Homozygous mutation in murine retrovirus integration site 1 gene associated with a non-syndromic form of isolated familial achalasia

Abstract

Background: Achalasia is a condition characterized by impaired function of esophageal motility and incomplete relaxation of the lower esophagus sphincter, causing dysphagia and regurgitation. Rare cases of early-onset achalasia appear often in combination with further symptoms in a syndromic form as an inherited disease. Methods: Whole genome sequencing was used to investigate the genetic basis of isolated achalasia in a family of Tunisian origin. We analyzed the function of the affected protein with immunofluorescence and affinity chromatography study. Key Results: A homozygous nonsense mutation was detected in murine retrovirus integration site 1 (MRVI1) gene (Human Genome Organisation Gene Nomenclature Committee (HGNC) approved gene symbol: IRAG1) encoding the inositol 1,4,5-trisphosphate receptor 1 (IP3R1)-associated cyclic guanosine monophosphate (cGMP) kinase substrate (IRAG). Sanger sequencing confirmed co-segregation of the mutation with the disease. Sequencing of the entire MRVI1 gene in 35 additional patients with a syndromic form of achalasia did not uncover further cases with MRVI1 mutations. Immunofluorescence analysis of transfected COS7 cells revealed GFP-IRAG with the truncating mutation p.Arg112* (transcript variant 1) or p.Arg121* (transcript variant 2) to be mislocalized in the cytoplasm and the nucleus. Co-transfection with cGMP-dependent protein kinase 1 isoform β (cGK1β) depicted a partial mislocalization of cGK1β due to mislocalized truncated IRAG. Isolation of protein complexes revealed that the truncation of this protein causes the loss of the interaction domain of IRAG with cGK1β. Conclusions & Inferences: In individuals with an early onset of achalasia without further accompanying symptoms, MRVI1 mutations should be considered as the disease-causing defect.