Melanoma phenotype plasticity underlies tumour dissemination and resistance to therapy, yet its regulation is incompletely understood. In vivo switching between a more differentiated, proliferative phenotype and a dedifferentiated, invasive phenotype is directed by the tumour microenvironment. We found that treatment of partially dedifferentiated, invasive A375M2 cells with two structurally unrelated p38 MAPK inhibitors, SB2021920 and BIRB796, induces a phenotype switch in 3D collagen, as documented by increased expression of melanocyte differentiation markers and a loss of invasive phenotype markers. The phenotype is accompanied by morphological change corresponding to amoeboid-mesenchymal transition. We performed RNA sequencing with an Illumina HiSeq platform to fully characterise transcriptome changes underlying the switch. Gene expression results obtained with RNA-seq were validated by comparing them with RT-qPCR. Transcriptomic data generated in the study will extend the present understanding of phenotype plasticity in melanoma and its contribution to invasion and metastasis.

• Klíčová slova
• amoeboid invasion, cancer, melanoma, metastasis, phenotype switch,
• MeSH
• buněčná diferenciace účinky léků   genetika   MeSH
• fenotyp MeSH
• genová ontologie MeSH
• imidazoly farmakologie   MeSH
• inhibitory proteinkinas farmakologie   MeSH
• kolagen metabolismus   MeSH
• lidé MeSH
• melanom genetika   patologie   MeSH
• mitogenem aktivované proteinkinasy p38 antagonisté a inhibitory   metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádorové mikroprostředí účinky léků   genetika   MeSH
• naftaleny farmakologie   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• proliferace buněk účinky léků   genetika   MeSH
• pyrazoly farmakologie   MeSH
• pyridiny farmakologie   MeSH
• regulace genové exprese u nádorů účinky léků   MeSH
• sekvenování transkriptomu metody   MeSH
• stanovení celkové genové exprese metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole MeSH Prohlížeč
• doramapimod MeSH Prohlížeč
• imidazoly MeSH
• inhibitory proteinkinas MeSH
• kolagen MeSH
• mitogenem aktivované proteinkinasy p38 MeSH
• naftaleny MeSH
• pyrazoly MeSH
• pyridiny MeSH

The invasive behaviour of cancer cells underlies metastatic dissemination; however, due to the large plasticity of invasion modes, it is challenging to target. It is now widely accepted that various secreted cytokines modulate the tumour microenvironment and pro-inflammatory signalling can promote tumour progression. Here, we report that cells after mesenchymal-amoeboid transition show the increased expression of genes associated with the type I interferon response. Moreover, the sustained activation of type I interferon signalling in response to IFNβ mediated by the Stat1/Stat2/IRF9 complex enhances the round amoeboid phenotype in melanoma cells, whereas its downregulation by various approaches promotes the mesenchymal invasive phenotype. Overall, we demonstrate that interferon signalling is associated with the amoeboid phenotype of cancer cells and suggest a novel role of IFNβ in promoting cancer invasion plasticity, aside from its known role as a tumour suppressor.

• Klíčová slova
• amoeboid, inflammation, interferon, invasion, melanoma, mesenchymal, plasticity,
• Publikační typ
• časopisecké články MeSH

The plasticity of cancer cell invasion represents substantial hindrance for effective anti-metastatic therapy. To better understand the cancer cells' plasticity, we performed complex transcriptomic and proteomic profiling of HT1080 fibrosarcoma cells undergoing mesenchymal-amoeboid transition (MAT). As amoeboid migratory phenotype can fully manifest only in 3D conditions, all experiments were performed with 3D collagen-based cultures. Two previously described approaches to induce MAT were used: doxycycline-inducible constitutively active RhoA expression and dasatinib treatment. RNA sequencing was performed with ribo-depleted total RNA. Protein samples were analysed with tandem mass tag (TMT)-based mass spectrometry. The data provide unprecedented insight into transcriptome and proteome changes accompanying MAT in true 3D conditions.

• MeSH
• fibrosarkom patologie   MeSH
• invazivní růst nádoru * MeSH
• kolagen chemie   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• pohyb buněk * MeSH
• proteom * MeSH
• rhoA protein vázající GTP MeSH
• sekvenční analýza RNA MeSH
• tandemová hmotnostní spektrometrie MeSH
• transkriptom * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• dataset MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kolagen MeSH
• proteom * MeSH
• rhoA protein vázající GTP MeSH
• RHOA protein, human MeSH Prohlížeč

Microtubule-targeting agents (MTAs) constitute a diverse group of chemical compounds that bind to microtubules and affect their properties and function. Disruption of microtubules induces various cellular responses often leading to cell cycle arrest or cell death, the most common effect of MTAs. MTAs have found a plethora of practical applications in weed control, as fungicides and antiparasitics, and particularly in cancer treatment. Here we summarize the current knowledge of MTAs, the mechanisms of action and their role in cancer treatment. We further outline the potential use of MTAs in anti-metastatic therapy based on inhibition of cancer cell migration and invasiveness. The two main problems associated with cancer therapy by MTAs are high systemic toxicity and development of resistance. Toxic side effects of MTAs can be, at least partly, eliminated by conjugation of the drugs with various carriers. Moreover, some of the novel MTAs overcome the resistance mediated by both multidrug resistance transporters as well as overexpression of specific β-tubulin types. In anti-metastatic therapy, MTAs should be combined with other drugs to target all modes of cancer cell invasion.

• Klíčová slova
• Cancer metastasis, Cancer therapy, Microtubule-targeting agents, Migrastatics,
• MeSH
• lidé MeSH
• mikrotubuly účinky léků   MeSH
• nádory farmakoterapie   MeSH
• protinádorové látky farmakologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• protinádorové látky MeSH

The ability of cancer cells to adopt various migration modes (the plasticity of cancer cell invasiveness) is a substantive obstacle in the treatment of metastasis, yet still an incompletely understood process. We performed a comparison of publicly available transcriptomic datasets from various cell types undergoing a switch between the mesenchymal and amoeboid migration modes. Strikingly, lncRNA MALAT1 (metastasis-associated lung adenocarcinoma transcript 1) was one of three genes that were found upregulated in all amoeboid cells analyzed. Accordingly, downregulation of MALAT1 in predominantly amoeboid cell lines A375m2 and A2058 resulted in decrease of active RhoA (Ras homolog family member A) and was accompanied by the amoeboid-mesenchymal transition in A375m2 cells. Moreover, MALAT1 downregulation in amoeboid cells led to increased cell proliferation. Our work is the first to address the role of MALAT1 in MAT/AMT (mesenchymal to amoeboid transition/amoeboid to mesenchymal transition) and suggests that increased MALAT1 expression is a common feature of amoeboid cells.

• Klíčová slova
• MALAT1, amoeboid invasion, cancer, invasion plasticity, lncRNA, melanoma, mesenchymal invasion,
• Publikační typ
• časopisecké články MeSH

M2-polarized macrophages have been shown to adapt their 3D migration mode to physical properties of surrounding extracellular matrix. They migrate in the integrin-mediated adhesion and proteolytic activity-dependent "mesenchymal" mode in stiff matrices and in the integrin and protease-independent "amoeboid" mode in low density, porous environments. To find out what impact the switching between the migration modes has on expression of both protein-coding and non-coding genes we employed RNA sequencing of total RNA depleted of ribosomal RNA isolated from macrophages migrating in either mode in 3D collagens. Differentially expressed genes from both categories have been detected and the changes in expression of selected genes were further validated with RT-qPCR. The acquired data will facilitate better understanding of how mechanical properties of tissue microenvironment reflect in macrophage immune function and how the transitions between mesenchymal and amoeboid migratory modes are regulated at the gene expression level.

• MeSH
• buněčné mikroprostředí * MeSH
• kolagen MeSH
• lidé MeSH
• makrofágy * cytologie   metabolismus   MeSH
• pohyb buněk * genetika   MeSH
• RNA ribozomální * MeSH
• sekvenční analýza RNA MeSH
• test migrace makrofágů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• dataset MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kolagen MeSH
• RNA ribozomální * MeSH

In solid cancers, invasion and metastasis account for more than 90% of mortality. However, in the current armory of anticancer therapies, a specific category of anti-invasion and antimetastatic drugs is missing. Here, we coin the term 'migrastatics' for drugs interfering with all modes of cancer cell invasion and metastasis, to distinguish this class from conventional cytostatic drugs, which are mainly directed against cell proliferation. We define actin polymerization and contractility as target mechanisms for migrastatics, and review candidate migrastatic drugs. Critical assessment of these antimetastatic agents is warranted, because they may define new options for the treatment of solid cancers.

• Klíčová slova
• contractility, invasion, metastasis, migrastatics, solid cancer, treatment,
• MeSH
• chemorezistence MeSH
• cílená molekulární terapie MeSH
• lidé MeSH
• metastázy nádorů farmakoterapie   MeSH
• nádorové biomarkery MeSH
• nádory farmakoterapie   etiologie   metabolismus   patologie   MeSH
• objevování léků * MeSH
• pohyb buněk účinky léků   MeSH
• protinádorové látky chemie   farmakologie   terapeutické užití   MeSH
• signální transdukce účinky léků   MeSH
• synergismus léků MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• nádorové biomarkery MeSH
• protinádorové látky MeSH

CAS is a docking protein, which was shown to act as a mechanosensor in focal adhesions. The unique assembly of structural domains in CAS is important for its function as a mechanosensor. The tension within focal adhesions is transmitted to a stretchable substrate domain of CAS by focal adhesion-targeting of SH3 and CCH domain of CAS, which anchor the CAS protein in focal adhesions. Mechanistic models of the stretching biosensor propose equal roles for both anchoring domains. Using deletion mutants and domain replacements, we have analyzed the relative importance of the focal adhesion anchoring domains on CAS localization and dynamics in focal adhesions as well as on CAS-mediated mechanotransduction. We confirmed the predicted prerequisite of the focal adhesion targeting for CAS-dependent mechanosensing and unraveled the critical importance of CAS SH3 domain in mechanosensing. We further show that CAS localizes to the force transduction layer of focal adhesions and that mechanical stress stabilizes CAS in focal adhesions.

• MeSH
• buněčná adheze MeSH
• buněčný převod mechanických signálů * MeSH
• fibroblasty cytologie   metabolismus   MeSH
• fokální adheze metabolismus   MeSH
• mechanický stres MeSH
• mutantní proteiny chemie   MeSH
• myši MeSH
• proteinové domény MeSH
• rekombinantní fúzní proteiny metabolismus   MeSH
• signální transdukce MeSH
• stabilita proteinů MeSH
• substrátový protein asociovaný s Crk chemie   metabolismus   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• zelené fluorescenční proteiny metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• Bcar1 protein, mouse MeSH Prohlížeč
• mutantní proteiny MeSH
• rekombinantní fúzní proteiny MeSH
• substrátový protein asociovaný s Crk MeSH
• zelené fluorescenční proteiny MeSH

BACKGROUND: The local invasion of tumor cells into the surrounding tissue is the first and most critical step of the metastatic cascade. Cells can invade either collectively, or individually. Individual cancer cell invasion can occur in the mesenchymal or amoeboid mode, which are mutually interchangeable. This plasticity of individual cancer cell invasiveness may represent an escape mechanism for invading cancer cells from anti-metastatic treatment. METHODS: To identify new signaling proteins involved in the plasticity of cancer cell invasiveness, we performed proteomic analysis of the amoeboid to mesenchymal transition with A375m2 melanoma cells in a 3D Matrigel matrix. RESULTS: In this screen we identified PKCα as an important protein for the maintenance of amoeboid morphology. We found that the activation of PKCα resulted in the mesenchymal-amoeboid transition of mesenchymal K2 and MDA-MB-231 cell lines. Consistently, PKCα inhibition led to the amoeboid-mesenchymal transition of amoeboid A375m2 cells. Next, we showed that PKCα inhibition resulted in a considerable decrease in the invading abilities of all analyzed cancer cell lines. CONCLUSIONS: Our results suggest that PKCα is an important protein for maintenance of the amoeboid morphology of cancer cells, and that downregulation of PKCα results in the amoeboid to mesenchymal transition. Our data also suggest that PKCα is important for both mesenchymal and amoeboid invasiveness, making it an attractive target for anti-metastatic therapies.

• MeSH
• invazivní růst nádoru genetika   patologie   MeSH
• lidé MeSH
• melanom genetika   patologie   MeSH
• mezoderm metabolismus   patologie   MeSH
• nádorové buněčné linie MeSH
• pohyb buněk genetika   MeSH
• proteinkinasa C-alfa biosyntéza   genetika   MeSH
• proteomika MeSH
• regulace genové exprese u nádorů MeSH
• signální transdukce MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• PRKCA protein, human MeSH Prohlížeč
• proteinkinasa C-alfa MeSH

The neural crest (NC) is a transient dynamic structure of ectodermal origin, found in early vertebrate embryos. The multipotential NC cells migrate along well defined routes, differentiate to various cell types including melanocytes and participate in the formation of various permanent tissues. As there is only limited information about the molecular mechanisms controlling early events in melanocyte specification and development, we exploited the AMV v-Myb transcriptional regulator, which directs differentiation of in vitro chicken NC cells to the melanocyte lineage. This activity is strictly dependent on v-Myb specifically binding to the Myb recognition DNA element (MRE). The two tamoxifen-inducible v-Myb alleles were constructed one which recognizes the MRE and one which does not. These were activated in ex ovo NC cells, and the expression profiles of resulting cells were analyzed using Affymetrix microarrays and RT-PCR. These approaches revealed up-regulation of the BMP antagonist Gremlin 2 mRNA, and down-regulation of mRNAs encoding several epithelial genes including KRT19 as very early events following the activation of melanocyte differentiation by v-Myb. The enforced v-Myb expression in neural tubes of chicken embryos resulted in detectable presence of Gremlin 2 mRNA. However, expression of Gremlin 2 in NC cells did not promote formation of melanocytes suggesting that Gremlin 2 is not the master regulator of melanocytic differentiation.

• Klíčová slova
• KRT19, Melanocyte development, PRDC, Tamoxifen-inducible v-Myb, v-Myb-dependent genes,
• MeSH
• aktivace transkripce * MeSH
• alely MeSH
• buněčná diferenciace * MeSH
• crista neuralis cytologie   MeSH
• keratin-19 genetika   metabolismus   MeSH
• kostní morfogenetický protein 5 genetika   metabolismus   MeSH
• kultivované buňky MeSH
• kuřecí embryo MeSH
• melanocyty fyziologie   MeSH
• mezibuněčné signální peptidy a proteiny genetika   metabolismus   MeSH
• onkogenní proteiny v-myb fyziologie   MeSH
• ptačí proteiny genetika   metabolismus   MeSH
• regulace genové exprese MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů MeSH
• transkriptom MeSH
• zvířata MeSH
• Check Tag
• kuřecí embryo MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• keratin-19 MeSH
• kostní morfogenetický protein 5 MeSH
• mezibuněčné signální peptidy a proteiny MeSH
• onkogenní proteiny v-myb MeSH
• ptačí proteiny MeSH