Well-orchestrated transcriptional regulation of pancreatic beta cells is essential for insulin production and glucose homeostasis. Pancreas duodenum homeobox-1 (PDX-1) is a key regulator of glucose-dependent insulin production and glucose metabolism. We find that PDX-1 interacts with the PDZ-domain coactivator Bridge-1 in yeast interaction trap assays. Rat Bridge-1 and PDX-1 interact directly in GST pull-down assays via Bridge-1 interactions with the amino-terminal transactivation domain of PDX-1. Bridge-1 also interacts with wild-type and mutant human PDX-1 (IPF-1) proteins and strongly interacts with the amino-terminal PDX-1 P63fsdelC (MODY4) mutant protein. Transcriptional activation by PDX-1 is increased by addition of Bridge-1 in multiple contexts, including synergistic activation of a Gal4 reporter by Gal4-Bridge-1 and Gal4-PDX-1 fusion proteins, activation of the somatostatin promoter TAAT1 enhancer, and synergistic activation of the rat insulin I promoter FarFlat enhancer by PDX-1, E12, and E47. We propose that the coactivator Bridge-1 modulates PDX-1 functions in the regulation of its target genes.