Cutinolytic enzymes are secreted by fungal pathogens attacking the aerial parts of the plant, to facilitate penetration of the outermost cuticular barrier of the host. The role of cutinases in soil-borne root pathogens has not been studied thus far. Here we report the characterization of the zinc finger transcription factor Ctf1 from the vascular wilt fungus Fusarium oxysporum, a functional orthologue of CTF1alpha that controls expression of cutinase genes and virulence in the pea stem pathogen Fusarium solani f. sp. pisi. Mutants carrying a Deltactf1 loss-of-function allele grown on inducing substrates failed to activate extracellular cutinolytic activity and expression of the cut1 and lip1 genes, encoding a putative cutinase and lipase, respectively, whereas strains harbouring a ctf1(C) allele in which the ctf1 coding region was fused to the strong constitutive Aspergillus nidulans gpdA promoter showed increased induction of cutinase activity and gene expression. These results suggest that F. oxysporum Ctf1 mediates expression of genes involved in fatty acid hydrolysis. However, expression of lip1 during root infection was not dependent on Ctf1, and virulence of the ctf1 mutants on tomato plants and fruits was indistinguishable from that of the wild-type. Thus, in contrast to the stem pathogen F. solani, Ctf1 is not essential for virulence in the root pathogen F. oxysporum.