Interaction of CRP1-7 on SVCV replication in EPC cells. (A) SVCV binding levels to EPC cell surfaces in the presence of CRP1-7. EPC cell-bound SVCV particles in the presence of CRP were quantified by the number of SVCV n gene copies determined by RT-qPCR, and the data are expressed, relative to the number of ef1a transcripts, as fold changes. (B) CRP1-7 inhibition of the fusogenic activity of SVCV G protein on the surface of SVCV-infected EPC cells. The levels of G protein-mediated syncytia of 5 or more cells in SVCV-infected EPC cell monolayers were determined by triggering cell fusion at pH 6 in the presence of CRP and are expressed as percentage of the counted syncytia. (C) The time course of SVCV replication in vitro at early stages post adsorption. EPC cell monolayers were incubated for 2 h with the CRP-mix before viral adsorption, and the SVCV replication was estimated by measuring the expression of SVCV n and g gene transcripts by RT-qPCR and is expressed as fold changes. (D) Modulation of the IFN system by CRP1-7. The transcript levels of the IFN-response reporter mx gene were quantified by RT-qPCR in EPC cells 20 h after treatment with CRP for 2 h and were normalized to the corresponding ef1a levels. The data are expressed as fold changes. (E) Presence of antiviral factors in supernatants from CRP1-7-treated EPC cell monolayers. SVCV neutralization was induced by supernatants collected from EPC cells previously treated for 2 h with CRP1-7 and was determined by the focus forming assay. The results are expressed relative to GFP treatments. All experiments were performed 3 times each in triplicate, except for (C,D), which were performed twice each in quadruplicate. The data are presented as the mean and s.d. The significantly different levels between them are indicated with symbols as in Fig. 1. Data were analysed by using one-way ANOVA (A,B,D,E) and two-way ANOVA (C) with Sidak’s multiple comparisons test.
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