A novel, basic region leucine zipper transcription factor (TH/bZIP) is dramatically up-regulated at the climax of metamorphosis in Xenopus laevis. It can be induced in tadpoles prematurely by thyroid hormone (TH) with kinetics that are intermediate between early and late Xenopus TH response genes. A small amount of early, cycloheximide-resistant up-regulation is observed, but the majority of TH/ bZIP mRNA accumulation occurs after 12 h of treatment in parallel with late response gene induction. There are two genomic TH/bZIP genes in the pseudotetraploid X. laevis genome that are coordinately regulated. They have highly conserved regulatory regions that contain two conserved, adjoining DR+4 thyroid response elements (TRE) in opposite orientation. The early/late TH induction kinetics has been reproduced in transient transfection assays. The secondary rise of transcriptional activity requires DNA regions other than the TREs and, therefore, the interaction of transcription factors other than the TH receptors. Finally, the regulatory region of the TH/bZIP gene has been used to drive green fluorescent protein in transgenic X. laevis tadpoles. Regulation of the transgene during spontaneous and induced metamorphosis mimics that of the endogenous TH/bZIP gene. The newly developed X. laevis transgenesis method has distinct advantages for the analysis of transcriptional regulatory elements over transient transfection assays and will be useful for further in vivo studies of TH-response gene regulation during development.