Updated project metadata. MicroRNAs are noncoding RNA species comprising 18–23 nucleotides that regulate host-virus interaction networks. Here, we showed that enterovirus A71 infection in human rhabdomyosarcoma (RD) involved miR-197 expression. miR-197 can regulate virus replication in the context of viral RNA synthesis via the transfection of its mimic into RD cells. We employed a mass spectrometry-based quantitative proteomic stable isotope labeling with amino acids in cell culture (SILAC) approach for the identification of miR-197 target genes by transfecting mimetic miR-197 into RD cells, and the differential expression of prospective target proteins was identified. A total of 1,822 genes were repeatedly and considerably downregulated in miR-197-transfected RD cells, 106 of which were predicted to have seed sites by TargetScan. Seven of the selected 8 genes potentially related to viral replication and immune response were confidently validated as direct miR-197 targets using a luciferase (untranslated region (3'-UTR)) reporter assay. The expression of three selected endogenous molecules (ITGAV, ETF1, and MAP2K1 (MEK1)) was significantly reduced when RD cells were transfected with an miR-197 mimic. Our results provide a database of miR-197 targets, which is potentially of interest in the area of viral pathogenesis and other research fields.