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Purpose: Noninvasive prenatal screening (NIPS) using cell-free DNA has been assimilated into prenatal care. Prior studies examined clinical validity and technical performance in high-risk populations. This systematic evidence review evaluates NIPS performance in a general-risk population., Methods: Medline (PubMed) and Embase were used to identify studies examining detection of Down syndrome (T21), trisomy 18 (T18), trisomy 13 (T13), sex chromosome aneuploidies, rare autosomal trisomies, copy number variants, and maternal conditions, as well as studies assessing the psychological impact of NIPS and the rate of subsequent diagnostic testing. Random-effects meta-analyses were used to calculate pooled estimates of NIPS performance (P < .05). Heterogeneity was investigated through subgroup analyses. Risk of bias was assessed., Results: A total of 87 studies met inclusion criteria. Diagnostic odds ratios were significant (P < .0001) for T21, T18, and T13 for singleton and twin pregnancies. NIPS was accurate (≥99.78%) in detecting sex chromosome aneuploidies. Performance for rare autosomal trisomies and copy number variants was variable. Use of NIPS reduced diagnostic tests by 31% to 79%. Conclusions regarding psychosocial outcomes could not be drawn owing to lack of data. Identification of maternal conditions was rare., Conclusion: NIPS is a highly accurate screening method for T21, T18, and T13 in both singleton and twin pregnancies., Competing Interests: Conflict of Interest N.C.R. is a consultant for The Jackson Laboratories and the ObG Project. E.S.B. and M.L.L. serve as directors in, and D.L. is employed by, clinical laboratories that perform a breadth of genetic and genomic analyses on a fee-for-service basis. All other authors declare no conflicts of interest., (Copyright © 2022 American College of Medical Genetics and Genomics. Published by Elsevier Inc. All rights reserved.)

Pathogenic variants in the IQSEC2 gene including nonsense, frameshift, splice-alterations, deletions, and missense changes have been identified in individuals with X-linked mental retardation. Although highly variable, clinical features may include hypotonia, moderate to severe delayed psychomotor development, intellectual disability, speech deficits, refractory seizures, autistic features, and stereotypical movements. Females with de novo variants have been described with classical features. In contrast, the phenotype in carrier females identified through an affected male may range from asymptomatic to mild intellectual disability. We present male (N = 2) and female (N = 3) probands ascertained via diagnostic exome sequencing with distinct variant types in the IQSEC2 gene encompassing a spectrum of phenotypic severity with patient sex, variant type and inheritance hypothesized to drive disease penetrance and expressivity. All of these patients demonstrated epilepsy, global developmental delays, intellectual disability, and constipation. Our data support that de novo, truncating variants correlate with severe disease in both female and male patients harboring an IQSEC2 alteration. Missense variants in male and female patients may account for a milder disease overall, with more severe symptoms in males than females. We also present the first confirmed case of parental mosaicism, which has implications regarding counseling for recurrence risk. These data further delineate a genotype-phenotype correlation of IQSEC2 variation., (Copyright © 2019 Elsevier Masson SAS. All rights reserved.)

Nonsyndromic hearing loss (NSHL), a common sensory disorder, is characterized by high clinical and genetic heterogeneity (i.e., approximately 115 genes and 170 loci so far identified). Nevertheless, almost half of patients submitted for genetic testing fail to receive a conclusive molecular diagnosis. We used next-generation sequencing to identify causal variants in PLS1 (c.805G>A, p.[E269K]; c.713G>T, p.[L238R], and c.383T>C, p.[F128S]) in three unrelated families of European ancestry with autosomal dominant NSHL. PLS1 encodes Plastin 1 (also called fimbrin), one of the most abundant actin-bundling proteins of the stereocilia. In silico protein modeling suggests that all variants destabilize the structure of the actin-binding domain 1, likely reducing the protein's ability to bind F actin. The role of PLS1 gene in hearing function is further supported by the recent demonstration that Pls1 -/ - mice show a hearing loss phenotype similar to that of our patients. In summary, we report PLS1 as a novel gene for autosomal dominant NSHL, suggesting that this gene is required for normal hearing in humans and mice., (© 2019 Wiley Periodicals, Inc.)

Wolf-Hirschhorn syndrome (WHS) is a rare but recurrent microdeletion syndrome associated with hemizygosity of an interstitial segment of Chromosome 4 (4p16.3). Consistent with historical reports in which overlapping deletions defined a minimal critical region in WHS patients, recent reports from exome sequence analysis have provided further evidence that haploinsufficiency of a specific gene within this critical region, NSD2 ( WHSC1 ), is causal for many features of the syndrome. In this report, we describe three unrelated patients with loss-of-function alterations in NSD2 who presented clinically with WHS features including intrauterine growth retardation and global developmental delay. Two of the three patients also had overlapping features of failure to thrive, short stature, constipation, and hypotonia. This series adds additional cases to expand the phenotypic spectrum of WHS and reports novel NSD2 variants., (© 2019 Barrie et al.; Published by Cold Spring Harbor Laboratory Press.)

Purpose: While chromosomal regions of homozygosity (ROH) may implicate genes in known recessive disorders, their correlation to disease pathogenicity remains unclear. ROH around the centromere of the X chromosome (pericentromeric, pROH) is regarded as benign, although this has not been empirically demonstrated., Methods: We examined microarray results from 122 female individuals harboring ROH bordering the X centromere., Results: Consecutive ROH was most frequently observed for regions Xp11.23 to Xp11.21 and Xq11.1 to Xq12, with an average total size of 16.5 Mb. X chromosome pROH was unlikely related to phenotype in 41% (50/122) of cases due to other explanations: likely pathogenic deletion/duplication (17%, 21/122), apparently unaffected female (7%, 8/122), other clinical explanation (7%, 9/122), or consanguinity (10%, 12/122). Of the remaining cases with pROH as the only finding, four genes were associated with recessive disorders that overlapped one or more clinical features reported in our probands (KDM5C, FGD1, ZC4H2, and LAS1L). X chromosome pROH observed in our cohort overlapped with previously reported regions., Conclusions: pROH on the X chromosome are commonly observed in both affected individuals with alternate causes of disease as well as in unaffected individuals, suggesting that X chromosome pROH has no clinically significant effect on phenotype., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Background: Frequent non-pathogenic genetic variants may act as moderators of phenotypic severity for complex disorders such as autism spectrum disorder (ASD). We previously identified polymorphisms affecting mRNA expression of candidate genes, including tryptophan hydroxylase 2 ( TPH2 ), dopamine beta hydroxylase ( DBH ), and dopamine transporter ( DAT, SLC6A3 )., Method: We compare genotypes and (1) clinical response to atomoxetine, (2) scores from the Autism Diagnostic Interview-Revised (ADI-R), and (3) severity of Attention Deficit Hyperactivity Disorder (ADHD) symptoms in a cohort of patients with ASD from multiple study sites., Results: There was no association between CYP2D6 metabolizer status and atomoxetine response. TPH2 rs7305115 genotype was associated with ADI-R Restrictive/Repetitive Behavior score ( p =0.03). DBH rs1611115 genotype was associated with ADI-R Social score ( p =0.002) and Restrictive/Repetitive Behavior score ( p =0.04). The DAT intron 8 5/6 repeat was associated with ADHD symptoms (ABC Hyperactivity p =0.01 and SNAP ADHD p =0.03), replicating a previous finding., Conclusions: We find associations between ASD phenotypes and regulatory variants in catecholamine biosynthesis genes. This work may help guide future genetics studies related to ASD., Competing Interests: Authors’ Conflicts of Interest Dr. Elizabeth Barrie declares that she has no conflict of interest. Ms. Emily Hansen-Kiss, LGC declares that she has no conflict of interest.

Background: Multiple variants in SNCA, encoding alpha-synuclein, a main component of Lewy bodies, are implicated in Parkinson's disease., Methods: We searched for cis-acting SNCA variants using allelic mRNA ratios in human brain tissues. In a SNCA 3'UTR (2,520 bp) luciferase reporter gene assay, translation in SH-SY5Y cells in the presence of the rs17016074 G/A alleles was measured. To assess clinical impact, we queried neurocognitive genome-wide association studies., Results: Allelic ratios deviated up to twofold, measured at a marker SNP in the middle of a long 3' untranslated region (3'UTR), but not at a marker at its start, suggesting regulation of 3'UTR processing. 3'UTR SNP rs17016074 G/A, minor allele frequency (MAF) <1% in Caucasians, 13% in Africans, strongly associates with large allelic mRNA expression imbalance (AEI), resulting in reduced expression of long 3'UTR isoforms. A second 3'UTR SNP (rs356165) associates with moderate AEI and enhances SNCA mRNA expression. The rs17016074 A allele reduces overall 3'UTR expression in luciferase reporter gene assays but supports more efficient translation, resolving previous contradictory results. We failed to detect significant genome-wide associations for rs17016074, possibly a result of low MAF in Caucasians or its absence from most genotyping panels. In the "Genome Wide Association Study of Yoruba in Nigeria," rs356165 was associated with reduced memory performance., Conclusions: Here, we identify two cis-acting regulatory variants affecting SNCA mRNA expression, measured by allelic ratios in the 3'UTR. The rs17016074 minor A allele is associated with higher expression of luciferase protein activity. Resolving the genetic influence of SNCA polymorphisms requires study of the interactions between multiple regulatory variants with distinct frequencies among populations., (© 2018 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.)

Functionally related genes often cluster into a genome region under coordinated regulation, forming a local regulome. To understand regulation of the CHRNA5/CHRNA3/CHRNB4 nicotinic receptor gene cluster, we integrate large-scale RNA expression data (brain and peripheral) from GTEx (Genotype Tissue Expression), clinical associations (GRASP), and linkage disequilibrium data (1000 Genomes) to find candidate SNPs representing independent regulatory variants. CHRNA3, CHRNA5, CHRNB4 mRNAs, and a well-expressed CHRNA5 antisense RNA (RP11-650L12.2) are co-expressed in many human tissues, suggesting common regulatory elements. The CHRNA5 enhancer haplotype tagged by rs880395 not only increases CHRNA5 mRNA expression in all tissues, but also enhances RP11-650L12.2 and CHRNA3 expression, suggesting DNA looping to multiple promoters. However, in nucleus accumbens and putamen, but not other brain regions, CHRNA3 expression associates uniquely with a haplotype tagged by rs1948 (located in the CHRNB4 3'UTR). Haplotype/diplotype analysis of rs880395 and rs1948 plus rs16969968 (a nonsynonymous CHRNA5 risk variant) in GWAS (COGEND, UW-TTURC, SAGE) yields a nicotine dependence risk profile only partially captured by rs16969968 alone. An example of local gene clusters, this nicotinic regulome is controlled by complex genetic variation, with broad implications for interpreting GWAS., (© 2016 WILEY PERIODICALS, INC.)

Dopamine β-hydroxylase (DBH) converts dopamine (DA) to norepinephrine (NE) in noradrenergic/adrenergic cells. DBH deficiency prevents NE production and causes sympathetic failure, hypotension and ptosis in humans and mice; DBH knockout (Dbh -/-) mice reveal other NE deficiency phenotypes including embryonic lethality, delayed growth, and behavioral defects. Furthermore, a single nucleotide polymorphism (SNP) in the human DBH gene promoter (-970C>T; rs1611115) is associated with variation in serum DBH activity and with several neurological- and neuropsychiatric-related disorders, although its impact on DBH expression is controversial. Phenotypes associated with DBH deficiency are typically treated with L-3,4-dihydroxyphenylserine (DOPS), which can be converted to NE by aromatic acid decarboxylase (AADC) in the absence of DBH. In this study, we generated transgenic mice carrying a human bacterial artificial chromosome (BAC) encompassing the DBH coding locus as well as ~45 kb of upstream and ~107 kb of downstream sequence to address two issues. First, we characterized the neuroanatomical, neurochemical, physiological, and behavioral transgenic rescue of DBH deficiency by crossing the BAC onto a Dbh -/- background. Second, we compared human DBH mRNA abundance between transgenic lines carrying either a "C" or a "T" at position -970. The BAC transgene drove human DBH mRNA expression in a pattern indistinguishable from the endogenous gene, restored normal catecholamine levels to the peripheral organs and brain of Dbh -/- mice, and fully rescued embryonic lethality, delayed growth, ptosis, reduced exploratory activity, and seizure susceptibility. In some cases, transgenic rescue was superior to DOPS. However, allelic variation at the rs1611115 SNP had no impact on mRNA levels in any tissue. These results indicate that the human BAC contains all of the genetic information required for tissue-specific, functional expression of DBH and can rescue all measured Dbh deficiency phenotypes, but did not reveal an impact of the rs11115 variant on DBH expression in mice.

Objective: CES1 encodes carboxylesterase-1, an important drug-metabolizing enzyme with high expression in the liver. Previous studies have reported a genomic translocation of the 5' region from the poorly expressed pseudogene CES1P1, to CES1, yielding the structural variant CES1VAR. The aim of this study was to characterize this translocation and its effect on CES1 expression in the human liver., Materials and Methods: Experiments were conducted in human liver tissues and cell culture (HepG2). The promoter and exon 1 of CES1 were sequenced by Sanger and Ion Torrent sequencing to identify gene translocations. The effects of CES1 5'UTRs on mRNA and protein expression were assessed by quantitative real-time PCR, allelic ratio mRNA analysis by primer extension (SNaPshot), quantitative targeted proteomics, and luciferase reporter gene assays., Results: Sequencing of CES1 identified two translocations: first, CES1VAR (17% minor allele frequency) comprising the 5'UTR, exon 1, and part of intron 1. A second shorter translocation, CES1SVAR, was observed excluding exon 1 and intron 1 regions (<0.01% minor allele frequency). CES1VAR is associated with 2.6-fold decreased CES1 mRNA and ∼1.35-fold lower allelic mRNA. Luciferase reporter constructs showed that CES1VAR decreases luciferase activity 1.5-fold, whereas CES1SVAR slightly increases activity. CES1VAR was not associated with CES1 protein expression or metabolism of the CES1 substrates enalapril, clopidogrel, or methylphenidate in the liver., Conclusion: The frequent translocation variant CES1VAR reduces mRNA expression of CES1 in the liver by ∼30%, but protein expression and metabolizing activity in the liver were not detectably altered - possibly because of variable CES1 expression masking small allelic effects. Whether drug therapies are affected by CES1VAR will require further in-vivo studies.

There is compelling evidence that autoreactive CD8(+)T cells play a central role in precipitating the development of autoimmune diabetes in non-obese diabetic (NOD) mice, but the underlying mechanisms remain unclear. Given that ITGAE (CD103) recognizes an islet-restricted ligand (E-cadherin), we postulated that its expression is required for initiation of disease. We herein use a mouse model of autoimmune diabetes (NOD/ShiLt mice) to test this hypothesis. We demonstrate that ITGAE is expressed by a discrete subset of CD8(+)T cells that infiltrate pancreatic islets before the development of diabetes. Moreover, we demonstrate that development of diabetes in Itgae-deficient NOD mice is significantly delayed at early but not late time points, indicating that ITGAE is preferentially involved in early diabetes development. To rule out a potential contribution by closely linked loci to this delay, we treated WT NOD mice beginning at 2 weeks of age through 5 weeks of age with a depleting anti-ITGAE mAb and found a decreased incidence of diabetes following anti-ITGAE mAb treatment compared with mice that received isotype control mAbs or non-depleting mAbs to ITGAE. Moreover, a histological examination of the pancreas of treated mice revealed that NOD mice treated with a depleting mAb were resistant to immune destruction. These results indicate that ITGAE(+) cells play a key role in the development of autoimmune diabetes and are consistent with the hypothesis that ITGAE(+)CD8(+)T effectors initiate the disease process., (© 2015 Society for Endocrinology.)

Rationale: Dopamine β-hydroxylase (DBH) catalyzes the conversion of dopamine to norepinephrine in the central nervous system and peripherally. DBH variants are associated with large changes in circulating DBH and implicated in multiple disorders; yet causal relationships and tissue-specific effects remain unresolved., Objective: To characterize regulatory variants in DBH, effect on mRNA expression, and role in modulating sympathetic tone and disease risk., Methods and Results: Analysis of DBH mRNA in human tissues confirmed high expression in the locus coeruleus and adrenal gland, but also in sympathetically innervated organs (liver>lung>heart). Allele-specific mRNA assays revealed pronounced allelic expression differences in the liver (2- to 11-fold) attributable to promoter rs1611115 and exon 2 rs1108580, but only small differences in locus coeruleus and adrenals. These alleles were also associated with significantly reduced mRNA expression in liver and lung. Although DBH protein is expressed in other sympathetically innervated organs, mRNA levels were too low for analysis. In mice, hepatic Dbh mRNA levels correlated with cardiovascular risk phenotypes. The minor alleles of rs1611115 and rs1108580 were associated with sympathetic phenotypes, including angina pectoris. Testing combined effects of these variants suggested protection against myocardial infarction in 3 separate clinical cohorts., Conclusions: We demonstrate profound effects of DBH variants on expression in 2 sympathetically innervated organs, liver and lung, but not in adrenals and brain. Preliminary results demonstrate an association of these variants with clinical phenotypes responsive to peripheral sympathetic tone. We hypothesize that in addition to endocrine effects via circulating DBH and norepinephrine, the variants act in sympathetically innervated target organs., (© 2014 American Heart Association, Inc.)

Most protein coding genes generate multiple RNA transcripts through alternative splicing, variable 3' and 5'UTRs, and RNA editing. Although drug design typically targets the main transcript, alternative transcripts can have profound physiological effects, encoding proteins with distinct functions or regulatory properties. Formation of these alternative transcripts is tissue-selective and context-dependent, creating opportunities for more effective and targeted therapies with reduced adverse effects. Moreover, genetic variation can tilt the balance of alternative versus constitutive transcripts or generate aberrant transcripts that contribute to disease risk. In addition, environmental factors and drugs modulate RNA splicing, affording new opportunities for the treatment of splicing disorders. For example, therapies targeting specific mRNA transcripts with splice-site-directed oligonucleotides that correct aberrant splicing are already in clinical trials for genetic disorders such as Duchenne muscular dystrophy. High-throughput sequencing technologies facilitate discovery of novel RNA transcripts and protein isoforms, applications ranging from neuromuscular disorders to cancer. Consideration of a gene's transcript diversity should become an integral part of drug design, development, and therapy.