Ebola virus VP35 hijacks the PKA-CREB1 pathway for replication and pathogenesis by AKIP1 association
Zhu L, Gao T, Huang Y, Jin J, Wang D, Zhang L, Jin Y, Li P, Hu Y, Wu Y, Liu H, Dong Q, Wang G, Zheng T, Song C, Bai Y, Zhang X, Liu Y, Yang W, Xu K, Zou G, Zhao L, Cao R, Zhong W, Xia X, Xiao G, Liu X, Cao C
Applications: WB
Reactive species: Ebola virus
"Abstract: Ebola virus (EBOV), one of the deadliest viruses, is the cause of fatal Ebola virus disease (EVD). The underlying mechanism of viral replication and EBOV-related hemorrhage is not fully understood. Here, we show that EBOV VP35, a cofactor of viral RNA-dependent RNA polymerase, binds human A kinase interacting protein (AKIP1), which consequently activates protein kinase A (PKA) and the PKA-downstream transcription factor CREB1. During EBOV infection, CREB1 is recruited into EBOV ribonucleoprotein complexes in viral inclusion bodies (VIBs) and employed for viral replication. AKIP1 depletion or PKA-CREB1 inhibition dramatically impairs EBOV replication. Meanwhile, the transcription of several coagulation-related genes, including THBD and SERPINB2, is substantially upregulated by VP35-dependent CREB1 activation, which may contribute to EBOV-related hemorrhage. The finding that EBOV VP35 hijacks the host PKA-CREB1 signal axis for viral replication and pathogenesis provides novel potential therapeutic approaches against EVD."
Article snippet: Immunoblotting was performed with anti-VP35 (Creative Diagnostics, CABT-B292 or CABT-B1099, 1:1000 dilution) and anti-L (Creative Diagnostics, CABT-B1095, 1:500 dilution) antibodies. 
Figure 1. Lysates of HEK293 cells cotransfected with the indicated plasmids were incubated with/without recombinant His-VP35 for 2h and then subjected to immunoprecipitation and immunoblotting analysis. 
Figure 2. HepG2 cells infected with Ad-VP35 or Ad-GFP (MOI=10) for 48h were treated with 25μM FSK or 10μM H89 for 4h and then analyzed using immunoblotting. 
Figure 3. Lysates of HepG2 cells transfected with the EBOV minigenome (p0) in the presence (upper panel) or absence (lower panel) of the T7 RNA polymerase expression plasmid pCAGGS-T7 were separated on a 25 to 60% (W/V) sucrose gradient. Fractions were collected and analyzed by immunoblotting.
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