FIG 5.
Construction and entry profile of KGNEreg. (A) Schematic representation of the genome structure of KGNEreg. UL, unique long segment; US, unique short segment; CMVp, HCMV IE promoter; gB:NT, D285N/A549T mutations in gB; gDschEREG, gD mutant that contains schEREG as depicted below; TM, transmembrane domain; N, amino terminus; C, carboxyl terminus. Numbers indicate amino acid positions relative to the start of the mature protein (position 1). (G4S)3, triple repeat of Gly-Gly-Gly-Gly-Ser; G4SGS, Gly-Gly-Gly-Gly-Ser-Gly-Ser; closed boxes, terminal and internal inverted repeats. (B) Cells shown above the panels were infected with KGNEreg at a multiplicity of infection (MOI) of 1 for 24 h, and EGFP fluorescence was recorded. Mock, mock-transduced. Bars, 1 mm. (C) U87 cells transfected for 3 d with the siRNAs shown above the panels were infected with KGNEreg at an MOI of 3 for 16 h, and EGFP fluorescence was recorded. siNC, negative-control siRNA; sihEREGs 1, 2, and 3, anti-hEREG siRNAs HSS176606, 176607, and 176608, respectively. Bars, 300 μm. (D) (Upper) Cells shown above the panels were infected with KGNEreg at an MOI of 3 for 16 h, and EGFP fluorescence was recorded. Bars, 300 μm. (Lower) Flow cytometry of cells shown above the panels was performed using U#1 (open histogram) or an isotype-matched negative-control Ab (MG3-35) (closed histogram) as a primary Ab and a goat anti-mouse IgG-Alexa Fluor 488 (AF488) conjugate as a secondary Ab.