Loss of PCLO function underlies pontocerebellar hypoplasia type III

Abstract

Objective: To identify the genetic cause of pontocerebellar hypoplasia type III (PCH3).

Methods: We studied the original reported pedigree of PCH3 and performed genetic analysis including genome-wide single nucleotide polymorphism genotyping, linkage analysis, whole-exome sequencing, and Sanger sequencing. Human fetal brain RNA sequencing data were then analyzed for the identified candidate gene.

Results: The affected individuals presented with severe global developmental delay and seizures starting in the first year of life. Brain MRI of an affected individual showed diffuse atrophy of the cerebrum, cerebellum, and brainstem. Genome-wide single nucleotide polymorphism analysis confirmed the linkage to chromosome 7q we previously reported, and showed no other genomic areas of linkage. Whole-exome sequencing of 2 affected individuals identified a shared homozygous, nonsense variant in the PCLO (piccolo) gene. This variant segregated with the disease phenotype in the pedigree was rare in the population and was predicted to eliminate the PDZ and C2 domains in the C-terminus of the protein. RNA sequencing data of human fetal brain showed that PCLO was moderately expressed in the developing cerebral cortex.

Conclusions: Here, we show that a homozygous, nonsense PCLO mutation underlies the autosomal recessive neurodegenerative disorder, PCH3. PCLO is a component of the presynaptic cytoskeletal matrix, and is thought to be involved in regulation of presynaptic proteins and synaptic vesicles. Our findings suggest that PCLO is crucial for the development and survival of a wide range of neuronal types in the human brain.

Figure 1. The pedigree studied and identification of a homozygous PCLO mutation

(A) The pedigree studied with genotype of the identified PCLO mutation in each individual. Only the affected individuals show the homozygous C to T change. (B) Brain MRIs of individual II:2 at age 11 years, 10 months. Sagittal T1-weighted image shows a thin pons and small cerebellar vermis. The corpus callosum is thin. Coronal T2-weighted image shows generalized reduction of the cerebral white matter and atrophic cerebellar hemispheres. Scale bars = 2 cm. (C) Genome-wide linkage analysis. A region on chromosome 7 shows a maximum multipoint LOD score of 2.958. (D) Schematic representation of the PCLO protein. X = the location of the identified mutation. C2 = C2 domain; LOD = logarithm of odds; PDZ = PDZ domain; Zn = zinc finger domain.

Figure 2. Expression of PCLO in the developing human brain

Expression of PCLO (top), a related presynaptic protein gene, BSN (middle), and another synaptic protein associated with intellectual disability, GRIK2 (bottom), seen in messenger RNA sequencing of the developing cerebral cortex of a 9-week human embryo. Y-axis units are in reads per million mapped reads (RPM). RNA isoforms as annotated in RefSeq are shown below each expression plot. The arrow indicates the location of the PCLO mutation.