ANALYSIS OF THE HEXOKINASE-II GENE IN SUBJECTS WITH INSULIN-RESISTANCE AND NIDDM AND DETECTION OF A GLN(142)-]HIS SUBSTITUTION

Hexokinase II (HKII) is the predominant hexokinase isozyme expressed i n insulin-responsive tissues, Since defects involving glucose transpor t and/or its phosphorylation to glucose-6-phosphate are present in mus cle of insulin-resistant humans, HKII should be viewed as a candidate gene for inherited insulin resistance and susceptibility to non-insuli n-dependent diabetes mellitus (NIDDM). To investigate the prevalence o f potential mutations in the gene encoding HKII, we used the polymeras e chain reaction (PCR) to amplify each of the 18 exons of the HKII gen e from genomic DNA derived from 59 subjects: 25 insulin-resistant prob ands with clinical features of the type A syndrome and 34 NIDDM subjec ts enrolled in the United Kingdom Prospective Study of Therapies of NI DDM (UKPDS) who represented the highest percentile of fasting hyperins ulinemia in the UKPDS population of 5,098 subjects, PCR products corre sponding to individual HKII: exons derived from each subject were scre ened for the presence of nucleotide variation using a sensitive nonrad ioactive single-strand conformation polymorphism (SSCP) protocol, Vari ant SSCP patterns indicative of genetic variation were detected only i n PCR amplimers containing exons 4-7, 10, 15, and 17. Direct sequencin g of amplified DNA from individuals affected with variant SSCP pattern s revealed the presence of the following silent polymorphisms: Asp(251 ) (GAT/C) in exon 7 and Asn(692) (AAT/C) in exon 15, SSCP variants det ected in PCR products containing exons 5, 10, and 17 were due to singl e base substitutions in flanking intronic sequences, A polymorphic GGA repeat was identified within intron 5, A common missense mutation (Gl n(142) --> His) was detected in exon 4; however, the prevalence of thi s variant allele was not increased in NIDDM versus control subjects. W e conclude that 1) His(142) is a Common variant HKII allele that is no t associated with insulin resistance or NIDDM; 2) other mutations affe cting the coding regions of the RW gene are very uncommon in patients with severe insulin resistance or insulin-resistant patients with NIDD M; and 3) inherited defects involving the HKII structural gene are unl ikely to contribute significantly to the genetic susceptibility to NID DM.