To determine the role of genetic defects in the
insulin receptor in the
insulin resistance of
lipoatrophic diabetes mellitus, we studied
insulin binding,
insulin receptor autophosphorylation, and
insulin receptor mRNA levels and performed Southern blot analysis of genomic
DNA in four siblings, all of whom have some degree of
insulin resistance and three of whom have lipoatrophy. The
insulin receptor concentration in Epstein-Barr virus-transformed lymphocytes was about 30% of normal in all three lipoatrophic siblings (LA1, LA2, and LA3) and was 55% of normal in the nonlipoatrophic sibling (LAS).
Insulin receptor mRNA concentrations in the lymphocytes paralleled
insulin binding and ranged from 15-67% of the mean normal level.
Insulin binding to fibroblasts was also reduced about 50% in the lipoatrophic siblings. In addition,
insulin binding to fibroblasts of LAS and LA2 exhibited a rightward shift of the competition curve, suggesting reduced receptor affinity [ED50, 35 and 50 ng/mL (5845 and 8350 pmol/L); normal, 1-3 ng/mL (167-501 pmol/L)]. Receptor autophosphorylation determined using
Triton X-100 extracts of the fibroblasts was decreased in LA1 and LA3, but normal in LA2 and LAS. Using restriction
enzyme digests of genomic
DNA and probes spanning the entire
cDNA of the
insulin receptor, no gross alterations in receptor gene structure were detected in any members of this family. In 2 of the lipoatrophic siblings (LA1 and LA3) and in the sibling with
insulin resistance but no lipoatrophy (LAS), a unique variant BamHI site was detected using a probe to the alpha-subunit region. This site was not found in 200 normal or diabetic
insulin receptor alleles. By use of probes 5' and 3' to the alpha-subunit probe and by genomic cloning analysis, this variant BamHI site was localized to an intron in the
insulin receptor gene downstream of exon 3 which encodes
amino acids 191-296 of the alpha-subunit of the receptor. These data indicate the complex nature of familial
lipoatrophic diabetes mellitus, with alterations in
insulin receptor expression and/or function in both clinically affected and non-affected siblings. Both the reduced
insulin binding and reduced levels of
insulin receptor mRNA in the lipoatrophic siblings suggest that an
insulin receptor gene defect contributes to this syndrome. Several members of this family also carry a unique variant
insulin receptor gene, which, however, could not be linked to a specific alteration in receptor expression or the presence of lipoatrophy.(ABSTRACT TRUNCATED AT 250 WORDS)