Genotype differences in cognitive functioning in Noonan syndrome.

Noonan syndrome (NS) is an autosomal-dominant genetic disorder associated with highly variable features, including heart disease, short stature, minor facial anomalies and learning disabilities. Recent gene discoveries have laid the groundwork for exploring whether variability in the NS phenotype is related to differences at the genetic level. In this study, we examine the influence of both genotype and nongenotypic factors on cognitive functioning. Data are presented from 65 individuals with NS (ages 4-18) who were evaluated using standardized measures of intellectual functioning. The cohort included 33 individuals with PTPN11 mutations, 6 individuals with SOS1 mutations, 1 individual with a BRAF mutation and 25 participants with negative, incomplete or no genetic testing. Results indicate that genotype differences may account for some of the variation in cognitive ability in NS. Whereas cognitive impairments were common among individuals with PTPN11 mutations and those with unknown mutations, all of the individuals with SOS1 mutations exhibited verbal and nonverbal cognitive skills in the average range or higher. Participants with N308D and N308S mutations in PTPN11 also showed no (or mild) cognitive delays. Additional influences such as hearing loss, motor dexterity and parental education levels accounted for significant variability in cognitive outcomes. Severity of cardiac disease was not related to cognitive functioning. Our results suggest that some NS-causing mutations have a more marked impact on cognitive skills than others.

An absence of cutaneous neurofibromas associated with a 3-bp inframe deletion in exon 17 of the NF1 gene (c.2970-2972 delAAT): evidence of a clinically significant NF1 genotype-phenotype correlation.

Neurofibromatosis type 1 (NF1) is characterized by cafe-au-lait spots, skinfold freckling, and cutaneous neurofibromas. No obvious relationships between small mutations (<20 bp) of the NF1 gene and a specific phenotype have previously been demonstrated, which suggests that interaction with either unlinked modifying genes and/or the normal NF1 allele may be involved in the development of the particular clinical features associated with NF1. We identified 21 unrelated probands with NF1 (14 familial and 7 sporadic cases) who were all found to have the same c.2970-2972 delAAT (p.990delM) mutation but no cutaneous neurofibromas or clinically obvious plexiform neurofibromas. Molecular analysis identified the same 3-bp inframe deletion (c.2970-2972 delAAT) in exon 17 of the NF1 gene in all affected subjects. The Delta AAT mutation is predicted to result in the loss of one of two adjacent methionines (codon 991 or 992) ( Delta Met991), in conjunction with silent ACA-->ACG change of codon 990. These two methionine residues are located in a highly conserved region of neurofibromin and are expected, therefore, to have a functional role in the protein. Our data represent results from the first study to correlate a specific small mutation of the NF1 gene to the expression of a particular clinical phenotype. The biological mechanism that relates this specific mutation to the suppression of cutaneous neurofibroma development is unknown.

Novel TFAP2B mutations that cause Char syndrome provide a genotype-phenotype correlation.

To elucidate further the role, in normal development and in disease pathogenesis, of TFAP2B, a transcription factor expressed in neuroectoderm, we studied eight patients with Char syndrome and their families. Four novel mutations were identified, three residing in the basic domain, which is responsible for DNA binding, and a fourth affecting a conserved PY motif in the transactivation domain. Functional analyses of the four mutants disclosed that two, R225C and R225S, failed to bind target sequence in vitro and that all four had dominant negative effects when expressed in eukaryotic cells. Our present findings, combined with data about two previously identified TFAP2B mutations, show that dominant negative effects consistently appear to be involved in the etiology of Char syndrome. Affected individuals in the family with the PY motif mutation, P62R, had a high prevalence of patent ductus arteriosus but had only mild abnormalities of facial features and no apparent hand anomalies, a phenotype different from that associated with the five basic domain mutations. This genotype-phenotype correlation supports the existence of TFAP2 coactivators that have tissue specificity and are important for ductal development but less critical for craniofacial and limb development.

A summary of 20 CACNA1F mutations identified in 36 families with incomplete X-linked congenital stationary night blindness, and characterization of splice variants.

Incomplete X-linked congenital stationary night blindness (CSNB) is a recessive, non-progressive eye disorder characterized by abnormal electroretinogram and psychophysical testing and can include impaired night vision, decreased visual acuity, myopia, nystagmus, and strabismus. Including the 20 families previously reported (Bech-Hansen et al. 1998b), we have now analyzed patients from a total of 36 families with incomplete CSNB and identified 20 different mutations in the calcium channel gene CACNA1F. Three of the mutations account for incomplete CSNB in two or more families, and a founder effect is clearly demonstrable for one of these mutations. Of the 20 mutations identified, 14 (70%) are predicted to cause premature protein truncation and six (30%) to cause amino acid substitutions or deletions at conserved positions in the alpha1F protein. In characterizing transcripts of CACNA1F we have identified several splice variants and defined a prototypical sequence based on the location of mutations in splice variants and comparison with the mouse orthologue, Cacnalf.

Evaluation of urinary carnitine and taurine excretion in 5 cystinuric dogs with carnitine and taurine deficiency.

Five client owned dogs with cystinuria were diagnosed with carnitine and taurine deficiency while participating in a clinical trial that used dietary management of their urolithiasis. Stored 24-hour urine samples collected from the cystinuric dogs before enrollment in the clinical diet trial were quantitatively evaluated for carnitine and taurine. These results were compared to those obtained from 18 healthy Beagles. Both groups of dogs were fed the same maintenance diet for a minimum of 2 weeks before 24-hour urine collection. The protocol used for 24-hour urine collections was the same for cystinuric dogs and healthy Beagles except that cystinuric dogs were catheterized at baseline, 8 hours, 12 hours, and at the end of the collection, whereas Beagles were catheterized at baseline, 8 hours, and at the end of the collection. Three of 5 dogs with cystinuria had increased renal excretion of carnitine. None of the cystinuric dogs had increased renal excretion of taurine, but cystinuric dogs excreted significantly less (P < .05) taurine in their urine than the healthy Beagles. Carnitinuria has not been recognized previously in either humans or dogs with cystinuria, and it may be 1 risk factor for developing carnitine deficiency. Cystinuric dogs in this study were not taurinuric; however, cystine is a precursor amino acid for taurine synthesis. Therefore, cystinuria may be 1 risk factor for developing taurine deficiency in dogs. We suggest that dogs with cystinuria be monitored for carnitine and taurine deficiency or supplemented with carnitine and taurine.

Effects of dietary fat and carnitine on urine carnitine excretion in healthy dogs.

Turnover of carnitine in the body is primarily the result of renal excretion, and high-fat (HF) diets have been shown to increase urine carnitine excretion in healthy people. Recently, increased renal excretion of carnitine was observed in dogs diagnosed with cystinuria and carnitine deficiency. Carnitine deficiency has been linked to dilated cardiomyopathy and lipid storage myopathies in dogs and humans, and low-fat (LF) diets have been beneficial in some human patients with carnitine deficiency. In addition, HF, protein-restricted diets are often recommended for management of cystinuria in dogs. However, whether HF diets increase renal carnitine excretion in dogs or whether dogs with carnitine deficiency would benefit from LF diets remains unknown. Therefore, the purpose of this study was to determine the influence of dietary fat and carnitine on renal carnitine excretion in healthy dogs. Results from this study revealed that an HF diet increased urine carnitine excretion in dogs; however, carnitine excretion with the HF diet was not significantly different from that in dogs consuming an LF diet. Nonetheless, these results raise the possibility that increased renal carnitine excretion associated with HF diets could be one risk factor for development of carnitine deficiency in dogs with an underlying disorder in carnitine metabolism, and some dogs with carnitine deficiency may benefit from an LF diet. Another important observation in this study was that renal excretion of carnitine exceeded dietary intake in all diet groups, confirming previous reports that concluded that canine renal tubular cells reabsorb carnitine poorly when compared with those of humans.

Unbalanced 4;6 translocation and progressive renal disease.

Two sibs are described with an unbalanced 4;6 translocation resulting in partial trisomy 6p and monosomy for distal 4p. Growth retardation, psychomotor retardation, and characteristic facial appearance are present. The facial anomalies include high prominent forehead, blepharoptosis, blepharophimosis, high nasal bridge, bulbous nose, long philtrum, small mouth with thin lips, and low-set ears. Both children have small kidneys and have had proteinuria since early childhood. The older boy developed progressive renal disease including hypertension and renal failure necessitating renal transplantation at age 18 years. Renal biopsy of the younger girl also indicates significant renal involvement. Progressive renal disease is likely an important part of the trisomy 6p phenotype.

Biochemical and molecular basis for mitochondrial cardiomyopathy in neonates and children.

Defects in myocardial bioenergetics have been reported in patients with cardiomyopathy but their molecular basis and role in pathophysiology remain unclear. We sought to establish a molecular basis for cardiac mitochondrial respiratory enzyme abnormalities frequently present (75%) in a group of 16 children (including 2 neonates) with end-stage cardiomyopathy. Decreased specific activity levels were found in complexes I, III, IV and V but not in II, the only complex that is entirely nuclear encoded. Sequence analysis of cardiac mtDNA revealed 4 patients harbouring heteroplasmic mtDNA mutations in cytb, tRNAArg, and ND5 at highly conserved positions. These mutations were present neither in controls nor in patients without enzymatic defect. In addition, 4 patients exhibited marked reduction in cardiac mtDNA levels. The basis for respiratory enzyme abnormalities can be explained in a subset of our patients as a result of either pathogenic mtDNA mutation or depletion. Patients harbouring both DNA and enzymatic defects fulfil rigorous criteria defining mitochondrial cardiomyopathy.

Metabolic aspects of myocardial disease and a role for L-carnitine in the treatment of childhood cardiomyopathy.

OBJECTIVES: A multicenter retrospective study was conducted to investigate the possible metabolic causes of pediatric cardiomyopathy and evaluate the outcome of patients treated with L-carnitine. METHODS: Seventy-six patients diagnosed with cardiomyopathy were treated with L-carnitine in addition to conventional cardiac treatment, and 145 patients were treated with conventional treatment only. There were 101 males and 120 females between 1 day and 18 years old. Cardiomyopathy diagnoses included dilated (148 patients), hypertrophic (42 patients), restrictive (16 patients), mixed diagnosis (11 patients), and 4 with an unknown type. Of 76 L-carnitine-treated patients, 29 (38%) had evidence to suggest a disorder of metabolism, and of 145 control patients, 15 (10%) were suspected to have a disorder of metabolism. These metabolic disorders were thought to be the cause for the cardiomyopathy of the patients. The duration of L-carnitine treatment ranged from 2 weeks to >1 year. Information was collected on length of survival (time-to-event), clinical outcome, echocardiogram parameters, and clinical assessments. Data were collected at intervals from baseline to study endpoint, death, transplant, or last known follow-up visit. RESULTS: L-Carnitine-treated patients were younger than control patients and had poorer clinical functioning at baseline, yet they demonstrated lower mortality and a level of clinical functioning and clinical severity comparable to control patients on conventional therapy by the end of the study. An analysis of the interaction between clinical outcome and concomitant medications unexpectedly revealed that the population of patients treated with angiotensin-converting enzyme (ACE) inhibitors (40% of patients) had significantly poorer survival (although their greater likelihood for poor survival may possibly have made them more likely to receive ACE inhibitors). CONCLUSION: Results suggest that L-carnitine provides clinical benefit in treating pediatric cardiomyopathy. There is a need for further exploration of potential explanatory factors for the higher mortality observed in the population of patients treated with ACE inhibitors.

Mutations in TFAP2B cause Char syndrome, a familial form of patent ductus arteriosus.

Char syndrome is an autosomal dominant trait characterized by patent ductus arteriosus, facial dysmorphism and hand anomalies. Using a positional candidacy strategy, we mapped TFAP2B, encoding a transcription factor expressed in neural crest cells, to the Char syndrome critical region and identified missense mutations altering conserved residues in two affected families. Mutant TFAP2B proteins dimerized properly in vitro, but showed abnormal binding to TFAP2 target sequence. Dimerization of both mutants with normal TFAP2B adversely affected transactivation, demonstrating a dominant-negative mechanism. Our work shows that TFAP2B has a role in ductal, facial and limb development and suggests that Char syndrome results from derangement of neural-crest-cell derivatives.

Familial carnitine transporter defect: A treatable cause of cardiomyopathy in children.

Carnitine transporter defect is characterized by severely reduced transport of carnitine into skeletal muscle, fibroblasts, and renal tubules. All children with dilated cardiomyopathy or hypoglycemia and coma should be evaluated for this transporter defect because it is readily amenable to therapy that results in prolonged prevention of cardiac failure. This article details the cases of 3 children who have carnitine transporter defect, 2 of whom had severe dilated cardiomyopathy. Plasma and skeletal muscle carnitine levels were extremely low and both children were treated with oral L-carnitine, resulting in resolution of severe cardiomyopathy and prevention of recurrence or cardiac enlargement for more than 5 years. The third child had hypoglycemia and coma as presenting findings of the transporter defect and had mild left ventricular hypertrophy but no cardiac failure. The prognosis for long-term survival in pediatric dilated cardiomyopathy is poor. Children with carnitine transporter defect can have a different outcome if their underlying condition is detected early and treated medically.

The complete sequence of mtDNA genes in idiopathic dilated cardiomyopathy shows novel missense and tRNA mutations.

BACKGROUND: Previous studies have shown that mitochondrial DNA (mtDNA) mutations are often present in patients with myocardial dysfunction. We sought to assess the prevalence and significance of heart mtDNA sequence changes in patients with idiopathic dilated cardiomyopathy (DCM). METHODS AND RESULTS: DNA sequence of all the transfer ribonucleic acid (tRNA), ribosomal RNA (rRNA), and structural genes in cardiac mtDNA of 28 patients with DCM was determined and compared with a control group that had no evidence of heart disease. An increased number of point mutations were found in DCM cardiac mtDNA when compared with controls. Both novel and previously reported mutations were found in mitochondrial tRNA and structural genes. One of these mutations was heteroplasmic and resulted in changing a highly conserved nucleotide in tRNAArg. Novel, heteroplasmic mtDNA mutations (n = 4) specifying changes in moderate to highly conserved amino acid residues were found in COII, COIII, ND5, and cytb. These novel mtDNA mutations were found only in patients with severe reduction in mitochondrial enzyme activities. CONCLUSIONS: Our results indicate that a high incidence of mtDNA nucleotide sequence changes in both tRNA and structural genes are present in DCM. Five heteroplasmic mutations were detected that both changed evolutionarily conserved residues (which may impair the function of proteins or tRNAs) and were associated with specific enzymatic defects. These mutations could play an important role in the pathogenesis of cardiomyopathy.

Genetic aspects of atrioventricular septal defects.

Formation of the atrioventricular canal (AVC) results from complex interactions of components of the extracellular matrix. In response to signaling molecules, endothelial/mesenchymal transformations are crucial to normal development of the AVC. Atrioventricular septal defects (AVSDs) can result from arrest or interruption of normal endocardial cushion development. The presence of AVSDs has been associated with chromosome abnormalities, laterality or left-right axis abnormalities, and a variety of syndromes. An AVSD susceptibility gene has been identified in a large kindred with many affected members. Studies of transcription factors and signaling molecules in heart development over the past decade are paving the way for our understanding of the heterogeneous mechanisms of causation of AVSDs.

The mutational spectrum of the sonic hedgehog gene in holoprosencephaly: SHH mutations cause a significant proportion of autosomal dominant holoprosencephaly.

Holoprosencephaly (HPE) is a common developmental anomaly of the human forebrain and midface where the cerebral hemispheres fail to separate into distinct left and right halves. We have previously reported haploinsufficiency for Sonic Hedgehog ( SHH ) as a cause for HPE. We have now performed mutational analysis of the complete coding region and intron-exon junctions of the SHH gene in 344 unrelated affected individuals. Herein, we describe 13 additional unrelated affected individuals with SHH mutations, including nonsense and missense mutations, deletions and an insertion. These mutations occur throughout the extent of the gene. No specific genotype-phenotype association is evident based on the correlation of the type or position of the mutations. In conjunction with our previous studies, we have identified a total of 23 mutations in 344 unrelated cases of HPE. They account for 14 cases of familial HPE and nine cases of sporadic HPE. Mutations in SHH were detected in 10 of 27 (37%) families showing autosomal dominant transmission of the HPE spectrum, based on structural anomalies. Interestingly, three of the patients with an SHH mutation also had abnormalities in another gene that is expressed during forebrain development. We suggest that the interactions of multiple gene products and/or environmental elements may determine the final phenotypic outcome for a given individual and that variations among these factors may cause the wide variability in the clinical features seen in HPE.

Cerebello-oculo-renal syndromes including Arima, Senior-Löken and COACH syndromes: more than just variants of Joubert syndrome.

Cerebellar vermis hypoplasia (CVH) is part of many different malformation syndromes, especially Joubert syndrome. However, the nosology of these disorders remains uncertain. We reviewed reports of 100 children with cerebellar vermis hypoplasia, and ocular or renal involvement. Although the status of the upper brainstem was not adequately documented in most of these patients, some had hypoplasia and dysplasia of the ponto-mesencephalic isthmus and the superior portion of the cerebellar vermis, which results in a "molar tooth" sign on MRI scan. Several distinct syndromes were apparent among this group. We conclude that (a) hypoplasia of the cerebellar vermis, especially the anterior vermis, is often associated with a complex brainstem malformation; (b) the latter comprises a "molar tooth" brainstem and vermis hypoplasia-dysplasia malformation complex; (c) this complex may include the Dandy-Walker malformation, occipital cephalocele, and some abnormalities of the cerebrum as evidenced by frequent mental retardation; and (d) the "molar tooth" sign or malformation is causally heterogeneous as it occurs in several distinct malformation syndromes including Joubert syndrome, Arima syndrome, Senior-Löken syndrome, COACH syndrome, and probably familial juvenile nephronophthisis.

Char syndrome, an inherited disorder with patent ductus arteriosus, maps to chromosome 6p12-p21.

BACKGROUND: Patent ductus arteriosus (PDA) is a relatively common form of congenital heart disease. Although polygenic inheritance has been implicated, no specific gene defects causing PDA have been identified to date. Thus, a positional cloning strategy was undertaken to determine the gene responsible for the Char syndrome, an autosomal dominant disorder characterized by PDA, facial dysmorphism, and hand anomalies. METHODS AND RESULTS: A genome scan was performed with 46 members of 2 unrelated families in which the disease was fully penetrant but the phenotype differed. Significant linkage was achieved with several polymorphic DNA markers mapping to chromosome 6p12-p21 (maximal 2-point LOD score of 8.39 with D6S1638 at theta=0.00). Haplotype analysis identified recombinant events that defined the Char syndrome locus with high probability to a 3. 1-cM region between D6S459/D6S1632/D6S1541 and D6S1024. CONCLUSIONS: A familial syndrome in which PDA is a common feature was mapped to a narrow region of chromosome 6p12-p21. Additional analysis with other families and polymorphic markers as well as evaluation of potential candidate genes should lead to the identification of the Char syndrome gene, which will provide insights into cardiogenesis as well as limb and craniofacial development.

Different TBX5 interactions in heart and limb defined by Holt-Oram syndrome mutations.

To better understand the role of TBX5, a T-box containing transcription factor in forelimb and heart development, we have studied the clinical features of Holt-Oram syndrome caused by 10 different TBX5 mutations. Defects predicted to create null alleles caused substantial abnormalities both in limb and heart. In contrast, missense mutations produced distinct phenotypes: Gly80Arg caused significant cardiac malformations but only minor skeletal abnormalities; and Arg237Gln and Arg237Trp caused extensive upper limb malformations but less significant cardiac abnormalities. Amino acids altered by missense mutations were located on the three-dimensional structure of a related T-box transcription factor, Xbra, bound to DNA. Residue 80 is highly conserved within T-box sequences that interact with the major groove of target DNA; residue 237 is located in the T-box domain that selectively binds to the minor groove of DNA. These structural data, taken together with the predominant cardiac or skeletal phenotype produced by each missense mutation, suggest that organ-specific gene activation by TBX5 is predicated on biophysical interactions with different target DNA sequences.

Reliability of taurine concentrations measured in single urine samples obtained from dogs eight hours after eating.

OBJECTIVE: To evaluate the reliability of taurine concentrations measured in a single urine sample obtained from dogs 8 hours after eating, compared with taurine concentrations measured in 24-hour urine samples. ANIMALS: 18 healthy Beagles. PROCEDURE: After emptying the urinary bladder by transurethral catheterization, dogs were fed a canned maintenance diet. Approximately 8 hours later, urine, plasma, and serum samples were obtained for determination of fractional urinary excretion of taurine and urine taurine-to-creatinine concentration ratios (Utaur:Ucr). Results were compared with 24-hour urinary taurine excretion rate. RESULTS: Unbound and total fractional urinary taurine excretion correlated well with unbound and total 24-hour urinary taurine excretion. However, bound fractional urinary taurine excretion correlated poorly with bound 24-hour urinary taurine excretion. Unbound and total Utaur:Ucr correlated well with unbound and total 24-hour urinary taurine excretion. However, bound Utaur:Ucr correlated poorly with bound 24-hour urinary taurine excretion. CONCLUSION AND CLINICAL RELEVANCE: Fractional urinary excretion of unbound and total taurine, and unbound and total Utaur:Ucr are reliable indicators of 24-hour urinary unbound and total taurine excretion in healthy dogs. However, determination of 24-hour urinary taurine excretion is recommended for evaluating urinary bound taurine concentrations of dogs.

Duplication 6q22-->qter: definition of the phenotype.

We report on a girl with duplication of 6q22.32 --> qter and microcephaly, frontal bossing, facial anomalies, and webbed neck. She has congenital heart disease, renal hypoplasia, and hearing loss along with severe developmental delay. Published reports of seven other patients are reviewed and compared. The most frequent anomalies include microcephaly, abnormal face, webbed neck, congenital heart disease, limb contractures, and developmental delay.