A Locus Encompassing the Epstein-Barr Virus bglf4 Kinase Regulates Expression of Genes Encoding Viral Structural Proteins
Figure 8
Knockdown of BGLF3 has no effect on expression of BGLF4.
To examine the effect of silencing BGLF3 expression on the mRNA and protein levels of BGLF4, we transfected 2089 cells with ZEBRA alone or together with siG3 or siG4. Cells were harvested after 48 h. RNA and protein samples were prepared for Northern (A and C) and Western (B) blot analyses. In cells transfected with ZEBRA, we detected four RNA species: 3.5, 3.3, 2.2 and 1.8 kb using the BGLF4 probe (A). The 4.3 kb was only detected using a probe unique to BGLF3.5 (C). Except for the 4.3 kb, the same RNA species were detected when siG3 was co-transfected. However, the 3.5, 3.3 and 2.2 kb RNA species disappeared in the sample co-transfected with siG4. These RNAs were replaced with two new species approximately 1.6 and 3 kb in size. Examining protein expression in the same samples demonstrated that siG4 and siG3 markedly reduced expression of the late FR3 protein (Fig. 8B). Nonetheless, similar to Fig. 9, siG3 had no effect on the level of the BGLF4 protein. The experiment was repeated twice and similar results were obtained. (D) A schematic diagram of the various transcripts expected to be synthesized from the late control locus. CMV, empty vector; Z, ZEBRA; siG3, siRNA to BGLF3; siG4, siRNA to BGLF4. The asterisks denote probable cleavage products.