Abstract Background The cerebellum is composed of a diverse array of neuronal subtypes. Here we have used a candidate approach to identify Zac1, a tumor suppressor gene encoding a zinc finger transcription factor, as a new player in the transcriptional network required for the development of a specific subset of cerebellar nuclei and a population of Golgi cells in the cerebellar cortex. Results We found that Zac1 has a complex expression profile in the developing cerebellum, including in two proliferating progenitor populations; the cerebellar ventricular zone and the external granular layer overlying posterior cerebellar lobules IX and X. Zac1 is also expressed in some postmitotic cerebellar neurons, including a subset of GABAergic interneurons in the medial cerebellar nuclei. Notably, GABAergic interneurons in the cerebellar nuclei are derived from the cerebellar ventricular zone, where Zac1 is also expressed, consistent with a lineage relationship between these two Zac1+ populations. Zac1 is also expressed in a small subset of cells in the posterior vermis, including some neurogranin-immunoreactive (NG+) Golgi cells, which, based on short-term birthdating, are derived from the EGL, where Zac1 is also expressed. However, Zac1+ cells and NG+ Golgi cells in the cerebellar cortex also display unique properties, as they are generated within different, albeit overlapping, time windows. Finally, consistent with the expression profile of Zac1, two conspicuous abnormalities were found in the cerebellum of Zac1 null mice: the medial cerebellar nuclei, and not the others, were significantly reduced in size; and the number of Golgi cells in cerebellar lobule IX was reduced by approximately 60% compared to wild-type littermates. Conclusions The data presented here indicate that the tumor suppressor gene Zac1 is expressed in a complex fashion in the developing cerebellum, including in two dividing progenitor populations and in specific subsets of postmitotic neurons, including Golgi cells and GABAergic neurons in the medial nuclei, which require Zac1 for their differentiation. We thus conclude that Zac1 is a critical regulator of normal cerebellar development, adding a new transcriptional regulator to the growing list of factors involved in generating neuronal diversity in the developing cerebellum.

The 22q13.3 deletion causes a neurodevelopmental syndrome, also known as Phelan-McDermid syndrome (MIM #606232), characterized by developmental delay and severe delay or absence of expressive speech. Two patients with hemizygous chromosome 22q13.3 telomeric deletion were referred to us when brain-imaging studies revealed cerebellar vermis hypoplasia (CBVH). To determine whether developmental abnormalities of the cerebellum are a consistent feature of the 22q13.3 deletion syndrome, we examined brain-imaging studies for 10 unrelated subjects with 22q13 terminal deletion. In seven cases where the availability of DNA and array technology allowed, we mapped deletion boundaries using comparative intensity analysis with single nucleotide polymorphism (SNP) microarrays. Approximate deletion boundaries for three additional cases were derived from clinical or published molecular data. We also examined brain-imaging studies for a patient with an intragenic SHANK3 mutation. We report the first brain-imaging data showing that some patients with 22q13 deletions have severe posterior CBVH, and one individual with a SHANK3 mutation has a normal cerebellum. This genotype-phenotype study suggests that the 22q13 deletion phenotype includes abnormal posterior fossa structures that are unlikely to be attributed to SHANK3 disruption. Other genes in the region, including PLXNB2 and MAPK8IP2, display brain expression patterns and mouse mutant phenotypes critical for proper cerebellar development. Future studies of these genes may elucidate their relationship to 22q13.3 deletion phenotypes.

[This corrects the article DOI: 10.1371/journal.pgen.1006425.].

Background

Autism is a complex childhood neurodevelopmental disorder with a strong genetic basis. Microdeletion or duplication of a approximately 500-700-kb genomic rearrangement on 16p11.2 that contains 24 genes represents the second most frequent chromosomal disorder associated with autism. The role of common and rare 16p11.2 sequence variants in autism etiology is unknown.

Methodology/principal findings

To identify common 16p11.2 variants with a potential role in autism, we performed association studies using existing data generated from three microarray platforms: Affymetrix 5.0 (777 families), Illumina 550 K (943 families), and Affymetrix 500 K (60 families). No common variants were identified that were significantly associated with autism. To look for rare variants, we performed resequencing of coding and promoter regions for eight candidate genes selected based on their known expression patterns and functions. In total, we identified 26 novel variants in autism: 13 exonic (nine non-synonymous, three synonymous, and one untranslated region) and 13 promoter variants. We found a significant association between autism and a coding variant in the seizure-related gene SEZ6L2 (12/1106 autism vs. 3/1161 controls; p = 0.018). Sez6l2 expression in mouse embryos was restricted to the spinal cord and brain. SEZ6L2 expression in human fetal brain was highest in post-mitotic cortical layers, hippocampus, amygdala, and thalamus. Association analysis of SEZ6L2 in an independent sample set failed to replicate our initial findings.

Conclusions/significance

We have identified sequence variation in at least one candidate gene in 16p11.2 that may represent a novel genetic risk factor for autism. However, further studies are required to substantiate these preliminary findings.

Gene expression levels vary across developmental stage, cell type, and region in the brain. Genomic variants also contribute to the variation in expression, and some neuropsychiatric disorder loci may exert their effects through this mechanism. To investigate these relationships, we present BrainVar, a unique resource of paired whole-genome and bulk tissue RNA sequencing from the dorsolateral prefrontal cortex of 176 individuals across prenatal and postnatal development. Here we identify common variants that alter gene expression (expression quantitative trait loci [eQTLs]) constantly across development or predominantly during prenatal or postnatal stages. Both "constant" and "temporal-predominant" eQTLs are enriched for loci associated with neuropsychiatric traits and disorders and colocalize with specific variants. Expression levels of more than 12,000 genes rise or fall in a concerted late-fetal transition, with the transitional genes enriched for cell-type-specific genes and neuropsychiatric risk loci, underscoring the importance of cataloging developmental trajectories in understanding cortical physiology and pathology.

WD40 repeat-containing proteins form a large family of proteins present in all eukaryotes. Here, we identified five pediatric probands with de novo variants in WDR37, which encodes a member of the WD40 repeat protein family. Two probands shared one variant and the others have variants in nearby amino acids outside the WD40 repeats. The probands exhibited shared phenotypes of epilepsy, colobomas, facial dysmorphology reminiscent of CHARGE syndrome, developmental delay and intellectual disability, and cerebellar hypoplasia. The WDR37 protein is highly conserved in vertebrate and invertebrate model organisms and is currently not associated with a human disease. We generated a null allele of the single Drosophila ortholog to gain functional insights and replaced the coding region of the fly gene CG12333/wdr37 with GAL4. These flies are homozygous viable but display severe bang sensitivity, a phenotype associated with seizures in flies. Additionally, the mutant flies fall when climbing the walls of the vials, suggesting a defect in grip strength, and repeat the cycle of climbing and falling. Similar to wall clinging defect, mutant males often lose grip of the female abdomen during copulation. These phenotypes are rescued by using the GAL4 in the CG12333/wdr37 locus to drive the UAS-human reference WDR37 cDNA. The two variants found in three human subjects failed to rescue these phenotypes, suggesting that these alleles severely affect the function of this protein. Taken together, our data suggest that variants in WDR37 underlie a novel syndromic neurological disorder.

Corpus callosum malformations are associated with a broad range of neurodevelopmental diseases. We report that de novo mutations in MAST1 cause mega-corpus-callosum syndrome with cerebellar hypoplasia and cortical malformations (MCC-CH-CM) in the absence of megalencephaly. We show that MAST1 is a microtubule-associated protein that is predominantly expressed in post-mitotic neurons and is present in both dendritic and axonal compartments. We further show that Mast1 null animals are phenotypically normal, whereas the deletion of a single amino acid (L278del) recapitulates the distinct neurological phenotype observed in patients. In animals harboring Mast1 microdeletions, we find that the PI3K/AKT3/mTOR pathway is unperturbed, whereas Mast2 and Mast3 levels are diminished, indicative of a dominant-negative mode of action. Finally, we report that de novo MAST1 substitutions are present in patients with autism and microcephaly, raising the prospect that mutations in this gene give rise to a spectrum of neurodevelopmental diseases.

PURPOSE: Nonmuscle myosin II complexes are master regulators of actin dynamics that play essential roles during embryogenesis with vertebrates possessing 3 nonmuscle myosin II heavy chain genes, MYH9, MYH10, and MYH14. As opposed to MYH9 and MYH14, no recognizable disorder has been associated with MYH10. We sought to define the clinical characteristics and molecular mechanism of a novel autosomal dominant disorder related to MYH10. METHODS: An international collaboration identified the patient cohort. CAS9-mediated knockout cell models were used to explore the mechanism of disease pathogenesis. RESULTS: We identified a cohort of 16 individuals with heterozygous MYH10 variants presenting with a broad spectrum of neurodevelopmental disorders and variable congenital anomalies that affect most organ systems and were recapitulated in animal models of altered MYH10 activity. Variants were typically de novo missense changes with clustering observed in the motor domain. MYH10 knockout cells showed defects in primary ciliogenesis and reduced ciliary length with impaired Hedgehog signaling. MYH10 variant overexpression produced a dominant-negative effect on ciliary length. CONCLUSION: These data presented a novel genetic cause of isolated and syndromic neurodevelopmental disorders related to heterozygous variants in the MYH10 gene with implications for disrupted primary cilia length control and altered Hedgehog signaling in disease pathogenesis.

The collapsin response mediator protein (CRMP) family proteins are intracellular mediators of neurotrophic factors regulating neurite structure/spine formation and are essential for dendrite patterning and directional axonal pathfinding during brain developmental processes. Among this family, CRMP5/DPYSL5 plays a significant role in neuronal migration, axonal guidance, dendrite outgrowth, and synapse formation by interacting with microtubules. Here, we report the identification of missense mutations in DPYSL5 in nine individuals with brain malformations, including corpus callosum agenesis and/or posterior fossa abnormalities, associated with variable degrees of intellectual disability. A recurrent de novo p.Glu41Lys variant was found in eight unrelated patients, and a p.Gly47Arg variant was identified in one individual from the first family reported with Ritscher-Schinzel syndrome. Functional analyses of the two missense mutations revealed impaired dendritic outgrowth processes in young developing hippocampal primary neuronal cultures. We further demonstrated that these mutations, both located in the same loop on the surface of DPYSL5 monomers and oligomers, reduced the interaction of DPYSL5 with neuronal cytoskeleton-associated proteins MAP2 and βIII-tubulin. Our findings collectively indicate that the p.Glu41Lys and p.Gly47Arg variants impair DPYSL5 function on dendritic outgrowth regulation by preventing the formation of the ternary complex with MAP2 and βIII-tubulin, ultimately leading to abnormal brain development. This study adds DPYSL5 to the list of genes implicated in brain malformation and in neurodevelopmental disorders.

Deadenylases are best known for degrading the poly(A) tail during mRNA decay. The deadenylase family has expanded throughout evolution and, in mammals, consists of 12 Mg²⁺-dependent 3'-end RNases with substrate specificity that is mostly unknown. Pontocerebellar hypoplasia type 7 (PCH7) is a unique recessive syndrome characterized by neurodegeneration and ambiguous genitalia. We studied 12 human families with PCH7, uncovering biallelic, loss-of-function mutations in TOE1, which encodes an unconventional deadenylase. toe1-morphant zebrafish displayed midbrain and hindbrain degeneration, modeling PCH-like structural defects in vivo. Surprisingly, we found that TOE1 associated with small nuclear RNAs (snRNAs) incompletely processed spliceosomal. These pre-snRNAs contained 3' genome-encoded tails often followed by post-transcriptionally added adenosines. Human cells with reduced levels of TOE1 accumulated 3'-end-extended pre-snRNAs, and the immunoisolated TOE1 complex was sufficient for 3'-end maturation of snRNAs. Our findings identify the cause of a neurodegenerative syndrome linked to snRNA maturation and uncover a key factor involved in the processing of snRNA 3' ends.