Key result
Inhibition of MEK1 using U0126, PD184352, or a dominant-negative mutant reduced hepatitis C virus RNA replication, suggesting low-level Ras-Erk signalling is required for replication.
Low level Ras-Erk signalling activity is required for HCV RNA replication, and its perturbation by NS5A may regulate genomic RNA replication.
MEK1 inhibition may curb HCV replication in models; hypothesis-generating for Ras-Erk targeting, with no clinical implications yet.
The hepatitis C virus NS5A protein has been previously demonstrated to partially attenuate activation of the Ras-Erk signalling pathway, via a conserved class II polyproline motif located towards the C terminus of the protein. However, the role of Ras-Erk signalling in the virus life cycle remains undetermined. To investigate this, levels of RNA replication were measured in genotypes 1 and 2 transient luciferase subgenomic replicon systems in the context of either pharmacological or genetic (dominant-negative) inhibition of MEK1, a kinase in the Ras-Erk signalling cascade. Incubation in the presence of two inhibitors (U0126 and PD184352) resulted in a decrease in the levels of RNA replication, conversely incubation with inhibitor PD98059 resulted in a modest increase in replication. The results obtained with PD98059 could not be explained by an off-target effect on Cox-2, stability of replicon RNA or stimulation of global translation levels, suggesting stimulation by a yet uncharacterized mechanism. To verify data obtained using pharmacological inhibitors the transient replicon RNA was co-electroporated with a dominant-negative mutant of MEK1. This resulted in a reduction in replication, confirming data seen with U0126 and PD184352. Our data are consistent with the hypothesis that a low level Ras-Erk signalling activity is required for RNA replication. However, complete inhibition of Ras-Erk signalling is inhibitory. These results suggest that perturbation of this signalling pathway by NS5A may be a mechanism to regulate levels of genomic RNA replication.
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Gretton et al. (2009) studied Hepatitis C virus infection (in vitro model). Pharmacological or genetic inhibition of MEK1 was evaluated on Levels of RNA replication. Inhibition of MEK1 using U0126, PD184352, or a dominant-negative mutant reduced hepatitis C virus RNA replication, suggesting low-level Ras-Erk signalling is required for replication.
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