DNAJC13 mutations in Parkinson disease

Abstract

A Saskatchewan multi-incident family was clinically characterized with Parkinson disease (PD) and Lewy body pathology. PD segregates as an autosomal-dominant trait, which could not be ascribed to any known mutation. DNA from three affected members was subjected to exome sequencing. Genome alignment, variant annotation and comparative analyses were used to identify shared coding mutations. Sanger sequencing was performed within the extended family and ethnically matched controls. Subsequent genotyping was performed in a multi-ethnic case-control series consisting of 2928 patients and 2676 control subjects from Canada, Norway, Taiwan, Tunisia, and the USA. A novel mutation in receptor-mediated endocytosis 8/RME-8 (DNAJC13 p.Asn855Ser) was found to segregate with disease. Screening of cases and controls identified four additional patients with the mutation, of which two had familial parkinsonism. All carriers shared an ancestral DNAJC13 p.Asn855Ser haplotype and claimed Dutch-German-Russian Mennonite heritage. DNAJC13 regulates the dynamics of clathrin coats on early endosomes. Cellular analysis shows that the mutation confers a toxic gain-of-function and impairs endosomal transport. DNAJC13 immunoreactivity was also noted within Lewy body inclusions. In late-onset disease which is most reminiscent of idiopathic PD subtle deficits in endosomal receptor-sorting/recycling are highlighted by the discovery of pathogenic mutations VPS35, LRRK2 and now DNAJC13. With this latest discovery, and from a neuronal perspective, a temporal and functional ecology is emerging that connects synaptic exo- and endocytosis, vesicular trafficking, endosomal recycling and the endo-lysosomal degradative pathway. Molecular deficits in these processes are genetically linked to the phenotypic spectrum of parkinsonism associated with Lewy body pathology.

Figure 1.

Pedigrees with DNAJC13 mutations. Individual pedigrees are labeled and numbered according to their geographic origin (SK, Saskatchewan, Canada; BC, British Columbia, Canada; TU, Tunisia). (A) Original family used for exome sequencing and identification of DNAJC13 p.Asn855Ser. (B) Additional kindreds presenting DNAJC13 p.Asn855Ser mutations. (C) Pedigree presenting the p.Arg2115Leu variant. Black-filled symbols indicate individuals diagnosed with PD, phenocopies are highlighted with a white circle, gray filled symbols indicate dementia, and gray circle indicates progressive supranuclear palsy. Heterozygote mutation carriers (M) with corresponding age at disease onset are indicated in gray and wild-type (wt) genotypes in black. An asterisk indicates an inferred mutation carrier.

Figure 2.

DNAJC13 mutations and cross-species conservation. Protein orthologs were aligned via ClustalW. Amino acid positions for DNAJC13 p.Asn855Ser and p.Arg2115Leu are highlighted in black. Protein orthologs with amino acid positions differing from those of the human sequence are indicated in gray. RefSeq accession numbers: Homo sapiens, NP_056083.3; Nomascus leucogenys, XP_003265273.1; Mus musculus, NP_001156498.1; Dasypus novemcinctus, ENSDNOP00000006627; Canis lupus familiaris, XP_542783.3; Ailuropoda melanoleuca, XP_002926293.1; Bos taurus, DAA33060.1; Loxodonta africana, XP_003420956.1; Monodelphis domestica, ENSMODP00000014684; Dicentrarchus labrax, CBN82117.1; Gallus gallus, ENSGALP00000019064; Anolis carolinensis, ENSACAG00000000850; Xenopus tropicalis, ENSXETG00000001850; Petromyzon marinus, ENSPMAP00000010330; Ciona savignyi, SINCSAVP00000001383; Drosophila melanogaster, NP_610467.1; Amphimedon queenslandica, XP_003383293.1; Caenorhabditis elegans, NP_492222.

Figure 3.

Immunological staining of DNAJC13 in brain sections from p.Asn855Ser mutation carriers SK1 II-7 (A–C) and SK1 II-9 (D–F). (A) Negative staining in hypoglossal motor neurons. (B) Finely granular cytoplasmic immunoreactivity in dorsal motor nucleus of vagus. A subset of Lewy bodies (<50%) were positively labeled (arrows). Some Lewy bodies were completely unstained (C) two in a locus coeruleus neuron; others had weak staining (D) dorsal motor nucleus and some had dark immunoreactivity (E and F) substantia nigra.

Figure 4.

Transferrin uptake in cells over expressing wild-type and mutant DNAJC13 protein. (A) COS7 cells were transiently transfected with either GFP, wild-type DNAJC13 (GFP-DNAJC13-WT) or DNAJC13 p.Asn855Ser (GFP-DNAJC13-MT). Transferrin uptake assay was performed and cells imaged at T0 (prior to uptake) or T40 (40 min after chase). (B) Average number of transferrin positive puncta, (C) positive puncta size and (D) mean integrated density of puncta per cell 40 min after uptake. Statistically significant Fisher's LSD post hoc values are provided.