Fig 5.

Comparison of siRNAs targeting the UL54-57 locus with siX3 targeting the UL123/122 IE genes. (A and B) HEL fibroblasts were untransfected or transfected with UL54A, UL54B, or CON siRNAs 24 h prior to infection with HCMV at an MOI of 0.1. Results from a similar experiment done at an MOI of 1.0 are shown in Fig. S3B to D in the supplemental material. (A) The levels of viral IE, E, and E/L protein expression were assessed by immunoblot analysis for IE1/IE2, pp65, and gB55, respectively. Results for the UL54B siRNA are shown in Fig. S3A in the supplemental material. (B) Culture supernatants were assayed for infectious virus production by plaque assay. (C and D) U373MG cells were untransfected or transfected with UL54A, UL54B, or CON siRNAs 24 h prior to infection with HCMV at an MOI of 0.3. (C) The levels of viral IE, E, and E/L protein expression were assessed by immunoblot analysis for IE1/IE2, pp65, and gB55, respectively. Results for the UL54B siRNA are shown in Fig. S4 in the supplemental material. (D) Culture supernatants and clarified cell lysates were combined and assayed for infectious virus by plaque assay. (E and F) Comparison of siUL54A with siX3 pretreatment in compromising HCMV replication. HEL fibroblast cells were untransfected or transfected with UL54A, X3, or CON siRNAs 24 h prior to infection with HCMV at an MOI of 0.1. (E) The levels of viral IE, E, and E/L protein expression were assessed by immunoblot analysis for IE1/IE2, pp65, and gB55, respectively. (F) Culture supernatants were assayed for infectious virus production by plaque assay. The boxed region in panel A labeled with an asterisk shows a longer exposure of that region of the membrane.