scholarly journals Golgi stabilization, not its front-rear bias, is associated with EMT-enhanced fibrillar migration

2018 ◽  
Author(s):  
Robert J. Natividad ◽  
Mark L. Lalli ◽  
Senthil K. Muthuswamy ◽  
Anand R. Asthagiri
Keyword(s):  
Cell Migration ◽  
Single Cell ◽  
Epithelial To Mesenchymal Transition ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Migration Speed ◽  
Mesenchymal Transition ◽  
Positional Bias ◽  
Persistent Cell ◽  
Posterior Position

ABSTRACTEpithelial-to-mesenchymal transition (EMT) and maturation of collagen fibrils in the tumor microenvironment play a significant role in cancer cell invasion and metastasis. Confinement along fiber-like tracks enhances cell migration. To what extent and in what manner EMT further promotes migration in a microenvironment already conducive to migration is poorly understood. Here, we show that TGFβ-mediated EMT significantly enhances migration on fiber-like micropatterned tracks of collagen, doubling migration speed and quadrupling persistence relative to untreated mammary epithelial cells. Thus, cell-intrinsic EMT and extrinsic fibrillar tracks have non-redundant effects on motility. To better understand EMT-enhanced fibrillar migration, we investigated the regulation of Golgi positioning, which is involved in front-rear polarization and persistent cell migration. Confinement along fiber-like tracks has been reported to favor posterior Golgi positioning, whereas anterior positioning is observed during 2d wound healing. While EMT also regulates cell polarity, little is known about its effect on Golgi positioning. Here, we show that EMT induces a 2:1 rearward bias in Golgi positioning; however, positional bias explains less than 5% of single-cell variability in migration speed and persistence. Meanwhile, EMT significantly stabilizes Golgi positioning. Cells that enhance migration in response to TGFβ maintain Golgi position for 3-4 fold longer than untreated counterparts, irrespective of whether the Golgi is ahead or behind the nucleus. In fact, 35% of cells that respond to TGFβ exhibit a fully-committed Golgi phenotype with the organelle either in the anterior or posterior position for over 90% of the time. Furthermore, single-cell differences in Golgi stability capture up to 30% of variations in migration speed and persistence. These results lead us to propose that the Golgi is part of a core physical scaffold that distributes cell-generated forces necessary for migration. A stable scaffold more consistently, and therefore more productively, distributes forces over time, leading to efficient migration.

scholarly journals The H1047R PIK3CA oncogene induces a senescence-like state, pleiotropy and acute HSP90 dependency in HER2+ mammary epithelial cells

2019 ◽  
Vol 40 (10) ◽  
pp. 1179-1190 ◽  
Author(s):  
Anindita Chakrabarty ◽  
Sreeraj Surendran ◽  
Neil E Bhola ◽  
Vishnu S Mishra ◽  
Tasaduq Hussain Wani ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Epithelial To Mesenchymal Transition ◽  
Mammary Epithelial Cells ◽  
Breast Tumors ◽  
Genetically Engineered ◽  
Mammary Epithelial ◽  
Hsp90 Inhibition ◽  
Mesenchymal Transition ◽  
Pik3ca Mutations

Abstract In pre-clinical models, co-existence of Human Epidermal Growth Factor Receptor-2 (HER2)-amplification and PI3K catalytic subunit (PIK3CA) mutations results in aggressive, anti-HER2 therapy-resistant breast tumors. This is not always reflected in clinical setting. We speculated that the complex interaction between the HER2 and PIK3CA oncogenes is responsible for such inconsistency. We performed series of biochemical, molecular and cellular assays on genetically engineered isogenic mammary epithelial cell lines and breast cancer cells expressing both oncogenes. In vitro observations were validated in xenografts models. We showed that H1047R, one of the most common PIK3CA mutations, is responsible for endowing a senescence-like state in mammary epithelial cells overexpressing HER2. Instead of imposing a permanent growth arrest characteristic of oncogene-induced senescence, the proteome secreted by the mutant cells promotes stem cell enrichment, angiogenesis, epithelial-to-mesenchymal transition, altered immune surveillance and acute vulnerability toward HSP90 inhibition. We inferred that the pleiotropism, as observed here, conferred by the mutated oncogene, depending on the host microenvironment, contributes to conflicting pre-clinical and clinical characteristics of HER2+, mutated PIK3CA-bearing tumor cells. We also came up with a plausible model for evolution of breast tumors from mammary epithelial cells harboring these two molecular lesions.

Erythropoietin Induces an Epithelial to Mesenchymal Transition-Like Process in Mammary Epithelial Cells MCF10A

2017 ◽  
Vol 118 (9) ◽  
pp. 2983-2992 ◽  
Author(s):  
Alejandra Ordoñez-Moreno ◽  
Cecilia Rodriguez-Monterrosas ◽  
Pedro Cortes-Reynosa ◽  
Julio Isael Perez-Carreon ◽  
Eduardo Perez Salazar
Keyword(s):  
Epithelial Cells ◽  
Epithelial To Mesenchymal Transition ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Mesenchymal Transition

scholarly journals A multi-tiered map of EMT defines major transition points and identifies vulnerabilities

2021 ◽  
Author(s):  
Indranil Paul ◽  
Dante Bolzan ◽  
Heather Hook ◽  
Ahmed Youssef ◽  
Gopal Karemore ◽  
...  
Keyword(s):  
Epithelial To Mesenchymal Transition ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Tgfβ Signaling ◽  
Mesenchymal Transition ◽  
Whole Cells ◽  
Major Transition ◽  
Transition Points ◽  
Acid Pathway ◽  
Arachidonic Acid Pathway

TGFβ mediated epithelial to mesenchymal transition (EMT) proceeds through hybrid "E/M" states. A deeper understanding of these states and events which regulate entry to and exit from the E/M states is needed for therapeutic exploitation. We quantified >60,000 molecules across ten time points and twelve omic layers in mammary epithelial cells. Proteomes of whole cells, phosphoproteins, nucleus, extracellular vesicles, secretome and membrane resolved major shifts, E→E/M and E/M→M during EMT, and defined state-specific signatures. Metabolomics identified early activation of arachidonic acid pathway and an enzyme-mediated switch from Cytochrome P450 to Cyclooxygenase / Lipoxygenase branches during E→E/M. Single-cell transcriptomics identified GLIS2 as an early modulator of EMT. Integrative modeling-predicted combinatorial inhibition of AURKB, PP2A and SRC exposed vulnerabilities at E→E/M juncture. Covariance analysis revealed remarkable discordance between proteins and transcripts, and between proteomic layers, implying insufficiency of current approaches. Overall, this dataset provides an unprecedented resource on TGFβ signaling, EMT and cancer.

scholarly journals Upregulated-flotillins and sphingosine kinase 2 derail vesicular trafic to stabilize AXL and promote epithelial-mesenchymal transition

2020 ◽  
Author(s):  
Mallory Genest ◽  
Franck Comunale ◽  
Damien Planchon ◽  
Pauline Govindin ◽  
Sophie Vacher ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Epithelial To Mesenchymal Transition ◽  
Epithelial Mesenchymal Transition ◽  
Mammary Epithelial Cells ◽  
Sphingosine Kinase ◽  
Vesicular Trafficking ◽  
Mammary Epithelial ◽  
Lipid Kinase ◽  
Mesenchymal Transition ◽  
Sphingosine Kinase 2

AbstractAltered endocytosis and vesicular trafficking are major players during tumorigenesis. Flotillin overexpression, a feature observed in many invasive tumors, and identified as a marker of poor prognosis, induces a deregulated endocytic and trafficking pathway called Upregulated Flotillin-Induced Trafficking (UFIT). Here, we found that, in non tumoral mammary epithelial cells, induction of the UFIT pathway promotes epithelial-to-mesenchymal transition (EMT) and accelerates the endocytosis of several transmembrane receptors, including AXL, in flotillin-positive late endosomes. AXL overexpression, frequently observed in cancer cells, is linked to EMT and metastasis formation. In flotillin-overexpressing non-tumoral mammary epithelial cells and in invasive breast carcinoma cells, we found that the UFIT-mediated AXL endocytosis allows its stabilization and depends on sphingosine-kinase 2, a lipid kinase recruited in flotillin-rich plasma membrane-domains and endosomes.Thus, the deregulation of vesicular trafficking following flotillin upregulation, and through sphingosine kinase 2, emerges as a new mechanism of AXL overexpression and EMT-inducing signaling pathway activation.

Molecular Characterization of Epithelial to Mesenchymal Transition in Human Prostatic Epithelial Cells

2010 ◽  
Vol 1 (2) ◽  
Author(s):  
Victor K. Lin ◽  
Shih-Ya Wang ◽  
Lanxiao Wu ◽  
Smitha M. Rao ◽  
J. C. Chiao ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cancer Metastasis ◽  
Epithelial To Mesenchymal Transition ◽  
Mammary Epithelial Cells ◽  
Critical Role ◽  
Three Dimensional ◽  
Mammary Epithelial ◽  
Mesenchymal Transition ◽  
Enhanced Mobility ◽  
E Cadherin

Epithelial to mesenchymal transition (EMT) has been believed to play a critical role in cancer metastasis. TGFβ has been described as an inducer of EMT in normal mammary epithelial cells by signaling through receptor serine/threonine kinase pathways to regulate epithelial cell plasticity and invasion. In this study, we investigated the EMT cellular responses, including morphologic changes, phenotype switches, invasiveness enhancement, and cellular contraction alteration, in TGFβ stimulated human prostate normal epithelial cells (PZ-HPV-7). Migration of TGFβ treated PZ-HPV-7 cells across matrigel was measured in invasion chambers (8 μm pore size). The cells were treated with or without TGFβ (2 ng/ml) in PrEGM media for 3 days. Immunoblot assay was conducted and it was demonstrated that the induction of vimentin when stimulated by TGFβ was accompanied by a downregulation of E-cadherin, though p-cadherin level was not altered. It was also observed that there was a decrease in cytokaretin 5/6 expression associated with the downregulation of E-cadherin during the induction of EMT. In order to study the cell contraction, three-dimensional collage lattice assay was performed. It was demonstrated that TGFβ-stimulated PZ-HPV-7 cells gained contractility. Our results showed that TGFβ stimulation induced PZ-HPV-7 cells to undergo epithelial to mesenchymal transition. EMT characteristics such as acquisition of mesenchymal markers and loss of epithelial markers were evident in the induction of vimentin and downregulation of E-cadherin and cytokeratins, as well as phenotypic alterations including increased contraction and enhanced mobility were detected.

scholarly journals ESX induces transformation and functional epithelial to mesenchymal transition in MCF-12A mammary epithelial cells

Oncogene ◽  
2004 ◽  
Vol 23 (9) ◽  
pp. 1766-1779 ◽  
Author(s):  
Pepper J Schedin ◽  
Kristin L Eckel-Mahan ◽  
Shauntae M McDaniel ◽  
Jason D Prescott ◽  
Kelley S Brodsky ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Epithelial To Mesenchymal Transition ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Mesenchymal Transition

scholarly journals The Numb/p53 circuitry couples replicative self-renewal and tumor suppression in mammary epithelial cells

2015 ◽  
Vol 211 (4) ◽  
pp. 845-862 ◽  
Author(s):  
Daniela Tosoni ◽  
Silvia Zecchini ◽  
Marco Coazzoli ◽  
Ivan Colaluca ◽  
Giovanni Mazzarol ◽  
...  
Keyword(s):  
Mammary Gland ◽  
Cell Fate ◽  
Epithelial To Mesenchymal Transition ◽  
Mammary Epithelial Cells ◽  
Ex Vivo ◽  
Mammary Epithelial ◽  
Mesenchymal Transition ◽  
Morphological Alterations ◽  
Cell Fate Determinant ◽  
Mammary Stem

The cell fate determinant Numb orchestrates tissue morphogenesis and patterning in developmental systems. In the human mammary gland, Numb is a tumor suppressor and regulates p53 levels. However, whether this function is linked to its role in fate determination remains unclear. Here, by exploiting an ex vivo system, we show that at mitosis of purified mammary stem cells (SCs), Numb ensures the asymmetric outcome of self-renewing divisions by partitioning into the progeny that retains the SC identity, where it sustains high p53 activity. Numb also controls progenitor maturation. At this level, Numb loss associates with the epithelial-to-mesenchymal transition and results in differentiation defects and reacquisition of stemness features. The mammary gland of Numb-knockout mice displays an expansion of the SC compartment, associated with morphological alterations and tumorigenicity in orthotopic transplants. This is because of low p53 levels and can be inhibited by restoration of Numb levels or p53 activity, which results in successful SC-targeted treatment.

scholarly journals Combinatorial perturbation analysis reveals divergent regulations of mesenchymal genes during epithelial-to-mesenchymal transition

2019 ◽  
Author(s):  
Kazuhide Watanabe ◽  
Nicholas Panchy ◽  
Shuhei Noguchi ◽  
Harukazu Suzuki ◽  
Tian Hong
Keyword(s):  
Epithelial To Mesenchymal Transition ◽  
Mammary Epithelial Cells ◽  
Phenotypic Diversity ◽  
Gene Clusters ◽  
Regulatory Elements ◽  
Mammary Epithelial ◽  
Breast Cancer Patients ◽  
M Gene ◽  
Mesenchymal Transition ◽  
Regulatory Circuits

AbstractEpithelial-to-mesenchymal transition (EMT), a fundamental transdifferentiation process in development, produces diverse phenotypes in different physiological or pathological conditions. Many genes involved in EMT have been identified to date, but mechanisms contributing to the phenotypic diversity and those governing the coupling between the dynamics of epithelial (E) genes and that of the mesenchymal (M) genes are unclear. In this study, we employed combinatorial perturbations to mammary epithelial cells to induce a series of EMT phenotypes by manipulating two essential EMT-inducing elements, namely TGF-β and ZEB1. By measuring transcriptional changes in more than 700 E- and M-genes, we discovered that the M-genes exhibit a significant diversity in their dependency to these regulatory elements and identified three groups of M-genes that are controlled by different regulatory circuits. Notably, functional differences were detected among the M-gene clusters in motility regulation and in survival of breast cancer patients. We computationally predicted and experimentally confirmed that the reciprocity and reversibility of EMT are jointly regulated by ZEB1. Our integrative analysis reveals the key roles of ZEB1 in coordinating the dynamics of a large number of genes during EMT, and it provides new insights into the mechanisms for the diversity of EMT phenotypes.

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