Volume 8, Number 12, December 2002

Induction of Inflammation by West Nile virus Capsid through the Caspase-9 Apoptotic Pathway

Joo-Sung Yang,* Mathura P. Ramanathan,*¹ Karuppiah Muthumani,* Andrew Y. Choo,* Sung-Ha Jin,* Qian-Chun Yu,* Daniel S. Hwang,* Daniel K. Choo,* Mark D. Lee,* Kesen Dang,* WaixingTang,* J. Joseph Kim,† and David B. Weiner*
*University of Pennsylvania, Philadelphia, Pennsylvania, USA; and †Viral Genomix, Inc., Philadelphia, Pennsylvania, USA

Figure 5. Apoptosis determining domain in WNV-Cp gene. a, Schematic diagram of pcWNV-Cp-DJY, pcWNV-CpWT, and pcWNV-Cp3'constructs. b, Immunoprecipitation of in vitro translated protein from pcWNV-Cp-DJY (Cp-DJY), pcWNV-CpWT (CpWT), and pcWNV-Cp-3'(Cp3') plasmids. As a negative control, pcDNA3.1 in vitro translated supernatants were analyzed. c , Colorimetric caspase-3 activity assay using pcWNV-Cp-DJY (Cp-DJY), pcWNV-CpWT (CpWT), or pcWNV-Cp3'(Cp3') plasmid–transfected cells. d, The cell lysates were assayed for caspase-9-like activity, and the pcDNA3.1-transfected cell lysate was used as the negative control. e, Inhibition of WNV-Cp–induced apoptosis by a dominant negative (DN) caspase-9 plasmid (DN Casp9) was assayed with equal amount of cell lysates from co-transfection of pcWNV-Cp-DJY (Cp-DJY) or pcWNV-CpWT (CpWT); an expression level of pro-caspase-9 cleavage products (35–37 kDa) was compared to 3'-terminal deletion mutant, pcWNV-Cp3'(Cp3') and pcDNA3.1 by Western blot analysis with anti-human caspase-9 mAb.

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National Center for Infectious Diseases
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