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. 2017 Feb 2;100(2):257-266.
doi: 10.1016/j.ajhg.2017.01.002. Epub 2017 Jan 26.

Biallelic Mutations in DNAJC12 Cause Hyperphenylalaninemia, Dystonia, and Intellectual Disability

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Free PMC article

Biallelic Mutations in DNAJC12 Cause Hyperphenylalaninemia, Dystonia, and Intellectual Disability

Yair Anikster et al. Am J Hum Genet. .
Free PMC article

Abstract

Phenylketonuria (PKU, phenylalanine hydroxylase deficiency), an inborn error of metabolism, can be detected through newborn screening for hyperphenylalaninemia (HPA). Most individuals with HPA harbor mutations in the gene encoding phenylalanine hydroxylase (PAH), and a small proportion (2%) exhibit tetrahydrobiopterin (BH4) deficiency with additional neurotransmitter (dopamine and serotonin) deficiency. Here we report six individuals from four unrelated families with HPA who exhibited progressive neurodevelopmental delay, dystonia, and a unique profile of neurotransmitter deficiencies without mutations in PAH or BH4 metabolism disorder-related genes. In these six affected individuals, whole-exome sequencing (WES) identified biallelic mutations in DNAJC12, which encodes a heat shock co-chaperone family member that interacts with phenylalanine, tyrosine, and tryptophan hydroxylases catalyzing the BH4-activated conversion of phenylalanine into tyrosine, tyrosine into L-dopa (the precursor of dopamine), and tryptophan into 5-hydroxytryptophan (the precursor of serotonin), respectively. DNAJC12 was undetectable in fibroblasts from the individuals with null mutations. PAH enzyme activity was reduced in the presence of DNAJC12 mutations. Early treatment with BH4 and/or neurotransmitter precursors had dramatic beneficial effects and resulted in the prevention of neurodevelopmental delay in the one individual treated before symptom onset. Thus, DNAJC12 deficiency is a preventable and treatable cause of intellectual disability that should be considered in the early differential diagnosis when screening results are positive for HPA. Sequencing of DNAJC12 may resolve any uncertainty and should be considered in all children with unresolved HPA.

Keywords: BH(4); DNAJC12; dystonia; hyperphenylalaninemia; neurotransmitter deficiency; newborn screening; phenylketonuria; tetrahydrobiopterin.

Figures

Figure 1
Figure 1
Pedigrees of the Investigated Families and Schematic Representation of the DNAJC12 Exon 4 Deletion (A) Pedigrees of four families with mutations in DNAJC12. The mutation status of affected (closed symbols) and healthy (open symbols) family members are noted. N.D., not determined. (B) Schematic representation of del6943 (c.298-968_503-2603del). Exome sequencing results (top) showing the alignment of sequencing reads to the reference genome for the controls and affected individuals (A-IV-2 and D-V-1), as visualized with the Integrative Genomics Viewer (IGV) tool. Breakpoint junction sequences (bottom) are shown, depicting the 6.943 kb deletion (del6943, c.298-968_503-2603del) containing exon 4. The breakpoint junction sequence (chromatogram) is aligned with reference sequences.
Figure 2
Figure 2
DNAJC12 Expression Studies (A) DNAJC12 mRNA expression in fibroblasts, measured by quantitative real-time polymerase chain reaction (qPCR) and standardized against GAPDH. The graph shows the normalized relative DNAJC12 mRNA levels in two affected individuals (B-IV-1 and B-IV-2) and three unaffected age-matched healthy control individuals. (B and C) Representative western blot (B) and densitometric quantification (C, left) of the DNAJC12 protein levels in fibroblasts from three unaffected age-matched healthy control individuals (C1–C3), in fibroblasts from the heterozygous unaffected father of B-IV-1 and B-IV-2 (Het), and in fibroblasts with the homozygous missense variant (c.215G>C, C-II-4), the homozygous del6943 (A-IV-2), and the homozygous canonical splice variant (c.158−2A>T, B-IV-1 and B-IV-2). Densitometric analysis of DNAJC12 protein (C, right) was performed in fibroblasts from individual C-II-4 (n = 3) and in fibroblasts from three unaffected age-matched healthy control individuals (n = 3). DNAJC12 level was normalized to β-actin. Results are presented as means ± SEM; p < 0.05.
Figure 3
Figure 3
Overview of the DNAJC12, PAH, TH, and TPH2 Protein Interaction Network, as Determined by Immunoprecipitation-Mass Spectrometry Analysis C-terminally tagged DNAJC12, PAH, TH, and TPH2 were immunopurified from HEK293 cells, and protein complexes were analyzed by liquid chromatography-mass spectrometry. CompPASS analysis was performed in technical duplicates. Proteins with NWD scores > 1, ASPM > 2, and entropy values > 0.7 that were reproducibly detected in the two CompPASS analyses were considered high-confidence interactors. Baits are represented in red. High-confidence interacting proteins (HCIPs) are represented in gray.
Figure 4
Figure 4
Phenylalanine Hydroxylase Enzyme Activity Is Reduced in the Presence of DNAJC12 Mutations (A) PAH activity was measured using liquid chromatography-electrospray ionization tandem mass spectrometry in fibroblasts with the homozygous missense variant (c.215G>C, individual C-II-4) and in fibroblasts from an unaffected age-matched healthy control after 5 μg or 10 μg PAH plasmids transfection. (B) Western blot assay of PAH, DNAJC12, and β-actin proteins in individual C-II-4 and control fibroblasts after 5 μg or 10 μg PAH plasmids transfection.

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