Background: Progressive cerebello-cerebral atrophy (PCCA) leading to profound mental retardation, progressive microcephaly, spasticity and early onset epilepsy, was diagnosed in four non-consanguineous apparently unrelated families of Jewish Moroccan ancestry. Common founder mutation(s) were assumed. Methods: Genome-wide linkage analysis and whole exome sequencing were done, followed by realtime PCR and immunofluorescent microscopy. Results: Genome-wide linkage analysis mapped the disease-associated gene to 0.5 Mb on chromosome 17p13.3. Whole exome sequencing identified only two mutations within this locus, which were common to the affected individuals: compound heterozygous mutations in VPS53, segregating as expected for autosomal recessive heredity within all four families, and common in Moroccan Jews (∼1:37 carrier rate). The Golgi-associated retrograde protein (GARP) complex is involved in the retrograde pathway recycling endocytic vesicles to Golgi; c.2084A>G and c.1556+5G>A VPS53 founder mutations are predicted to affect the C-terminal domain of VPS53, known to be critical to its role as part of this complex. Immunofluorescent microscopy demonstrated swollen and abnormally numerous CD63 positive vesicular bodies, likely intermediate recycling/late endosomes, in fibroblasts of affected individuals. Conclusions: Autosomal recessive PCCA type 2 is caused by VPS53 mutations.

Primary microcephaly is a congenital neurodevelopmental disorder of reduced head circumference and brain volume, with fewer neurons in the cortex of the developing brain due to premature transition between symmetrical and asymmetrical cellular division of the neuronal stem cell layer during neurogenesis. We now show through linkage analysis and whole exome sequencing, that a dominant mutation in ALFY, encoding an autophagy scaffold protein, causes human primary microcephaly. We demonstrate the dominant effect of the mutation in drosophila: transgenic flies harboring the human mutant allele display small brain volume, recapitulating the disease phenotype. Moreover, eye-specific expression of human mutant ALFY causes rough eye phenotype. In molecular terms, we demonstrate that normally ALFY attenuates the canonical Wnt signaling pathway via autophagy-dependent removal specifically of aggregates of DVL3 and not of Dvl1 or Dvl2. Thus, autophagic attenuation of Wnt signaling through removal of Dvl3 aggregates by ALFY acts in determining human brain size. One of the major events in human evolution is the significant increase in brain volume in the transition from primates to humans. The molecular pathways determining the larger size of the human brain are not fully understood. Hereditary primary microcephaly, a neurodevelopmental disorder in which infants are born with small head circumference and reduced brain volume with intellectual disability, offers insights to the embryonic molecular pathways determining human brain size. Previous studies have shown that human microcephaly can be caused by mutations in genes affecting cell division processes, such as cell cycle regulation, DNA replication, primary cilia formation and centriole and centrosome duplication. We now show a novel molecular pathway determining human brain size: human primary microcephaly can be caused by a mutation in ALFY, a gene that encodes an autophagy scaffold protein. In fact, transgenic flies over expressing the mutant form of human ALFY recapitulate the human disease phenotype of microcephaly. We show the molecular pathway through which ALFY regulates cell division and differentiation: we demonstrate that ALFY normally controls removal of aggregate of DVL3, and through this regulates Wnt signaling, a major molecular pathway in embryogenesis. Thus, Wnt signaling, controlled by ALFY-mediated aggregate removal of DVL3, determines human brain size and human microcephaly.

Foveal hypoplasia, always accompanied by nystagmus, is found as part of the clinical spectrum of various eye disorders such as aniridia, albinism and achromatopsia. However, the molecular basis of isolated autosomal recessive foveal hypoplasia is yet unknown. Individuals of apparently unrelated non consanguineous Israeli families of Jewish Indian (Mumbai) ancestry presented with isolated foveal hypoplasia associated with congenital nystagmus and reduced visual acuity. Genome-wide homozygosity mapping followed by fine mapping defined a 830 Kb disease-associated locus (LOD score 3.5). Whole-exome sequencing identified a single missense mutation in the homozygosity region: c.95T>G, p.(Ile32Ser), in a conserved amino acid within the first predicted transmembrane domain of SLC38A8. The mutation fully segregated with the disease-associated phenotype, demonstrating an ∼10% carrier rate in Mumbai Jews. SLC38A8 encodes a putative sodium-dependent amino-acid/proton antiporter, which we showed to be expressed solely in the eye. Thus, a homozygous SLC38A8 mutation likely underlies isolated foveal hypoplasia.