A syndromic neurodevelopmental disorder caused by rare variants in PPFIA3.

PPFIA3 encodes the protein-tyrosine phosphatase, receptor-type, F-polypeptide-interacting-protein-alpha-3 (PPFIA3), which is a member of the LAR-protein-tyrosine phosphatase-interacting-protein (liprin) family involved in synapse formation and function, synaptic vesicle transport, and presynaptic active zone assembly. The protein structure and function are evolutionarily well conserved, but human diseases related to PPFIA3 dysfunction are not yet reported in OMIM. Here, we report 20 individuals with rare PPFIA3 variants (19 heterozygous and 1 compound heterozygous) presenting with developmental delay, intellectual disability, hypotonia, dysmorphisms, microcephaly or macrocephaly, autistic features, and epilepsy with reduced penetrance. Seventeen unique PPFIA3 variants were detected in 18 families. To determine the pathogenicity of PPFIA3 variants in vivo, we generated transgenic fruit flies producing either human wild-type (WT) PPFIA3 or five missense variants using GAL4-UAS targeted gene expression systems. In the fly overexpression assays, we found that the PPFIA3 variants in the region encoding the N-terminal coiled-coil domain exhibited stronger phenotypes compared to those affecting the C-terminal region. In the loss-of-function fly assay, we show that the homozygous loss of fly Liprin-α leads to embryonic lethality. This lethality is partially rescued by the expression of human PPFIA3 WT, suggesting human PPFIA3 function is partially conserved in the fly. However, two of the tested variants failed to rescue the lethality at the larval stage and one variant failed to rescue lethality at the adult stage. Altogether, the human and fruit fly data reveal that the rare PPFIA3 variants are dominant-negative loss-of-function alleles that perturb multiple developmental processes and synapse formation.

Rare variants in PPFIA3 cause delayed development, intellectual disability, autism, and epilepsy.

PPFIA3 encodes the Protein-Tyrosine Phosphatase, Receptor-Type, F Polypeptide-Interacting Protein Alpha-3 (PPFIA3), which is a member of the LAR protein-tyrosine phosphatase-interacting protein (liprin) family involved in synaptic vesicle transport and presynaptic active zone assembly. The protein structure and function are well conserved in both invertebrates and vertebrates, but human diseases related to PPFIA3 dysfunction are not yet known. Here, we report 14 individuals with rare mono-allelic PPFIA3 variants presenting with features including developmental delay, intellectual disability, hypotonia, autism, and epilepsy. To determine the pathogenicity of PPFIA3 variants in vivo , we generated transgenic fruit flies expressing either human PPFIA3 wildtype (WT) or variant protein using GAL4-UAS targeted gene expression systems. Ubiquitous expression with Actin-GAL4 showed that the PPFIA3 variants had variable penetrance of pupal lethality, eclosion defects, and anatomical leg defects. Neuronal expression with elav-GAL4 showed that the PPFIA3 variants had seizure-like behaviors, motor defects, and bouton loss at the 3 rd instar larval neuromuscular junction (NMJ). Altogether, in the fly overexpression assays, we found that the PPFIA3 variants in the N-terminal coiled coil domain exhibited stronger phenotypes compared to those in the C-terminal region. In the loss-of-function fly assay, we show that the homozygous loss of fly Liprin- α leads to embryonic lethality. This lethality is partially rescued by the expression of human PPFIA3 WT, suggesting human PPFIA3 protein function is partially conserved in the fly. However, the PPFIA3 variants failed to rescue lethality. Altogether, the human and fruit fly data reveal that the rare PPFIA3 variants are dominant negative loss-of-function alleles that perturb multiple developmental processes and synapse formation.

PAK1 c.1409 T > a (p. Leu470Gln) de novo variant affects the protein kinase domain, leading to epilepsy, macrocephaly, spastic quadriplegia, and hydrocephalus: Case report and review of the literature.

The p-21-activated kinase 1 (PAK1) protein, encoded by the PAK1 gene, is an evolutionarily conserved serine/threonine-protein kinase that regulates key cellular developmental processes. To date, seven de novo PAK1 variants have been reported to cause the Intellectual Developmental Disorder with Macrocephaly, Seizures, and Speech Delay (IDDMSSD). In addition to the namesake features, other common characteristics include structural brain anomalies, delayed development, hypotonia, and dysmorphic features. Here, we report a de novo PAK1 NM_002576.5: c.1409 T > A variant (p.Leu470Gln) identified by trio genome sequencing (GS) in a 13-year-old boy with postnatal macrocephaly, obstructive hydrocephalus, medically refractory epilepsy, spastic quadriplegia, white matter hyperintensities, profound developmental disabilities, and a horseshoe kidney. This is the first recurrently affected residue identified in the protein kinase domain. Combined assessment of the eight pathogenic PAK1 missense variants reveal that the variants cluster in either the protein kinase or autoregulatory domains. Although interpretation of the phenotypic spectrum is limited by the sample size, neuroanatomical alterations were found more often in individuals with PAK1 variants in the autoregulatory domain. In contrast, non-neurological comorbidities were found more often in individuals with PAK1 variants in the protein kinase domain. Together, these findings expand the clinical spectrum of PAK1-associated IDDMSSD and reveal potential correlations with the affected protein domains.

An Integrated Phenotypic and Genotypic Approach Reveals a High-Risk Subtype Association for EBF3 Missense Variants Affecting the Zinc Finger Domain.

OBJECTIVE: Collier/Olf/EBF (COE) transcription factors have distinct expression patterns in the developing and mature nervous system. To date, a neurological disease association has been conclusively established for only the Early B-cell Factor-3 (EBF3) COE family member through the identification of heterozygous loss-of-function variants in individuals with autism spectrum/neurodevelopmental disorders (NDD). Here, we identify a symptom severity risk association with missense variants primarily disrupting the zinc finger domain (ZNF) in EBF3-related NDD. METHODS: A phenotypic assessment of 41 individuals was combined with a literature meta-analysis for a total of 83 individuals diagnosed with EBF3-related NDD. Quantitative diagnostic phenotypic and symptom severity scales were developed to compare EBF3 variant type and location to identify genotype-phenotype correlations. To stratify the effects of EBF3 variants disrupting either the DNA-binding domain (DBD) or the ZNF, we used in vivo fruit fly UAS-GAL4 expression and in vitro luciferase assays. RESULTS: We show that patient symptom severity correlates with EBF3 missense variants perturbing the ZNF, which is a key protein domain required for stabilizing the interaction between EBF3 and the target DNA sequence. We found that ZNF-associated variants failed to restore viability in the fruit fly and impaired transcriptional activation. However, the recurrent variant EBF3 p.Arg209Trp in the DBD is capable of partially rescuing viability in the fly and preserved transcriptional activation. INTERPRETATION: We describe a symptom severity risk association with ZNF perturbations and EBF3 loss-of-function in the largest reported cohort to date of EBF3-related NDD patients. This analysis should have potential predictive clinical value for newly identified patients with EBF3 gene variants. ANN NEUROL 2022;92:138-153.