Detection of 9 different point mutations of <i>PCDH19</i> in 11 female patients by direct sequencing.

<p>A) Sequence electropherograms of the mutations and the missense variant (c.3319C>G/p.Arg1107Gly) identified in association with the c.859G>T/p.Glu287X nonsense mutation. The mutation nomenclature is based on the <i>PCDH19</i> transcript reference EF676096. Nucleotides are numbered according to the cDNA with +1 corresponding to the A of the ATG translation initiation codon in the reference sequence, according to the journal guidelines (<a href="http://www.hgvs.org/mutnomen" target="_blank">www.hgvs.org/mutnomen</a>). B) Alignment of the regions surrounding the mutations (indicated by an arrow) in orthologous and paralogous proteins, showing the high conservation of each affected amino-acid in vertebrates and in the delta protocadherin paralogous genes.</p

Clinical characteristics of patients with <i>PCDH19</i> mutations.

<p>2*: patient N 06 1258 is the sister of patient N 06 1257 (index case); PMD = psychomotor development, Nl = normal, F = febrile, unit = unilateral, GTC = generalized tonic-clonic, W-S = words-sentences, abs = absent, AED = anti epileptic drugs.</p

Schematic illustration of the cellular interference mechanism associated with <i>PCDH19</i> mutations.

<p>A) In normal individuals, characterized by a homogeneous population of <i>PCDH19</i>-positive cells, neurons are able to form normal neuronal networks; B) In mutated male patients, hemizygosity leads to a homogeneous population of <i>PCDH19</i>-negative cells; in this condition, neurons preserve the ability to form normal neuronal networks; C) In heterozygous mutated females, random X inactivation leads to the co-existence of two <i>PCDH19</i>-positive and <i>PCDH19</i>-negative cell populations. These two cell populations cause divergent cell sorting and migration (due to attractive or repulsive interactions) and lead to abnormal neuronal networks. Somatic mosaicism in mutated males gives rise to the same pathological situation. The precise mechanisms by which the neuronal networks are altered are still unknown.</p

Pedigrees of the families and segregation analysis of the <i>PCDH19</i> deletion and point mutations.

<p>del/+, m/+ or v/+ denote individuals heterozygous for the deletion, mutation or variant, respectively; +/+ denotes individuals carrying homozygous wild-type alleles. Squares represent males, circles females; filled black symbols: patients diagnosed as having Dravet syndrome; right black half: Cognitive delay or impairment; left grey half: adolescence-onset idiopathic epilepsy. Dots in the middle of the squares indicate unaffected mutation carriers. The arrows indicate the index cases.</p

FISH analysis of the <i>PCDH19</i> deletion in the male patient showing somatic mosaicism in fibroblasts.

<p>(A) Absence of the specific Xq22.1 probe site on metaphase chromosomes in peripheral blood lymphocytes (PBL); (B) In fibroblasts, presence of one hybridization spot in 53% of the cells and absence of signal in the remaining 47%; C) and D) FISH analysis on PBL (C) and fibroblasts (D) of a female control. <i>PCDH19</i>-specific signals (red) are indicated by arrowheads. Magnification ×1000.</p

Identification of a deletion encompassing <i>PCDH19</i> in a male patient.

<p>A) Identification of a hemizygous Xq22.1 deletion with a 370 K SNP microarray (Illumina): Y-axes represent Log R ratio (above) and B allele frequency (below); the X-axis indicates the position on the X chromosome. The red line (log R ratio profile) corresponds to the median smoothing series (Beadstudio). B) Analysis of the patient and his mother with CGH microarrays (Nimblegen), showing that the deletion occurred de novo. Indicated genomic positions are based upon the Ensembl Genome Browser. Black horizontal bars (below) represent the gene (<i>PCDH19</i>) and pseudogenes comprised in the deleted region.</p

Additional file 5: of Uncommon nucleotide excision repair phenotypes revealed by targeted high-throughput sequencing

Clinical pictures of patients #8 (A) and #9 (B) (mutated in ERCC8(CSA)). (PPTX 403 kb

Table1_The different clinical facets of SYN1-related neurodevelopmental disorders.xlsx

Synapsin-I (SYN1) is a presynaptic phosphoprotein crucial for synaptogenesis and synaptic plasticity. Pathogenic SYN1 variants are associated with variable X-linked neurodevelopmental disorders mainly affecting males. In this study, we expand on the clinical and molecular spectrum of the SYN1-related neurodevelopmental disorders by describing 31 novel individuals harboring 22 different SYN1 variants. We analyzed newly identified as well as previously reported individuals in order to define the frequency of key features associated with these disorders. Specifically, behavioral disturbances such as autism spectrum disorder or attention deficit hyperactivity disorder are observed in 91% of the individuals, epilepsy in 82%, intellectual disability in 77%, and developmental delay in 70%. Seizure types mainly include tonic-clonic or focal seizures with impaired awareness. The presence of reflex seizures is one of the most representative clinical manifestations related to SYN1. In more than half of the cases, seizures are triggered by contact with water, but other triggers are also frequently reported, including rubbing with a towel, fever, toothbrushing, fingernail clipping, falling asleep, and watching others showering or bathing. We additionally describe hyperpnea, emotion, lighting, using a stroboscope, digestive troubles, and defecation as possible triggers in individuals with SYN1 variants. The molecular spectrum of SYN1 variants is broad and encompasses truncating variants (frameshift, nonsense, splicing and start-loss variants) as well as non-truncating variants (missense substitutions and in-frame duplications). Genotype-phenotype correlation revealed that epileptic phenotypes are enriched in individuals with truncating variants. Furthermore, we could show for the first time that individuals with early seizures onset tend to present with severe-to-profound intellectual disability, hence highlighting the existence of an association between early seizure onset and more severe impairment of cognitive functions. Altogether, we present a detailed clinical description of the largest series of individuals with SYN1 variants reported so far and provide the first genotype-phenotype correlations for this gene. A timely molecular diagnosis and genetic counseling are cardinal for appropriate patient management and treatment.</p

DataSheet1_The different clinical facets of SYN1-related neurodevelopmental disorders.PDF

Synapsin-I (SYN1) is a presynaptic phosphoprotein crucial for synaptogenesis and synaptic plasticity. Pathogenic SYN1 variants are associated with variable X-linked neurodevelopmental disorders mainly affecting males. In this study, we expand on the clinical and molecular spectrum of the SYN1-related neurodevelopmental disorders by describing 31 novel individuals harboring 22 different SYN1 variants. We analyzed newly identified as well as previously reported individuals in order to define the frequency of key features associated with these disorders. Specifically, behavioral disturbances such as autism spectrum disorder or attention deficit hyperactivity disorder are observed in 91% of the individuals, epilepsy in 82%, intellectual disability in 77%, and developmental delay in 70%. Seizure types mainly include tonic-clonic or focal seizures with impaired awareness. The presence of reflex seizures is one of the most representative clinical manifestations related to SYN1. In more than half of the cases, seizures are triggered by contact with water, but other triggers are also frequently reported, including rubbing with a towel, fever, toothbrushing, fingernail clipping, falling asleep, and watching others showering or bathing. We additionally describe hyperpnea, emotion, lighting, using a stroboscope, digestive troubles, and defecation as possible triggers in individuals with SYN1 variants. The molecular spectrum of SYN1 variants is broad and encompasses truncating variants (frameshift, nonsense, splicing and start-loss variants) as well as non-truncating variants (missense substitutions and in-frame duplications). Genotype-phenotype correlation revealed that epileptic phenotypes are enriched in individuals with truncating variants. Furthermore, we could show for the first time that individuals with early seizures onset tend to present with severe-to-profound intellectual disability, hence highlighting the existence of an association between early seizure onset and more severe impairment of cognitive functions. Altogether, we present a detailed clinical description of the largest series of individuals with SYN1 variants reported so far and provide the first genotype-phenotype correlations for this gene. A timely molecular diagnosis and genetic counseling are cardinal for appropriate patient management and treatment.</p