Two novel mutations identified in familial cases with Donohue syndrome

Abstract

Donohue syndrome (DS) is a rare and lethal autosomal recessive disease caused by mutations in the insulin receptor (INSR) gene, manifesting marked insulin resistance, severe growth retardation, hypertrichosis, and characteristic dysmorphic features. We report the clinical, molecular, and biochemical characterization of three new patients with DS, and address genotype-phenotype issues playing a role in the pathophysiology of DS. A female infant born to first-degree cousins Muslim Arab parents and two brothers born to first-degree cousins Druze parents presented classical features of DS with hypertrophic cardiomyopathy and died in infancy. Each patient was found homozygous for one missense mutation within the extracellular domain of the INSR gene. Western blot analysis identified the proreceptor of INSR, but not its mature subunits alpha and beta. Of 95 healthy Muslims, no heterozygous was found and of 52 healthy Druze from the same village, one was heterozygous. This study presents two novel familial mutations in the alpha subunit of the INSR which appear to impair post-translational processing of the INSR, resulting loss of its function. Both mutations cause DS with hypertrophic cardiomyopathy and early death. Identification of the causative mutation enables prevention of this devastating disease.

Keywords: Cardiomyopathy; Donohue syndrome; genotype–phenotype; insulin receptor..

Figure 1

Clinical features of patients. See description in text.

Figure 2

Mutation analysis. Genomic DNA sequence analysis revealed a dHPLC abnormal pattern of exon 3 of patient ISR1; the novel c.858G>A mutation was detected (A). The mutation was further confirmed by allele-specific oligonucleotide hybridization (B). High conservation of p.C286 throughout the phylogenetic tree (C). 3 genomic DNA sequence analysis of the 22 exons of INSR gene revealed a homozygous T > C transition at nucleotide 167 in exon 2 (D). The family pedigree was drawn according to restriction digestion with TaqI enzyme (E). Fragmented polymerase chain reaction (PCR) products were visualized by ethidium bromide-stained acrylamide gel. P, PCR product (E). The amino acid residues Cysteine at position 286 (C) and Isoleucine at position 56 (F) in the INSR are shown to be conserved throughout the phylogenetic tree. Amino acid conservation was analyzed by NCBI Basic Local Alignment Search Tool, using protein blast.

Figure 3

INSR determination and functionality. The proreceptor but not the mature subunits in the ISR1 and ISR2 fibroblasts (labeled P) is detected, compared with both forms identified in the healthy control (labeled C) (A). Fibroblasts were treated with 1 μmol/L of insulin (B) or IGF1(C), immunoprecipitated with an antibody against phosphorylated tyrosine, and immunoblotted with an antibody against INSR or IGF receptor, respectively.