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M Goossens

Publications and source records attributed to M Goossens.

At least 37 records · Page 2Linked to original sources
PubMed2000

Two mild cystic fibrosis-associated mutations result in severe cystic fibrosis when combined in cis and reveal a residue important for cystic fibrosis transmembrane conductance regulator processing and function.

The number of complex cystic fibrosis transmembrane conductance regulator (CFTR) genotypes identified as having double-mutant alleles with two mutations inherited in cis has been growing. We investigated the structure-function relationships of a severe cystic fibrosis (CF)-associated double mutant (R347H-D979A) to evaluate the contribution of each mild mutation to the phenotype. CFTR mutants expressed in HeLa cells were analyzed for protein biosynthesis and Cl(-) channel activity. Our data show that R347H is associated with mild defective Cl(-) channel activity and that the D979A defect leads to misprocessing. The mutant R347H-D979A combines both defects for a dramatic decrease in Cl(-) current. To decipher the molecular mechanism of this phenotype, single and double mutants with different charge combinations at residues 347 and 979 were constructed as charged residues were involved in this complex genotype. These studies revealed that residue 979, located in the third cytoplasmic loop, is critical for CFTR processing and Cl(-) channel activity highlighting the role of charged residues. These results have also important implications for CF, as they show that two mutations in cis can act in concert to alter dramatically CFTR function contributing to the wide phenotypic variability of CF disease.

Cystic Fibrosis↗
PubMed2000

Interaction among SOX10, PAX3 and MITF, three genes altered in Waardenburg syndrome.

Waardenburg syndrome (WS) is an autosomal dominant disorder with an incidence of 1 in 40 000 that manifests with sensorineural deafness and pigmentation defects. It is classified into four types depending on the presence or absence of additional symptoms. WS1 and WS3 are due to mutations in the PAX3 gene whereas some WS2 cases are associated with mutations in the microphthalmia-associated transcription factor (MITF) gene. The WS4 phenotype can result from mutations in the endothelin-B receptor gene (EDNRB), in the gene for its ligand, endothelin-3 (EDN3), or in the SOX10 gene. PAX3 has been shown to regulate MITF gene expression. The recent implication of SOX10 in WS4 prompted us to test whether this transcription factor, known to cooperate in vitro with PAX3, is also able to regulate expression from the MITF promoter. Here we show that SOX10, in synergy with PAX3, strongly activates MITF expression in transfection assays. Analyses revealed that PAX3 and SOX10 interact directly by binding to a proximal region of the MITF promoter containing binding sites for both factors. Moreover, SOX10 or PAX3 mutant proteins fail to transactivate this promoter, providing further evidence that the two genes act in concert to directly regulate expression of MITF. In situ hybridization experiments carried out in the dominant megacolon (DOM:) mouse, confirmed that SOX10 dysfunction impairs MITF: expression as well as melanocytic development and survival. These experiments, which demonstrate an interaction between three of the genes that are altered in WS, could explain the auditory-pigmentary symptoms of this disease.

Animals↗
PubMed2000

Species-specific alternative splice mimicry at the growth hormone receptor locus revealed by the lineage of retroelements during primate evolution.

In humans, growth hormone receptor (GHR) transcripts exist in two isoforms differing by the retention (GHRfl) or exclusion (GHRd3) of exon 3, whereas in mice GHRfl is solely expressed. This species-specific expression pattern is believed to result from an alternative splice event that, on the basis of conflicting data obtained in humans, has been considered to be tissue-, developmentally, and/or individual-specific. To decipher the molecular basis of this unusual trait, we isolated a 6.8-kilobase fragment spanning exon 3 from individuals expressing GHRfl. Sequence analysis revealed the existence of two 99% identical retroelements flanking this exon. Unexpectedly, individuals expressing GHRd3 displayed a 2.7-kilobase deletion involving exon 3, which most likely results from an ancestral homologous recombination between the two retroelements. The lineage of these retroelements during primate evolution revealed the species specificity of the GHRd3 allele. These findings led us to propose a model underlying the existence of the sole GHRfl allele in most species. Such a retrovirus-mediated alternative splice mimicry, which clears up several as yet unexplained phenomena (i.e. the above-mentioned expression data, the Mendelian inheritance of GHR expression patterns, and the deletion of nonconsecutive exons in growth hormone resistant patients), represents a novel physiological mechanism accounting for protein diversity between and within species.

Alternative Splicing↗
PubMed2000

Mutations in the MEFV gene in a large series of patients with a clinical diagnosis of familial Mediterranean fever.

Familial Mediterranean fever (FMF) is an autosomal recessively inherited disease affecting patients of the Mediterranean basin. FMF is characterized by recurrent episodes of fever accompanied with topical signs of inflammation. Some patients can develop a renal amyloidosis associated (AA) amyloidosis. The administration of colchicine is an effective preventive treatment of both the attacks and amyloidosis. The FMF gene (MEFV) was cloned and missense mutations were found to be responsible for the disease. We investigated a large series of 303 unselected and unrelated patients of various ethnic backgrounds with a clinical suspicion of FMF to confirm or invalidate the diagnosis of FMF and to determine the spectrum of MEFV mutations. Molecular analysis focused on all the most frequent mutations identified so far, and an exhaustive analysis of exon 10, containing the mutational hotspot, was performed through DNA sequencing. Sixty-two percent of Sephardic, North African Arabs, Armenian and Turkish patients were either homozygous or compound heterozygous for MEFV mutations. In other populations surrounding the Mediterranean Sea such as Greek, Italian, Portuguese, Kurdish and Lebanese populations, mutations were also found. In general, patients without Mediterranean origin had no mutations in the MEFV gene. Two new mis-sense mutations were identified in exon 10 of the MEFV gene: the S675N in an Italian patient and the M680L in a French patient without any known at-risk ethnic ancestry.

Amino Acid Substitution↗
PubMed2000

Neurological phenotype in Waardenburg syndrome type 4 correlates with novel SOX10 truncating mutations and expression in developing brain.

Waardenburg syndrome type 4 (WS4), also called Shah-Waardenburg syndrome, is a rare neurocristopathy that results from the absence of melanocytes and intrinsic ganglion cells of the terminal hindgut. WS4 is inherited as an autosomal recessive trait attributable to EDN3 or EDNRB mutations. It is inherited as an autosomal dominant condition when SOX10 mutations are involved. We report on three unrelated WS4 patients with growth retardation and an as-yet-unreported neurological phenotype with impairment of both the central and autonomous nervous systems and occasionally neonatal hypotonia and arthrogryposis. Each of the three patients was heterozygous for a SOX10 truncating mutation (Y313X in two patients and S251X [corrected] in one patient). The extended spectrum of the WS4 phenotype is relevant to the brain expression of SOX10 during human embryonic and fetal development. Indeed, the expression of SOX10 in human embryo was not restricted to neural-crest-derived cells but also involved fetal brain cells, most likely of glial origin. These data emphasize the important role of SOX10 in early development of both neural-crest-derived tissues, namely melanocytes, autonomic and enteric nervous systems, and glial cells of the central nervous system.

Autonomic Nervous System↗
PubMed2000

Screening practices for mutations in the CFTR gene ABCC7.

Cystic fibrosis transmembrane conductance regulator (CFTR) gene studies are now one of the most frequent activities in clinical molecular genetics laboratories. The number of requests is growing, owing to the increasingly wide range of recognized CFTR gene diseases (cystic fibrosis, congenital bilateral absence of the vas deferens, disseminated bronchiectasis, allergic bronchopulmonary aspergillosis and chronic pancreatitis), and the availability of efficient molecular tools for detecting mutations. A growing number of tests capable of simultaneously detecting several frequent CF mutations are being developed, and commercial kits are now available. The most recent kits detect nearly 90% of defective alleles in Caucasians, a rate high enough for carrier screening and for the majority of diagnostic requests. However, because of the wide variety of molecular defects documented in the CFTR gene, only a limited number of laboratories have mastered the entire panoply of necessary techniques, while other laboratories have to refer certain cases to specialized centers with complementary and/or scanning tools at their disposal. A good knowledge of CFTR diseases and their molecular mechanisms, together with expertise in the various techniques, is crucial for interpreting the results. Diagnostic strategies must take into account the indication, the patient's ethnic origin, and the time available in the framework of genetic counseling. This review presents the methods most frequently used for detecting CFTR gene mutations, and discusses the strategies most suited to the different clinical settings.

Cystic Fibrosis↗
PubMed2000

NF1 gene analysis focused on CpG-rich exons in a cohort of 93 patients with neurofibromatosis type 1.

We studied the NF1 gene in 93 unrelated patients with neurofibromatosis type1, focusing the analysis on four exons that contain the highest number of possible mutations occurring at CpG sites. We used denaturing gradient gel electrophoresis to analyse exons 16, 28, 29 and 49, which contain 45 (25%) of the 183 possible mutations that could occur at the 120 CpG dinucleotides of the coding sequence. Six different mutations were identified, five of which are novel: two truncating mutations, W1810X and 5448insG, located in exon29; two splice defects leading to exon29 skipping, 5206-2A>G and 5546G>A; and one missense mutation, L844F, located in exon16. The already described R1748X mutation located in exon29 was found in two unrelated patients. The 5546G>A and R1748X mutations are located at CpG sites, whereas the W1810X involves a CpNpG site. Four novel polymorphisms, which may be helpful for family studies, were also identified. Overall, all but one mutations were found in exon29, a result which suggests that all the CpG sites of the NF1 coding sequence do not have the same mutability, and that exon29, the most CpG-rich exon, contains mutational hotspots associated with NF1.

Adolescent↗
PubMed2000

The human dynein intermediate chain 2 gene (DNAI2): cloning, mapping, expression pattern, and evaluation as a candidate for primary ciliary dyskinesia.

Primary ciliary dyskinesia (PCD) is an autosomal recessive disease characterized by chronic sinusitis and bronchiectasis, and usually associated with hypofertility. Half of the patients present a situs inversus, defining the Kartagener's syndrome. This phenotype results from axonemal abnormalities of respiratory cilia and sperm flagella, i.e., mainly an absence of dynein arms. Recently, a candidate-gene approach, based on documented abnormalities of immotile strains of Chlamydomonas reinhardtii, allowed us to identify the first gene involved in PCD. Following the same strategy, we have characterized DNAI2, a human gene related to Chlamzydomonas IC69, and evaluated its possible involvement in a PCD population characterized by an absence of outer dynein arms. DNAI2, which is composed of 14 exons located at 17q25, is highly expressed in trachea and testis. No mutation was found in the DNAI2 coding sequence of the twelve patients investigated. However, ten intragenic polymorphic sites and an EcoRI RFLP have been identified, allowing the exclusion of DNAI2 in three consanguineous families.

Adolescent↗
PubMed2000

Mutations in LHX3 result in a new syndrome revealed by combined pituitary hormone deficiency.

Combined pituitary hormone deficiency (CPHD) has been linked with rare abnormalities in genes encoding transcription factors necessary for pituitary development. We have isolated LHX3, a gene involved in a new syndrome, using a candidate-gene approach developed on the basis of documented pituitary abnormalities of a recessive lethal mutation in mice generated by targeted disruption of Lhx3 (ref. 2). LHX3, encoding a member of the LIM class of homeodomain proteins, consists of at least six exons located at 9q34. We identified a homozygous LHX3 defect in patients of two unrelated consanguineous families displaying a complete deficit in all but one (adrenocorticotropin) anterior pituitary hormone and a rigid cervical spine leading to limited head rotation. Two of these patients also displayed a severe pituitary hypoplasia, whereas one patient presented secondarily with an enlarged anterior pituitary. These LHX3 mutations consist of a missense mutation (Y116C) in the LIM2 domain at a phylogenetically conserved residue and an intragenic deletion predicting a severely truncated protein lacking the entire homeodomain. These data are consistent with function of LHX3 in the proper development of all anterior pituitary cell types, except corticotropes, and extrapituitary structures.

Abnormalities, Multiple↗
PubMed2000

Recommendations for quality improvement in genetic testing for cystic fibrosis. European Concerted Action on Cystic Fibrosis.

These recommendations for quality improvement of cystic fibrosis genetic diagnostic testing provide general guidelines for the molecular genetic testing of cystic fibrosis in patients/individuals. General strategies for testing as well as guidelines for laboratory procedures, internal and external quality assurance, and for reporting the results, including the requirements of minimal services in mutation testing, the nomenclature for describing mutations, procedures to control false-positive amplification reactions and to validate tests, and guidelines to implement a quality system in a molecular diagnostic laboratory are reviewed.

Accreditation↗
PubMed2000

Clinical versus genetic diagnosis of familial Mediterranean fever.

The diagnosis of familial Mediterranean fever (FMF) has until recently been based on clinical signs alone. Discovery of the MEFV gene has enabled a molecular approach to diagnosis, which is already well established for diagnosing typical clinical forms of FMF. We evaluated the utility of this molecular approach in a large series of patients with various clinical presentations and ethnic origins. We looked for mutations in the MEFV gene in 303 unselected consecutive patients with a variable (from high to low) clinical suspicion of FMF. Two mutations were found in 133 patients (44%). In 22 patients (7%), the clinical diagnosis of FMF was unlikely according to the Tel Hashomer clinical criteria. Our results suggest that the spectrum of FMF-associated signs is broader than previously believed. Wider indications for genotyping should lead to more frequent diagnosis of FMF.

Adolescent↗
PubMed2000

Peripheral neuropathy with hypomyelination, chronic intestinal pseudo-obstruction and deafness: a developmental "neural crest syndrome" related to a SOX10 mutation.

We describe the case of a girl with an unusual congenital phenotype, combining peculiar peripheral nerve lesions with hypomyelination, chronic intestinal pseudoobstruction, and deafness. She was found to have a de novo heterozygous frameshift mutation in the gene encoding the SOX10 transcription factor. The likely role of SOX10 in determining the fate of Schwann cells during early embryogenesis may explain the peripheral nervous system developmental disorder observed in this patient.

Amino Acid Sequence↗
PubMed1999

Structure-function analysis of a double-mutant cystic fibrosis transmembrane conductance regulator protein occurring in disorders related to cystic fibrosis.

A number of disorders related to cystic fibrosis have been described since the cloning of the cystic fibrosis gene, including infertility due to the congenital bilateral absence of the vas deferens. We have identified, in several patients, complex cystic fibrosis transmembrane conductance regulator genotypes like double-mutant alleles. We have now analyzed the structure-function relationships of one of these mutants, R74W-D1270N cystic fibrosis transmembrane conductance regulator, expressed in HeLa cells, to evaluate the contribution of each mutation in the phenotype. We found that R74W cystic fibrosis transmembrane conductance regulator appears to be a polymorphism, while D1270N cystic fibrosis transmembrane conductance regulator could be responsible for the congenital bilateral absence of the vas deferens phenotype. The combination of the two produced a more severe effect on the chloride conductance pathway as well as on the phenotype.

Alleles↗