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. 2014 Nov;51(5):652–659. doi: 10.1165/rcmb.2014-0125OC

Figure 6.

Figure 6.

GMF-γ phosphorylation at Tyr-104 modulates actin polymerization and contraction in smooth muscle without affecting myosin activation. (A) Cells expressing WT GMF-γ or mutant Y104F GMF-γ were stimulated with 10−4 M ACh for 5 minutes or left unstimulated. F/G-actin ratios were evaluated as described in Materials and Methods (*P < 0.05; n = 6). (B) Myosin light chain (MLC) phosphorylation at Ser-19 was determined for cells expressing the recombinant proteins. (C) Verification of recombinant proteins in human bronchial tissues. Tissues were transfected with EGFP plasmid or plasmid encoding WT GMF-γ or mutant Y104F GMF-γ or left untransfected. Protein extracts of these cells were immunoblotted with EGFP antibody or GMF-γ antibody. Immunoblots are representative of four identical experiments. (D) Contractile responses of human bronchial rings to ACh (10−4 M) were determined, after which plasmid encoding WT or mutant Y104F GMF-γ was transduced into the tissues by reversible permeabilization and incubated for 3 days. Force development in bronchial rings was then determined. All values are means ± SE (*P < 0.05; n = 4).