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Loss of γ-cytoplasmic actin triggers myofibroblast transition of human epithelial cells.

Lechuga S, Baranwal S, Li C, Naydenov NG, Kuemmerle JF, Dugina V, Chaponnier C, Ivanov AI - Mol. Biol. Cell (2014)

Bottom Line: Such epithelial plasticity is accompanied by dramatic reorganizations of the actin cytoskeleton, although mechanisms underlying cytoskeletal effects on epithelial transdifferentiation remain poorly understood.Induction of EMyT in γ-CYA-depleted cells depended on activation of serum response factor and its cofactors, myocardial-related transcriptional factors A and B.Loss of γ-CYA stimulated formin-mediated actin polymerization and activation of Rho GTPase, which appear to be essential for EMyT induction.

View Article: PubMed Central - PubMed

Affiliation: Department of Human and Molecular Genetics, Virginia Commonwealth University, Richmond, VA 23298.

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EMyT induction in γ-CYA–depleted epithelial cells is controlled by actin polymerization, and γ-CYA preferentially interacts with MRTF. (A, B) γ-CYA–depleted A549 cells were treated for 24 h with either vehicle or the F-actin–depolymerizing drug Lat B (1 μM), and expression of EMyT markers was determined by immunoblotting. Data are presented as mean ± SE (n = 3); *p < 0.05. (C) Association of MRTF-A with γ-CYA and β-CYA in control and γ-CYA–depleted A549 cells was examined using immunoprecipitation with anti–MRTF-A antibody.
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Figure 6: EMyT induction in γ-CYA–depleted epithelial cells is controlled by actin polymerization, and γ-CYA preferentially interacts with MRTF. (A, B) γ-CYA–depleted A549 cells were treated for 24 h with either vehicle or the F-actin–depolymerizing drug Lat B (1 μM), and expression of EMyT markers was determined by immunoblotting. Data are presented as mean ± SE (n = 3); *p < 0.05. (C) Association of MRTF-A with γ-CYA and β-CYA in control and γ-CYA–depleted A549 cells was examined using immunoprecipitation with anti–MRTF-A antibody.

Mentions: To gain functional insight into the role of monomeric actin in EMyT induction, we treated γ-CYA–depleted A549 cells with either vehicle or an actin-depolymerizing drug, latrunculin B (LatB; Morton et al., 2000). LatB treatment dramatically attenuated γ-CYA–dependent expressional up-regulation of contractile proteins, highlighting the significant role of G-actin sequestration in EMyT (Figure 6, A and B). Finally, we sought to compare interactions of two cytoplasmic actins with MRTF. MRTF-A was immunoprecipitated from total A549 epithelial cell lysates and subsequently probed with monoclonal antibodies selectively recognizing either γ-CYA or β-CYA. Remarkably, only γ-CYA coprecipitated with MRTF-A, whereas β-CYA was not precipitated with this transcriptional regulator (Figure 6C). Similarly, γ-CYA but not β-CYA was detected in immunoprecipitates obtained with SRF antibody (unpublished data). In contrast, MRTF-A antibody pulled down β-CYA from γ-CYA–depleted A549 cell lysates (Figure 6C). These data suggest that γ-CYA and β-CYA can compete for MRTF binding and that MRTF preferentially binds to γ-CYA in control A549 cells. Of interest, loss of γ-CYA appears to decrease total expression of MRTF-A protein (Figures 4C and 6C). This provides additional indirect arguments in support of preferential MRTF–γ-CYA interactions that not only inactivate but might also stabilize this transcriptional regulator in epithelial cells.


Loss of γ-cytoplasmic actin triggers myofibroblast transition of human epithelial cells.

Lechuga S, Baranwal S, Li C, Naydenov NG, Kuemmerle JF, Dugina V, Chaponnier C, Ivanov AI - Mol. Biol. Cell (2014)

EMyT induction in γ-CYA–depleted epithelial cells is controlled by actin polymerization, and γ-CYA preferentially interacts with MRTF. (A, B) γ-CYA–depleted A549 cells were treated for 24 h with either vehicle or the F-actin–depolymerizing drug Lat B (1 μM), and expression of EMyT markers was determined by immunoblotting. Data are presented as mean ± SE (n = 3); *p < 0.05. (C) Association of MRTF-A with γ-CYA and β-CYA in control and γ-CYA–depleted A549 cells was examined using immunoprecipitation with anti–MRTF-A antibody.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Figure 6: EMyT induction in γ-CYA–depleted epithelial cells is controlled by actin polymerization, and γ-CYA preferentially interacts with MRTF. (A, B) γ-CYA–depleted A549 cells were treated for 24 h with either vehicle or the F-actin–depolymerizing drug Lat B (1 μM), and expression of EMyT markers was determined by immunoblotting. Data are presented as mean ± SE (n = 3); *p < 0.05. (C) Association of MRTF-A with γ-CYA and β-CYA in control and γ-CYA–depleted A549 cells was examined using immunoprecipitation with anti–MRTF-A antibody.
Mentions: To gain functional insight into the role of monomeric actin in EMyT induction, we treated γ-CYA–depleted A549 cells with either vehicle or an actin-depolymerizing drug, latrunculin B (LatB; Morton et al., 2000). LatB treatment dramatically attenuated γ-CYA–dependent expressional up-regulation of contractile proteins, highlighting the significant role of G-actin sequestration in EMyT (Figure 6, A and B). Finally, we sought to compare interactions of two cytoplasmic actins with MRTF. MRTF-A was immunoprecipitated from total A549 epithelial cell lysates and subsequently probed with monoclonal antibodies selectively recognizing either γ-CYA or β-CYA. Remarkably, only γ-CYA coprecipitated with MRTF-A, whereas β-CYA was not precipitated with this transcriptional regulator (Figure 6C). Similarly, γ-CYA but not β-CYA was detected in immunoprecipitates obtained with SRF antibody (unpublished data). In contrast, MRTF-A antibody pulled down β-CYA from γ-CYA–depleted A549 cell lysates (Figure 6C). These data suggest that γ-CYA and β-CYA can compete for MRTF binding and that MRTF preferentially binds to γ-CYA in control A549 cells. Of interest, loss of γ-CYA appears to decrease total expression of MRTF-A protein (Figures 4C and 6C). This provides additional indirect arguments in support of preferential MRTF–γ-CYA interactions that not only inactivate but might also stabilize this transcriptional regulator in epithelial cells.

Bottom Line: Such epithelial plasticity is accompanied by dramatic reorganizations of the actin cytoskeleton, although mechanisms underlying cytoskeletal effects on epithelial transdifferentiation remain poorly understood.Induction of EMyT in γ-CYA-depleted cells depended on activation of serum response factor and its cofactors, myocardial-related transcriptional factors A and B.Loss of γ-CYA stimulated formin-mediated actin polymerization and activation of Rho GTPase, which appear to be essential for EMyT induction.

View Article: PubMed Central - PubMed

Affiliation: Department of Human and Molecular Genetics, Virginia Commonwealth University, Richmond, VA 23298.

Show MeSH
Related in: MedlinePlus