Summary Nicotiana benthamiana has been described as non‐host for Melon necrotic spot virus (MNSV). We investigated the basis of this resistance using the unique opportunity provided by strain MNSV‐264, a recombinant virus that is able to overcome the resistance. Analysis of chimeric MNSV mutants showed that virulence in N. benthamiana is conferred by a 49 nucleotide section of the MNSV‐264 3′‐UTR, which acts in this host as a cap‐independent translational enhancer (3′‐CITE). Although the 3′‐CITE of non‐adapted MNSV‐Mα5 is active in susceptible melon, it does not promote efficient translation in N. benthamiana, thus preventing expression of proteins required for virus replication. However, MNSV‐Mα5 gains the ability to multiply in N. benthamiana cells if eIF4E from a susceptible melon variety (Cm‐eIF4E‐S) is supplied in trans. These data show that N. benthamiana resistance to MNSV‐Mα5 results from incompatibility between the MNSV‐Mα5 3′‐CITE and N. benthamiana eIF4E in initiating efficient translation of the viral genome. Therefore, non‐host resistance conferred by the inability of a host susceptibility factor to support viral multiplication may be a possible mechanism for this type of resistance to viruses.