'Salt & Pepper' syndrome is an autosomal recessive condition characterized by severe intellectual disability, epilepsy, scoliosis, choreoathetosis, dysmorphic facial features and altered dermal pigmentation. High-density SNP array analysis performed on siblings first described with this syndrome detected four shared regions of loss of heterozygosity (LOH). Whole-exome sequencing narrowed the candidate region to chromosome 2p11.2. Sanger sequencing confirmed a homozygous c.994G>A transition (p.E332K) in the ST3GAL5 gene, which encodes for a sialyltransferase also known as GM3 synthase. A different homozygous mutation of this gene has been previously associated with infantile-onset epilepsy syndromes in two other cohorts. The ST3GAL5 enzyme synthesizes ganglioside GM3, a glycosophingolipid enriched in neural tissue, by adding sialic acid to lactosylceramide. Unlike disorders of glycosphingolipid (GSL) degradation, very little is known regarding the molecular and pathophysiologic consequences of altered GSL biosynthesis. Glycolipid analysis confirmed a complete lack of GM3 ganglioside in patient fibroblasts, while microarray analysis of glycosyltransferase mRNAs detected modestly increased expression of ST3GAL5 and greater changes in transcripts encoding enzymes that lie downstream of ST3GAL5 and in other GSL biosynthetic pathways. Comprehensive glycomic analysis of N-linked, O-linked and GSL glycans revealed collateral alterations in response to loss of complex gangliosides in patient fibroblasts and in zebrafish embryos injected with antisense morpholinos that targeted zebrafish st3gal5 expression. Morphant zebrafish embryos also exhibited increased apoptotic cell death in multiple brain regions, emphasizing the importance of GSL expression in normal neural development and function.

Chloride intracellular channel 2 (CLIC2) protein is a member of the glutathione transferase class of proteins. Its' only known function is the regulation of ryanodine receptor (RyR) intracellular Ca(2+) release channels. These RyR proteins play a major role in the regulation of Ca(2+) signaling in many cells. Utilizing exome capture and deep sequencing of genes on the X-chromosome, we have identified a mutation in CLIC2 (c.303C>G, p.H101Q) which is associated with X-linked intellectual disability (ID), atrial fibrillation, cardiomegaly, congestive heart failure (CHF), some somatic features and seizures. Functional studies of the H101Q variant indicated that it stimulated rather than inhibited the action of RyR channels, with channels remaining open for longer times and potentially amplifying Ca(2+) signals dependent on RyR channel activity. The overly active RyRs in cardiac and skeletal muscle cells and neuronal cells would result in abnormal cardiac function and trigger post-synaptic pathways and neurotransmitter release. The presence of both cardiomegaly and CHF in the two affected males and atrial fibrillation in one are consistent with abnormal RyR2 channel function. Since the dysfunction of RyR2 channels in the brain via 'leaky mutations' can result in mild developmental delay and seizures, our data also suggest a vital role for the CLIC2 protein in maintaining normal cognitive function via its interaction with RyRs in the brain. Therefore, our patients appear to suffer from a new channelopathy comprised of ID, seizures and cardiac problems because of enhanced Ca(2+) release through RyRs in neuronal cells and cardiac muscle cells.

Microdeletions of chromosomal region 2q23.1 that disrupt MBD5 contribute to a spectrum of neurodevelopmental phenotypes, however the impact of this locus in human psychopathology has not been described. To characterize the structural variation landscape of MBD5 disruptions and the associated psychopathology, 22 individuals with genomic disruption of MBD5 (translocation, point mutation, and deletion) were identified through whole-genome sequencing or cytogenomic microarray at 11 molecular diagnostic centers. The genomic impact ranged from a single base pair to 5.4 Mb. Parents were available for 11 cases, all of which confirmed the rearrangement arose de novo. Phenotypes were largely indistinguishable between patients with full-segment 2q23.1 deletions and those with intragenic MBD5 rearrangements, including alterations confined entirely to the 5′UTR, confirming the critical impact of non-coding sequence at this locus. We found heterogeneous, multi-system pathogenic effects of MBD5 disruption and characterized the associated spectrum of psychopathology, which includes sensory integration disorder, anxiety, self-hugging, bipolar disorder and others. Importantly, unique features of the oldest assessed patient were early-onset dementia and behavioral regression. Analyses also revealed phenotypes that distinguish MBD5 disruptions from seven well-established syndromes with significant diagnostic overlap. This study indicates that haploinsufficiency of MBD5 causes diverse phenotypes, yields insight into the spectrum of resulting neurodevelopmental and behavioral psychopathology, and provides clinical context for interpretation of MBD5 structural variations. Empirical evidence also suggests that disruption of non-coding MBD5 regulatory regions is sufficient for clinical manifestation, highlighting the limitations of exon-focused assessments. These results suggest an ongoing perturbation of neurological function throughout the lifespan, including risks for neurobehavioral regression and early-onset dementia.

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Global medical associations (ACOG, ISUOG, ACMG) recommend diagnostic prenatal testing for the detection and prevention of genetic disorders. Historically, cytogenetic methods such as karyotype analysis, fluorescent in situ hybridization (FISH) and chromosomal microarray (CMA) are utilized worldwide to diagnose common syndromes. However, the limitations of each of these methods, either performed in tandem or simultaneously, demonstrates the need of a revolutionary technology that can alleviate the need for multiple technologies. Optical genome mapping (OGM) is a novel method that fills this void by being able to detect all classes of structural variations (SVs), including copy number variations (CNVs). OGM is being adopted by laboratories as a tool for both postnatal constitutional genetic disorders and hematological malignancies. This commentary highlights the potential for OGM to become a standard of care in prenatal genetic testing based on its capability to comprehensively identify large balanced and unbalanced SVs (currently the strength of karyotyping and metaphase FISH), CNVs (by CMA), repeat contraction disorders (by Southern blotting) and multiple repeat expansion disorders (by PCR-based methods or Southern blotting). Next-generation sequencing (NGS) methods are excellent at detecting sequence variants, but they are unable to accurately resolve repeat regions of the genome, which limits their ability to detect all classes of SVs. Notably, multiple molecular methods are used to identify repeat expansion and contraction disorders in routine clinical laboratories around the world. With non-invasive prenatal testing (NIPT) becoming the standard of care screening assay for all global pregnancies, we anticipate that OGM can provide a high-resolution, cytogenomic assay to be employed following a positive NIPT screen or for high-risk pregnancies with an abnormal ultrasound. Accurate detection of all types of genetic disorders by OGM, such as liveborn aneuploidies, sex chromosome anomalies, microdeletion/microduplication syndromes, repeat expansion/contraction disorders is key to reducing the global burden of genetic disorders.