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C Cotinot

Publications and source records attributed to C Cotinot.

At least 19 recordsLinked to original sources
PubMed2003

Expression studies of the PIS-regulated genes suggest different mechanisms of sex determination within mammals.

In mammals, the Y-located SRY gene is known to induce testis formation from the indifferent gonad. A related gene, SOX9, also plays a critical role in testis differentiation in mammals, in birds and reptiles. It is now assumed that SRY acts upstream of SOX9 in the sex determination cascade, but the regulatory link which should exist between these two genes remains unknown. Studies on XX sex reversal in polled goats (PIS mutation: Polled Intersex Syndrome) have led to the discovery of a female-specific locus crucial for ovarian differentiation. This genomic region is composed of at least two genes, FOXL2 and PISRT1, which share a common transcriptional regulatory region, PIS. In this review, we present the expression pattern of these PIS-regulated genes in mice. The FOXL2 expression profile of mice is similar to that described in goats in accordance with a conserved role of this ovarian differentiating gene in mammals. On the contrary, the PISRT1 expression profile is different between mice and goats, suggesting different mechanisms of the primary switch in the testis determination process within mammals. A model based on two different modes of SOX9 regulation in mice and other mammals is proposed in order to integrate our results into the current scheme of gonad differentiation.

Animals↗
PubMed2002

[Human infertility: meiotic genes as potential candidates].

Up to now, the identification of gene mutations causing infertility in humans remains poorly investigated. Temporal progression through meiosis and meiosis specific genes had been extensively characterized in yeast. Recently some mammalian homologous were found. The molecular mechanisms regulating entry into and progression through meiosis in mammals are still unknown. However, disruption of some meiotic genes in mouse showed an essential role of them in meiotic chromosome synapsis and gametogenesis. Moreover, the phenotype of gonads in null mutant mice for some meiotic genes (failure to initiate or blockage in meiosis, lack of gametes or small size of gonads...) could be strikingly similar to clinical observations found in human infertility. The aim of this study was to identify putative mutations in 5 meiotic genes of several clinically well-characterized patients who present unexplained infertility (normal karyotype, women with premature ovarian failure, men with azospermia and without Y micro-deletion). For this purpose, the exons of these 5 genes (DMC1, SPO11, MSH4, MSH5, CCNA1) were all amplified by PCR with specific primers and each amplified-exon was sequenced. Sequences were aligned in comparison to the human corresponding gene available in Genbank. Many heterozygous mutations were found in different genes. Two homozygous mutations were found in MSH4 and DMC1 genes in a young man presenting a testis vanishing syndrome and a woman presenting a premature ovarian failure, respectively. Consequences of such mutations will be examined and verified in model organisms (yeast, mouse) to check the relevance of the mutations in clinical setting.

Adenosine Triphosphatases↗
PubMed2002

A case of intersexuality in pigs associated with a de novo paracentric inversion 9 (p1.2; p2.2).

In several mammalian species, genetic defects can be responsible for the interruption of and/or the deviation from the sequential steps of normal gonadal differentiation, leading to a sex-reversal syndrome. In pigs, female-to-male sex-reversal conditions are particularly frequent, but their aetiologies remain unclear. Chromosomal abnormalities that co-occur with sex-reversal disorders can be useful in the identification of loci containing responsible or susceptibility genes. This report describes a female-to-male SRY-negative intersex pig with a de novo paracentric inversion of the short arm of one chromosome 9 (p1.2; p2.2). We have fine mapped the proximal chromosomal breakpoint of this rearrangement because it corresponded to a region potentially involved in the pig intersexuality. Fluorescent in situ hybridization (FISH) experiments carried out with Bacterial Artificial Chromosome (BAC) clones located within the critical region defined by genetic linkage analysis and ordered on the porcine RH map allowed us to locate the proximal breakpoint between markers SW2571 and SW539. Further investigations are currently in progress to find new markers inside this interval, in order to determine the BAC in which the break occurred.

Animals↗
PubMed2001

Contribution of domestic animals to the identification of new genes involved in sex determination.

Among farm animals, two species present an intersex condition at a relatively high frequency: pig and goat. Both are known to contain XX sex-reversed individuals which are genetically female but with a true hermaphrodite or male phenotype. It has been clearly demonstrated that the SRY gene is not involved in these phenotypes. Consequently, autosomal or X-linked mutations in the sex-determining pathway may explain these sex-reversed phenotypes. A mutation referred to as "polled" has been characterized in goats by the suppression of horn formation and abnormal sexual differentiation. The Polled Intersex Syndrome locus (PIS) was initially located in the distal region of goat chromosome 1. The homologous human region has been precisely identified as an HSA 3q23 DNA segment containing the Blepharophimosis Ptosis Epicanthus locus (BPES), a syndrome combining Premature Ovarian Failure (POF) and an excess of epidermis of the eyelids. In order to isolate genes involved in pig intersexuality, a similar genetic approach was attempted in pigs using genome scanning of resource families. Genetic analyses suggest that pig intersexuality is controlled multigenically. Parallel to this work, gonads of fetal intersex animals have been studied during development by light and electron microscopy. The development of testicular tissue and reduction of germ cell number by apoptosis, which simultaneously occurs as soon as 50 days post coïtum, also suggests that several separate genes could be involved in pig intersexuality.

Animals↗
PubMed2001

Testis determination in mammals: more questions than answers.

In humans, testis development depends on a regulated genetic hierarchy initiated by the Y-linked SRY gene. Failure of testicular determination results in the condition termed 46,XY gonadal dysgenesis (GD). Several components of the testis determining pathway have recently been identified though it has been difficult to articulate a cascade with the known elements of the system. It seems, however, that early gonadal development is the result of a network of interactions instead of the outcome of a linear cascade. Accumulating evidence shows that testis formation in man is sensitive to gene dosage. Haploinsufficiency of SF1, WT1 and SOX9 is responsible for 46,XY gonadal dysgenesis. Besides, data on SRY is consistent with possible dosage anomalies in certain cases of male to female sex reversal. 46,XY GD due to monosomy of distal 9p and 10q might also be associated with an insufficient gene dosage effect. Duplications of the locus DSS can lead to a failure of testicular development and a duplication of the region containing SOX9 has been implicated in XX sex reversal. Transgenic studies in mouse have shown, however, that this mammal is less sensitive to gene dosage than man. Here, we will try to put in place the known pieces of the jigsaw puzzle that is sex determination in mammals, as far as current knowledge obtained from man and animal models allows. We are certain that from this attempt more questions than answers will arise.

Animals↗
PubMed2001

Time course of female-to-male sex reversal in 38,XX fetal and postnatal pigs.

In an attempt to understand the etiology of intersexuality in pigs, we thoroughly analyzed the gonads of 38,XX (SRY negative) female to male sex-reversed animals at different developmental stages: during fetal life [50 and 70 days postcoitum (dpc)], just after birth [35 days postpartum (dpp)] and during adulthood. For each animal studied, we performed parallel histological and ultrastructural analyses on one gonad and RT-PCR analysis on the other gonad in order to define the expression profiles of sexually regulated genes: SOX9, 3beta-HSD, P450 aromatase, AMH, FOXL2, and Wnt4. Light and electron microscopic examination showed that testicular cords differentiated in XX sex-reversed gonads but were hypoplastic. Although the testicular cords contained gonia at the fetal stages, the germ cells had all died through apoptosis within a few weeks after birth. Ultrastructurally normal Leydig cells also differentiated, but later, and enclosed whorl-like residual bodies. At the fetal stages, three of the six genes studied in the intersex gonads presented, as early as 50 dpc, a modified expression profile corresponding to an elevated expression of SOX9 and the beginning of AMH and P450 aromatase gene transcription. In addition to genes involved in the testicular pathway, the same gonads expressed FOXL2, an ovarian-specific factor. The ovaries of true hermaphrodites were ineffective in ensuring correct folliculogenesis and presented abnormal expression profiles of ovarian specific genes after birth. These results indicate that the genes involved in this pathology act very early during gonadogenesis and affect the ovary-differentiating pathway with variable expressivity from ovarian germ cell depletion through to trans-differentiation into testicular structures.

Animals↗
PubMed2001

A 11.7-kb deletion triggers intersexuality and polledness in goats.

Mammalian sex determination is governed by the presence of the sex determining region Y gene (SRY) on the Y chromosome. Familial cases of SRY-negative XX sex reversal are rare in humans, often hampering the discovery of new sex-determining genes. The mouse model is also insufficient to correctly apprehend the sex-determination cascade, as the human pathway is much more sensitive to gene dosage. Other species might therefore be considered in this respect. In goats, the polled intersex syndrome (PIS) mutation associates polledness and intersexuality. The sex reversal affects exclusively the XX individuals in a recessive manner, whereas the absence of horns is dominant in both sexes. The syndrome is caused by an autosomal gene located at chromosome band 1q43 (ref. 9), shown to be homologous to human chromosome band 3q23 (ref. 10). Through a positional cloning approach, we demonstrate that the mutation underlying PIS is the deletion of a critical 11.7-kb DNA element containing mainly repetitive sequences. This deletion affects the transcription of at least two genes: PISRT1, encoding a 1.5-kb mRNA devoid of open reading frame (ORF), and FOXL2, recently shown to be responsible for blepharophimosis ptosis epicanthus inversus syndrome (BPES) in humans. These two genes are located 20 and 200 kb telomeric from the deletion, respectively.

Animals↗
PubMed2001

Establishment and characterization of immortalized ovine Sertoli cell lines.

The objective of this study was to generate immortalized Sertoli cell lines from prepubertal lamb testes to facilitate investigations during the course of testicular differentiation. The Sertoli cells were enzymatically isolated and immortalized by transfection, with the sequences coding for the SV40 large T-antigen fused downstream of regulatory elements from the human vimentin gene. The different cell lines were positively stained with antibodies to vimentin and transferrin, in agreement with their Sertoli origin. Reverse transcriptase polymerase chain reaction was used to analyze the specific expression of molecular markers (clusterin/sulfated glycoprotein ISGP-2], follicle-stimulating hormone [rFSH], alpha-inhibin, anti-Müllerian hormone, Wilms' tumor gene [WT-1], steroidogenic factor 1 [SF-1], SRY-related HMG box gene g [SOX9], and sex-determining region of Y chromosome) normally expressed in this cellular type. All were shown to express messenger ribonucleic acids for SGP-2, alpha-inhibin, WT-1, SOX9, and SF-1 (except SF-1 for clone no. 1). Moreover, we performed alkaline phosphatase and receptor tyrosine kinase p145 (c-kit) detection to ensure the absence of contamination by peritubular, germ cells, and Leydig cells. Both tests were negative for all the seven cell lines. These ovine Sertoli cell lines are the first ones obtained from livestock that exhibit specific Sertoli cell characteristics resembling different stages of phenotypic development. They provide useful in vitro model systems for toxicological investigations, coculture, and transfection experiments, making it possible to study signal transduction pathways, cell-cell interactions, and gene expression in species other than rodents.

Animals↗
PubMed2001

Mammalian gonadal differentiation: the pig model.

In mammals, testicular differentiation is initiated by SRY (the sex-determining region of the Y chromosome) gene expression in Sertoli cell precursors, followed by upregulation of the SOX9 gene (SRY-related HMG box gene 9). Subsequently, differentiated testis produces two hormones that induce sexual differentiation of the internal and external genital tract. Knowledge of the molecular mechanisms involved in gonadal differentiation has increased greatly over the past decade. Several genes are involved in genital ridge formation in both sexes, and others act specifically in testicular or ovarian developmental pathways. As for other mammals, relatively few data are available on the first steps of ovarian differentiation in pigs. In this review, the expression profiles of most genes known to be involved in gonadal differentiation in pigs will be presented and compared with those observed in mice. The main feature of gonadal differentiation in the pig is fetal steroidogenesis, especially cytochrome P450 aromatase gene organization and expression. Another specific feature of gonadal differentiation in pigs is the appearance of numerous cases of XX sex-reversed animals. This intersex condition occurs as early as day 50 after coitus, during embryogenesis, and appears to be triggered genetically. It leads to a wide range of phenotypes, strikingly similar to those observed in humans. Identification of the genes involved in this pathology will improve our knowledge of mammalian gonadal differentiation and may allow the eradication of this genetic disease in pigs.

Animals↗
PubMed1999

The xenotransplantation of goat and human hematopoietic cells to sheep fetuses.

Hematopoietic xenografts were carried out in three experiments using goat fetal liver (44-48 days, experiments I and II) or purified human CD 34+ cells (experiment III) as the donor cells. Recipients were sheep fetuses at 41-47 days of gestation. Goat fetal liver cells were either injected without any pretreatment or stimulated by preincubation in a culturing in goat phytohemagglutinin-stimulated lymphocyte supernatant. Human CD 34+ myeloid progenitor cells were purified from bone marrow by minimacs immunomagnetic purification and cultured in medium supplemented with stem cell factor, IL3, and IL6. Goat-sheep chimerism was assessed by flow cytometry analysis (FCA) of peripheral blood and bone marrow cells using a mouse anti-goat CD 45 monoclonal antibody and by karyotype analysis of peripheral blood from goat/sheep chimeras. Human cell engraftment was assessed by polymerase chain reaction amplification of the human DAX1 gene in blood and bone marrow DNA from sheep which had received human cells. In the three experiments, a mean of 76% (26 of 34) of injected fetuses were born alive without any clinical evidence of graft-versus-host disease. Three lambs were found to be goat/sheep chimeric after flow cytometry analysis (peripheral blood and bone marrow) and karyotype (peripheral blood) analysis. Both tissues continued to express goat cells at 6 or 12 months (last assessment) depending on the experiment. No human chimerism was detected using polymerase chain reaction amplification in peripheral blood and bone marrow of any of the six sheep grafted with human cells. These data and those also obtained on other species (human, pig/sheep) show that it is possible to carry out hematopoietic xenografts using the sheep fetus as recipient provided both donor and recipient fetal cells are processed during the period of tolerance to foreign antigens.

Aging↗
PubMed1999

High-resolution human/goat comparative map of the goat polled/intersex syndrome (PIS): the human homologue is contained in a human YAC from HSA3q23.

The genetic and cytogenetic map around the chromosome 1 region shown to be linked with polledness and intersexuality (PIS) in the domestic goat (Capra hircus) was refined. For this purpose, a goat BAC library was systematically screened with primers from human coding sequences, scraped chromosome 1 DNA, bovine microsatellites from the region, and BAC ends. All the BACs (n = 30) were mapped by fluorescence in situ hybridization (FISH) on goat chromosome 1q41-q45. The genetic mapping of 30 new goat polymorphic markers, isolated from these BACs, made it possible to reduce the PIS interval to a region of less than 1 cM on goat chromosome 1q43. The PIS locus is now located between the two genes ATP1B and COP, which both map to 3q23 in humans. Genetic, cytogenetic, and comparative data suggest that the PIS region is now probably circumscribed to an approximately 1-Mb DNA segment for which construction of a BAC contig is in progress. In addition, a human YAC contig encompassing the blepharophimosis-ptosis-epicanthus-inversus region was mapped by FISH to goat chromosome 1q43. This human disease, mapped to HSA 3q23 and affecting the development and maintenance of ovarian function, could be a potential candidate for goat PIS.

Animals↗
PubMed1999

The same mutation affecting the splicing of WT1 gene is present on Frasier syndrome patients with or without Wilms' tumor.

Denys-Drash and Frasier syndromes are rare human disorders that associate nephropathy with gonadal and genital abnormalities. In DDS there is a predisposition to Wilms' tumor. Heterozygous point mutations in the Wilms' tumor, type1 gene (WT1), particularly those altering the zinc finger (ZF) encoding exons, have been reported in most DDS patients, while mutations in intron 9 of the same gene cause FS. This paper describes two cases of DDS, one FS and one patient with Wilm's tumor and intersex genitalia, in which mutations were searched by sequencing the exons 8 and 9 of WT1 gene. Patient 1 carried a missense point mutation in exon 8 (ZF2), converting a CGA-Arg codon to a TGA-stop codon. Patient 2 presented a single nucleotide deletion within exon 9 (ZF3) introducing a premature chain termination at codon 398. Patients 3 and 4 had a C-->T transition at position +4 of the second alternative splice donor site of exon 9 (this mutation was detected in peripheral blood and in tumor derived DNA of patient 3). However, patient 3 had previously developed a Wilms' tumor. This is the first case of Wilms' tumor development in a phenotypically and genetically confirmed case of FS.

Adolescent↗
PubMed1999

Reverse transcription-polymerase chain reaction analysis of genes involved in gonadal differentiation in pigs.

In mammals, testis development is initiated in the embryo as a response to the expression of the sex-determining gene, SRY. The time course of SRY expression during gonadal differentiation in the male has been described in detail only in mice and sheep. In this study, we used reverse transcription-polymerase chain reaction analysis to define the SRY transcription profile in pig genital ridges. SRY transcripts were first detectable from 23 days postcoitum (dpc), then declined sharply after 35 dpc. None were detected at 60 dpc. In addition, we analyzed temporal expression of other genes known to be involved in mammalian sex determination: WT-1, SF-1, SOX9, and AMH. A key stage seems to be 28 dpc, in which SOX9 expression switches between the male and female, and AMH expression begins to attest to Sertoli cell differentiation and to correspond to seminiferous cord formation in the male. Expression of gonadotropin receptors and aromatase was also investigated in porcine gonads, and we showed that their transcripts were detected very early on, especially in the male: 25 dpc for the LH receptor (rLH) and aromatase, and 28 dpc for the FSH receptor (rFSH). In the female, aromatase transcripts were not detected until 70 dpc, and rFSH expression occurred later: at 45 dpc at the onset of meiosis. Moreover, no difference was observed between the sexes for the onset of rLH transcription at 25 dpc. Such a thorough study has never been performed on pigs; developmental analysis will be useful for investigating sex-reversed gonads and determining ontogeny in intersexuality, a common pathology in pigs.

Animals↗
PubMed1997

Porcine Dax-1 gene: isolation and expression during gonadal development.

The identification of XY females carrying a duplication of a region of the X chromosome (Xp21) led to the hypothesis that a double dose of a gene in the duplicated region causes sex reversal (DSS; dosage sensitive sex reversal). A gene isolated from this region, named DAX-1 (DSS-AHC critical region on the X), encodes a new member of the nuclear hormone receptor family. Here, we describe the isolation of porcine Dax-1 and the analysis of its pattern of expression both during foetal development and in several adult tissues. Dax-1 is expressed in the adrenals, the pituitary gland and the gonads at various stages of differentiation. In gonads, Dax-1 expression starts between 21 and 23 days post coitum in both XX and XY urogenital ridges then continues to be expressed until adult age. The expression in these tissues indicates the involvement of DAX-1 in the development and the function of the reproductive system at multiple levels.

Amino Acid Sequence↗
PubMed1997

The ovine SOX2 gene: sequence, chromosomal localization and gonadal expression.

The SOX gene family consists of a large number of embryonically expressed genes capable of encoding putative transcription factors and related by a DNA-binding domain, the HMG-box. We cloned and characterized the ovine SOX2 transcript using the screening of a testis (12dpp) cDNA library with a probe containing the SRY-HMG-box and we performed 3'RACE experiments. The ovine SOX2 sequence is strongly conserved in comparison to the human, mouse and chicken homologues and is located on sheep chromosome 1q33. The SOX2 expression pattern in developing gonads is consistent with the hypothesis that this gene plays a role in the germ cell line.

Amino Acid Sequence↗