DLG4-related synaptopathy: a new rare brain disorder

PURPOSE: Postsynaptic density protein-95 (PSD-95), encoded by DLG4, regulates excitatory synaptic function in the brain. Here we present the clinical and genetic features of 53 patients (42 previously unpublished) with DLG4 variants.METHODS: The clinical and genetic information were collected through GeneMatcher collaboration. All the individuals were investigated by local clinicians and the gene variants were identified by clinical exome/genome sequencing.RESULTS: The clinical picture was predominated by early onset global developmental delay, intellectual disability, autism spectrum disorder, and attention deficit-hyperactivity disorder, all of which point to a brain disorder. Marfanoid habitus, which was previously suggested to be a characteristic feature of DLG4-related phenotypes, was found in only nine individuals and despite some overlapping features, a distinct facial dysmorphism could not be established. Of the 45 different DLG4 variants, 39 were predicted to lead to loss of protein function and the majority occurred de novo (four with unknown origin). The six missense variants identified were suggested to lead to structural or functional changes by protein modeling studies.CONCLUSION: The present study shows that clinical manifestations associated with DLG4 overlap with those found in other neurodevelopmental disorders of synaptic dysfunction; thus, we designate this group of disorders as DLG4-related synaptopathy.Genetics of disease, diagnosis and treatmen

Whole thorax irradiation of non-human primates induces persistent nuclear damage and gene expression changes in peripheral blood cells.

We investigated the cytogenetic and gene expression responses of peripheral blood cells of non-human primates (NHP, Macaca mulatta) that were whole-thorax irradiated with a single dose of 10 Gy. In this model, partial irradiation of NHPs in the thoracic region (Whole Thorax Lung Irradiation, WTLI) allows the study of late radiation-induced lung injury, while avoiding acute radiation syndromes related to hematopoietic and gastrointestinal injury. A transient drop in circulating lymphocytes and platelets was seen by 9 days, followed by elevations in respiratory rate, circulating neutrophils, lymphocytes, and monocytes at 60-100 days, corresponding to computed tomography (CT) and histologic evidence of pneumonitis, and elective euthanasia of four animals. To evaluate long-term DNA damage in NHP peripheral blood lymphocytes after 10 Gy WTLI, we used the cytokinesis-block micronucleus (CBMN) assay to measure chromosomal aberrations as post-mitotic micronuclei in blood samples collected up to 8 months after irradiation. Regression analysis showed significant induction of micronuclei in NHP blood cells that persisted with a gradual decline over the 8-month study period, suggesting long-term DNA damage in blood lymphocytes after WTLI. We also report transcriptomic changes in blood up to 30 days after WTLI. We isolated total RNA from peripheral blood at 3 days before and then at 2, 5 and 30 days after irradiation. We identified 1187 transcripts that were significantly changed across the 30-day time course. From changes in gene expression, we identified biological processes related to immune responses, which persisted across the 30-day study. Response to oxygen-containing compounds and bacteria were implicated by gene-expression changes at the earliest day 2 and latest, day 30 time-points. Gene expression changes suggest a persistent altered state of the immune system, specifically response to infection, for at least a month after WTLI

Micronucleus frequency of individual animals.

<p>Logistic regression analysis of the fractions of binucleated cells with micronuclei at various times after irradiation across different NHPs. Error bars are 95% confidence intervals.</p

Microarray results of gene expression changes of 25 genes, common across all time points.

<p>Gene expression changes of the 25 genes that were differentially expressed at all the time points assayed, the intersection of all groups taken from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0191402#pone.0191402.g004" target="_blank">Fig 4</a>. These genes were selected based on being significantly changed at all times in the study. In each comparison, gene expression signal intensities of these 25 genes at days 2, 5 and 30 were compared with the pre-exposure levels in the matched control to determine fold changes, the bars in the histogram are the mean fold change in five independent biological replicates, error bars are SEM. The dashed line represents the normalized pre-exposure expression level.</p

Differential blood cell counts.

<p>Total leukocyte counts (A) were most elevated at 66 days, reflecting neutrophilia (D). Transient reductions of lymphocyte counts (B) and platelets (C) were seen at day 9. Day 0 values are the overall average and SEM of the day -72 baseline measurements. * = significantly different from baseline at p<0.05 using Dunnett’s method. Error bars are 1 standard error of the mean.</p

qRT-PCR validation of gene expression changes.

<p>Gene expression was measured in blood cell RNA using qRT-PCR to validate the microarray measurements. Two consistently up regulated genes are shown here, <i>IL1R1</i> (A) and <i>LY96</i> (B); and two downregulated genes, <i>MARCKSL1</i> (C) and <i>FAIM3</i> (D). Relative expression are mean fold changes at 2, 5 and 30 days in at least three biological replicates. Microarray changes are solid symbols (●) and qRT-PCR fold changes are shown in open symbols (o), representing the mean, error bars are SEM.</p

PANTHER biological processes related to the immune response, which are significant at 2 days after WTLI and not significant at later time points.

<p>PANTHER biological processes related to the immune response, which are significant at 2 days after WTLI and not significant at later time points.</p

Micronucleus frequency in NHP peripheral blood lymphocytes measured up to 249 days post exposure.

<p>Micronucleus frequency in NHP peripheral blood lymphocytes measured up to 249 days post exposure.</p

Venn diagram of differentially expressed genes.

<p>Differentially expressed genes in blood at 2, 5 and 30 days after WTLI in NHP are shown in the Venn diagram. 25 genes were significantly differentially expressed at all the time points in the study.</p