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Myopathy with lactic acidosis is linked to chromosome 12q23.3-24.11 and caused by an intron mutation in the ISCU gene resulting in a splicing defect
Umeå University, Faculty of Medicine, Department of Medical Biosciences, Medical and Clinical Genetics.
Umeå University, Faculty of Medicine, Department of Medical Biosciences, Medical and Clinical Genetics.
Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB), Anatomy.
Umeå University, Faculty of Medicine, Department of Medical Biosciences, Medical and Clinical Genetics.
2008 (English)In: Human Molecular Genetics, ISSN 0964-6906, E-ISSN 1460-2083, Vol. 17, no 11, 1666-1672 p.Article in journal (Refereed) Published
Abstract [en]

We describe the mapping and identification of the gene for hereditary myopathy with lactic acidosis (HML). HML is characterized by low physical performance, resulting in physical exertion that causes early exhaustion, dyspnoea and palpitations. Using an autosomal recessive mode of inheritance, we mapped the trait to chromosome 12q23.3-24.11, with a maximum lod score of 5.26. The 1.6-Mb disease-critical region contained one obvious candidate gene-ISCU-specifying a protein involved in iron-sulphur cluster assembly. IscU is produced in two isoforms; one cytosolic and one mitochondrial, coded for by different splice variants of the ISCU gene. Mutational analysis of all exon and intron sequences as well as 1000 bp of the promoter of the ISCU gene revealed one intron mutation that was specific for the disease haplotype. The mutation is located in a region with homology to the interferon-stimulated response element (ISRE), but we could not see any effect of the mutation on expression levels in vitro or in vivo. We did, however, observe a drastic difference in the splicing pattern between patients and controls. In controls the mRNA was, as expected, mainly in the mitochondrial form, while in the patients a larger mRNA transcript was predominant. Sequencing of the product revealed that the mutation activates cryptic splice sites in intron 5 resulting in aberrant mRNA containing 100 bp of the intron. To conclude, our data strongly suggest that an intron mutation in the ISCU gene, leading to incorrectly spliced mRNA, is the cause of myopathy with lactic acidosis in this family.

Place, publisher, year, edition, pages
2008. Vol. 17, no 11, 1666-1672 p.
Keyword [en]
succinate-dehydrogenase; hereditary myopathy; paroxysmal myoglobinuria; aconitase deficiency; cluster; identification; exercise
National Category
Medical Genetics
Identifiers
URN: urn:nbn:se:umu:diva-19316DOI: 10.1093/hmg/ddn057PubMedID: 18296749OAI: oai:DiVA.org:umu-19316DiVA: diva2:201705
Available from: 2009-03-05 Created: 2009-03-05 Last updated: 2017-12-13Bibliographically approved
In thesis
1. Genetic and functional studies of hereditary myopathy with lactic acidosis
Open this publication in new window or tab >>Genetic and functional studies of hereditary myopathy with lactic acidosis
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Alternative title[sv]
Genetiska och funktionella studier av hereditär myopati med laktacidos
Abstract [en]

Hereditary myopathy with lactic acidosis (HML, OMIM#255125) is an autosomal recessive disorder which originates from Västerbotten and Ångermanland in the Northern part of Sweden. HML is characterized by severe exercise intolerance which manifests with tachycardia, dyspnea, muscle pain, cramps, elevated lactate and pyruvate levels, weakness and myoglobinuria. The symptoms arise from malfunction of the energy metabolism in skeletal muscles with defects in several important enzymes involved in the TCA cycle and the electron transport chain. All affected proteins contain iron-sulfur (Fe-S) clusters, which led to the suggestion that the disease was caused by malfunctions in either the transportation, assembly or processing of Fe-S clusters.

The aim of my thesis was to identify the disease causing gene of HML and to investigate the underlying disease-mechanisms. In paper I we identified a disease-critical region on chromosome 12; a region containing 16 genes. One of the genes coded for the Fe-S cluster assembly protein ISCU and an intronic base pair substitution (g.7044G>C) was identified in the last intron of this gene. The mutation gave rise to the insertion of intron sequence into the mRNA, leading to a protein containing 15 abberant amino acids and a premature stop. In paper II we investigated why a mutation in an evolutionary well conserved protein with a very important cellular role, which in addition is expressed in almost all tissues, gives rise to a muscle-restricted phenotype. Semi-quantitative RT-PCR analysis showed that the mutant transcript constituted almost 80% of total ISCU mRNA in muscle, while in both heart and liver the normal splice form was dominant. We could also show that, in mice, complete absence of Iscu protein was coupled with early embryonic death, further emphasizing the importance of the protein in all tissues. These data strongly suggested that tissue-specific splicing was the main mechanism responsible for the muscle-specific phenotype of HML. In paper III the splicing mechanisms that give rise to the mutant ISCU transcript was further investigated. We identified three proteins; PTBP1, IGF2BP1 and RBM39, that could bind to the region containing the mutation and could affect the splicing pattern of ISCU in an in vitro system. PTBP1 repressed the inclusion of the intronic sequence, while IGF2BP1 and RBM39 repressed the total ISCU mRNA level though the effect was more pronounced for the normal transcript. Moreover, IGF2BP1 and RBM39 were also able to reverse the effect of PTBP1. IGF2BP1, though not a splicing factor, had higher affinity for the mutant sequence. This suggested that the mutation enables IGF2BP1 binding, thereby preventing the PTBP1 induced repression seen in the normal case.

In conclusion, we have determined the genetic cause of HML, identifying a base pair substitution in the last intron of the ISCU gene that gives rise to abnormally spliced transcript. The muscle-specific phenotype was also analyzed and tissue-specific splicing was identified as the main disease-mechanism. Furthermore, nuclear factors with ability to affect the splicing pattern of the mutant ISCU gene were identified. This work has thoroughly investigated the fundamental disease mechanisms, thus providing deeper understanding for this hereditary myopathy.

Place, publisher, year, edition, pages
Umeå: Umeå Universitet, 2011. 39 p.
Series
Umeå University medical dissertations, ISSN 0346-6612 ; 1454
Keyword
Hereditary myopathy with lactic acidosis, ISCU, intron mutation, mouse model, tissue-specific splicing
National Category
Medical Genetics
Research subject
Medical Genetics
Identifiers
urn:nbn:se:umu:diva-50592 (URN)978-91-7459-308-2 (ISBN)
Public defence
2012-01-27, Sal B, By 1D 9 tr, Norrlands Universitetssjukhus, Umeå, 09:00 (English)
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Supervisors
Available from: 2012-01-05 Created: 2011-12-14 Last updated: 2012-01-05Bibliographically approved

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