Mechanisms of Mendelian dominance.

Genetic dominance has long been considered as a qualitative reflection of interallelic interactions. Dominance arises from many multiple sources whose unifying theme is the existence of non-linear relationships between the genotypic and phenotypic values. One of the clearest examples are dominant negative mutations (DNMs) in which a defective subunit poisons a macromolecular complex. Dominance can also be due to the presence of a heterozygous null allele, as is the case of haploinsufficiency. Dominance can also be influenced by epistatic (interloci) interactions. For instance, a pre-existing genetic variant can make possible the expression of a pathogenic variant in a seemingly "dominant" fashion. Such interactions, which can make an individual more or less sensitive to a particular pathogenic variant, will also be discussed here.

The genetic make-up of ovarian development and function: the focus on the transcription factor FOXL2.

In a 46 XY individual, the presence of the Y chromosome harboring the testis-determining factor (SRY) triggers testis determination and differentiation. In a 46 XX individual, the absence of SRY and the activation of genes associated with the female pathway lead to ovarian development. The latter process has long been considered as a default pathway. However, recent studies have cast doubts on this dogma. Here, after a brief overview of the main steps of ovarian development, we focus on three genes WNT4, RSPO1 and FOXL2 that are essential for ovarian determination, differentiation and/or maintenance. Special attention is paid to FOXL2 whose mutations are responsible for the blepharophimosis syndrome, often associated with female infertility, and for cancer. We highlight the cooperation of WNT4, RSPO1 and FOXL2 within a regulatory network and the need for further research to better understand their role in defining and maintaining ovarian identity.

A non-sense MCM9 mutation in a familial case of primary ovarian insufficiency.

Primary ovarian insufficiency (POI) results in an early loss of ovarian function, and remains idiopathic in about 80% of cases. Here, we have performed a complete genetic study of a consanguineous family with two POI cases. Linkage analysis and homozygosity mapping identified 12 homozygous regions with linkage, totalling 84 Mb. Whole-exome sequencing of the two patients and a non-affected sister allowed us to detect a homozygous causal variant in the MCM9 gene. The variant c.1483G>T [p.E495*], confirmed using Sanger sequencing, introduced a premature stop codon in coding exon 8 and is expected to lead to the loss of a functional protein. MCM9 belongs to a complex required for DNA repair by homologous recombination, and its impairment in mouse is known to induce meiotic recombination defects and oocyte degeneration. A previous study recently described two consanguineous families in which homozygous mutations of MCM9 were responsible for POI and short stature. Interestingly, the affected sisters in the family described here had a normal height. Altogether, our results provide the confirmation of the implication of MCM9 variants in POI and expand their phenotypic spectrum.

GermOnline, a cross-species community knowledgebase on germ cell differentiation.

GermOnline provides information and microarray expression data for genes involved in mitosis and meiosis, gamete formation and germ line development across species. The database has been developed, and is being curated and updated, by life scientists in cooperation with bioinformaticists. Information is contributed through an online form using free text, images and the controlled vocabulary developed by the GeneOntology Consortium. Authors provide up to three references in support of their contribution. The database is governed by an international board of scientists to ensure a standardized data format and the highest quality of GermOnline's information content. Release 2.0 provides exclusive access to microarray expression data from Saccharomyces cerevisiae and Rattus norvegicus, as well as curated information on approximately 700 genes from various organisms. The locus report pages include links to external databases that contain relevant annotation, microarray expression and proteome data. Conversely, the Saccharomyces Genome Database (SGD), S.cerevisiae GeneDB and Swiss-Prot link to the budding yeast section of GermOnline from their respective locus pages. GermOnline, a fully operational prototype subject-oriented knowledgebase designed for community annotation and array data visualization, is accessible at http://www.germonline.org. The target audience includes researchers who work on mitotic cell division, meiosis, gametogenesis, germ line development, human reproductive health and comparative genomics.

A recurrent polyalanine expansion in the transcription factor FOXL2 induces extensive nuclear and cytoplasmic protein aggregation.

Blepharophimosis syndrome is an autosomal dominant disease characterised by eyelid malformations, associated or not with premature ovarian failure. It is caused by mutations in the FOXL2 gene, which encodes a forkhead transcription factor containing a polyalanine (polyAla) domain of 14 alanines. Expansions of the polyAla tract from 14 to 24 residues account for 30% of the reported mutations and lead mainly to isolated palpebral defects. We have transfected COS-7 cells with DNA constructs driving the expression of the wildtype and mutant FOXL2 proteins fused to the green fluorescent protein. The polyAla expansion was found to induce the formation of intranuclear aggregates and a mislocalisation of the protein due to extensive cytoplasmic aggregation. These findings were confirmed by immunofluorescence. Co-transfection experiments suggest that the wildtype and mutant proteins can co-aggregate. We propose that the mechanism for the molecular pathogenesis of the polyAla expansions of FOXL2 may be its mislocalisation concomitant with its inclusion into nuclear aggregates. This may diminish the pool of active protein. Potential effects of aggregation on cell viability are under study.

Targeting the molecular mechanism of DNA replication.

Genome stability is crucial for the complete maintenance of the cellular pathways that govern the cell cycle. As a result of irregularities in DNA replication occurring throughout the S phase, key genes that regulate cell cycle pathways are damaged, giving rise to single-base mutations and chromosomal aberrations. Thus, the efficient replication of the genome, which depends on a precise temporal and spatial pattern of activation of origins of replication, is greatly impaired. The approach discussed below aims at monitoring the replication pattern and the kinetics of replication throughout the entire genome of living cells. It could shed light on the mechanisms by which drugs act on DNA replication and, moreover, it might assist the discovery and design of novel drugs that inhibit cell proliferation under pathophysiological conditions.

Molecular combing.

This unit describes an important advance in fiber-FISH technology called molecular combing, in which single DNA molecules are bound by one or both ends to a surface and stretched in a uniform and parallel manner by a receding meniscus. This technique is gentle on the molecules, rapid, and easy to perform. Reliable, quantitative information for genome-wide studies can be obtained without the need for other techniques and a large number of accurate measurements can be made in a single experiment. The authors provide detailed protocols for basic molecular combing, high-resolution physical mapping, and gene-dosage approaches as well as support protocols outlining surface preparation, DNA solution preparation, and probe labeling.