De Novo DNM1L Mutation in a Patient with Encephalopathy, Cardiomyopathy and Fatal Non-Epileptic Paroxysmal Refractory Vomiting

Abstract

Mitochondrial fission and fusion are vital dynamic processes for mitochondrial quality control and for the maintenance of cellular respiration; they also play an important role in the formation and maintenance of cells with high energy demand including cardiomyocytes and neurons. The DNM1L (dynamin-1 like) gene encodes for the DRP1 protein, an evolutionary conserved member of the dynamin family that is responsible for the fission of mitochondria; it is ubiquitous but highly expressed in the developing neonatal heart. De novo heterozygous pathogenic variants in the DNM1L gene have been previously reported to be associated with neonatal or infantile-onset encephalopathy characterized by hypotonia, developmental delay and refractory epilepsy. However, cardiac involvement has been previously reported only in one case. Next-Generation Sequencing (NGS) was used to genetically assess a baby girl characterized by developmental delay with spastic-dystonic, tetraparesis and hypertrophic cardiomyopathy of the left ventricle. Histochemical analysis and spectrophotometric determination of electron transport chain were performed to characterize the muscle biopsy; moreover, the morphology of mitochondria and peroxisomes was evaluated in cultured fibroblasts as well. Herein, we expand the phenotype of DNM1L-related disorder, describing the case of a girl with a heterozygous mutation in DNM1L and affected by progressive infantile encephalopathy, with cardiomyopathy and fatal paroxysmal vomiting correlated with bulbar transitory abnormal T2 hyperintensities and diffusion-weighted imaging (DWI) restriction areas, but without epilepsy. In patients with DNM1L mutations, careful evaluation for cardiac involvement is recommended.

Keywords: DNM1L; cardiomyopathy; mitochondrial disorders; mitochondrial dynamics; mitochondrial fission; paroxysmal vomiting.

Figure 1

Brain MRI pattern. (a) Brain MRI performed in the acute phase at 10 years of age. Coronal and Sagittal T2 weighted images shows global cerebral and cerebellar atrophy. Red arrow shows hyperintensity of the right bulbar pyramid; (b) brain MRI performed 20 days later shows reduction in the right corresponding bulbar pyramid lesion.

Figure 2

Echocardiogram pattern. Echocardiogram performed during acute phase that showed moderate hypertrofic cardiomyopathy of the left ventricle (a) without outflow obstruction and with normal EF (b).

Figure 3

Genetic and structural analysis. (a) The c.116G>A variant identified by NGS in DNM1L was confirmed by Sanger sequencing in the patient and appeared absent in both parents; (b) cDNA, obtained by retrotranscription of mRNA from patient’s fibroblasts, revealed balanced expression of the two alleles; (c) DRP1 structure and multiple protein sequence alignment around the site of the p.Ser39Asn replacement. Crystal structure of the human dynamin-1-like protein in complex with GDP-AlF4 (Protein Data Bank code 3W6P) and protein sequence alignment around Ser39, highlighting the role of this serine in the binding of the GTP/GDP ligand and its conservation among different organisms.

Figure 4

Western blotting and immunostaining analysis. (a) Immunoblot analysis of total lysates from controls (Ct) and patient (Pt) fibroblasts using DRP1 and GAPDH antibodies. The latter was used as loading control. The steady-state level of DRP1 protein in the patient is in the low range of normal controls. Fibroblasts from a patient with recessive DNM1L mutations (Pt-AR) were used as “positive” control. Values in the graph are given as the mean ± SD (n = 4–5). PT vs. CTs: p-value = 0.018. (b) Characterization of the mitochondrial network: representative images of mitochondrial morphology in control (Ct) and patient (Pt) fibroblasts grown in galactose-supplemented medium. Mitochondrial network of DNM1L-mutant fibroblasts showed an altered mitochondria morphology, with swollen, dots, and “chain-like” structures. (c) Characterization of the peroxisomal network: immunofluorescence staining with the anti-PMP70 antibody of fibroblasts from controls (Ct) and patient (Pt). Fibroblasts from patient displayed organelles longer and larger compared with control. (Scale bar = 10 μm).