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. 2021 Sep 27;95(20):e00837-21. doi: 10.1128/JVI.00837-21

FIG 2.

FIG 2

Characterization of A549 cells with inducible IFITM1, IFITM2, or IFITM3 overexpression. (A) Lentiviral transduction was used to generate A549 cell lines with DOX-inducible expression of IFITM1, IFITM2, or IFITM3 proteins, each with an N-terminal FLAG tag (IFITM1i, IFITM2i, and IFITM3i cell lines). Constitutive expression of mCherry was used to determine transduction efficiency during the generation of each cell line. Control A549 cells express an irrelevant inducible protein without a FLAG-tag. (B) Flow cytometric analysis of FLAG expression in IFITM1i, IFITM2i, and IFITM3i A549 cell lines following culture for 24 hours in media supplemented with (+DOX) or without (−DOX) doxycycline. (C, D, E, and F) IFITMs were induced in A549 cells by DOX treatment for 24 hours, and expression of each specific IFITM (approximately 15 kDa in size) in cell lysates was confirmed by Western blotting. Proteins were detected using anti-FLAG antibody (C) or antibodies specific for IFITM1 (D), IFITM2 (E), or IFITM3 (F). Beta-actin (42 kDa) was used as a protein loading control.