Discussion
In this study, we described two novel heterozygous mutations ofTMEM126B(c.82-2A>G and
c.290dupT) from a Chinese patient manifested with LS.In silico predictions,
minigene splicing assays and patients’ RNA analyses were combined to
determine that the
c.82-2A>G mutation resulted in exon 2 completely skipped,
and that the c.290dupT mutation caused an increase of partial and
complete exon 3 deletion transcripts, which would lead to
frameshift and premature termination.
Patient-derived immortalized lymphocytes carrying biallelic mutations
exhibited complex I content and assembly defect and mitochondrial
dysfunction. To the best of our knowledge, this is the first report thatTMEM126B mutations cause LS.
TMEM126B was identified as the part of MCIA complex to co-migrate with
other MCIA complex components (NDUFAF1, ECSIT, and ACAD9) by complexome
profiling (Heide et al., 2012). The defect of each components can cause
heterogeneous clinical phenotypes. For example, mutations inNDUFAF1 mostly associated with cardiological symptoms (Dunning et
al., 2007; Elisa Fassone et al., 2011), including Wolff-Parkinson-White
syndrome and hypertrophic cardiomyopathy. Genetically deficit inACAD9 commonly linked to cardiac symptoms, neurological symptoms,
and severe lactic acidosis (E. Fassone et al., 2011; Schiff et al.,
2015). Mendelian mutations in ECSIT gene has not yet reported.
Individuals carrying TMEM126B mutations mainly presented with
exercise intorlance, muscle weakness, hyperlactic acidemia, pure
myopathy, chronic renal failure
and cardiomyopathy (Alston et al., 2016; Sánchez-Caballero et al., 2016;
Theunissen et al., 2017). Notably, the clinical phenotypes of our
patient are consistent with those of patients carrying TMEM126Bmutations previously reported, except for chronic renal failure and
cardiomyopathy (Table 1). Chronic renal failure is not a necessary
symptom to diagnose mitochondrial diseases. Significantly, our patient
show a more severe neurological symptoms with clinical presentation
consistent with LS. Overall, we believe that patients withTMEM126B mutations may exhibit high clinical heterogenicity. The
combination of clinical and molecular diagnosis are required for the
diagnosis of TMEM126B mutation‐related mitochondrial diseases.
In silico analysis indicateed that the c.82-2A>G
mutation
located
2bp before exon 2 could lead to the loss of the original 3’splice
acceptor site, while the
c.290dupT mutation located exon 3
might create a new constitutive splice acceptor site. In vitroexperiments verified that c.82-2A>G mutation led to
complete exon 2 skipping and c.290dupT caused an increase of transcripts
with partial and complete deletion of TMEM126B exon 3.
Mutations that located in exonic
splicing enhancer (ESE) region are thought to prevent serine and
arginine-rich (SR) proteins from binding to ESE sequence motifs, which
induced exon skipping. Results of ESE finder software suggested that
c.290dupT mutation may disrupt a putative SRp55 binding site, thus
increasing the proportion of abnormal splicing transcripts. Further
functional study indicated that patient-derived immortalized lymphocytes
exhibited a global mitochondrial dysfunction with decreased
mitochondrial respiratory capacity, reduced ATP content and increased
mitochondrial ROS levels due to the complex I content and assembly
defect. Unlike previous reports, biallelic mutations in our patient
caused the complete deletion of exon 2 and the partial truncation of
exon 3 of TMEM126B , which resulted in a more severeTMEM126B defect, leading to a more severe complex I deficiency
and brain phenotypes.
In summary, we identify TMEM126B as a novel disease-causing gene
resulting in LS with obvious neurological symptoms, and report two novelTMEM126B mutations (c.82-2A>G and c.290dupT) that
cause splicing defects and lead to mitochondrial dysfunction due to the
severe complex I deficiency. Our study expands the genetic mutation
spectrum of LS and the clinical spectrum caused by TMEM126Bmutations.