A new member of the tobacco (Nicotiana tabacum) AP2/ERF (ethylene response factor) transcription factor family, designated NtERF5, has been isolated by yeast one-hybrid screening. In vitro, recombinant NtERF5 protein weakly binds GCC box cis-elements, which mediate pathogen-regulated transcription of several PR (pathogenesis related) genes. NtERF5 transcription is transiently activated by wounding, by infection with the bacterial pathogen Pseudomonas syringae, as well as by inoculation with Tobacco mosaic virus (TMV). In contrast, NtERF5 transcription is not enhanced after application of salicylic acid, jasmonic acid, or ethylene. Constitutive overexpression of NtERF5 (ERF5-Oex) under control of the 35S promoter results in no visible alterations in plant growth or enhanced resistance to Pseudomonas infection. Furthermore, no constitutive expression of PR genes has been observed. In contrast, ERF5-Oex plants show enhanced resistance to TMV with reference to reduced size of local hypersensitive-response lesions and impaired systemic spread of the virus. Since, in TMV-infected ERF5-Oex plants, the viral RNA accumulates only up to 10 to 30% of the wild-type level, we suggest that NtERF5-regulated gene expression is controlling resistance to viral propagation. Previous research has demonstrated that overexpression of ERF genes enhances resistance to bacterial and fungal pathogens. Here, we provide further evidence that resistance to viral infection can be engineered by overexpression of ERF transcription factors.