Protein-protein interaction analysis for TIPE1.

<p>STRING platform is used to predict protein interactions. FBXW5, caspase8, caspase10, POMP and et al. were predicted to be interacted with TIPE1.</p

Homology modeling and evaluation of model stability.

<p>(A) Homology modeling was performed by Swiss-Model Server and the predicted 3D structure of TIPE1 protein was shown. (B) Model quality was evaluated using the method of Ramachandran plot and the results represent the acceptable stability of 3D structure of TIPE1 protein.</p

Post-translational modifications of TIPE1.

<p>(A) NetPhosK Server was performed to predict kinase specific phosphorylation sites in TIPE1. (B) NetPhos Server was performed to predict generic phosphorylation sites in TIPE1. X axis represents amino acid sequence from N- to C- terminal. Y axis represents values computed by the software. The values above the threshold mean the most potential of phosphorylation.</p

Prediction of signal peptide and transmembrane domain of TIPE1.

<p>Signal peptide (A) and transmembrane domain (B) of TIPE1 are predicted by SignaIP Server and TMpred Server, respectively. X axis represents amino acid sequence from N- to C- terminal. Y axis represents scores computed by each server.</p

Prediction of hydrophilicity, accessibility, polarity, flexibility, mutability and bulkiness of TIPE1.

<p>The hydrophilicity (A), accessibility (B), polarity (C), flexibility (D), mutability (E) and bulkiness (F) of TIPE1 are predicted using Protscale Server of expasy platform. X axis represents amino acid sequence from N- to C- terminal. Y axis represents scores computed by each algorithm.</p

Sequences of the synthesized siRNA molecules and their positions in the EV71 2A^pro genomic region.

<p>Sequences of the synthesized siRNA molecules and their positions in the EV71 2A^pro genomic region.</p

siRNAs protect cells against EV71-induced cytopathic effects.

<p>Morphological changes in RD cells were observed after infection. Cells were transfected with each siRNA at a final concentration of 60 nM and then infected with strain EV71-2006-52-9 at an MOI of 0.01. Micrographs were taken at 48 h postinfection under an inverted microscope. The tests were performed in three independent experiments.</p

RNAi inhibits viral protein.

<p>RD cells transfected with siRNA were infected with strain EV71-2006-52-9 at an MOI of 0.01. At 36 h postinfection, total protein was extracted and analyzed with western blotting. β-Actin was used as the internal loading control. The protein measurements were made in three independent experiments.</p

Transfection efficiency of siRNA in RD cells.

<p>(A) Cellular distribution of BLOCK-iT Fluorescent Oligo in transfected RD cells. RD cells were transfected with different concentrations of BLOCK-iT Fluorescent Oligo and 2 μl of Lipofectamine 2000. At 24 h after transfection, the cells were observed under a fluorescence microscope. (B) RD cells transfected with BLOCK-iT Fluorescent Oligo were quantified with flow cytometry. The cells were assayed in three independent experiments.</p

RNAi inhibits viral RNA.

<p>RD cells were treated with each siRNA and then infected with strain EV71-2006-52-9 at an MOI of 0.01. At 24 h postinfection, the viral RNA was extracted and analyzed with real-time RT–PCR. The data shown represent the means ± SD of three independent experiments (**p < 0.01).</p