P < 0.05). Compared with the normal control group and solvent control group, the exposure group had significantly decreased mRNA and protein expression of p53 and significantly increased mRNA and protein expression of cyclin E in BEAS-2B cells at passages 20 and 30 (P < 0.05). Centrosome abnormalities occur before the malignant transformation in BEAS-2B cells treated with coal tar pitch smoke extracts, and they may be mediated by the p53/p21/cyclin E signaling pathway.

Coevolution pattern analysis of centrosomal proteins CEP63 and CEP152 using Bayesian Algorithm

Coevolution pattern analysis of centrosomal proteins CEP63 and CEP152 using Bayesian Algorithm

Molecular mechanism of oncogenic B-RafV600E induces chromosome instability in melanoma Sbcl2 and SK-MEL31 cells

Objective To explore the molecular mechanism of BRAFV600E inducing chromosome instability in Sbcl2 and SK-MEL31 melanoma cells.Methods The endogenous Mps1 in stable Sbcl2-and SK-MEL31-B-RafV600E expression cells were depleted by siRNA approach.To test the effect of B-RafV600E on the centrosome amplification and the formation of multipolar spindles,cells at S-phase with HU-treatment were arrested and then the centrosomes and mitotic spindles structure were detected through immunofluoresence.Results The percentage of B-RafV600E expressing Sbcl2 and SK-MEL31 cells (Sbcl2-B-RafV600E and SKMEL31-B-RafV600E) with centrosome amplification and multipolar spindle was reduced from 36 ％ to 6 ％ when Mps1 was absent.Conclusion B-RafV600E leads to centrosome amplification and multipolar spindle through Mps1,thus results in chromosome instability in Sbcl2 and SK-MEL31 melanoma cells. Key words: Genes, B-RafV600E; Genes, Mps1; Centrosome; Mitotic spindle apparatus; Chromosome

Clathrin helps centrosomes come of age

Clathrin helps centrosomes come of age

TIG3 Is a Multifunctional Regulator of Keratinocyte Differentiation and Survival

Tazarotene induced gene 3 (TIG3) is a tumor suppressor protein that regulates epidermal keratinocyte differentiation and survival. TIG3 is expressed in the upper layers of normal epidermis where it participates in halting cell proliferation and driving terminal differentiation. Reduced levels of TIG3 are observed in hyperproliferative diseases and restoration of TIG3 decreases cell survival and correlates with disease normalization. In normal epidermis, TIG3 binds to and activates type I transglutaminase resulting in cornified envelope formation and also localizes at the centrosome to halt cell proliferation by inhibiting centrosome function.

Regulation of cyclin E and cancers

Cyclin E is expressed starting from the middle G1 phase of the cell cycle,and is accumulated in the G1/S boundry.Cyclin E binds to and activates the cyclin-dependent kinase CDK2.Cyclin E-CDK2 complex initiates a cascade of events that leads to DNA replication by phosphorylating its substrates,such as Rb,CDC6,NPAT and P107,etc.Additionally,cyclin E plays an important role in the regulation of genomic stability,spindle-organizing structure and centrosome cycle.Cyclin E expression is trans-activated by members of the transcription factor E2F family and degrades via the ubiquitin-proteasome pathway.At the same time,it is also negatively regulated by the CIP/KIP proteins.Cyclin E highly expressed in the initiation and progression of different human cancers,such as breast cancers,lung cancers,leukemia,lymphomas and others. Key words: Cyclin E; Cell cycle regulation; Cyclins; Neoplasms

Centrosome Center Stage?

Centrosome Center Stage?

No centrosome, no problem

No centrosome, no problem

29 Centrosomes in Mouse Development and PKD

29 Centrosomes in Mouse Development and PKD

The centrosome neither persistently leads migration nor determines the site of axonogenesis in migrating neurons in vivo

The centrosome neither persistently leads migration nor determines the site of axonogenesis in migrating neurons in vivo

Expressions and significances of Nek2 and β-catenin in breast invasive ductal carcinoma

Objective To investigate the expression and significance of NIMA related kinases 2 ( Nek2) and β-catenin (β-cat) in breast invasive ductal carcinoma (IDC) and concomitant ductal carcinoma in situ (DCIS). The roles of Nek2 and β-acat in the development and progression of breast cancer were explored. Methods The protein expression of Nek2 and β-cat in the tissues from 186 patients with IDC, 75 concomitant DCIS, 80 normal tissue adjacent to carcinoma and 40 fibroadenoma of breast was detected by using immunohistochemistry. Their relationship between clinicopathologic parameters was analyzed. Results There were correlations between the Nek2 expression in cytoplasm and grade(x2=8. 756;P 0. 05), the Nek2 expression in cytoplasm had a correlation with the β-cat expression in cytoplasm (r=0. 226,P<0.05) in concomitant DCIS, and the same relationship could also be seen in IDC (r=0. 368,P<0.01). There was significant difference in the β-cat expression on the cytomembrane between IDC and concomitant DCIS (Z=-3. 804,P<0. 01). In the normal tissue adjacent to carcinoma and fibroadenoma, the Nek2 expression in cytoplasm and nuclei Was low or negative;the β-cat expression on the cytomembrane was high but that Was low in cytoplasm.Conclusion Centrosome regulatory factor Nek2 and β-cat can support a new way to explore the mechanisms of breast tumorigrenesis.And they may become a new antitumor drug target in the future. Key words: Breast invasive ductal carcinoma; Nek2; β-catenin