Enterovirus A71 (EV-A71) is one of the main pathogens causing hand-foot-mouth disease (HFMD). There are no specific and effective antiviral drugs for the treatment of HFMD. Studies have shown that cell membrane transport molecules are involved in virus invasion, replication and the release of infectious virus particles. In order to find the host factors that can effectively inhibit EV-A71 infection, human colon cancer cells (Caco-2) were used as target cells, and RNA interference was used to downregulate the expression of transport-related membrane proteins. The data showed that ATP-binding cassette transport subfamily C member 3 (ABCC3) and solute carrier family 7 member 7 (SLC7A7) played important roles in EV-A71 infection in Caco-2 cells, which provided potential targets for the prevention and treatment of EV-A71 infection.