Mutation Analysis and Characterization of Alternative Splice Variants of the Wilson Disease Gene ATP7B

Abstract

Wilson disease is a copper metabolism disorder caused by mutations in ATP7B, a copper‐transporting adenosine triphosphatase. A molecular diagnosis was performed on 135 patients with Wilson disease in Taiwan. We identified 36 different mutations, eight of which were novel: five missense mutations (Ser986Phe, Ile1348Asn, Gly1355Asp, Met1392Lys, and Ala1445Pro), one deletion (2810delT) in the coding region, and two nucleotide substitutions (−133A→C and −215A→T) in the promoter region. These mutations were not observed in 100 control subjects and reduced the activity of the mutated protein by at least 50% when compared with wild‐type ATP7B. In addition to exon 8, our data indicate another mutation hotspot in exon 12 where 9.62% of all mutations occurred. An alternative splice variant of ATP7B lacking exon 12 was observed in one patient who had a homozygous 2810delT mutation and very mild clinical symptoms. Clinical examination and functional characterization of alternative splice variants of ATP7B lacking exon 12 showed that they retained 80% of their biological activity. The 2810delT mutation increased the expression of these variants, which may have explained the mild symptoms in the patient with the 2810delT mutation. We also discovered that treating liver cancer cells with a Na+/H+ exchanger inhibitor, 5‐(N‐ethyl‐N‐isopropyl)‐amiloride, significantly enhanced the expression of the alternative splice variant of ATP7B lacking exon 12. Conclusion: This study suggests a novel therapeutic strategy for patients with mutations in exon 12. (Hepatology 2010;52:1662‐1670)

Identification of Sequence Variants in Genetic Disease-Causing Genes Using Targeted Next-Generation Sequencing

“…The known homozygous SPG11 gene mutation (c.2163_2164insT, MIM# 610844) was detected in patient P1 and was supported by 234 out of the 235 reads [20]. Patient P4, with the rare inherited disorder Wilson's disease, presented two known heterozygous ATP7B mutations (c.2333G>T, p.R778L; c.2810detT, MIM# 606882); 127 of the 254 reads at this locus carried the prevalent substitution mutation, and almost half of the reads included the deletion mutation [21], [22]. In the two patients suffering from β-thalassemia, we identified two different mutations in the β-subunit of the hemoglobin gene (HBB, MIM# 141900): 126_129delCTTT (269 variants and 421 wild types reads) in patient P5 and IVS2+654C>T (130 of 255 reads) in patient P6 [23], [24].…”

Evolving Perspectives in Wilson Disease: Diagnosis, Treatment and Monitoring

“…In vitro studies suggested that certain ATP7B mutations could be rescued by medical treatment with drugs influencing splicing [22] or protein processing [23].…”

Wilson's Disease: Changes in Methionine Metabolism and Inflammation Affect Global DNA Methylation in Early Liver Disease

“…The earliest phases of hepatic involvement in Wilson's disease (WD) include portal inflammation that may present as lymphocyte and neutrophil infiltrations,1 and microvesicular and macrovesicular steatosis,2 which is exhibited both clinically2 and in animal models of WD 3, 4. Previous studies on the pathogenesis of WD explored the possibilities of genetic polymorphisms in the ATP7B copper (Cu) transporter,5 alternative ATP7B gene splice variants,6 alterations in the RNA processing machinery,7 and the presence of gene modifiers 8. The mechanisms connecting Cu accumulation to hepatocyte damage are poorly understood and may include oxidative damage,9 apoptosis,10 and mitochondrial membrane cross‐linking 11…”