FIG 1.

gB/gp64 and gB/VSV-G chimeras are capable of forming cell syncytia. (A and B) HEK293T cells transiently expressing native gB, the CTD mutants gB-mutEndo and gB-HSVmutLL, the gB/gp64 chimera gB703/461gp64, and the gB-VSV-G chimeras gB771/483VSV-G, gB750/463VSV-G, and gB749/468VSV-G (alternatively termed gB/VSV-G). For indirect immunofluorescence analysis, cells were stained with anti-gB MAb 27-287, and nuclei were visualized by counterstaining with DAPI. The schematic representations on the right-hand side illustrate the domain architecture of native HCMV gB and the respective mutants. The individual protein domains are displayed in different colors; signal peptide (SP), membrane proximal region (MPR), transmembrane (TM) domain, and C-terminal domain (CTD/CT) are indicated. (A) gB-mutEndo harbors the endocytosis motif mutations Y845A LL883/884AA Y894A in the CTD. The gB-HSVmutLL chimera contains the TM and CTD of HSV-1 including the LL871/872AA mutation of hyperfusogenic HSV-1 gB. (B) Domains derived from gp64 and VSV-G are displayed in blue and magenta, respectively. Numbers indicate the first and last amino acids of gB (black), gp64 (blue), and VSV-G (magenta), respectively. (C) Box plot of the number of nuclei per syncytium as counted upon transfection of HEK293T cells with the individual gB constructs indicated in panel B and in Fig. 2B. At about 2 days posttransfection, cells were fixed and stained for expression of the gB/chimera. At least 50 gB-positive cells/syncytium were counted for each construct.