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The proximal region of chromosome 15 is one of the genomic hotspots for copy number variants ( CNVs). Among the rearrangements observed in this region, CNVs from the interval between the common breakpoints 1 and 2 ( BP1 and BP2) have been reported cosegregating with autism spectrum disorder ( ASD). Although evidence supporting an association between BP1- BP2 CNVs and autism accumulates, the magnitude of the effect of BP1- BP2 CNVs remains elusive, posing a great challenge to recurrence-risk counseling. To gain further insight into their pathogenicity for ASD, we estimated the penetrance of the BP1- BP2 CNVs for ASD as well as their effects on ASD-related phenotypes in a well-characterized ASD sample (n = 2525 families). Transmission disequilibrium test revealed significant preferential transmission only for the duplicated chromosome in probands (20 T:9 NT). The penetrance of the BP1- BP2 CNVs for ASD was low, conferring additional risks of 0.3% (deletion) and 0.8% (duplication). Stepwise regression analyses suggest a greater effect of the CNVs on ASD-related phenotype in males and when maternally inherited. Taken together, the results are consistent with BP1- BP2 CNVs as risk factors for autism. However, their effect is modest, more akin to that seen for common variants. To be consistent with the current American College of Medical Genetics guidelines for interpretation of postnatal CNV, the BP1- BP2 deletion and duplication CNVs would probably best be classified as variants of uncertain significance ( VOUS): they appear to have an impact on risk, but one so modest that these CNVs do not merit pathogenic status. Autism Res 2014, 7: 355-362. © 2014 International Society for Autism Research, Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Background and aim Martin-Probst syndrome (MPS) is a rare X-linked disorder characterised by deafness, cognitive impairment, short stature and distinct craniofacial dysmorphisms, among other features. The authors sought to identify the causative mutation for MPS. Methods and results Massively parallel sequencing in two affected, related male subjects with MPS identified a RAB40AL (also called RLGP) missense mutation (chrX:102,079,078-102,079,079AC→GA p.D59G; hg18). RAB40AL encodes a small Ras-like GTPase protein with one suppressor of cytokine signalling box. The p.D59G variant is located in a highly conserved region of the GTPase domain between β-2 and β-3 strands. Using RT-PCR, the authors show that RAB40AL is expressed in human fetal and adult brain and kidney, and adult lung, heart, liver and skeletal muscle. RAB40AL appears to be a primate innovation, with no orthologues found in mouse, Xenopus or zebrafish. Western analysis and fluorescence microscopy of GFP-tagged RAB40AL constructs from transiently transfected COS7 cells show that the D59G missense change renders RAB40AL unstable and disrupts its cytoplasmic localisation. Conclusions This is the first study to show that mutation of RAB40AL is associated with a human disorder. Identification of RAB40AL as the gene mutated in MPS allows for further investigations into the molecular mechanism(s) of RAB40AL and its roles in diverse processes such as cognition, hearing and skeletal development. [ABSTRACT FROM AUTHOR]

Background: Autism spectrum disorders (ASD) are early onset neurodevelopmental syndromes typified by impairments in reciprocal social interaction and communication, accompanied by restricted and repetitive behaviors. While rare and especially de novo genetic variation are known to affect liability, whether common genetic polymorphism plays a substantial role is an open question and the relative contribution of genes and environment is contentious. It is probable that the relative contributions of rare and common variation, as well as environment, differs between ASD families having only a single affected individual (simplex) versus multiplex families who have two or more affected individuals. Methods: By using quantitative genetics techniques and the contrast of ASD subjects to controls, we estimate what portion of liability can be explained by additive genetic effects, known as narrow-sense heritability. We evaluate relatives of ASD subjects using the same methods to evaluate the assumptions of the additive model and partition families by simplex/multiplex status to determine how heritability changes with status. Results: By analyzing common variation throughout the genome, we show that common genetic polymorphism exerts substantial additive genetic effects on ASD liability and that simplex/multiplex family status has an impact on the identified composition of that risk. As a fraction of the total variation in liability, the estimated narrow-sense heritability exceeds 60% for ASD individuals from multiplex families and is approximately 40% for simplex families. By analyzing parents, unaffected siblings and alleles not transmitted from parents to their affected children, we conclude that the data for simplex ASD families follow the expectation for additive models closely. The data from multiplex families deviate somewhat from an additive model, possibly due to parental assortative mating. Conclusions: Our results, when viewed in the context of results from genome-wide association studies, demonstrate that a myriad of common variants of very small effect impacts ASD liability. [ABSTRACT FROM AUTHOR]

Holoprosencephaly (HPE) is the most common developmental forebrain anomaly in humans. Both environmental and genetic factors have been identified to play a role in the HPE phenotype. Previous studies of the genetic bases of HPE have taken a phenotype-first approach by examining groups of patients with HPE for specific mutations or deletions in known or candidate HPE genes. In this study, we characterized the presence or absence of HPE or a microform in 136 individuals in which microarray-based comparative genomic hybridization (aCGH) identified a deletion of one of 35 HPE loci. Frank holoprosencephaly was present in 11 individuals with deletions of one of the common HPE genes SHH, ZIC2, SIX3, and TGIF1, in one individual with a deletion of the HPE8 locus at 14q13, and in one individual with a deletion of FGF8, whereas deletions of other HPE loci and candidate genes ( FOXA2 and LRP2) expressed microforms of HPE. Although individuals with deletions of other HPE candidates ( DISP1, LSS, HHIP, SMO, BMP4, CDON, CDC42, ACVR2A, OTX2, and WIF1) had clinically significant features, none had frank HPE or a microform. A search for significant aCGH findings in individuals referred for testing for HPE revealed a novel association of a duplication involving GSK3B at 3q13.33 with HPE or a microform, seen in two unrelated individuals. [ABSTRACT FROM AUTHOR]

PITX2, β-catenin and lymphoid enhancer factor (LEF-1) are required for the inductive formation of several epithelial-derived organs, including teeth. Lef-1 is expressed in the dental epithelium after Pitx2, and both factors have overlapping expression patterns in the tooth bud and cap stages. Our analysis of Pitx2-/- mutant mice showed reduced Lef-1 expression in facial tissues by RT-PCR and quantitative RT-PCR. Consistent with these results we show that the human 2.5 kb LEF-1 promoter is activated by PITX2. Furthermore, the LEF-1 promoter is differentially activated by PITX2 isoforms, which are coexpressed in dental epithelium. The 2.5 kb LEF-1 promoter contains two regions that act to inhibit its transcription in concert with PITX2. The proximal region contains a Wnt-responsive element (WRE) that attenuates PITX2 activation. LEF-1 cannot autoregulate LEF-1 expression; however co-transfection of PITX2 and LEF-1 result in a synergistic activation of the 2.5 kb LEF-1 promoter. LEF-1 specifically interacts with the PITX2 C-terminal tail. Deletion of a distal 800 bp segment of the LEF-1 promoter resulted in enhanced PITX2 activation, and increased synergistic activation in the presence of LEF-1. Furthermore, β-catenin in combination with PITX2 synergistically activates the LEF-1 promoter and this activation is independent of the Wnt-responsive element. β-catenin directly interacts with PITX2 to synergistically regulate LEF-1 expression. We show a new mechanism where LEF-1 expression is regulated through PITX2, LEF-1 and β-catenin direct physical interactions. LEF-1 and β-catenin interactions with PITX2 provide new mechanisms for the regulation of PITX2 transcriptional activity. [ABSTRACT FROM AUTHOR]

Pitx2, a homeodomain transcription factor, is essential for normal development of the pituitary gland, craniofacial region, eyes, heart, abdominal viscera, and limbs. Complete loss of Pitx2 in mice (Pitx2−/−) results in embryonic lethality by approximately e15 due to cardiac defects, whereas embryos with partial loss of function (Pitx2neo/− or Pitx2neo/neo) survive until later in development (e17–e19). Pitx2 is expressed in discrete populations of postmitotic neurons in the mouse brain, but its role in mammalian central nervous system (CNS) development is not known. We undertook an analysis of Pitx2-deficient embryos to determine whether loss of Pitx2 affects CNS development. The CNS is normal in hypomorphic e16.5 Pitx2neo/− and e18.5 Pitx2neo/neo embryos, with no evidence of midline or other defects. Midgestation (e10.5) Pitx2−/− embryos have normally formed neural tube structures and cerebral vesicles, whereas older (e14.5) Pitx2−/− embryos exhibit loss of gene expression and axonal projections in the subthalamic nucleus (a group of cells in the ventrolateral thalamus) and in the developing superior colliculus of dorsal midbrain. Our results suggest a role for Pitx2 in regulating regionally specific terminal neuronal differentiation in the developing ventrolateral thalamus and midbrain. [Copyright &y& Elsevier]

Pitx2, a homeodomain transcription factor, is essential for normal development of pituitary, eyes, heart, and teeth. In the developing mouse brain, Pitx2 (Rieg, Ptx2, Otlx2, Brx1) mRNA is expressed in discrete regions of the diencephalon, mesencephalon, and rhombencephalon. While prior reports have provided an overview of the temporal and regional specificity of Pitx2 mRNA expression in the brain, the precise cell types that express PITX2 are not known. In this study, we analyzed Pitx2 mRNA and PITX2 protein expression in individual cells of the developing e10.5–e14.5 mouse CNS using multiple markers of cellular proliferation and differentiation. We identified Pitx2 expression in nestin-positive neural progenitors and in postmitotic, developing neurons. In the diencephalon, PITX2 is expressed in neurons of the zona limitans intrathalamica and mammillary region and in γ-aminobutyric acid (GABA)-producing neurons of the zona incerta. In the mesencephalon, PITX2-labeled nuclei also appear in differentiated neurons, some of which are GABAergic and destined to occupy superior colliculus. Our results suggest that PITX2 expression in postmitotic neurons may contribute to development of GABAergic and other differentiated neuronal phenotypes. [Copyright &y& Elsevier]

CHARGE syndrome, a rare multiple congenital anomaly condition, is caused by haploinsufficiency of the chromatin remodeling protein gene CHD7 (Chromodomain helicase DNA binding protein 7). Brain abnormalities and intellectual disability are commonly observed in individuals with CHARGE, and neuronal differentiation is reduced in CHARGE patient-derived iPSCs and conditional knockout mouse brains. However, the mechanisms of CHD7 function in nervous system development are not well understood. In this study, we asked whether CHD7 promotes gene transcription in neural progenitor cells via changes in chromatin accessibility. We used Chd7 null embryonic stem cells (ESCs) derived from Chd7 mutant mouse blastocysts as a tool to investigate roles of CHD7 in neuronal and glial differentiation. Loss of Chd7 significantly reduced neuronal and glial differentiation. Sholl analysis showed that loss of Chd7 impaired neuronal complexity and neurite length in differentiated neurons. Genome-wide studies demonstrated that loss of Chd7 leads to modified chromatin accessibility (ATAC-seq) and differential nascent expression (Bru-Seq) of neural-specific genes. These results suggest that CHD7 acts preferentially to alter chromatin accessibility of key genes during the transition of NPCs to neurons to promote differentiation. Our results form a basis for understanding the cell stage-specific roles for CHD7-mediated chromatin remodeling during cell lineage acquisition. [ABSTRACT FROM AUTHOR]

Inner ear morphogenesis requires tightly regulated epigenetic and transcriptional control of gene expression. CHD7, an ATP-dependent chromodomain helicase DNA-binding protein, and SOX2, an SRY-related HMG box pioneer transcription factor, are known to contribute to vestibular and auditory system development, but their genetic interactions in the ear have not been explored. Here, we analyzed inner ear development and the transcriptional regulatory landscapes in mice with variable dosages of Chd7 and/or Sox2. We show that combined haploinsufficiency for Chd7 and Sox2 results in reduced otic cell proliferation, severe malformations of semicircular canals, and shortened cochleae with ectopic hair cells. Examination of mice with conditional, inducible Chd7 loss by Sox2CreER reveals a critical period (~E9.5) of susceptibility in the inner ear to combined Chd7 and Sox2 loss. Data from genome-wide RNA-sequencing and CUT&Tag studies in the otocyst show that CHD7 regulates Sox2 expression and acts early in a gene regulatory network to control expression of key otic patterning genes, including Pax2 and Otx2. CHD7 and SOX2 directly bind independently and cooperatively at transcription start sites and enhancers to regulate otic progenitor cell gene expression. Together, our findings reveal essential roles for Chd7 and Sox2 in early inner ear development and may be applicable for syndromic and other forms of hearing or balance disorders. [ABSTRACT FROM AUTHOR]

Enlargement of the endolymphatic sac, duct, and vestibular aqueduct (EVA) is the most common inner ear malformation identified in patients with sensorineural hearing loss. EVA is associated with pathogenic variants in SLC26A4. However, in European–Caucasian populations, about 50% of patients with EVA carry no pathogenic alleles of SLC26A4. We tested for the presence of variants in CHD7, a gene known to be associated with CHARGE syndrome, Kallmann syndrome, and hypogonadotropic hypogonadism, in a cohort of 34 families with EVA subjects without pathogenic alleles of SLC26A4. In two families, NM_017780.4: c.3553A > G [p.(Met1185Val)] and c.5390G > C [p.(Gly1797Ala)] were detected as monoallelic CHD7 variants in patients with EVA. At least one subject from each family had additional signs or potential signs of CHARGE syndrome but did not meet diagnostic criteria for CHARGE. In silico modeling of these two missense substitutions predicted detrimental effects upon CHD7 protein structure. Consistent with a role of CHD7 in this tissue, Chd7 transcript and protein were detected in all epithelial cells of the endolymphatic duct and sac of the developing mouse inner ear. These results suggest that some CHD7 variants can cause nonsyndromic hearing loss and EVA. CHD7 should be included in DNA sequence analyses to detect pathogenic variants in EVA patients. Chd7 expression and mutant phenotype data in mice suggest that CHD7 contributes to the formation or function of the endolymphatic sac and duct. [ABSTRACT FROM AUTHOR]

A correction to the article "The 20th International Mammalian Genome Conference Meeting Report" that was published in the August 2007 issue is presented.

• Development of the mammalian inner ear is orchestrated by epigenetic factors. • DNA methylation, histone modifications, and chromatin remodeling all influence various aspects of inner ear development. • Emerging genomic technologies reveal epigenetic mechanisms of inner ear development at single cell resolution. Epigenetic factors are critically important for embryonic and postnatal development. Over the past decade, substantial technological advancements have occurred that now permit the study of epigenetic mechanisms that govern all aspects of inner ear development, from otocyst patterning to maturation and maintenance of hair cell stereocilia. In this review, we highlight how three major classes of epigenetic regulation (DNA methylation, histone modification, and chromatin remodeling) are essential for the development of the inner ear. We highlight open avenues for research and discuss how new tools enable the employment of epigenetic factors in regenerative and therapeutic approaches for hearing and balance disorders. [ABSTRACT FROM AUTHOR]

Failure of the amygdala to habituate, or decrease response intensity, to repeatedly presented faces may be one mechanism by which individuals with autism spectrum disorders (ASD) develop and maintain social symptoms. However, genetic influences on habituation in ASD have not been examined. We hypothesized that serotonin transporter-linked promoter region (5-HTTLPR) genotype affects change in amygdala response to repeated sad faces differently in individuals with ASD vs healthy controls. Forty-four youth with ASD and 65 controls aged 8–19 years were genotyped and underwent an event-related functional magnetic resonance imaging scan where they identified the gender of emotional faces presented for 250 ms. The first half of the run was compared with the second half to assess habituation. 5-HTTLPR genotype influences amygdala habituation to sad faces differently for individuals with ASD vs controls. The genotype-by-diagnosis-by-run half interaction was driven by individuals with ASD and low expressing genotypes (S/S, S/LG and LG/LG), who trended toward sensitization (increase in amygdala activation) and whose habituation scores significantly differed from individuals with ASD and higher expressing genotypes (LA/LA, S/LA and LA/LG) as well as controls with low expressing genotypes. Our results show that amygdala response to social stimuli in ASD, which may contribute to social symptoms, is genetically influenced. [ABSTRACT FROM PUBLISHER]

Epigenetic regulation of gene transcription by chromatin remodeling proteins has recently emerged as an important contributing factor in inner ear development. Pathogenic variants in CHD7 , the gene encoding Chromodomain Helicase DNA binding protein 7, cause CHARGE syndrome, which presents with malformations in the developing ear. Chd7 is broadly expressed in the developing mouse otocyst and mature auditory epithelium, yet the pathogenic effects of Chd7 loss in the cochlea are not well understood. Here we characterized cochlear epithelial phenotypes in mice with deletion of Chd7 throughout the otocyst (using Foxg1 Cre/+ and Pax2Cre), in the otic mesenchyme (using TCre), in hair cells (using Atoh1Cre), in developing neuroblasts (using NgnCre), or in spiral ganglion neurons (using Shh Cre/+ ). Pan-otic deletion of Chd7 resulted in shortened cochleae with aberrant projections and axonal looping, disorganized, supernumerary hair cells at the apical turn and a narrowed epithelium with missing hair cells in the middle region. Deletion of Chd7 in the otic mesenchyme had no effect on overall cochlear morphology. Loss of Chd7 in hair cells did not disrupt their formation or organization of the auditory epithelium. Similarly, absence of Chd7 in spiral ganglion neurons had no effect on axonal projections. In contrast, deletion of Chd7 in developing neuroblasts led to smaller spiral ganglia and disorganized cochlear neurites. Together, these observations reveal dosage-, tissue-, and time-sensitive cell autonomous roles for Chd7 in cochlear elongation and cochlear neuron organization, with minimal functions for Chd7 in hair cells. These studies provide novel information about roles for Chd7 in development of auditory neurons. [Display omitted] • Chd7 is required in the mouse otocyst for cochlear elongation. • Chd7 is required in the mouse otocyst for hair cell patterning. • Chd7 is not required in peri-otic mesenchyme for inner ear morhphogenesis • Spiral ganglion neurons and cochlear hair cells are sensitive to Chd7 dosage. [ABSTRACT FROM AUTHOR]

Oligodendrocyte precursor cells (OPCs) constitute the main proliferative cells in the adult brain, and deregulation of OPC proliferation-differentiation balance results in either glioma formation or defective adaptive (re)myelination. OPC differentiation requires significant genetic reprogramming, implicating chromatin remodeling. Mounting evidence indicates that chromatin remodelers play important roles during normal development and their mutations are associated with neurodevelopmental defects, with CHD7 haploinsuficiency being the cause of CHARGE syndrome and CHD8 being one of the strongest autism spectrum disorder (ASD) high-risk-associated genes. Herein, we report on uncharacterized functions of the chromatin remodelers Chd7 and Chd8 in OPCs. Their OPC-chromatin binding profile, combined with transcriptome and chromatin accessibility analyses of Chd7-deleted OPCs, demonstrates that Chd7 protects nonproliferative OPCs from apoptosis by chromatin closing and transcriptional repression of p53. Furthermore, Chd7 controls OPC differentiation through chromatin opening and transcriptional activation of key regulators, including Sox10, Nkx2.2, and Gpr17. However, Chd7 is dispensable for oligodendrocyte stage progression, consistent with Chd8 compensatory function, as suggested by their common chromatin-binding profiles and genetic interaction. Finally, CHD7 and CHD8 bind in OPCs to a majority of ASD risk-associated genes, suggesting an implication of oligodendrocyte lineage cells in ASD neurological defects. Our results thus offer new avenues to understand and modulate the CHD7 and CHD8 functions in normal development and disease. [ABSTRACT FROM AUTHOR]

CHARGE syndrome--which stands for coloboma of the eye, heart defects, atresia of choanae, retardation of growth/development, genital abnormalities, and ear anomalies--is a severe developmental disorder with wide phenotypic variability, caused mainly by mutations in CHD7 (chromodomain helicase DNA-binding protein 7), known to encode a chromatin remodeler. The genetic lesions responsible for CHD7 mutation-negative cases are unknown, at least in part because the pathogenic mechanisms underlying CHARGE syndrome remain poorly defined. Here, we report the characterization of a mouse model for CHD7 mutation-negative cases of CHARGE syndrome generated by insertional mutagenesis of Fam172a (family with sequence similarity 172, member A). We show that Fam172a plays a key role in the regulation of cotranscriptional alternative splicing, notably by interacting with Ago2 (Argonaute-2) and Chd7. Validation studies in a human cohort allow us to propose that dysregulation of cotranscriptional alternative splicing is a unifying pathogenic mechanism for both CHD7 mutation-positive and CHD7 mutation-negative cases. We also present evidence that such splicing defects can be corrected in vitro by acute rapamycin treatment. [ABSTRACT FROM AUTHOR]

Mutations in the chromatin remodeling factor CHD7 are the predominant cause of CHARGE syndrome, a congenital disorder that frequently includes ocular coloboma. Although CHD7 is known to be required for proper ocular morphogenesis, its role in retinal development has not been thoroughly investigated. Given that individuals with CHARGE syndrome can experience visual impairment even in the absence of coloboma, a better understanding of CHD7 function in the retina is needed. In this study, we characterized the expression pattern of Chd7 in the developing zebrafish and mouse retina and documented ocular and retinal phenotypes in Chd7 loss-of-function mutants. Zebrafish Chd7 was expressed throughout the retinal neuroepithelium when retinal progenitor cells were actively proliferating, and later in subsets of newly post-mitotic retinal cells. At stages of retinal development when most retinal cell types had terminally differentiated, Chd7 expression remained strong in the ganglion cell layer and in some cells in the inner nuclear layer. Intriguingly, strong expression of Chd7 was also observed in the outer nuclear layer where it was co-expressed with markers of post-mitotic cone and rod photoreceptors. Expression of mouse CHD7 displayed a similar pattern, including expression in the ganglion cells, subsets of inner nuclear layer cells, and in the distal outer nuclear layer as late as P15. Two different mutant chd7 zebrafish lines were characterized for ocular and retinal defects. These mutants displayed microphthalmia, reduced numbers of cone photoreceptors, and truncated rod and cone photoreceptor outer segments. Reduced cone photoreceptor number and abnormal outer segments were also observed in heterozygous Chd7 mutant mice. Taken together, our results in zebrafish and mouse reveal a conserved, previously undescribed role for Chd7 in retinal development and photoreceptor outer segment morphogenesis. Moreover, our work suggests an avenue of future investigation into the pathogenesis of visual system defects in CHARGE syndrome. • Chd7 is expressed in both retinal progenitor cells and in differentiated retinal neurons, including post-mitotic rod and cone photoreceptors. • Loss of Chd7 results in a significant decrease in cone photoreceptors in both zebrafish and mouse. • Photoreceptor outer segments are truncated in chd7 mutants, suggesting a heretofore unappreciated role for Chd7 in outer segment morphogenesis. [ABSTRACT FROM AUTHOR]

• Chd7Gt/+ mice exhibit reduced satellite glia in adult spiral ganglia. • Hypermyelination of Chd7Gt/+ peripheral spiral ganglion neuronal projections. • Chd7 is transiently expressed in glial precursor cells in the developing inner ear. • This study provides the first evidence of CHD7 in peripheral glial cell development. CHARGE syndrome is a multiple anomaly developmental disorder characterized by a variety of sensory deficits, including sensorineural hearing loss of unknown etiology. Most cases of CHARGE are caused by heterozygous pathogenic variants in CHD7 , the gene encoding Chromodomain DNA-binding Protein 7 (CHD7), a chromatin remodeler important for the development of neurons and glial cells. Previous studies in the Chd7Gt/+ mouse model of CHARGE syndrome showed substantial neuron loss in the early stages of the developing inner ear that are compensated for by mid-gestation. In this study, we sought to determine if early developmental delays caused by Chd7 haploinsufficiency affect neurons, glial cells, and inner hair cell innervation in the mature cochlea. Analysis of auditory brainstem response recordings in Chd7Gt/+ adult animals showed elevated thresholds at 4 kHz and 16 kHz, but no differences in ABR Wave I peak latency or amplitude compared to wild type controls. Proportions of neurons in the Chd7Gt/+ adult spiral ganglion and densities of nerve projections from the spiral ganglion to the organ of Corti were not significantly different from wild type controls. Inner hair cell synapse formation also appeared unaffected in mature Chd7Gt/+ cochleae. However, histological analysis of adult Chd7Gt/+ cochleae revealed diminished satellite glial cells and hypermyelinated Type I spiral ganglion axons. We characterized the expression of CHD7 in developing inner ear glia and found CHD7 to be expressed during a tight window of inner ear development at the Schwann cell precursor stage at E9.5. While cochlear neurons appear to differentiate normally in the setting of Chd7 haploinsufficiency, our results suggest an important role for CHD7 in glial cells in the inner ear. This study highlights the dynamic nature of CHD7 activity during inner ear development in mice and contributes to understanding CHARGE syndrome pathology. [ABSTRACT FROM AUTHOR]

Background Phenotypic heterogeneity in autism has long been conjectured to be a major hindrance to the discovery of genetic risk factors, leading to numerous attempts to stratify children based on phenotype to increase power of discovery studies. This approach, however, is based on the hypothesis that phenotypic heterogeneity closely maps to genetic variation, which has not been tested. Our study examines the impact of subphenotyping of a well-characterized autism spectrum disorder (ASD) sample on genetic homogeneity and the ability to discover common genetic variants conferring liability to ASD. Methods Genome-wide genotypic data of 2576 families from the Simons Simplex Collection were analyzed in the overall sample and phenotypic subgroups defined on the basis of diagnosis, IQ, and symptom profiles. We conducted a family-based association study, as well as estimating heritability and evaluating allele scores for each phenotypic subgroup. Results Association analyses revealed no genome-wide significant association signal. Subphenotyping did not increase power substantially. Moreover, allele scores built from the most associated single nucleotide polymorphisms, based on the odds ratio in the full sample, predicted case status in subsets of the sample equally well and heritability estimates were very similar for all subgroups. Conclusions In genome-wide association analysis of the Simons Simplex Collection sample, reducing phenotypic heterogeneity had at most a modest impact on genetic homogeneity. Our results are based on a relatively small sample, one with greater homogeneity than the entire population; if they apply more broadly, they imply that analysis of subphenotypes is not a productive path forward for discovering genetic risk variants in ASD. [ABSTRACT FROM AUTHOR]

CHARGE syndrome is a multiple anomaly disorder in which patients present with a variety of phenotypes, including ocular coloboma, heart defects, choanal atresia, retarded growth and development, genitourinary hypoplasia and ear abnormalities. Despite 70-90% of CHARGE syndrome cases resulting from mutations in the gene CHD7, which encodes an ATP-dependent chromatin remodeller, the pathways underlying the diverse phenotypes remain poorly understood. Surprisingly, our studies of a knock-in mutant mouse strain that expresses a stabilized and transcriptionally dead variant of the tumour-suppressor protein p53 (p5325,26,53,54), along with a wild-type allele of p53 (also known as Trp53), revealed late-gestational embryonic lethality associated with a host of phenotypes that are characteristic of CHARGE syndrome, including coloboma, inner and outer ear malformations, heart outflow tract defects and craniofacial defects. We found that the p5325,26,53,54 mutant protein stabilized and hyperactivated wild-type p53, which then inappropriately induced its target genes and triggered cell-cycle arrest or apoptosis during development. Importantly, these phenotypes were only observed with a wild-type p53 allele, as p5325,26,53,54/− embryos were fully viable. Furthermore, we found that CHD7 can bind to the p53 promoter, thereby negatively regulating p53 expression, and that CHD7 loss in mouse neural crest cells or samples from patients with CHARGE syndrome results in p53 activation. Strikingly, we found that p53 heterozygosity partially rescued the phenotypes in Chd7-null mouse embryos, demonstrating that p53 contributes to the phenotypes that result from CHD7 loss. Thus, inappropriate p53 activation during development can promote CHARGE phenotypes, supporting the idea that p53 has a critical role in developmental syndromes and providing important insight into the mechanisms underlying CHARGE syndrome. [ABSTRACT FROM AUTHOR]

The S and LG alleles of the serotonin transporter-linked polymorphic region (5-HTTLPR) lower serotonin transporter expression. These low-expressing alleles are linked to increased risk for depression and brain activation patterns found in depression (increased amygdala activation and decreased amygdala-prefrontal cortex connectivity). Paradoxically, serotonin transporter blockade relieves depression symptoms. Rodent models suggest that decreased serotonin transporter in early life produces depression that emerges in adolescence, whereas decreased serotonin transporter that occurs later in development ameliorates depression. However, no brain imaging research has yet investigated the moderating influence of human development on the link between 5-HTTLPR and effect-related brain function. We investigated the age-related effect of 5-HTTLPR on amygdala activation and amygdala-prefrontal cortex connectivity using a well-replicated probe, an emotional face task, in children and adolescents aged 9-19 years. A significant genotype-by-age interaction predicted amygdala activation, such that the low-expressing genotype (S/S and S/LG) group showed a greater increase in amygdala activation with age compared to the higher expressing (LA/LA and S/LA) group. Additionally, compared to the higher expressing group, the low-expressing genotype group exhibited decreased connectivity between the right amygdala and ventromedial prefrontal cortex with age. Findings indicate that low-expressing genotypes may not result in the corticolimbic profile associated with depression risk until later adolescence. Hum Brain Mapp 35:646-658, 2014. © 2012 Wiley-Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Background: Brain development follows a different trajectory in children with autism spectrum disorders (ASD) than in typically developing children. A proxy for neurodevelopment could be head circumference (HC), but studies assessing HC and its clinical correlates in ASD have been inconsistent. This study investigates HC and clinical correlates in the Simons Simplex Collection cohort. Methods: We used a mixed linear model to estimate effects of covariates and the deviation from the expected HC given parental HC (genetic deviation). After excluding individuals with incomplete data, 7225 individuals in 1891 families remained for analysis. We examined the relationship between HC/genetic deviation of HC and clinical parameters. Results: Gender, age, height, weight, genetic ancestry, and ASD status were significant predictors of HC (estimate of the ASD effect = .2 cm). HC was approximately normally distributed in probands and unaffected relatives, with only a few outliers. Genetic deviation of HC was also normally distributed, consistent with a random sampling of parental genes. Whereas larger HC than expected was associated with ASD symptom severity and regression, IQ decreased with the absolute value of the genetic deviation of HC. Conclusions: Measured against expected values derived from covariates of ASD subjects, statistical outliers for HC were uncommon. HC is a strongly heritable trait, and population norms for HC would be far more accurate if covariates including genetic ancestry, height, and age were taken into account. The association of diminishing IQ with absolute deviation from predicted HC values suggests HC could reflect subtle underlying brain development and warrants further investigation. [Copyright &y& Elsevier]

Abstract: Transcriptional regulation of gene expression during development is critical for proper neuronal differentiation and migration. Alternative splicing and differential isoform expression have been demonstrated for most mammalian genes, but their specific contributions to gene function are not well understood. In mice, the transcription factor gene Pitx2 is expressed as three different isoforms (PITX2A, PITX2B, and PITX2C) which have unique amino termini and common DNA binding homeodomains and carboxyl termini. The specific roles of these isoforms in neuronal development are not known. Here we report the onset of Pitx2ab and Pitx2c isoform-specific expression by E9.5 in the developing mouse brain. Using isoform-specific Pitx2 deletion mouse strains, we show that collicular neuron migration requires PITX2AB and that collicular GABAergic differentiation and targeting of hypothalamic projections require unique Pitx2 isoform dosage. These results provide insights into Pitx2 dosage and isoform-specific requirements underlying midbrain and hypothalamic development. [Copyright &y& Elsevier]

Background: Some copy-number variants are associated with genomic disorders with extreme phenotypic heterogeneity. The cause of this variation is unknown, which presents challenges in genetic diagnosis, counseling, and management. Methods: We analyzed the genomes of 2312 children known to carry a copy-number variant associated with intellectual disability and congenital abnormalities, using array comparative genomic hybridization. Results: Among the affected children, 10.1% carried a second large copy-number variant in addition to the primary genetic lesion. We identified seven genomic disorders, each defined by a specific copy-number variant, in which the affected children were more likely to carry multiple copy-number variants than were controls. We found that syndromic disorders could be distinguished from those with extreme phenotypic heterogeneity on the basis of the total number of copy-number variants and whether the variants are inherited or de novo. Children who carried two large copy-number variants of unknown clinical significance were eight times as likely to have developmental delay as were controls (odds ratio, 8.16; 95% confidence interval, 5.33 to 13.07; P=2.11×10−38). Among affected children, inherited copy-number variants tended to co-occur with a second-site large copy-number variant (Spearman correlation coefficient, 0.66; P<0.001). Boys were more likely than girls to have disorders of phenotypic heterogeneity (P<0.001), and mothers were more likely than fathers to transmit second-site copy-number variants to their offspring (P=0.02). Conclusions: Multiple, large copy-number variants, including those of unknown pathogenic significance, compound to result in a severe clinical presentation, and secondary copy-number variants are preferentially transmitted from maternal carriers. (Funded by the Simons Foundation Autism Research Initiative and the National Institutes of Health.) [ABSTRACT FROM PUBLISHER]

Abstract: A fundamental component of brain development is the formation of large-scale networks across the cortex. One such network, the default network, undergoes a protracted development, displaying weak connectivity in childhood that strengthens in adolescence and becomes most robust in adulthood. Little is known about the genetic contributions to default network connectivity in adulthood or during development. Alterations in connectivity between posterior and frontal portions of the default network have been associated with several psychological disorders, including anxiety, autism spectrum disorders, schizophrenia, depression, and attention-deficit/hyperactivity disorder. These disorders have also been linked to variants of the serotonin transporter linked polymorphic region (5-HTTLPR). The LA allele of 5-HTTLPR results in higher serotonin transporter expression than the S allele or the rarer LG allele. 5-HTTLPR may influence default network connectivity, as the superior medial frontal region has been shown to be sensitive to changes in serotonin. Also, serotonin as a growth factor early in development may alter large-scale networks such as the default network. The present study examined the influence of 5-HTTLPR variants on connectivity between the posterior and frontal structures and its development in a cross-sectional study of 39 healthy children and adolescents. We found that children and adolescents homozygous for the S allele (S/S, n=10) showed weaker connectivity in the superior medial frontal cortex compared to those homozygous for the LA allele (LA/LA, n=13) or heterozygotes (S/LA, S/LG, n=16). Moreover, there was an age-by-genotype interaction, such that those with LA/LA genotype had the steepest age-related increase in connectivity between the posterior hub and superior medial frontal cortex, followed by heterozygotes. In contrast, individuals with the S/S genotype had the least age-related increase in connectivity strength. This preliminary report expands our understanding of the genetic influences on the development of large-scale brain connectivity and lays down the foundation for future research and replication of the results with a larger sample. [Copyright &y& Elsevier]

Although fixation of the stapes is usually progressive and secondary to otosclerosis, it may present congenitally, with other skeletal manifestations, as an autosomal dominant syndrome—such as proximal symphalangism (SYM1) or multiple-synostoses syndrome (SYNS1), both of which are caused by mutations in NOG, the gene encoding noggin. We describe a family that was ascertained to have nonsyndromic otosclerosis but was subsequently found to have a congenital stapes ankylosis syndrome that included hyperopia, a hemicylindrical nose, broad thumbs and great toes, and other minor skeletal anomalies but lacked symphalangism. A heterozygous nonsense NOG mutation—c.328C→T (Q110X), predicted to truncate the latter half of the protein—was identified, and a heterozygous insertion in NOG—c.252-253insC, in which the frameshift is predicted to result in 96 novel amino acids before premature truncation—was identified in a previously described second family with a similar phenotype. In contrast to most NOG mutations that have been reported in kindreds with SYM1 and SYNS1, the mutations observed in these families with stapes ankylosis without symphalangism are predicted to disrupt the cysteine-rich C-terminal domain. These clinical and molecular findings suggest that (1) a broader range of conductive hearing-loss phenotypes are associated with NOG mutations than had previously been recognized, (2) patients with sporadic or familial nonsyndromic otosclerosis should be evaluated for mild features of this syndrome, and (3) NOG alterations should be considered in conductive hearing loss with subtle clinical and skeletal features, even in the absence of symphalangism. [ABSTRACT FROM AUTHOR]