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G-Alpha Subunit Abundance and Activity Differentially Regulate ?-Catenin Signaling.

ABSTRACT: Heterotrimeric G proteins are signal transduction proteins involved in regulating numerous signaling events. In particular, previous studies have demonstrated a role for G-proteins in regulating ?-catenin signaling. However, the link between G-proteins and ?-catenin signaling is controversial and appears to depend on G-protein specificity. We describe a detailed analysis of a link between specific G-alpha subunits and ?-catenin using G-alpha subunit genetic knockout and knockdown approaches. The Pasteurella multocida toxin was utilized as a unique tool to activate G-proteins, with LiCl treatment serving as a ?-catenin signaling agonist. The results show that Pasteurella multocida toxin (PMT) significantly enhanced LiCl-induced active ?-catenin levels in HEK293T cells and mouse embryo fibroblasts. Evaluation of the effect of specific G-alpha proteins on the regulation of ?-catenin showed that Gq/11 and G12/13 knockout cells had significantly higher levels of active and total ?-catenin than wild-type cells. The stimulation of active ?-catenin by PMT and LiCl was lost upon both constitutive and transient knockdown of G12 and G13 but not Gq Based on our results, we conclude that endogenous G-alpha proteins are negative regulators of active ?-catenin; however, PMT-activated G-alpha subunits positively regulate LiCl-induced ?-catenin expression in a G12/13-dependent manner. Hence, G-alpha subunit regulation of ?-catenin is context dependent.

SUBMITTER: Banu A

PROVIDER: S-EPMC6379582 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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G-Alpha Subunit Abundance and Activity Differentially Regulate β-Catenin Signaling.

Banu Arshiya A Liu Karen J KJ Lax Alistair J AJ Grigoriadis Agamemnon E AE

Molecular and cellular biology 20190215 5

Heterotrimeric G proteins are signal transduction proteins involved in regulating numerous signaling events. In particular, previous studies have demonstrated a role for G-proteins in regulating β-catenin signaling. However, the link between G-proteins and β-catenin signaling is controversial and appears to depend on G-protein specificity. We describe a detailed analysis of a link between specific G-alpha subunits and β-catenin using G-alpha subunit genetic knockout and knockdown approaches. The ...[more]

PMID: 30559307

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Project description:BackgroundTumor necrosis factor-α (TNF-α) is an immunostimulatory cytokine that is consistently high in the breast tumor microenvironment (TME); however, its differential role in mitochondrial functions and cell survival in ER/PR +ve and ER/PR -ve breast cancer cells is not well understood.MethodsIn the current study, we investigated TNF-α modulated mitochondrial proteome using high-resolution mass spectrometry and identified the differentially expressed proteins in two different breast cancer cell lines, ER/PR positive cell line; luminal, MCF-7 and ER/PR negative cell line; basal-like, MDA-MB-231 and explored its implication in regulating the tumorigenic potential of breast cancer cells. We also compared the activity of mitochondrial complexes, ATP, and ROS levels between MCF-7 and MDA-MB-231 in the presence of TNF-α. We used Tumor Immune Estimation Resource (TIMER) webserver to analyze the correlation between TNF-α and mitochondrial proteins in basal and luminal breast cancer patients. Kaplan-Meier method was used to analyze the correlation between mitochondrial protein expression and survival of breast cancer patients.ResultsThe proteome analysis revealed that TNF-α differentially altered the level of critical proteins of mitochondrial respiratory chain complexes both in MCF-7 and MDA-MB-231, which correlated with differential assembly and activity of mitochondrial ETC complexes. The inhibition of the glycolytic pathway in the presence of TNF-α showed that glycolysis is indispensable for the proliferation and clonogenic ability of MDA-MB-231 cells (ER/PR -ve) as compared to MCF-7 cells (ER/PR +ve). The TIMER database showed a negative correlation between the expressions of TNF-α and key regulators of mitochondrial OXPHOS complexes in basal breast vs lobular carcinoma. Conversely, patient survival analysis showed an improved relapse-free survival with increased expression of identified proteins of ETC complexes and survival of the breast cancer patients.ConclusionThe evidence presented in our study convincingly demonstrates that TNF-α regulates the survival and proliferation of aggressive tumor cells by modulating the levels of critical assembly factors and subunits involved in mitochondrial respiratory chain supercomplexes organization and function. This favors the rewiring of mitochondrial metabolism towards anaplerosis to support the survival and proliferation of breast cancer cells. Collectively, the results strongly suggest that TNF-α differentially regulates metabolic adaptation in ER/PR +ve (MCF-7) and ER/PR -ve (MDA-MB-231) cells by modulating the mitochondrial supercomplex assembly and activity.

Dataset Information

G-Alpha Subunit Abundance and Activity Differentially Regulate ?-Catenin Signaling.

Publications

G-Alpha Subunit Abundance and Activity Differentially Regulate β-Catenin Signaling.

Similar Datasets

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