Imaging the Impact of Chemically Inducible Proteins on Cellular Dynamics In Vivo

Leong, Hon S.; Lizardo, Michael M.; Ablack, Amber; McPherson, Victor; Wandless, Thomas J.; Chambers, Ann F.; Lewis, John D.

doi:10.1371/journal.pone.0030177

Cited by 12 publications

(12 citation statements)

References 29 publications

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“…Paraffin sections of breast cancer and pancreatic cancer xenografts were used as positive controls for GPC1 staining. GPC1-positive breast cancer cells (MDA-MB 231) and pancreatic cancer cells (PANC-1) were onplanted onto chorioallantoic membrane of chicken embryos [31, 32]. After 7 days of growth in vivo , tumors formed were extracted and embedded in paraffin or OCT for immunohistochemistry analysis.…”

Section: Methodsmentioning

confidence: 99%

Glypican-1 and glycoprotein 2 bearing extracellular vesicles do not discern pancreatic cancer from benign pancreatic diseases

et al. 2019

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Pancreatic ductal adenocarcinoma (PDAC) is a lethal disease that is clinically asymptomatic in its early stages of development. Non-invasive testing for pancreatic cancer biomarkers would significantly improve early detection and patient care. Extracellular vesicles (EVs) are circulating tumor fragments present in the blood and may express cancer specific biomarkers that would enable early detection of pancreatic cancer. We tested the utility of a blood test enumerating EVs positive for the pancreas-specific marker Glycoprotein 2 (GP2) and the putative pancreatic cancer marker Glypican-1 (GPC1) in patients with PDAC. Various levels of GPC1-positive and GP2/GPC1-positive EVs were detected in PDAC patients but were not significantly higher than benign pancreatic disease (BPD) patients. The sensitivity and specificity of the GPC1 EV test was 26.67% and 87.50% respectively, whereas the sensitivity and specificity for the GPC1+GP2 EV test was 23.33% and 90.00% respectively. Immunohistochemistry of GPC1 expression in a tissue microarray of PDAC and various controls also did not demonstrate specificity of GPC1 to PDAC. Hence, enumeration of GPC1-positive EVs, solely or in conjunction with GP2, was unable to effectively distinguish between BPD and pancreatic cancer.

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Section: Methodsmentioning

confidence: 99%

Glypican-1 and glycoprotein 2 bearing extracellular vesicles do not discern pancreatic cancer from benign pancreatic diseases

et al. 2019

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“…This tunable E-cad-zsGreen cell line gains E-cad expression when activated by the cell-permeant high-affinity ligand Shield-1. 20 First, we sought to determine if E-cad influenced extravasation rates, which would be an indicator of differential survival and colonization ability. In all cell lines (MDA-MB-231 E-cad + /E-cadcell line pair and the MDA-MB-231 E-cad-zsGreen cell line treated with Shield-1), we did not observe a statistically significant difference in the percentage of cells that successfully extravasated into the stroma of the CAM (Fig.…”

Section: E-cadherin Promotes Growth At the Distant Metastatic Sitementioning

confidence: 99%

“…This tunable E-cad-zsGreen cell line gains E-cad expression when activated by the cell-permeant high-affinity ligand Shield-1. 20…”

Section: E-cadherin Promotes Growth At the Distant Metastatic Sitementioning

confidence: 99%

E-cadherin interacts with EGFR resulting in hyper-activation of ERK in multiple models of breast cancer

Russo

Karl

Clark

et al. 2020

Preprint

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The loss of the intercellular adhesion molecule E-cadherin is a hallmark of the epithelial-mesenchymal transition (EMT), which promotes a transition of cancer cells to a migratory and invasive phenotype. E-cadherin is associated with a decrease in cell proliferation in normal cells. Here, using physiologically relevant 3D in vitro models, we find that E-cadherin induces hyper-proliferation in breast cancer cells through activation of the Raf/MEK/ERK signaling pathway. These results were validated and consistent across multiple in vivo models of primary tumor growth and metastatic outgrowth. E-cadherin expression dramatically increases tumor growth and, without affecting the ability of cells to extravasate and colonize the lung, significantly increases macrometastasis formation via cell proliferation at the distant site. Pharmacological inhibition of MEK1/2, blocking phosphorylation of ERK in E-cadherin-expressing cells, significantly depresses both tumor growth and macrometastasis. This work suggests a novel role of E-cadherin in tumor progression and identifies a potential new target to treat hyper-proliferative breast tumors.

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“…The addition of a specific small molecule ligand, Shield-1, can rescue the fusion protein from degradation in a rapid, dose-dependent, and reversible manner. This system has been widely applied in variety of cell types and organisms [3] , [4] , [5] , [6] , [7] , [8] , [9] , [10] , [11] .…”

Section: Introductionmentioning

confidence: 99%

Intracellular Context Affects Levels of a Chemically Dependent Destabilizing Domain

Sellmyer

Chen²,

Egeler³

et al. 2012

PLoS ONE

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The ability to regulate protein levels in live cells is crucial to understanding protein function. In the interest of advancing the tool set for protein perturbation, we developed a protein destabilizing domain (DD) that can confer its instability to a fused protein of interest. This destabilization and consequent degradation can be rescued in a reversible and dose-dependent manner with the addition of a small molecule that is specific for the DD, Shield-1. Proteins encounter different local protein quality control (QC) machinery when targeted to cellular compartments such as the mitochondrial matrix or endoplasmic reticulum (ER). These varied environments could have profound effects on the levels and regulation of the cytoplasmically derived DD. Here we show that DD fusions in the cytoplasm or nucleus can be efficiently degraded in mammalian cells; however, targeting fusions to the mitochondrial matrix or ER lumen leads to accumulation even in the absence of Shield-1. Additionally, we characterize the behavior of the DD with perturbants that modulate protein production, degradation, and local protein QC machinery. Chemical induction of the unfolded protein response in the ER results in decreased levels of an ER-targeted DD indicating the sensitivity of the DD to the degradation environment. These data reinforce that DD is an effective tool for protein perturbation, show that the local QC machinery affects levels of the DD, and suggest that the DD may be a useful probe for monitoring protein quality control machinery.

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Imaging the Impact of Chemically Inducible Proteins on Cellular Dynamics In Vivo

Cited by 12 publications

References 29 publications

Glypican-1 and glycoprotein 2 bearing extracellular vesicles do not discern pancreatic cancer from benign pancreatic diseases

Glypican-1 and glycoprotein 2 bearing extracellular vesicles do not discern pancreatic cancer from benign pancreatic diseases

E-cadherin interacts with EGFR resulting in hyper-activation of ERK in multiple models of breast cancer

Intracellular Context Affects Levels of a Chemically Dependent Destabilizing Domain

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