Insulin is expressed exclusively in the adult beta-cells of the islets of Langerhans. Pancreatic Duodenum Homeobox-1 (PDX-1) is a major regulator of transcription in these cells. It transactivates the insulin gene by binding to a specific DNA motif in its promoter region. Glucose, the main physiological regulator of insulin secretion, also regulates insulin gene transcription through PDX-1. While acute exposure to high glucose concentrations causes an increase in PDX-1 binding, and consequently in insulin mRNA levels, chronic hyperglycemia (toxic to the beta-cell) leads to a decrease in PDX-1 and insulin levels. PDX-1 is absolutely required for pancreas development. In view of the selective expression in adult beta-cells, pancreatic agenesis in both the pdx-1 null mouse and a human carrying a homozygous mutation of PDX-1 was an unexpected and remarkable finding. The homozygous clinical phenotype was neonatal diabetes mellitus (DM) and exocrine insufficiency. Heterozygosity for PDX-1 mutations was found in some individuals with a newly characterized subtype of maturity-onset diabetes of the young (MODY4) and in others with type 2 DM. This review underlines the unique role of PDX-1 in maintaining adult beta-cell-specific functions in normal and disease-related states.