Journal: Current Research in Microbial Sciences

Article Title: The lncRNA GAS5-encoded micropeptide facilitates influenza virus replication through modulation of the Wnt/β-catenin signaling pathway

doi: 10.1016/j.crmicr.2026.100559

Figure Lengend Snippet: GAS5-P50 interacts with NOTUM to enhance Wnt/β-catenin activation. (A) Proteins interacting with GAS5-P50 were identified through immunoprecipitation (IP) and subsequent silver staining. (B) Molecular docking of GAS5-P50 and NOTUM protein. (C, D) 293T cells were co-transfected with GAS5-P50-Flag and NOTUM-HA plasmids, followed by infection with PR8 influenza virus. GAS5-P50-Flag was immunoprecipitated from the lysates with anti-Flag antibody and the precipitates were blotted with anti-HA antibody (C). NOTUM-HA was immunoprecipitated from the lysates with anti-HA antibody and the precipitates were blotted with anti-Flag antibody (D). (E) 293T cells were transfected with either EV or GAS5-P50-Flag plasmids, followed by infection with PR8 influenza virus. GAS5-P50-Flag was immunoprecipitated from the lysates with anti-Flag antibody and the precipitates were subjected to immunoblotting with anti-NOTUM antibody. (F) 293T cells were transfected with EV, GAS5-P50, NOTUM, or GAS5-P50 together with NOTUM, respectively. After 16 h of infection with PR8 influenza virus, viral titers in the supernatants were determined by plaque assay. (G) 293T cells were transfected with EV, GAS5-P50, NOTUM, or GAS5-P50 together with NOTUM, followed by treatment with Wnt3a (1 μg/mL). The expression of β-catenin was examined by Western blotting. (H, I) 293T cells were transfected with EV, GAS5-P50, NOTUM, or GAS5-P50 together with NOTUM, respectively. After 16 h of infection with PR8 influenza virus, the expression of AXIN2 (H) and LEF1 (I) was examined by RT-qPCR. (J, K) A549 cells were treated with Wnt3a (1 μg/mL) (J) or IWP-2 (50 μM) (K), followed by infection with PR8 influenza virus for 16 h. Viral titers in the supernatants were determined by plaque assay. Data are represented as mean ± SD; Shown are representative data from three biologically independent experiments; * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, and ns represents no significance.

Article Snippet: Wnt3a and IWP-2 were purchased from MedChemExpress (New Jersey, USA).

Techniques: Activation Assay, Immunoprecipitation, Silver Staining, Transfection, Infection, Virus, Western Blot, Plaque Assay, Expressing, Quantitative RT-PCR