Specific mosaic KRAS mutations affecting codon 146 cause oculoectodermal syndrome and encephalocraniocutaneous lipomatosis.

Oculoectodermal syndrome (OES) and encephalocraniocutaneous lipomatosis (ECCL) are rare disorders that share many common features, such as epibulbar dermoids, aplasia cutis congenita, pigmentary changes following Blaschko lines, bony tumor-like lesions, and others. About 20 cases with OES and more than 50 patients with ECCL have been reported. Both diseases were proposed to represent mosaic disorders, but only very recently whole-genome sequencing has led to the identification of somatic KRAS mutations, p.Leu19Phe and p.Gly13Asp, in affected tissue from two individuals with OES. Here we report the results of molecular genetic studies in three patients with OES and one with ECCL. In all four cases, Sanger sequencing of the KRAS gene in DNA from lesional tissue detected mutations affecting codon 146 (p.Ala146Val, p.Ala146Thr) at variable levels of mosaicism. Our findings thus corroborate the evidence of OES being a mosaic RASopathy and confirm the common etiology of OES and ECCL. KRAS codon 146 mutations, as well as the previously reported OES-associated alterations, are known oncogenic KRAS mutations with distinct functional consequences. Considering the phenotype and genotype spectrum of mosaic RASopathies, these findings suggest that the wide phenotypic variability does not only depend on the tissue distribution but also on the specific genotype.

Skirting the pitfalls: a clear-cut nomenclature for H3K4 methyltransferases.

To unravel the system of epigenetic control of transcriptional regulation is a fascinating and important scientific pursuit. Surprisingly, recent successes in gene identification using high-throughput sequencing strategies showed that, despite their ubiquitous role in transcriptional control, dysfunction of chromatin-modifying enzymes can cause very specific human developmental phenotypes. An intriguing example is the identification of de novo dominant mutations in MLL2 as a cause of Kabuki syndrome, a well-known congenital syndrome that is associated with a very recognizable facial gestalt. However, the existing confusion in the nomenclature of the human and mouse MLL gene family impedes correct interpretation of scientific findings for these genes and their encoded proteins. This Review aims to point out this nomenclature pitfall, to explain its historical background, and to promote an unequivocal nomenclature system for chromatin-modifying enzymes as proposed by Allis et al. (2007).

Unmasking Kabuki syndrome.

The identification of de novo dominant mutations in KMT2D (MLL2) as the main cause of Kabuki syndrome (KS) has shed new light on the pathogenesis of this well-delineated condition consisting of a peculiar facial appearance, short stature, organ malformations and a varying degree of intellectual disability. Mutation screening studies have confirmed KMT2D as the major causative gene for KS and have at the same time provided evidence for its genetic heterogeneity. In this review, we aim to summarize the current clinical and molecular genetic knowledge on KS, provide genotype-phenotype correlations and propose a strategic clinical and molecular diagnostic approach for patients with suspected KS.

The R110C mutation in Notch3 causes variable clinical features in two Turkish families with CADASIL syndrome.

Mutations in Notch3 gene are responsible for the cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL). It is a late onset neurological disorder recognized by recurrent strokes and dementia. We describe here the clinical and molecular findings of three unrelated Turkish families with CADASIL syndrome. Two of the families were identified to have the same mutation, p.R110C (c.C328T), located in exon 3 of the Notch3 gene. Interestingly, the phenotypic expression of the disease in these two families was markedly different in severity and age of onset implicating additional genetic and/or non-genetic modulating factors involved in the pathogenesis. In addition, we identified the novel p.C201R (c.T601C) mutation in exon 4 of the Notch3 gene in a proband of the third family with two consecutive stroke-like episodes and typical MRI findings. Mutations described here cause an odd number of cysteines in the N-terminal of the EGF domain of Notch3 protein, which seems to have an important functional effect in the pathophysiology of CADASIL. The phenotypic variability in families carrying the same molecular defect as presented here makes the prediction of prognosis inconceivable. Although DNA analysis is effective and valuable in diagnosing approximately 90% of the CADASIL patients, lack of genotype-phenotype correlation and prognostic parameters makes the presymptomatic genetic counseling very difficult.

The novel R75Q mutation in the GJB2 gene causes autosomal dominant hearing loss and palmoplantar keratoderma in a Turkish family.

Dominant mutations in the GJB2 gene encoding connexin 26 (Cx26) can cause non-syndromic hearing impairment alone or in association with palmoplantar keratoderma (PPK). We have identified the novel G224A (R75Q) mutation in the GJB2 gene in a four-generation family from Turkey with autosomal dominant inherited hearing impairment and PPK. The age of onset and progression of hearing loss were found to be variable among affected family members, but all of them had more severe impairment at higher hearing frequencies. Interestingly, the novel R75Q mutation affects the same amino acid residue as described recently in a small family (R75W) with profound prelingual hearing loss and PPK. However, the R75W mutation was also observed in a control individual without PPK and unknown hearing status. Therefore, the nature of the R75W mutation remains ambiguous. Our molecular findings provide further evidence for the importance of the conserved R75 in Cx26 for the physiological function of the inner ear and the epidermal cells of the skin.

CADASIL syndrome in a large Turkish kindred caused by the R90C mutation in the Notch3 receptor.

Mutations in the Notch3 gene are the cause of the autosomal dominant disorder CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy). The CADASIL is an adult-onset neurologic disorder (average age of onset is 45 years) characterized by recurrent strokes and dementia. Clinical features combined with cerebral magnetic resonance imaging (MRI), showing a diffuse leukoencephalopathy with subcortical infarcts in the basal ganglia and white matter, are highly contributive to the diagnosis. We present a Turkish family with CADASIL, in which 12 individuals in four generations were affected showing the typical clinical features of recurrent strokes. Mutation analysis of the Notch3 receptor gene identified the recently described R90C mutation in the N-terminal part of the gene in affected individuals. Interestingly, migraine without aura was found as an initial symptom of the disease in two young mutation carriers (22 and 25 years, respectively), who did not show any additional clinical features or any MRI abnormalities. This indicates that migraine without aura in the absence of MRI abnormalities may represent an early initial symptom of CADASIL, which is difficult to diagnose in the absence of molecular diagnosis. Therefore, the used molecular screening method for Notch3 mutations provides a rapid and accurate diagnostic tool in addition to the standard diagnostic procedures.

Physical map of the chromosome 6q22 region containing the oculodentodigital dysplasia locus: analysis of thirteen candidate genes and identification of novel ESTs and DNA polymorphisms.

Oculodentodigital dysplasia (ODDD) is an autosomal dominant condition with congenital anomalies of the craniofacial and limb regions and neurodegeneration. Genetic anticipation for the dysmorphic and neurologic features has been inferred in a few families. Our previous linkage studies have refined the ODDD candidate region to chromosome 6q22-->q23. In an attempt to clone the ODDD gene, we created a yeast artificial chromosome contig with 31 redundant clones spanning the region and identified and ordered candidate genes and markers. Fluorescent IN SITU hybridization mapped two of these YAC clones to chromosome 6q22.2 telomeric to a known 6q21 fragile site, excluding it as a possible cause of the suggested anticipation. We performed mutation analysis on thirteen candidate genes - GRIK2, HDAC2, COL10A1, PTD013, KPNA5, PIST, ROS1, BRD7, PLN, HSF2, PKIB, FABP7, and HEY2. Although no mutations were found, we identified 44 polymorphisms, including 28 single nucleotide polymorphisms. Direct cDNA selection was performed and fifty-five clones were found to contain sequences that were not previously reported as known genes or ESTs. These clones and polymorphisms will assist in the further characterization of this region and identification of disease genes.

hKChIP2 is a functional modifier of hKv4.3 potassium channels: cloning and expression of a short hKChIP2 splice variant.

OBJECTIVE: The Ca(2+) independent transient outward K(+) current (I(to1)) in the heart is responsible for the initial phase of repolarization. The hKv4.3 K(+) channel alpha-subunit contributes to the I(to1) current in many regions of the human heart. Consistently, downregulation of hKv4.3 transcripts in heart failure and atrial fibrillation is linked to reduction in I(to1) conductance. The recently cloned KChIP family of calcium sensors has been shown to modulate A-type potassium channels of the Kv4 K(+) channel subfamily. METHODS AND RESULTS: We describe the cloning and tissue distribution of hKChIP2, as well as its functional interaction with hKv4.3 after expression in Xenopus oocytes. Furthermore, we isolated a short splice variant of the hKChIP2 gene (hKCNIP2), which represents the major hKChIP2 transcript. Northern blot analyses revealed that hKChIP2 is expressed in the human heart and occurs in the adult atria and ventricles but not in the fetal heart. Upon coexpression with hKv4.3 both hKChIP2 isoforms increased the current amplitude, slowed the inactivation and increased the recovery from inactivation of hKv4.3 currents. For the first time we analyzed the influence of a KChIP protein on the voltage of half-maximal inactivation of Kv4 channels. We demonstrate that the hKChIP2 isoforms shifted the half-maximal inactivation to more positive potentials, but to a different extent. By elucidating the genomic structure, we provide important information for future analysis of the hKCNIP2 gene in candidate disorders. In the course of this work we mapped the hKCNIP2 gene to chromosome 10q24. CONCLUSIONS: Heteromeric hKv4.3/hKChIP2 currents more closely resemble native epicardial I(to1), suggesting that hKChIP2 is a true beta-subunit of human cardiac I(to1). As a result hKChIP2 might play a role in cardiac diseases, where a contribution of I(to1) has been shown.

Campomelic dysplasia without sex reversal in a Turkish patient is due to mutation Ala119Val within the SOX9 gene.

Campomelic dysplasia is a rare neonatal skeletal malformation syndrome mainly characterized by congenital bowing and angulation of long bones in combination with other skeletal and extraskeletal defects. Two thirds of karyotypic males exhibit male-to-female sex reversal. Point mutations within SOX9 in 17q24-25 or rearrangements upstream to SOX9 as well as a deletion of a complete gene, causing haploinsufficiency of the gene product, have been detected in some patients. Recurrent mutations appear to be rare and most mutations detected in campomelic dysplasia are family specific. Here, we report on a Turkish patient with a 46,XY karyotype affected by campomelic dysplasia without sex reversal. Sequencing the SOX9 gene revealed a heterozygous Ala119Val mutation in exon 1, coding for the highly conserved HMG-box of the gene. This mutation is not present in the parents' lymphocyte DNAs. The same mutation was recently reported in a patient with 46,XX karyotype. Additionally, our patient is homozygous for the common polymorphism c507C-->T, while both parents are heterozygous.

Immediate-early gene induction by repetitive mechanical but not electrical activity in adult rat cardiomyocytes.

Mechanical factors are thought to play an important role in the induction of myocardial hypertrophy. Yet, it is not known whether active contraction induces genes that probably represent initial steps in the hypertrophic response in the adult myocardium--and if so, whether the mechanical or the electrical component of the twitch governs this response. We therefore investigated whether electrical stimulation of contraction was able to induce the immediate-early genes (IEGs) egr-1 and c-fos in adult rat cardiomyocytes. Cyclical contraction led to an increase in egr-1 and c-fos mRNA levels within 30 min. Full inhibition of contraction during electrostimulation by the Ca(2+)-desensitizer 2,3-butanedione monoxime (BDM) totally blocked this IEG-response without altering membrane potential. These data suggest that in adult myocardium, the mechanical rather than the electrical activity is responsible for the IEG-response during active twitch.

Hyperosmotic stress induces immediate-early gene expression in ventricular adult cardiomyocytes.

Mammalian cells possessing osmosensors have long been described in brain and kidney. The genetic basis of the response to hyperosmotic stress has been well characterized in prokaryotes. In contrast, the genetic response of eukaryotic cells is poorly understood. Therefore we investigated the effect of hypertonic NaCl and sucrose solutions on the transcriptional activation of the immediate-early genes (IEGs) egr-1 and c-fos in isolated ventricular adult rat cardiomyocytes. We observed that even small increases in osmolarity to 315 +/- 5 mosmol/l and 370 +/- 8 mosmol/l by hypertonic NaCl solution resulted in dose-dependent induction of egr-1 (4-and 5-fold) and c-fos (3-and 4-fold), respectively. Hypertonic sucrose solution had the same effect on egr-1 and c-fos mRNA levels while increased sucrose concentration under isotonic conditions had no effect. Cardiomyocytes exposed to hypertonic media did not significantly shrink as shown by a cell length measurement. We conclude that isolated adult cardiomyocytes possess an osmoreceptor mechanism which is able to sense even slight changes in osmolarity and to translate these into a transcriptional response of the myocardial IEG program.