Véronique Satre

From Lowe syndrome to Dent disease: correlations between mutations of the OCRL1 gene and clinical and biochemical phenotypes

Mutations of OCRL1 are associated with both the Lowe oculocerebrorenal syndrome, a multisystemic and Dent‐2 disease, a renal tubulopathy. We have identified a mutation in 130 Lowe syndrome families and 6 affected by Dent‐2 disease with 51 of these mutations being novel. No founding effect was evidenced for recurrent mutations. Two mutations initially reported as causing Dent‐2 disease were identified in patients, including two brothers, presenting with Lowe syndrome thus extending the clinical variability of OCRL1 mutations. mRNA levels, protein content, and PiP2‐ase activities were analyzed in patients fibroblasts. Although mRNA levels were normal in cells harboring a missense mutation, the OCRL1 content was markedly lowered, suggesting that enzymatic deficiency resulted mainly from protein degradation rather than from a catalytic inactivation. Analysis of a splicing mutation that led to the elimination of the initiation codon evidenced the presence of shortened forms of OCRL1 that might result from the use of alternative initiation codons. The specific mapping of the frameshift and nonsense mutations, exclusively identified in exons 1–7 and exons 8–23, respectively, for Dent disease and Lowe syndrome together with the possible use of alternative initiation codons might be related to their clinical expression, that is, Lowe syndrome or Dent‐2 disease. Hum Mutat 32:1–10, 2011.

OCRL1 mutation analysis in French Lowe syndrome patients: Implications for molecular diagnosis strategy and genetic counseling

The oculocerebrorenal syndrome of Lowe (OCRL) is a rare X‐linked recessively inherited disease characterized by a severe pleiotropic phenotype including mental retardation, bilateral congenital cataract, and renal Fanconi syndrome. The gene responsible for OCRL encodes an inositol polyphosphate‐5‐phosphatase. We performed mutation analysis in 36 families and characterized 27 new mutations with two of them being recurrent mutations. The panel of mutations consisted of 27 truncating mutations (frameshift, nonsense, splice site mutations, and large genomic deletions), one in‐frame deletion, and six missense mutations. The four large genomic deletions occurred in the first half of the gene, whereas all the remaining mutations took place in the second part of the gene and were concentrated in a few exons. This distribution may be of interest in terms of screening strategy when looking for unknown mutations. Haplotyping of the families was performed to analyze segregation of the mutated loci, and revealed a somatic mosaicism in one family. This is the second case of mosaicism we characterized in a total panel of 44 unrelated families affected by Lowes syndrome. Considering the low number of families investigated, it appeared that somatic and germinal mosaicisms are quite common in this disease and must be taken into account for genetic counseling. Hum Mutat 16:157–165, 2000.

Characterization of a Germline Mosaicism in Families with Lowe Syndrome, and Identification of Seven Novel Mutations in the OCRL1 Gene

The oculocerebrorenal syndrome of Lowe (OCRL) is an X-linked disorder characterized by major abnormalities of eyes, nervous system, and kidneys. Mutations in the OCRL1 gene have been associated with the disease. OCRL1 encodes a phosphatidylinositol 4, 5-biphosphate (PtdIns[4,5]P2) 5-phosphatase. We have examined the OCRL1 gene in eight unrelated patients with OCRL and have found seven new mutations and one recurrent in-frame deletion. Among the new mutations, two nonsense mutations (R317X and E558X) and three other frameshift mutations caused premature termination of the protein. A missense mutation, R483G, was located in the highly conserved PtdIns(4,5)P2 5-phosphatase domain. Finally, one frameshift mutation, 2799delC, modifies the C-terminal part of OCRL1, with an extension of six amino acids. Altogether, 70% of missense mutations are located in exon 15, and 52% of all mutations cluster in exons 11-15. We also identified two new microsatellite markers for the OCRL1 locus, and we detected a germline mosaicism in one family. This observation has direct implications for genetic counseling of Lowe syndrome families.

SPINK2 deficiency causes infertility by inducing sperm defects in heterozygotes and azoospermia in homozygotes

Azoospermia, characterized by the absence of spermatozoa in the ejaculate, is a common cause of male infertility with a poorly characterized etiology. Exome sequencing analysis of two azoospermic brothers allowed the identification of a homozygous splice mutation in SPINK2, encoding a serine protease inhibitor believed to target acrosin, the main sperm acrosomal protease. In accord with these findings, we observed that homozygous Spink2 KO male mice had azoospermia. Moreover, despite normal fertility, heterozygous male mice had a high rate of morphologically abnormal spermatozoa and a reduced sperm motility. Further analysis demonstrated that in the absence of Spink2, protease‐induced stress initiates Golgi fragmentation and prevents acrosome biogenesis leading to spermatid differentiation arrest. We also observed a deleterious effect of acrosin overexpression in HEK cells, effect that was alleviated by SPINK2 coexpression confirming its role as acrosin inhibitor. These results demonstrate that SPINK2 is necessary to neutralize proteases during their cellular transit toward the acrosome and that its deficiency induces a pathological continuum ranging from oligoasthenoteratozoospermia in heterozygotes to azoospermia in homozygotes.

17p13.1 microduplication in a boy with Silver-Russell syndrome features and intellectual disability

Many deletions of chromosome 17p13.1 have been described, but very few 17p13.1 duplications have been reported yet. Here, we describe the genotype and phenotype of a boy with a duplication of this region. The main clinical features are mild intellectual deficiency, growth retardation, and a typical Silver–Russell syndrome (SRS) appearance with small triangular face, prominent forehead, micrognathia, low‐set ears, and clinodactyly. Array‐CGH revealed a 586 kb duplication containing many genes with a high neuronal expression. Interestingly, this region covers the minimal critical region including all candidate genes suggested to explain the 17p13.1 microdeletion syndrome. In the neighboring region 17p13.3, deletions and duplications of the same region are each responsible of a specific phenotype. Future case descriptions will show if a similar mechanism applies to the region 17p13.1. The 17p13.1 region contains interesting putative candidate genes that might be involved in the SRS etiology. Additional data are needed to verify the significance of this aberration.

PBX1 haploinsufficiency leads to syndromic congenital anomalies of the kidney and urinary tract (CAKUT) in humans

Background Congenital anomalies of the kidney and urinary tract (CAKUT) represent a significant healthcare burden since it is the primary cause of chronic kidney in children. CNVs represent a recurrent molecular cause of CAKUT but the culprit gene remains often elusive. Our study aimed to define the gene responsible for CAKUT in patients with an 1q23.3q24.1 microdeletion. Methods We describe eight patients presenting with CAKUT carrying an 1q23.3q24.1 microdeletion as identified by chromosomal microarray analysis (CMA). Clinical features were collected, especially the renal and urinary tract phenotype, and extrarenal features. We characterised PBX1 expression and localisation in fetal and adult kidneys using quantitative RT-PCR and immunohistochemistry. Results We defined a 276-kb minimal common region (MCR) that only overlaps with the PBX1 gene. All eight patients presented with syndromic CAKUT. CAKUT were mostly bilateral renal hypoplasia (75%). The most frequent extrarenal symptoms were developmental delay and ear malformations. We demonstrate that PBX1 is strongly expressed in fetal kidneys and brain and expression levels decreased in adult samples. In control fetal kidneys, PBX1 was localised in nuclei of medullary, interstitial and mesenchymal cells, whereas it was present in endothelial cells in adult kidneys. Conclusions Our results indicate that PBX1 haploinsufficiency leads to syndromic CAKUT as supported by the Pbx1-null mice model. Correct PBX1 dosage appears to be critical for normal nephrogenesis and seems important for brain development in humans. CMA should be recommended in cases of fetal renal anomalies to improve genetic counselling and pregnancy management.

Interphase fluorescent in situ hybridization detection of the 7q11.23 chromosomal inversion in a clinical laboratory: automated versus manual scoring.

*Corresponding author: Gwena ë l Nadeau , Unit é Fonctionnelle de Cytog é n é tique, Centre Hospitalier de Valence, 179 Boulevard du Mar é chal Juin, 26953 Valence Cedex 9, France, Phone: + 33 475757297, Fax: + 33-475757161, E-mail: gnadeau@ch-valence.fr Gwena ë l Nadeau, Charles Coutton, Florence Amblard, Gabrielle Michalowicz, Sylvie Frasca , Fran ç oise Devillard , V é ronique Satre and Pierre-Simon Jouk: Centre Hospitalier Universitaire de Grenoble , D é partement de G é n é tique et Procr é ation, Grenoble , France Charles Coutton and V é ronique Satre: Laboratoire AGIM , CNRS FRE3405, Equipe “ G é n é tique, Infertilit é et Th é rapeutiques ” , La Tronche, France, Universit é Joseph Fourier, Grenoble , France Gabrielle Michalowicz , Arnold Fertin , Yves Usson and Pierre-Simon Jouk: UJF-Grenoble1 , TIMC-IMAG, DyCTIM Team, CNRS-UJF UMR 5525, Grenoble , France

Further delineation of the MECP2 duplication syndrome phenotype in 59 French male patients, with a particular focus on morphological and neurological features

The Xq28 duplication involving the MECP2 gene (MECP2 duplication) has been mainly described in male patients with severe developmental delay (DD) associated with spasticity, stereotypic movements and recurrent infections. Nevertheless, only a few series have been published. We aimed to better describe the phenotype of this condition, with a focus on morphological and neurological features. Through a national collaborative study, we report a large French series of 59 affected males with interstitial MECP2 duplication. Most of the patients (93%) shared similar facial features, which evolved with age (midface hypoplasia, narrow and prominent nasal bridge, thick lower lip, large prominent ears), thick hair, livedo of the limbs, tapered fingers, small feet and vasomotor troubles. Early hypotonia and global DD were constant, with 21% of patients unable to walk. In patients able to stand, lower limbs weakness and spasticity led to a singular standing habitus: flexion of the knees, broad-based stance with pseudo-ataxic gait. Scoliosis was frequent (53%), such as divergent strabismus (76%) and hypermetropia (54%), stereotypic movements (89%), without obvious social withdrawal and decreased pain sensitivity (78%). Most of the patients did not develop expressive language, 35% saying few words. Epilepsy was frequent (59%), with a mean onset around 7.4 years of age, and often (62%) drug-resistant. Other medical issues were frequent: constipation (78%), and recurrent infections (89%), mainly lung. We delineate the clinical phenotype of MECP2 duplication syndrome in a large series of 59 males. Pulmonary hypertension appeared as a cause of early death in these patients, advocating its screening early in life.

Microduplication of the ARID1A gene causes intellectual disability with recognizable syndromic features

Purpose:To determine whether duplication of the ARID1A gene is responsible for a new recognizable syndrome.Methods:We describe four patients with a 1p36.11 microduplication involving ARID1A as identified by array–comparative genomic hybridization . We performed comparative transcriptomic analysis of patient-derived fibroblasts using RNA sequencing and evaluated the impact of ARID1A duplication on the cell cycle using fluorescence-activated cell sorting. Functional relationships between differentially expressed genes were investigated with ingenuity pathway analysis (IPA).Results:Combining the genomic data, we defined a small (122 kb), minimally critical region that overlaps the full ARID1A gene. The four patients shared a strikingly similar phenotype that included intellectual disability and microcephaly. Transcriptomic analysis revealed the deregulated expression of several genes previously linked to microcephaly and developmental disorders as well as the involvement of signaling pathways relevant to microcephaly, among which the polo-like kinase (PLK) pathway was especially notable. Cell-cycle analysis of patient-derived fibroblasts showed a significant increase in the proportion of cells in G1 phase at the expense of G2-M cells.Conclusion:Our study reports a new microduplication syndrome involving the ARID1A gene. This work is the first step in clarifying the pathophysiological mechanism that links changes in the gene dosage of ARID1A with intellectual disability and microcephaly.Genet Med advance online publication 01 December 2016