Lake NJ, Webb BD, Stroud DA, Richman TR, Ruzzenente B, Compton AG, Mountford HS, Pulman J, Zangarelli C, Rio M, Bodaert N, Assouline Z, Sherpa MD, Schadt EE, Houten SM, Byrnes J, McCormick EM, Zolkipli-Cunningham Z, Haude K, Zhang Z, Retterer K, Bai R, Calvo SE, Mootha VK, Christodoulou J, Rötig A, Filipovska A, Cristian I, Falk MJ, Metodiev MD, Thorburn DR. Biallelic Mutations in MRPS34 Lead to Instability of the Small Mitoribosomal Subunit and Leigh Syndrome. Am J Hum Genet. 2017 Aug 3;101(2):239-254.
The synthesis of all 13 mitochondrial DNA (mtDNA)-encoded protein subunits of the human oxidative phosphorylation (OXPHOS) system is carried out by mitochondrial ribosomes (mitoribosomes). Defects in the stability of mitoribosomal proteins or mitoribosome assembly impair mitochondrial protein translation, causing combined OXPHOS enzyme deficiency and clinical disease. Here we report four autosomal-recessive pathogenic mutations in the gene encoding the small mitoribosomal subunit protein, MRPS34, in six subjects from four unrelated families with Leigh syndrome and combined OXPHOS defects. Whole-exome sequencing was used to independently identify all variants. Two splice-site mutations were identified, including homozygous c.321+1G>T in a subject of Italian ancestry and homozygous c.322-10G>A in affected sibling pairs from two unrelated families of Puerto Rican descent. In addition, compound heterozygous MRPS34 mutations were identified in a proband of French ancestry; a missense (c.37G>A [p.Glu13Lys]) and a nonsense (c.94C>T [p.Gln32∗]) variant. We demonstrated that these mutations reduce MRPS34 protein levels and the synthesis of OXPHOS subunits encoded by mtDNA. Examination of the mitoribosome profile and quantitative proteomics showed that the mitochondrial translation defect was caused by destabilization of the small mitoribosomal subunit and impaired monosome assembly. Lentiviral-mediated expression of wild-type MRPS34 rescued the defect in mitochondrial translation observed in skin fibroblasts from affected subjects, confirming the pathogenicity of MRPS34 mutations. Our data establish that MRPS34 is required for normal function of the mitoribosome in humans and furthermore demonstrate the power of quantitative proteomic analysis to identify signatures of defects in specific cellular pathways in fibroblasts from subjects with inherited disease.
Scientists have developed a new strategy for diagnosing mitochondrial diseases and identified the Mrps34 gene as a cause for Leigh syndrome.
The study, “Biallelic Mutations in MRPS34 Lead to Instability of the Small Mitoribosomal Subunit and Leigh Syndrome,” was published in The American Journal of Human Genetics.
Diagnosing mitochondrial diseases is often a hard task, and while genetic screening of a person’s entire genome has helped, additional strategies are in need to identify the most difficult cases.
Now, a team of scientists at the Murdoch Children’s Research Institute (MCRI) in Australia found a new cause underlying Leigh syndrome, the most common form of childhood mitochondrial disease.
They found mutations in a gene called Mrps34 in six patients with Leigh syndrome from different parts of the world, including Australia, France, and the United States. The findings were only possible with researchers applying a different technique called quantitative proteomics.
“A key approach was using quantitative proteomics. This process involves sampling all the proteins in a cell at once to identify any problems with the cellular machinery,” Nicole Lake, a Murdoch PhD student and the study’s first author, said in a press release. “Using this technique, you get a snapshot of what’s happening in cells.”
The MRPS34 is a component of mitochondrial ribosomes, called mitoribosomes, which are the protein synthesis machinery responsible for the production of proteins involved in the mitochondrial oxidative phosphorylation (OXPHOS) system. This system, located in the inner membrane of our mitochondria, is where the majority of the energy used by cells is produced.
Researchers performed quantitative proteomics in skin cells extracted from the six patients as well as in healthy skin cells, called controls. They found that the mutations led to a reduction in MRPS34 protein levels. Examining the mitoribosome profile by quantitative proteomics showed that its assembly was impaired, with half of its members falling apart.
These results establish the Mrps34 gene as an important cause for mitochondrial diseases and that employing quantitative proteomics could help improve the diagnosis of these fatal diseases.
“This approach will therefore help to end the diagnostic odyssey for families with children suspected of mitochondrial and other inherited diseases,” said Murdoch chief investigator and Prof. David Thorburn, the study’s lead author.
“Early diagnosis improves the chance for early intervention. It can also provide the opportunity to enroll patients with mitochondrial diseases into clinical trials to test many new promising therapies that are in the pipeline, but not yet proven,” Thorburn added.
Sean Murray, CEO of the Australian Mitochondrial Disease Foundation (AMDF) said he was thrilled to see “an outcome like this, as it has been partly supported by the hard work of AMDF fund-raisers through activities such as our ‘Bloody Long Walks’ and demonstrates the value of researchers engaging with the patient community.”