The actin cytoskeleton in rapid steroid hormone actions

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Background/Aims: The epigenetic factor KDM2B is a histone demethylase expressed in various tumors. Recently, we have shown that KDM2B regulates actin cytoskeleton organization, small Rho GTPases signaling, cell-cell adhesion and migration of prostate tumor cells. In the present study, we addressed its role in regulating EMT and small GTPases expression in colon tumor cells. Methods: We used RT-PCR for the transcriptional analysis of various genes, Western blotting for the assessment of protein expression and immunofluorescence microscopy for visualization of fluorescently labeled proteins. Results: We report here that KDM2B regulates EZH2 and BMI1 in HCT116 colon tumor cells. Knockdown of this epigenetic factor induced potent up-regulation of the protein levels of the epithelial markers E-cadherin and ZO-1, while the mesenchymal marker N-cadherin was downregulated. On the other hand, KDM2B overexpression downregulated the levels of both epithelial markers and upregulated the mesenchymal marker, suggesting control of EMT by KDM2B. In addition, RhoA, RhoB and RhoC protein levels diminished upon KDM2B-knockdown, while all three small GTPases became upregulated in KDM2B-overexpressing HCT116 cell clones. Interestingly, Rac1 GTPase level increased upon KDM2B-knockdown and diminished in KDM2B-overexpressing HCT116 colon tumor- and DU-145 prostate cancer cells. Conclusions: These results establish a clear functional role of the epigenetic factor KDM2B in the regulation of EMT and small-GTPases expression in colon tumor cells and further support the recently postulated oncogenic role of this histone demethylase in various tumors.

Section: Discussionmentioning

confidence: 99%

The Epigenetic Factor KDM2B Regulates EMT and Small GTPases in Colon Tumor Cells

Zacharopoulou

Tsapara

Kallergi

et al. 2018

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“…Candidate mediators of mAR-induced beta-catenin association to actin could be E-cadherin [25], although interactions via additional cytoskeletal components such as the microfilament-bundling protein fascin [24]) cannot be excluded. Since actin reorganization is a main cellular event in mAR-induced downstream signaling that regulates pro-apoptotic responses [5,39] the mechanisms linking beta-catenin to TAC induced actin cytoskeleton restructuring are of particular interest. We are currently addressing this issue in our laboratory.…”

Section: Discussionmentioning

confidence: 99%

Membrane Androgen Receptor Down-Regulates c-Src-Activity and Beta-Catenin Transcription and Triggers GSK-3beta-Phosphorylation in Colon Tumor Cells

Honisch

Kounenidakis

et al. 2014

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Background/Aims: Functional membrane androgen receptors (mARs) have recently been described in colon tumor cells and tissues. Their activation by specific testosterone albumin conjugates (TAC) down-regulates the PI-3K/Akt pro-survival signaling and triggers potent pro-apoptotic responses both, in vitro and in vivo. The present study explored the mAR-induced regulation of gene products implicated in the tumorigenic activity of Caco2 colon cancer cells. Methods: In Caco2 human colon cancer cells transcript levels were determined by RT-PCR, protein abundance and phosphorylation by Western blotting and confocal microscopy, as well as cytoskeletal architecture by confocal microscopy. Results: We report time dependent significant decrease in Tyr-416 phosphorylation of c-Src upon mAR activation. In line with the reported late down-regulation of the PI-3K/Akt pathway in testosterone-treated colon tumors, GSK-3beta was phosphorylated at Tyr-216 after long term stimulation of the cells with TAC, a finding supporting the role of this kinase to promote apoptosis. PCR analysis revealed significant decrease of beta-catenin and cyclin D1 transcript levels following TAC treatment. Moreover, confocal laser scanning microscopic analysis disclosed co-localization of beta-catenin with actin cytoskeleton. It is thus conceivable that beta-catenin may participate in the reported modulation of cytoskeletal dynamics in mAR stimulated Caco2 cells. Conclusions: Our results provide strong evidence that mAR activation regulates late expression and/or activity of the tumorigenic gene products c-Src, GSK-3beta, and beta-catenin thus facilitating the pro-apoptotic response in colon tumor cells.

“…The molecular mechanism governing this structural reorganisation revealed the implication of FAK/PI-3K signaling [1,18] and the specific actin regulating molecules Rac1 GTPase and PAK1 [38]. In addition, this extensive actin reorganization was related to differential stiffness behaviour of endothelial cells upon chorein silencing [18], a finding well correlated with the significant role of diminished actin cytoskeleton stability in cellular functions [39,40,41], including migration [30,42,43], adhesion [44] and apoptosis [32,45,46]. Based on these reports, our finding shows depolymerised actin cytoskeleton both in erythrocytes and fibroblasts isolated from ChAc-patients implying destabilized microfilaments.…”

Section: Discussionmentioning

confidence: 93%

“…Impaired cytoskeletal dynamics and disarranged structural organization underlie several human diseases, including muscular dystrophy [24,25,26], cardiac disorders [27], liver diseases [28] and malignancies [29,30,31,32]. Since ChAc is a genetic disease affecting multiple organs including brain and blood, but also muscle and liver cells [4,7], disarranged cytoskeletal structures may not be limited to actin microfilaments.…”

Section: Introductionmentioning

confidence: 99%

Chorein Sensitive Arrangement of Cytoskeletal Architecture

Honisch

Hagen

et al. 2015

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Background/Aims: Chorein is a protein expressed in various cell types. Loss of function mutations of the chorein encoding gene VPS13A lead to chorea-acanthocytosis, an autosomal recessive genetic disease characterized by movement disorder and behavioral abnormalities. Recent observations revealed that chorein is a powerful regulator of actin cytoskeleton in erythrocytes, platelets, K562 and endothelial HUVEC cells. Methods: In the present study we have used Western blotting to study actin polymerization dynamics, laser scanning microscopy to evaluate in detail the role of chorein in microfilaments, microtubules and intermediate filaments cytoskeleton architecture and RT-PCR to assess gene transcription of the cytoskeletal proteins. Results: We report here powerful depolymerization of actin microfilaments both, in erythrocytes and fibroblasts isolated from chorea-acanthocytosis patients. Along those lines, morphological analysis of fibroblasts from chorea-acanthocytosis patients showed disarranged microtubular network, when compared to fibroblasts from healthy donors. Similarly, the intermediate filament networks of desmin and cytokeratins showed significantly disordered organization with clearly diminished staining in patient's fibroblasts. In line with this, RT-PCR analysis revealed significant downregulation of desmin and cytokeratin gene transcripts. Conclusion: Our results provide for the first time evidence that defective chorein is accompanied by significant structural disorganization of all cytoskeletal structures in human fibroblasts from chorea-acanthocytosis patients.