Detection rates and phenotypic spectrum of m.3243A > G in the MT-TL1 gene: A molecular diagnostic laboratory perspective

Highlights

• We report on the experience of screening 745 patients for the m.3243A > G mutation in the MT-TL1 gene.

• The detection rate for this specific mutation over a period of 11 years is found to be 4.7%.

• We presented the clinical presentations, detection methods and the mutant heteroplasmic level of 35 patients harbouring this point mutation.

• It is important that screening for the mutation m.3243A > G is tested in more than one tissue type.

• Majority of patients harbouring this point mutation exhibit oligo-symptomatic or atypical presentation.

Abstract

The nucleotide change A to G at position m.3243 in the mitochondrial tRNA leucine (UUR) gene (MT-TL1) is the most common point mutation reported in association with the Mitochondrial Encephalomyopathy, Lactic Acidosis and Stroke-like episodes (MELAS) syndrome. Since the original description of this disorder, factors including random mitochondrial segregation and consequent variable tissue heteroplasmy are recognised to contribute to a much broader phenotypic spectrum associated with the MT-TL1 m.3243A > G mutation, often rendering the process of making a diagnosis complex. Reliance on clinicians’ referral patterns means that for most molecular diagnostic laboratories, their positive identification rates for the common pathogenic mitochondrial DNA (mtDNA) mutations, including MT-TL1 m.3243A > G, is often relatively low compared to those reported in clinically targeted research studies. Herein, we report our results of consecutive prospective screening of 745 patients with a clinically suspected mitochondrial syndrome encompassing features associated with MT-TL1 m.3243A > G mutation.

Introduction

The disorder characterised by Mitochondrial Encephalomyopathy, Lactic Acidosis and Stroke-like episodes (MELAS) is a well-recognised syndrome in the field of mitochondrial diseases. Goto and colleagues (Goto et al., 1990) were among the first to identify the most common point mutation associated with MELAS m.3243A > G, located in the tRNA leucine (UUR) gene (MT-TL1) of the mitochondrial genome. In their study, genetic testing on skeletal muscle biopsies of patients with MELAS revealed 80% of these patients harboured the point mutation (Goto et al., 1990, Goto et al., 1991). The consequent invariant criteria established for the clinical diagnosis of MELAS includes (1) stroke-like episodes before the age of 40 years; (2) encephalopathy characterized by seizures, dementia, or both; and (3) lactic acidosis, ragged-red fibres (RRF) or both. Diagnosis would be considered certain if there are also at least two of the following: normal early development, recurrent headache or recurrent vomiting (Ciafaloni et al., 1992, Hirano et al., 1992, Pavlakis et al., 1984).

Subsequent to the seminal reports, the clinical spectrum of the m.3243A > G mutation has been recognised to be much more diverse and very heterogeneous (Ciafaloni et al., 1992, Jean-Francois et al., 1994, Nesbitt et al., 2013) and there are many other distinct syndromes reported in association with MT-TL1 m.3243A > G, such as Myoclonic Epilepsy and Ragged Red Fibres (MERRF) (Fabrizi et al., 1996), MERRF/MELAS overlap syndrome (Campos et al., 1996), Progressive External Ophthalmoplegia (PEO) (Moraes et al., 1993), Chronic Progressive External Ophthalmoplegia (CPEO) (Bosbach et al., 2003, Koga et al., 2000, Mariotti et al., 1995), MERRF/CPEO overlap syndrome (Verma et al., 1996), Maternally Inherited Diabetes and Deafness (MIDD) (Van den Ouweland et al., 1992) and Leigh’s syndrome (Koga et al., 2000). Non-syndromic phenotypes associated with this point mutation include hypertrophic cardiomyopathy (Koga et al., 2000, Silvestri et al., 1997), cluster headache (Shimomura et al., 1994), pancreatitis (Kishnani et al., 1996) subacute dementia with myoclonus mimicking Creutzfeldt-Jakob Disease (Isozumi et al., 1994) and myoclonous with ragged red fibre phenotype (Nesbitt et al., 2013). According to the studies by Chinnery et al. (1999) and Frederiksen et al. (2006), there appears to be a uniform distribution of mutant mtDNA throughout the three germ layers in embryogenesis; however, there are significant differences between heteroplasmic levels of the individual tissue types, indicating tissue-specific segregation of MT-TL1 m.3243A > G later in embryogenesis, which is believed to explain the diversity of clinical phenotypes.

Therefore, a series of ancillary clinical and laboratory investigations including biochemical assays (lactic acid levels and respiratory chain complex activities), neuroimaging (computed tomographic (CT) or magnetic resonance imaging (MRI) for evidence of focal brain abnormalities) (Sparaco et al., 2003), tissue histochemistry (to detect the presence of ragged red fibres or cytochrome-oxidase deficient fibres), and molecular mitochondrial DNA analysis need to be considered in order to provide a definitive diagnosis for MELAS syndrome.

In this report, we describe the detection rate of the MT-TL1 m.3243A > G mutation and the associated clinical features for 745 adult patients referred for mitochondrial genetic screening specifically directed at this mutation in the setting of a suspected mitochondrial disorder. A detailed summary of the associated clinical features is discussed, including evaluation of the percentage of patients fulfilling the invariant criteria for MELAS as compared to those who manifest only some of the symptoms or show other distinct clinical features associated with this point mutation.