FIG. 2.

Effect of RBL2 expression on influenza virus replication and growth. (A) ΔN-qRBL2 increases influenza virus replication in a minireplicon assay. 293T cells were transfected with plasmids expressing HK483 PB1, PB2-627K, PA, and NP proteins, pPolI-Luc, and ΔN-qRBL2. Twenty-four hours after incubation at 33°C, luciferase expression was detected. (B) Viral polymerase activity in cells expressing hRBL2 or qRBL2. 293 cells were transfected with plasmids expressing qRBL2 or hRBL2 or a control vector. Twenty-four hours later, cells were infected with strain WSN (MOI of 1). Three hours later, RNA was extracted and quantified by real-time RT-PCR with primer sets specific for NP vRNA, cRNA, or mRNA. These values were normalized to beta-actin. The error bars represent standard deviations (n = 3). (C) Viral M1 protein production in cells overexpressing hRBL2. 293 cells expressing hRBL2 or a control vector were infected with strain WSN (MOI of 3). At the indicated hours postinfection (hpi), cell lysates were subjected to Western blot analysis with antibodies against M1 and beta-actin. The values show the ratio of M1 to actin normalized to control cells 4 h after infection. (D) Influenza virus titers in 293 cells overexpressing hRBL2 or qRBL2. 293 cells overexpressing hRBL2 (square), qRBL2 (triangle), or a control vector (circle) were infected with strain WSN (MOI of 0.05). The cells were incubated at 37°C for the indicated time periods. Virus titers in the supernatant were determined by plaque assays in MDCK cells. The error bars represent standard deviations (n = 3). α, anti.