Inhibition of MLC Phosphorylation Restricts Replication of Influenza Virus—A Mechanism of Action for Anti-Influenza Agents
Figure 9
Induction of MLC phosphorylation reverses the inhibitory effects of HRas\ERK on influenza proliferation.
The anti-influenza effects of Raf kinase and MEK/ERK inhibitors were reversed when HUVECs were treated with an MLC phosphatase inhibitor. (A) MDCK cells were treated as indicated GW5074 and U0126 at 10 µM each overnight; calyculin A at 0.5 µM for 1 h] and then infected with influenza (MOI, 0.01). After 24 h, virus yield was measured in the medium by using RT-PCR. HRas-enhanced influenza proliferation is attenuated by inhibition of MLC kinase, ERK and Raf-1. (B) MDCK cells were seeded on 24-well plates, transduced with the indicated adenovirus, and transfected 24 h later with the indicated constructs. The cells were infected with influenza A virus (MOI, 0.01) and 24 h after the infection, RT-PCR was used to measure the virus yield in the medium. Inhibition of Rho kinase 1 in human bronchial epithelial cells leads to inhibition of the nuclear translocation of influenza RNP complexes. (C) Human bronchial epithelial cells were seeded on chamber slides, transfected with the indicated siRNA, and infected 24 h later with influenza virus (MOI, 10). After 8 h, the cells were stained for influenza nucleoprotein (NP) protein. Hoechst 33342 was used as a nuclear counterstain (blue). Bar, 5 µm; *P<0.01; N = 4 for each experiment.