Glucose responsiveness is a fundamental metabolic feature of pancreatic P-cells. Glucose-regulated transcription of the insulin gene is in part mediated via the homeobox transcription factor PDX-1. Another islet protein and diabetes autoantigen, glutamic acid decarboxylase (GAD), has been shown to be subject to regulation by glycemia. We have studied the mRNA level of two isoforms of GAD, GAD(65) and GAD(67), and found that GAD(67) but not GAD65 mRNA steady-state level is regulated by glucose. By transfection of a rat GAD67 promoter-driven luciferase reporter gene into primary rat islet cells, we demonstrate glucose-regulated expression of the reporter gene. We show that PDX-1 is able to bind to two TAAT-boxes in the GAD67 promoter and that functional disruption of these two PDX-1 binding elements has an additive effect in severely impairing glucose responsiveness of the GAD67 promoter. These data strongly suggest that PDX-1 is involved in glucose-regulated expression of GAD(67) (C) 2002 Elsevier Science (USA). All rights reserved.