Although the majority of genes required for the transfer of T-DNA from Agrobacterium tumefaciens to plant nuclei are located on the Ti plasmid, some chromosomal genes, including the recently described acvB gene, are also required. We show that AcvB shows 50% identity with the product of an open reading frame, designated virJ, that is found between the virA and virB genes in the octopine-type Ti plasmid pTiA6. This reading frame is not found in the nopaline-type Ti plasmid pTiC58. acvB is required for tumorigenesis by a strain carrying a nopaline-type Ti plasmid, and virJ complements this nontumorigenic phenotype, indicating that the products of these genes have similar functions. A virJ-phoA fusion expressed enzymatically active alkaline phosphatase, indicating that VirJ is at least partially exported. virJ is induced in a VirA/VirG-dependent fashion by the vir gene inducer acetosyringone. Primer extension analysis and subcloning of the virJ-phoA fusion indicate that the acetosyringone-inducible promoter lies directly upstream of the virJ structural gene. Although the roles of the two homologous genes in tumorigenesis remain to be elucidated, strains lacking acvB and virJ (i) are proficient for induction of the vir regulon, (ii) are able to transfer their Ti plasmids by conjugation, and (iii) are resistant to plant wound extracts. Finally, mutations in these genes cannot be complemented extracellularly.