We have developed a plant virus-mediated transgene activation (VMTA) system that utilizes a viral expression vector to present the inducer. The concept was tested using two well characterized components: (i) an artificial promoter based on the yeast GAL4 upstream activating sequence and the minimal TATA element of Cauliflower Mosaic Virus 35S RNA promoter, and (ii) a transcriptional activator (TA) consisting of a fusion between the GAL4 DNA binding domain and the Herpes simplex virus VP16 activation domain. The TA was expressed under the control of the subgenomic promoter of a Tobacco Mosaic Virus-based expression vector. The VMTA system was functional in transient Agroinfiltration assays with the reporter gene beta-glucuronidase, the intracellular domain of the diabetes associated autoimmune antigen, IA-2ic, and with the anti-tetanus antibody 9F12. Transgenic lines harboring the reporter gene were also examined. The VMTA system displayed tight transcriptional control in both transient assays and in transgenic Nicotiana benthamiana plants carrying the TA-inducible reporter.