MOLECULAR SCREENING OF BOTH THE PROMOTER AND THE PROTEIN-CODING REGIONS IN THE HUMAN OB GENE IN JAPANESE OBESE SUBJECTS WITH NON-INSULIN-DEPENDENT DIABETES-MELLITUS

Objective: Although the molecular mechanism of obesity has been poorly understood, recent studies indicate that leptin plays a critical role in regulating both food intake and body weight, Because obesity decre ases the sensitivity to insulin, the human ob gene is presumed to be o ne of the candidate genes for non-insulin-dependent diabetes mellitus (NIDDM) associated with obesity, Although the protein coding region in the ob gene has been screened for mutations, the promoter region and the noncoding first exon have not yet been studied, We investigated th e involvement of the human ob gene, especially mutations at the promot er region and the non-coding first exon, in the development of NIDDM a ssociated with obesity. Subjects: The study group comprised 60 Japanes e obese subjects with NIDDM (body mass index (BMI) 43.6 greater than o r equal to BMI greater than or equal to 26.4, 29.0 +/- 0.41 (mean +/- S.E.M.)) and 24 obese individuals with impaired glucose tolerance (IGT ) (30 greater than or equal to BMI greater than or equal to 26.4, 27.1 +/- 0.22). Methods: Mutations at both the promoter region and all thr ee exons in the human ob gene were screened by the single-stranded con formational polymorphism analysis, When aberrantly migrated bands were recognized, the PCR-amplified DNA fragment was directly sequenced, Re sults: In the protein coding region a silent mutation in the second ex on was detected, The noncoding first exon and the about 100 bp 5'-flan king region of the gene which contains a proximal CCAAT/enhancer-bindi ng protein site were screened, but no mutations were found, Conclusion : These results suggest that no mutations in either the promoter regio n at the about 100 bp 5'-flanking region of the gene, or in any of the three exons, are involved in the development of NIDDM or IGT associat ed with obesity.