Visualizing Dynamic Changes During TGF-β-Induced Epithelial to Mesenchymal Transition

Sinha, Abhishek; Mehta, Pranav; Fan, Chuannan; Zhang, Jing; Marvin, Dieuwke L.; Dinther, Maarten van; Ritsma, Laila; Boukany, Pouyan E.; Dijke, Peter ten

doi:10.1007/978-1-0716-2277-3_5

Cited by 4 publications

(3 citation statements)

References 26 publications

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“…For detection of SMURF2, cells were incubated first with a primary antibody against SMURF2 (1:100 dilution; Santa Cruz; sc‐393848) or MYC (1:100 dilution; Santa Cruz; sc40) for 45 min at room temperature and then with a secondary antibody (Invitrogen; A21428) for 1 h at room temperature. For F‐actin immunofluorescent staining, cells were incubated with Phalloidin conjugated with Alexa Fluor 488 (1:500 dilution; Thermo Fisher Scientific; A12379) for 30 min at room temperature as described before (Sinha et al , 2022 ). VECTASHIELD Antifade Mounting Medium with DAPI (Vector Laboratories; H‐1200) was used to mount coverslips, and images were acquired with a Leica SP8 confocal microscope (Leica Microsystems).…”

Section: Methodsmentioning

confidence: 99%

LncRNA LITATS1 suppresses TGF‐β‐induced EMT and cancer cell plasticity by potentiating TβRI degradation

et al. 2023

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Epithelial cells acquire mesenchymal phenotypes through epithelial-mesenchymal transition (EMT) during cancer progression. However, how epithelial cells retain their epithelial traits and prevent malignant transformation is not well understood. Here, we report that the long noncoding RNA LITATS1 (LINC01137, ZC3H12A-DT) is an epithelial gatekeeper in normal epithelial cells and inhibits EMT in breast and non-small cell lung cancer cells. Transcriptome analysis identified LITATS1 as a TGF-b target gene. LITATS1 expression is reduced in lung adenocarcinoma tissues compared with adjacent normal tissues and correlates with a favorable prognosis in breast and non-small cell lung cancer patients. LITATS1 depletion promotes TGF-b-induced EMT, migration, and extravasation in cancer cells. Unbiased pathway analysis demonstrated that LITATS1 knockdown potently and selectively potentiates TGF-b/SMAD signaling. Mechanistically, LITATS1 enhances the polyubiquitination and proteasomal degradation of TGF-b type I receptor (TbRI). LITATS1 interacts with TbRI and the E3 ligase SMURF2, promoting the cytoplasmic retention of SMURF2. Our findings highlight a protective function of LITATS1 in epithelial integrity maintenance through the attenuation of TGF-b/SMAD signaling and EMT.

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Section: Methodsmentioning

confidence: 99%

LncRNA LITATS1 suppresses TGF‐β‐induced EMT and cancer cell plasticity by potentiating TβRI degradation

et al. 2023

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“…To evaluate the expression and localization of F-actin, fluorescent staining was performed as previously described ( 74 , 75 ). Briefly, A549 cells were stimulated with SB431542 (SB; 10 μM) or TGF-β (1 ng/ml) or the corresponding vehicle for 48 hours.…”

Section: Methodsmentioning

confidence: 99%

The lncRNA LETS1 promotes TGF-β–induced EMT and cancer cell migration by transcriptionally activating a TβR1-stabilizing mechanism

et al. 2023

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Transforming growth factor–β (TGF-β) signaling is a critical driver of epithelial-to-mesenchymal transition (EMT) and cancer progression. In SMAD-dependent TGF-β signaling, activation of the TGF-β receptor complex stimulates the phosphorylation of the intracellular receptor-associated SMADs (SMAD2 and SMAD3), which translocate to the nucleus to promote target gene expression. SMAD7 inhibits signaling through the pathway by promoting the polyubiquitination of the TGF-β type I receptor (TβRI). We identified an unannotated nuclear long noncoding RNA (lncRNA) that we designated LETS1 (lncRNA enforcing TGF-β signaling 1) that was not only increased but also perpetuated by TGF-β signaling. Loss of LETS1 attenuated TGF-β–induced EMT and migration in breast and lung cancer cells in vitro and extravasation of the cells in a zebrafish xenograft model. LETS1 potentiated TGF-β–SMAD signaling by stabilizing cell surface TβRI, thereby forming a positive feedback loop. Specifically, LETS1 inhibited TβRI polyubiquitination by binding to nuclear factor of activated T cells (NFAT5) and inducing the expression of the gene encoding the orphan nuclear receptor 4A1 (NR4A1), a component of a destruction complex for SMAD7. Overall, our findings characterize LETS1 as an EMT-promoting lncRNA that potentiates signaling through TGF-β receptor complexes.

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“…38,39 A549-VIM-RFP cells have been previously reported to exhibit EMT with an upregulation in VIM-RFP fluorescence upon exogenous TGF-β stimulation. 40 The dual-reporter A549 cells were maintained in Dulbecco's modified eagle medium high glucose (DMEM, Sigma) containing 4.5 g L −1 glucose, L-glutamine without sodium pyruvate, and supplemented with 10% fetal bovine serum (FBS, Sigma) and 1% antibiotic-antimycotic solution (Gibco). All cells were incubated at 37 °C with 5% CO 2 and sub-cultured 2 times per week.…”

Section: Cell Culturementioning

confidence: 99%

Interstitial flow potentiates TGF-β/Smad-signaling activity in lung cancer spheroids in a 3D-microfluidic chip

Rahman,

Bordoloi,

Rouhana

et al. 2024

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Visualizing Dynamic Changes During TGF-β-Induced Epithelial to Mesenchymal Transition

Cited by 4 publications

References 26 publications

LncRNA LITATS1 suppresses TGF‐β‐induced EMT and cancer cell plasticity by potentiating TβRI degradation

LncRNA LITATS1 suppresses TGF‐β‐induced EMT and cancer cell plasticity by potentiating TβRI degradation

The lncRNA LETS1 promotes TGF-β–induced EMT and cancer cell migration by transcriptionally activating a TβR1-stabilizing mechanism

Interstitial flow potentiates TGF-β/Smad-signaling activity in lung cancer spheroids in a 3D-microfluidic chip

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