Genetic etiology of progressive pediatric neurological disorders

Background The aim of the study was to characterize molecular diagnoses in patients with childhood-onset progressive neurological disorders of suspected genetic etiology. Methods We studied 48 probands (age range from newborn to 17 years old) with progressive neurological disorders of unknown etiology from the largest pediatric neurology clinic in Finland. Phenotypes included encephalopathy (54%), neuromuscular disorders (33%), movement disorders (11%), and one patient (2%) with hemiplegic migraine. All patients underwent whole-exome sequencing and disease-causing genes were analyzed. Results We found 20 (42%) of the patients to have variants in genes previously associated with disease. Of these, 12 were previously reported disease-causing variants, whereas eight patients had a novel variant on a disease-causing gene: ATP7A, CHD2, PURA, PYCR2, SLC1A4, SPAST, TRIT1, and UPF3B. Genetics also enabled us to define atypical clinical presentations of Rett syndrome (MECP2) and Menkes disease (ATP7A). Except for one deletion, all findings were single-nucleotide variants (missense 72%, truncating 22%, splice-site 6%). Nearly half of the variants were de novo. Conclusions The most common cause of childhood encephalopathies are de novo variants. Whole-exome sequencing, even singleton, proved to be an efficient tool to gain specific diagnoses and in finding de novo variants in a clinically heterogeneous group of childhood encephalopathies. Impact Whole-exome sequencing is useful in heterogeneous pediatric neurology cohorts. Our article provides further evidence for and novel variants in several genes. De novo variants are an important cause of childhood encephalopathies.


List of primers 1.1 Primers used in Sanger sequencing
the parents were unaffected, while both sisters had CMT.D) Sanger sequencing was inconclusive for the mother, the father had the wild type allele, while the sisters were heterozygotes for the c.1072G>A variant.E) Minisequencing confirmed the mother to be a mosaic carrier, with a mutation load ranging from 20 to 32 % in urine, saliva, and blood.

Supplementary figure 2: CHD2
Figure 2. A) CHD2 protein with previously reported disease-causing variants, the novel p.Q438X variant is depicted below the linear graph.B) Protein conservation of the Gln438 amino acid residue (yellow shade) through species and previously reported mutations (blue shade).C) Family pedigree, the proband had a de novo variant.D) Sanger sequencing results confirm the inheritance pattern of c.1312C>T.

Supplementary figure 3: UPF3B
Figure 3. A) UPF3B protein with previously reported disease-causing variants, the novel p.I154MfsX variant is depicted below the linear graph.B) Protein conservation of the Ile154 amino acid residue (yellow shade) through species and previously described mutations (blue shade).C) Family pedigree, the proband and affected brother had the four-nucleotide deletion as hemizygote, while the mother was an unaffected carrier.D) Sanger sequencing results confirm the inheritance pattern of c.462_465delCGAT.

Supplementary figure 5: PYCR2
Figure 5. A) PYCR2 protein with previously reported disease-causing variants, the novel p.R199Q variant is depicted below the linear graph.B) Family pedigree, the parents were first degree cousins and heterozygote carriers of the variant, the proband homozygote.C) Sanger sequencing results confirm the inheritance pattern of c.596G>A.D) Conservation of the Arg199 amino acid residue (yellow shade) through species.

Figure 1 .
Figure1.A) DNM2 protein with previously reported disease-causing variants.The sister-pair, P16 and P17, had the p.G358R variant.B) Conservation of the Gly358 amino acid residue (yellow shade) through species.C) Family pedigree; the parents were unaffected, while both sisters had CMT.D) Sanger sequencing was inconclusive for the mother, the father had the wild type allele, while the sisters were heterozygotes for the c.1072G>A variant.E) Minisequencing confirmed the mother to be a mosaic carrier, with a mutation load ranging from 20 to 32 % in urine, saliva, and blood.

Figure 4 .
Figure 4. A) PURA protein with previously reported disease-causing variants, the novel p.L100R variant is depicted below the linear graph.B) Protein conservation of the Leu100 amino acid residue (yellow shade) through species.C) Family pedigree, the proband had a de novo variant.D) Sanger sequencing results confirm the inheritance pattern of c.299T>G.

Primers for minisequencing DNM2 exon 8
1. Yarham JW et al.Defective i6A37 modification of mitochondrial and cytosolictRNAs results from pathogenic mutations in TRIT1 and its substrate tRNA.