Karin Segers

The callipyge mutation enhances the expression of coregulated imprinted genes in cis without affecting their imprinting status

The callipyge (CLPG) phenotype (from καλι, “beautiful,” and πιγɛ, “buttocks”) described in sheep is an inherited muscular hypertrophy that is subject to an unusual parent-of-origin effect referred to as polar overdominance: only heterozygous individuals having inherited the CLPG mutation from their sire exhibit the muscular hypertrophy. The callipyge (clpg) locus was mapped to a chromosome segment of approximately 400 kb (refs. 2–4), which was shown to contain four genes (DLK1, GTL2, PEG11 and MEG8) that are preferentially expressed in skeletal muscle and subject to parental imprinting in this tissue. Here we describe the effect of the CLPG mutation on the expression of these four genes, and demonstrate that callipyge individuals have a unique expression profile that may account for the observed polar overdominance.

The callipyge mutation and other genes that affect muscle hypertrophy in sheep

Genetic strategies to improve the profitability of sheep operations have generally focused on traits for reproduction. However, natural mutations exist in sheep that affect muscle growth and development, and the exploitation of these mutations in breeding strategies has the potential to significantly improve lamb-meat quality. The best-documented mutation for muscle development in sheep is callipyge (CLPG), which causes a postnatal muscle hypertrophy that is localized to the pelvic limbs and loin. Enhanced skeletal muscle growth is also observed in animals with the Carwell (or rib-eye muscling) mutation, and a double-muscling phenotype has been documented for animals of the Texel sheep breed. However, the actual mutations responsible for these muscular hypertrophy phenotypes in sheep have yet to be identified, and further characterization of the genetic basis for these phenotypes will provide insight into the biological control of muscle growth and body composition.

Fine-mapping and construction of a bovine contig spanning the ovine callipyge locus

Abstract. The callipyge (CLPG) gene was fine-mapped by linkage analysis to a 4.6-cM chromosome interval on distal ovine OAR18q, flanked by microsatellite markers IDVGA30 and OY3. The OAR18q linkage map and human HSA14q transcript map were aligned by genotyping two bovine-hamster whole-genome radiation hybrid panels with the microsatellite markers, as well as with sequences corresponding to HSA14q genes. Using Type I loci mapping to the IDVGA30–OY3 interval as anchor points, we have constructed a 1.4-Mb bovine BAC contig containing the IDVGA30–OY3 interval. We demonstrate that the IDVGA30– OY3 interval spans approximately 770 kb and contains at least four genes: YY1, WARS, DLK1, and GTL2.

Novel SACS mutation in a Belgian family with sacsin-related ataxia.

The authors describe the four patients in the first known Belgian family with autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS). A novel homozygous missense mutation, NM_014363.3: c.3491T>A in exon 9, of the SACS gene was identified in the present family, which results in an original amino acid of methionine to lysine substitution at amino acid residue 1164 (p.M1164K). Although the cardinal clinical features, i.e., spastic ataxia with peripheral neuropathy, in our patients were similar to those in Quebec patients, our patients exhibited some atypical clinical features, e.g., teenage-onset and absence of retinal hypermyelination. The present family is from Wallonia, and there could be shared ethnicity with the families of Charlevoix-Saguenay.

Germline PTPN11 missense mutation in a case of Noonan syndrome associated with mediastinal and retroperitoneal neuroblastic tumors

Noonan syndrome (NS) is an autosomal dominant disorder characterized by short stature, typical craniofacial dysmorphism, skeletal anomalies, congenital heart defects, and predisposition to malignant tumors. In approximately 50% of cases, the disease is caused by missense mutations in the PTPN11 gene. To date, solid tumors, and particularly brain tumors and rhabdomyosarcomas, have been documented in patients with NS; however, few cases of neuroblastoma associated with NS have been reported. Here we report an unusual case of neuroblastoma with mediastinal, retroperitoneal, and medullar locations associated in a NS patient carrying a PTPN11 germline missense mutation (p.G60A). This missense mutation occurs within the N-SH2 domain of the PTPN11 gene and has been reported to be associated with acute leukemia in NS patients. The association of this p.G60A PTPN11 mutation with neuroblastoma provides new evidence that gain of function PTPN11 mutations may play an important role in the pathogenesis of solid tumors associated with Noonan syndrome.

3-years experience review of neonatal screening for hemoglobin disorders using tandem mass spectrometry.

BACKGROUND Neonatal screening programs for sickle cell disease are common in North America and in some European countries. Isoelectric Focusing or High Performance Liquid Chromatography is the main technique used for hemoglobin variant detection. METHODS Since tandem mass spectrometry is being used for screening of inherited metabolic disorders and allows protein identification, we had developed an application to identify the most relevant hemoglobin mutations with this technology. RESULTS This approach had been previously validated and has been routinely applied in our laboratory for the last three years. We report here our experience with this new method in the field, applied to our East-Belgian population. CONCLUSIONS To conclude, mass spectrometry provides an efficient alternative approach for laboratories performing neonatal screening of hemoglobin disorders.

Breakpoint mapping positions the callipyge gene within a 450-kilobase chromosome segment containing the DLK1 and GTL2 genes.

We have previously mapped the ovine callipyge (CLPG) gene,causing a muscular hypertrophy with parent-of-origin-dependentexpression referred to as polar overdominance, to a 4.6-cM chro-mosome interval on distal OAR18q flanked by microsatellitesIDVGA30 and OY3 (Cockett et al. 1996; Shay et al. 2000). BACcontigs spanning this interval were constructed by using bovine(Shay et al. 2000) and subsequently ovine (Segers et al. 2000)reagents. We herein report the isolation of eight novel microsat-ellite markers from these contigs, yielding a marker density of onemicrosatellite per 68 kilobases and the use of these novel markersto position the CLPG gene by breakpoint analysis within a ≈450-kilobase chromosome segment.Five BAC clones jointly spanning most of the IDVGA30–OY3interval were selected from the ovine BAC contig (BACs: 724D11,239G7, 218E10, 497C1, 265F11). BAC DNA was digested tocompletion with three four-cutters yielding blunt-ended restrictionfragments (AluI, HaeIII, and RsaI) used either separately or com-bined. Restriction fragments containing microsatellites were de-tected by standard Southern blotting and hybridization with a(CA)

Spinocerebellar Ataxia Type 2 (SCA2): Clinical Features and Genetic Analysis

Spinocerebellar ataxia type 2 (SCA2) is an autosomal dominant neurodegenerative disease that results from the expansion of an unstable trinucleotide CAG repeat encoding for a polyglutamine tract. In normal individuals, alleles contain between 14 and 31 CAG repeats, whereas the pathological alleles have more than 35 CAG repeats. The clinical phenotype of SCA2 includes a progressive cerebellar ataxia with additional features such as ophthalmoplegia, extra-pyramidal or pyramidal signs and peripheral neuropathy. We report a SCA2 large African family with several affected individuals. A major pathological allele carrying 43 CAG repeats was identified in the proband. To our knowledge, this is a first report of a SCA disorder described in Central African patients, thus indicating the need to consider this diagnosis in young African ataxic patients.

Breast cancer in a male-to-female transsexual patient with a BRCA2 mutation

Breast cancer is rare in male patients. Certain predisposing factors, be they genetic (e.g., BRCA2 gene mutations) or hormonal (imbalance between estrogen and androgen levels), have been implicated in male breast cancer pathophysiology. Male-to-female (MtF) transsexualism is a condition that generally involves cross-sex hormone therapy. Anti-androgens and estrogens are used to mimic the female hormonal environment and induce the cross-sex secondary characteristics. In certain situations, the change in the hormonal milieu can be disadvantageous and favor the development of hormone-dependent pathologies, such as cancer. We report a case of a MtF transgender patient who developed breast cancer after 7 years of cross-sex hormonal therapy. The patient was found to be BRCA2 positive, and suffered recurrent disease. The patient was unaware of being a member of an established BRCA2 mutation-positive kindred. This represents the first case of a BRCA2 mutation predisposing to breast cancer in a MtF transgender patient.

Evaluation of BRCA1-related molecular features and microRNAs as prognostic factors for triple negative breast cancers.

BackgroundThe BRCA1 gene plays a key role in triple negative breast cancers (TNBCs), in which its expression can be lost by multiple mechanisms: germinal mutation followed by deletion of the second allele; negative regulation by promoter methylation; or miRNA-mediated silencing. This study aimed to establish a correlation among the BRCA1-related molecular parameters, tumor characteristics and clinical follow-up of patients to find new prognostic factors.MethodsBRCA1 protein and mRNA expression was quantified in situ in the TNBCs of 69 patients. BRCA1 promoter methylation status was checked, as well as cytokeratin 5/6 expression. Maintenance of expressed BRCA1 protein interaction with BARD1 was quantified, as a marker of BRCA1 functionality, and the tumor expression profiles of 27 microRNAs were determined.ResultsmiR-548c-5p was emphasized as a new independent prognostic factor in TNBC. A combination of the tumoral expression of miR-548c and three other known prognostic parameters (tumor size, lymph node invasion and CK 5/6 expression status) allowed for relapse prediction by logistic regression with an area under the curve (AUC) = 0.96.BRCA1 mRNA and protein in situ expression, as well as the amount of BRCA1 ligated to BARD1 in the tumor, lacked any associations with patient outcomes, likely due to high intratumoral heterogeneity, and thus could not be used for clinical purposes.ConclusionsIn situ BRCA1-related expression parameters could be used for clinical purposes at the time of diagnosis. In contrast, miR-548c-5p showed a promising potential as a prognostic factor in TNBC.