FIG 4.

G3BP1 and HuR modulate ZIKV replication. (A) Schematic of the ZIKV MR766 Gaussia Luciferase reporter genome within the pCDNA6.2 MR766 clGLuc Intron3127 HDVr plasmid showing the 5ʹ and 3ʹ UTRs, the mature viral proteins within the single open reading frame, and the position of the Gaussia Luciferase gene within the MR766 genome. Two elements within the plasmid, namely, the CMV promoter and HDVr, which creates an authentic 3ʹ UTR in the genome, are also denoted. (B and D) Huh7 cells were first transfected with either the control or target-specific siRNAs and then transfected with the same siRNAs and the pCDNA6.2 MR766 clGLuc Intron3127 HDVr WT replication-competent or Pol(−) replication-deficient plasmid. GLuc activity in the medium of transfected cells was assayed at 6, 24, 48, and 96 h posttransfection. (C and E) Huh7 cells were transfected with p3×Flag-BAP, pG3BP1-Flag, pHuR-Flag, and WT and Pol(−) pCDNA6.2 MR766 clGLuc Intron3127 HDVr. At 6, 24, 48, and 96 h posttransfection, medium from the transfected cells was collected and GLuc activity measured. (B) Effect of G3BP1 knockdown on ZIKV-GLuc genome expression. (C) Effect on ZIKV-GLuc reporter genome expression following overexpression of 3×Flag-BAP (control) and G3BP1-Flag. (D) Effect of depleting HuR on ZIKV-GLuc genome expression. (E) Effect of overexpressing 3×Flag-BAP and HuR-Flag on ZIKV-GLuc genome expression. The data shown are from a single experiment and are representative of at least three independent experiments. Error bars indicate means ± SD.