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. Author manuscript; available in PMC: 2012 Sep 4.
Published in final edited form as: Cell Host Microbe. 2011 Jan 20;9(1):32–45. doi: 10.1016/j.chom.2010.12.002

Fig. 6. Role of NS5A domain I in recruitment of PI4KIIIα and induction of PI4P.

Fig. 6

(A) Huh7-Lunet/T7 cells were cotransfected with HA-tagged PI4KIIIα and expression constructs encoding individual viral proteins specified in the top. After 8 h cell proteins were radiolabeled for 16 h with [35S] methionine/cysteine-containing medium. Cells were lysed and immunoprecipitation was performed using antibodies indicated in the bottom. Filled arrowheads point to HCV proteins; the open arrowhead marks PI4KIIIα. (B) Huh7-Lunet/T7 cells were cotransfected with HA-tagged PI4KIIIα and pTM-based NS3 to NS5B polyprotein constructs containing given deletions of individual NS5A domains. Immunoprecipitation was performed as described in (A) using an NS5A-specific antibody. Filled arrowheads on the right refer to NS5A variants; the open arrowhead marks PI4KIIIα. (C) Huh7-Lunet/T7 cells were transfected with constructs encoding NS3 to NS5B harboring wildtype NS5A or NS5A deletions mutants specified in the top. Twenty four hours after transfection NS5A (red) and PI4P (green) were detected by immunofluorescence. Nuclear DNA was stained with DAPI (blue). Images were acquired with a confocal microscope. Numbers in the bottom refer to percentage and SEM of NS5A positive dot-like structures costaining with PI4P. Values are derived from analysis of 20 individual cells.