Unveiling the crucial neuronal role of the proteasomal ATPase subunit gene PSMC5 in neurodevelopmental proteasomopathies.

Neurodevelopmental proteasomopathies represent a distinctive category of neurodevelopmental disorders (NDD) characterized by genetic variations within the 26S proteasome, a protein complex governing eukaryotic cellular protein homeostasis. In our comprehensive study, we identified 23 unique variants in PSMC5 , which encodes the AAA-ATPase proteasome subunit PSMC5/Rpt6, causing syndromic NDD in 38 unrelated individuals. Overexpression of PSMC5 variants altered human hippocampal neuron morphology, while PSMC5 knockdown led to impaired reversal learning in flies and loss of excitatory synapses in rat hippocampal neurons. PSMC5 loss-of-function resulted in abnormal protein aggregation, profoundly impacting innate immune signaling, mitophagy rates, and lipid metabolism in affected individuals. Importantly, targeting key components of the integrated stress response, such as PKR and GCN2 kinases, ameliorated immune dysregulations in cells from affected individuals. These findings significantly advance our understanding of the molecular mechanisms underlying neurodevelopmental proteasomopathies, provide links to research in neurodegenerative diseases, and open up potential therapeutic avenues.

Role of CAMK2D in neurodevelopment and associated conditions.

The calcium/calmodulin-dependent protein kinase type 2 (CAMK2) family consists of four different isozymes, encoded by four different genes-CAMK2A, CAMK2B, CAMK2G, and CAMK2D-of which the first three have been associated recently with neurodevelopmental disorders. CAMK2D is one of the major CAMK2 proteins expressed in the heart and has been associated with cardiac anomalies. Although this CAMK2 isoform is also known to be one of the major CAMK2 subtypes expressed during early brain development, it has never been linked with neurodevelopmental disorders until now. Here we show that CAMK2D plays an important role in neurodevelopment not only in mice but also in humans. We identified eight individuals harboring heterozygous variants in CAMK2D who display symptoms of intellectual disability, delayed speech, behavioral problems, and dilated cardiomyopathy. The majority of the variants tested lead to a gain of function (GoF), which appears to cause both neurological problems and dilated cardiomyopathy. In contrast, loss-of-function (LoF) variants appear to induce only neurological symptoms. Together, we describe a cohort of individuals with neurodevelopmental disorders and cardiac anomalies, harboring pathogenic variants in CAMK2D, confirming an important role for the CAMK2D isozyme in both heart and brain function.

PSMC3 proteasome subunit variants are associated with neurodevelopmental delay and type I interferon production.

A critical step in preserving protein homeostasis is the recognition, binding, unfolding, and translocation of protein substrates by six AAA-ATPase proteasome subunits (ATPase-associated with various cellular activities) termed PSMC1-6, which are required for degradation of proteins by 26S proteasomes. Here, we identified 15 de novo missense variants in the PSMC3 gene encoding the AAA-ATPase proteasome subunit PSMC3/Rpt5 in 23 unrelated heterozygous patients with an autosomal dominant form of neurodevelopmental delay and intellectual disability. Expression of PSMC3 variants in mouse neuronal cultures led to altered dendrite development, and deletion of the PSMC3 fly ortholog Rpt5 impaired reversal learning capabilities in fruit flies. Structural modeling as well as proteomic and transcriptomic analyses of T cells derived from patients with PSMC3 variants implicated the PSMC3 variants in proteasome dysfunction through disruption of substrate translocation, induction of proteotoxic stress, and alterations in proteins controlling developmental and innate immune programs. The proteostatic perturbations in T cells from patients with PSMC3 variants correlated with a dysregulation in type I interferon (IFN) signaling in these T cells, which could be blocked by inhibition of the intracellular stress sensor protein kinase R (PKR). These results suggest that proteotoxic stress activated PKR in patient-derived T cells, resulting in a type I IFN response. The potential relationship among proteosome dysfunction, type I IFN production, and neurodevelopment suggests new directions in our understanding of pathogenesis in some neurodevelopmental disorders.

Unexpected moves: a conformational change in MutSα enables high-affinity DNA mismatch binding.

The DNA mismatch repair protein MutSα recognizes wrongly incorporated DNA bases and initiates their correction during DNA replication. Dysfunctions in mismatch repair lead to a predisposition to cancer. Here, we study the homozygous mutation V63E in MSH2 that was found in the germline of a patient with suspected constitutional mismatch repair deficiency syndrome who developed colorectal cancer before the age of 30. Characterization of the mutant in mouse models, as well as slippage and repair assays, shows a mildly pathogenic phenotype. Using cryogenic electron microscopy and surface plasmon resonance, we explored the mechanistic effect of this mutation on MutSα function. We discovered that V63E disrupts a previously unappreciated interface between the mismatch binding domains (MBDs) of MSH2 and MSH6 and leads to reduced DNA binding. Our research identifies this interface as a 'safety lock' that ensures high-affinity DNA binding to increase replication fidelity. Our mechanistic model explains the hypomorphic phenotype of the V63E patient mutation and other variants in the MBD interface.

The MAP3K7 gene: Further delineation of clinical characteristics and genotype/phenotype correlations.

Mitogen-activated protein 3 kinase 7 (MAP3K7) encodes the ubiquitously expressed transforming growth factor ß-activated kinase 1, which plays a crucial role in many cellular processes. Mutationsin the MAP3K7 gene have been linked to two distinct disorders: frontometaphyseal dysplasia type 2 (FMD2) and cardiospondylocarpofacial syndrome (CSCF). The fact that different mutations can induce two distinct phenotypes suggests a phenotype/genotype correlation, but no side-by-side comparison has been done thus far to confirm this. Here, we significantly expand the cohort and the description of clinical phenotypes for patients with CSCF and FMD2 who carry mutations in MAP3K7. Our findings support that in contrast to FMD2-causing mutations, CSCF-causing mutations in MAP3K7 have a loss-of-function effect. Additionally, patients with pathogenic mutations in MAP3K7 are at risk for (severe) cardiac disease, have symptoms associated with connective tissue disease, and we show overlap in clinical phenotypes of CSCF with Noonan syndrome (NS). Together, we confirm a molecular fingerprint of FMD2- versus CSCF-causing MAP3K7 mutations and conclude that mutations in MAP3K7 should be considered in the differential diagnosis of patients with syndromic congenital cardiac defects and/or cardiomyopathy, syndromic connective tissue disorders, and in the differential diagnosis of NS.

Pro-mutagenic effects of the gut microbiota in a Lynch syndrome mouse model.

The gut microbiota strongly impacts the development of sporadic colorectal cancer (CRC), but it is largely unknown how the microbiota affects the pathogenesis of mismatch-repair-deficient CRC in the context of Lynch syndrome. In a mouse model for Lynch syndrome, we found a nearly complete loss of intestinal tumor development when animals were transferred from a conventional "open" animal facility to specific-pathogen-free (SPF) conditions. Using 16S sequencing we detected large changes in microbiota composition between the two facilities. Transcriptomic analyses of tumor-free intestinal tissues showed signs of strong intestinal inflammation in conventional mice. Whole exome sequencing of tumors developing in Msh2-Lynch mice revealed a much lower mutational load in the single SPF tumor than in tumors developing in conventional mice, suggesting reduced epithelial proliferation in SPF mice. Fecal microbiota transplantations with conventional feces altered the immune landscape and gut homeostasis, illustrated by increased gut length and elevated epithelial proliferation and migration. This was associated with drastic changes in microbiota composition, in particular increased relative abundances of different mucus-degrading taxa such as Desulfovibrio and Akkermansia, and increased bacterial-epithelial contact. Strikingly, transplantation of conventional microbiota increased microsatellite instability in untransformed intestinal epithelium of Msh2-Lynch mice, indicating that the composition of the microbiota influences the rate of mutagenesis in MSH2-deficient crypts.

Loss of CAMK2G affects intrinsic and motor behavior but has minimal impact on cognitive behavior.

Introduction: The gamma subunit of calcium/calmodulin-dependent protein kinase 2 (CAMK2G) is expressed throughout the brain and is associated with neurodevelopmental disorders. Research on the role of CAMK2G is limited and attributes different functions to specific cell types. Methods: To further expand on the role of CAMK2G in brain functioning, we performed extensive phenotypic characterization of a Camk2g knockout mouse. Results: We found different CAMK2G isoforms that show a distinct spatial expression pattern in the brain. Additionally, based on our behavioral characterization, we conclude that CAMK2G plays a minor role in hippocampus-dependent learning and synaptic plasticity. Rather, we show that CAMK2G is required for motor function and that the loss of CAMK2G results in impaired nest-building and marble burying behavior, which are innate behaviors that are associated with impaired neurodevelopment. Discussion: Taken together, our results provide evidence for a unique function of this specific CAMK2 isozyme in the brain and further support the role of CAMK2G in neurodevelopment.

TAOK1 is associated with neurodevelopmental disorder and essential for neuronal maturation and cortical development.

Thousand and one amino-acid kinase 1 (TAOK1) is a MAP3K protein kinase, regulating different mitogen-activated protein kinase pathways, thereby modulating a multitude of processes in the cell. Given the recent finding of TAOK1 involvement in neurodevelopmental disorders (NDDs), we investigated the role of TAOK1 in neuronal function and collected a cohort of 23 individuals with mostly de novo variants in TAOK1 to further define the associated NDD. Here, we provide evidence for an important role for TAOK1 in neuronal function, showing that altered TAOK1 expression levels in the embryonic mouse brain affect neural migration in vivo, as well as neuronal maturation in vitro. The molecular spectrum of the identified TAOK1 variants comprises largely truncating and nonsense variants, but also missense variants, for which we provide evidence that they can have a loss of function or dominant-negative effect on TAOK1, expanding the potential underlying causative mechanisms resulting in NDD. Taken together, our data indicate that TAOK1 activity needs to be properly controlled for normal neuronal function and that TAOK1 dysregulation leads to a neurodevelopmental disorder mainly comprising similar facial features, developmental delay/intellectual disability and/or variable learning or behavioral problems, muscular hypotonia, infant feeding difficulties, and growth problems.