4 (HNF-4/MODY 1), glucokinase (GCK/MODY 2), and hepatocyte nuclear factor-1 (HNF 1/MODY 3) on chromosomes 20q, 7p, and 12q, respectively (3–5). Investigation of 67 MODY pedigrees collected in France showed that 63%(42 families) have GCK/MODY 2 subtype and 21%(14 families) have HNF 1/MODY 3 subtype (4, 6, 7). However, linkage and mutation searches in the 11 remaining MODY pedigrees did not give evidence for a pathogenic role of the three known MODY loci, implying that at least one additional MODY locus is associated with MODY (14; J.-CC, unpublished observations). Recently, a heterozygous single nucleotide deletion within codon 63 of insulin promoter factor 1 (IPF 1) gene (also known as STF 1, IDX 1, and PDX 1) has been reported to cosegregate with early-onset NIDDM in a MODY-like pedigree (8). IPF 1 is a homeodomain-containing transcription factor critically required for pancreatic islet development as well as for expression of insulin and other-cell–speci fic genes (9). The recently described IPF1 deletion results in a frame shift at the COOH-terminal border of the transactivation domain of IPF 1, resulting in a protein that lacks a domain that is crucial for DNA binding. The proband was homozygous for this mutation and completely lacked pancreatic tissue (10). Her heterozygous relatives presented with early-onset NIDDM. These data prompted us to evaluate the role of IPF 1 in our 11 unlinked MODY families. Thus, we directly sequenced (on both strands) IPF 1 exons with their flanking introns and the proximal 170-bp promoter region, and a 110-bp potential enhancer sequence. The latter sequence, located~ 1.8-kb upstream from the transcriptional start site of IPF 1, is highly homologous to its murine counterpart and contains regulatory elements that may be important for pancreas-speci fic IPF 1expression (DAS, unpublished observations). Notably, the 110-bp sequence contains sequence homologies, over short stretches of 7–10 nucleotides, to the rat elastase I pancreas-specific enhancer (11). Probands of 11 MODY families were investigated. All these families met the criteria of MODY, with onset of diabetes before age 25 years and familial hyperglycemia consistent with an autosomal-dominant inheritance (12). Genomic DNA was extracted from peripheral blood leukocytes using the Puregene kit (Gentra, Minneapolis, MN). The enhancer, the proximal promoter region, and the two exons with their flanking intronic sequences of IPF 1 gene of the 11 probands were amplified by PCR using the primers 5-GCCGCAGACA AT GGACTC-3 and 5-AG AT G CCCTTGCTGTCACC-3 for the enhancer, 5-GCC TA G CCTCT TA GTGCG-3 and 5-TGGGTCCTTG TA AAGCTG-3 for the minimal promoter, 5-CC AT GAACGGCGAGGAGC-3 and 5-CAGGCT TA CCTGCCCAC T-3 for exon 1, 5-GCCC TGTGTCGCCCGCAG-3 and 5-TTGAAGC CC CTCAGCCAG-3 for exon 2. The primers used for amplifying the promotor region and the two exons were designed based on the sequence published by Inoue et al.(13). The primers used to amplify the 110-bp enhancer sequence were designed based on the genomic organization of the human IPF1 (DAS, unpublished observations). Polymerase chain reaction (PCR) was performed in a 50-µl volume containing 100 ng of genomic DNA, 0.25 mmol/l of each primer, and 1.5 U of A mpli TaqGold (Perkin-Elmer, Applied Biosystems, Foster C ity, CA). For all fragments, PCR cycling consisted in an initial denaturation at 94 C for 12 min followed by 30 cycles of denaturation at 94 C for 30 s, annealing at 57 C for 30 s, and extension at 72 C for 45 s, with a final extension at 72 C for 10 min. PCR products were …