Gene expression regulation in the context of mouse interspecific mosaic genomes

The testis transcriptome of mouse strains containing homozygous segments of Mus spretus origin in a Mus musculus background was analyzed

Fidgetin-Like1 Is a Strong Candidate for a Dynamic Impairment of Male Meiosis Leading to Reduced Testis Weight in Mice

Chantier qualité GAInternational audienceBACKGROUND: In a previous work, using an interspecific recombinant congenic mouse model, we reported a genomic region of 23 Mb on mouse chromosome 11 implicated in testis weight decrease and moderate teratozoospermia (∼20-30%), a Quantitative Trait Locus (QTL) called Ltw1. The objective of the present study is to identify the gene underlying this phenotype. RESULTS: In the present study, we refined the QTL position to a 5 Mb fragment encompassing only 11 genes. We showed that the low testis weight phenotype was due to kinetic alterations occurring during the first wave of the spermatogenesis where we could point out to an abnormal lengthening of spermatocyte prophase. We identify Fidgetin-like 1 (Fignl1) as the gene underlying the phenotype, since if fulfilled both the physiological and molecular characteristics required. Indeed, amongst the 11 positional candidates it is the only gene that is expressed during meiosis at the spermatocyte stage, and that presents with non-synonymous coding variations differentiating the two mouse strains at the origin of the cross. CONCLUSIONS: This work prompted us to propose Fignl1 as a novel actor in mammal's male meiosis dynamics which has fundamental interest. Besides, this gene is a new potential candidate for human infertilities caused by teratozoospermia and blockades of spermatogenesis. In addition this study demonstrates that interspecific models may be useful for understanding complex quantitative traits

Functional Exploration of the Adult Ovarian Granulosa Cell Tumor-Associated Somatic FOXL2 Mutation p.Cys134Trp (c.402C>G)

International audienceBACKGROUND: The somatic mutation in the FOXL2 gene c.402C>G (p.Cys134Trp) has recently been identified in the vast majority of adult ovarian granulosa cell tumors (OGCTs) studied. In addition, this mutation seems to be specific to adult OGCTs and is likely to be a driver of malignant transformation. However, its pathogenic mechanisms remain elusive. METHODOLOGY/PRINCIPAL FINDINGS: We have sequenced the FOXL2 open reading frame in a panel of tumor cell lines (NCI-60, colorectal carcinoma cell lines, JEG-3, and KGN cells). We found the FOXL2 c.402C>G mutation in the adult OGCT-derived KGN cell line. All other cell lines analyzed were negative for the mutation. In order to gain insights into the pathogenic mechanism of the p.Cys134Trp mutation, the subcellular localization and mobility of the mutant protein were studied and found to be no different from those of the wild type (WT). Furthermore, its transactivation ability was in most cases similar to that of the WT protein, including in conditions of oxidative stress. A notable exception was an artificial promoter known to be coregulated by FOXL2 and Smad3, suggesting a potential modification of their interaction. We generated a 3D structural model of the p.Cys134Trp variant and our analysis suggests that homodimer formation might also be disturbed by the mutation. CONCLUSIONS/SIGNIFICANCE: Here, we confirm the specificity of the FOXL2 c.402C>G mutation in adult OGCTs and begin the exploration of its molecular significance. This is the first study demonstrating that the p.Cys134Trp mutant does not have a strong impact on FOXL2 localization, solubility, and transactivation abilities on a panel of proven target promoters, behaving neither as a dominant-negative nor as a loss-of-function mutation. Further studies are required to understand the specific molecular effects of this outstanding FOXL2 mutation

SUMOylation of the Forkhead Transcription Factor FOXL2 Promotes Its Stabilization/Activation through Transient Recruitment to PML Bodies

International audienceBACKGROUND: FOXL2 is a transcription factor essential for ovarian development and maintenance. It is mutated in the genetic condition called Blepharophimosis Ptosis Epicantus inversus Syndrome (BPES) and in cases of isolated premature ovarian failure. We and others have previously shown that FOXL2 undergoes several post-translational modifications. METHODS AND PRINCIPAL FINDINGS: Here, using cells in culture, we show that interference with FOXL2 SUMOylation leads to a robust inhibition of its transactivation ability, which correlates with a decreased stability. Interestingly, FOXL2 SUMOylation promotes its transient recruitment to subnuclear structures that we demonstrate to be PML (Promyelocytic Leukemia) Nuclear Bodies. Since PML bodies are known to be sites where post-translational modifications of nuclear factors take place, we used tandem mass spectrometry to identify new post-translational modifications of FOXL2. Specifically, we detected four phosphorylated, one sulfated and three acetylated sites. CONCLUSIONS: By analogy with other transcription factors, we propose that PML Nuclear Bodies might transiently recruit FOXL2 to the vicinity of locally concentrated enzymes that could be involved in the post-translational maturation of FOXL2. FOXL2 acetylation, sulfation, phosphorylation as well as other modifications yet to be discovered might alter the transactivation capacity of FOXL2 and/or its stability, thus modulating its global intracellular activity

Interspecific resources: a major tool for quantitative trait locus cloning and speciation research.

International audiencePositional cloning of the quantitative trait locus (QTL) still encounters numerous difficulties, which explains why thousands of QTL have been mapped, while only a few have been identified at the molecular level. Here, we focus on a specific mapping tool that exists in plant and animal model species: interspecific recombinant congenic strains (IRCSs) or interspecific nearly isogenic lines (NILs). Such panels exhibit a much higher sequence diversity than intraspecific sets, thus enhancing the contrasts between phenotypes. In animals, it allows statistical significance to be reached even when using a limited number of individuals. Therefore, we argue that interspecific resources may constitute a major genetic tool for positional cloning and for understanding some bases of speciation mechanisms

Interspecific resources: A major tool for quantitative trait locus cloning and speciation research

Positional cloning of the quantitative trait locus (QTL) still encounters numerous difficulties, which explains why thousands of QTL have been mapped, while only a few have been identified at the molecular level. Here, we focus on a specific mapping tool that exists in plant and animal model species: interspecific recombinant congenic strains (IRCSs) or interspecific nearly isogenic lines (NILs). Such panels exhibit a much higher sequence diversity than intraspecific sets, thus enhancing the contrasts between phenotypes. In animals, it allows statistical significance to be reached even when using a limited number of individuals. Therefore, we argue that interspecific resources may constitute a major genetic tool for positional cloning and for understanding some bases of speciation mechanisms. © 2010 Wiley Periodicals, Inc

Interspecific resources: A major tool for quantitative trait locus cloning and speciation research

Positional cloning of the quantitative trait locus (QTL) still encounters numerous difficulties, which explains why thousands of QTL have been mapped, while only a few have been identified at the molecular level. Here, we focus on a specific mapping tool that exists in plant and animal model species: interspecific recombinant congenic strains (IRCSs) or interspecific nearly isogenic lines (NILs). Such panels exhibit a much higher sequence diversity than intraspecific sets, thus enhancing the contrasts between phenotypes. In animals, it allows statistical significance to be reached even when using a limited number of individuals. Therefore, we argue that interspecific resources may constitute a major genetic tool for positional cloning and for understanding some bases of speciation mechanisms. © 2010 Wiley Periodicals, Inc

Identification of a pituitary ERα-activated enhancer triggering the expression of Nr5a1, the earliest gonadotrope lineage-specific transcription factor

International audienceBackground: Gonadotrope lineage differentiation is a stepwise process taking place during pituitary development. The early step of gonadotrope lineage specification is characterized by the expression of the Nr5a1 transcription factor , a crucial factor for gonadotrope cell fate determination. Abnormalities affecting Nr5a1 expression lead to hypo-gonadotropic hypogonadism and infertility. Although significant knowledge has been gained on the signaling and transcriptional events controlling gonadotrope differentiation, epigenetic mechanisms regulating Nr5a1 expression during early gonadotrope lineage specification are still poorly understood. Results: Using ATAC chromatin accessibility analyses on three cell lines recapitulating gradual stages of gonadotrope differentiation and in vivo on developing pituitaries, we demonstrate that a yet undescribed enhancer is transiently recruited during gonadotrope specification. Using CRISPR/Cas9, we show that this enhancer is mandatory for the emergence of Nr5a1 during gonadotrope specification. Furthermore, we identify a highly conserved estrogen-binding element and demonstrate that the enhancer activation is dependent upon estrogen acting through ERα. Lastly, we provide evidence that binding of ERα is crucial for chromatin remodeling of Nr5a1 enhancer and promoter, leading to RNA polymerase recruitment and transcription. Conclusion: This study identifies the earliest regulatory sequence involved in gonadotrope lineage specification and highlights the key epigenetic role played by ERα in this differentiation process

The transcription factor FOXL2: At the crossroads of ovarian physiology and pathology.

International audienceFOXL2 is a gene encoding a forkhead transcription factor. Its mutations or misregulation have been shown to cause the blepharophimosis-ptosis-epicanthus inversus (BPES) syndrome and more recently have been associated with the development of Ovarian Granulosa Cell Tumors (OGCT). BPES is a genetic disorder involving mild craniofacial abnormalities often associated with premature ovarian failure. OGCTs are endocrine malignancies, accounting for 2-5% of ovarian cancers, the treatment of which is still challenging. In this review we summarize recent data concerning FOXL2 transcriptional targets and molecular partners, its post-translational modifications, its mutations and its involvement in newly discovered pathophysiological processes. In the ovary, FOXL2 is involved in the regulation of cholesterol and steroid metabolism, apoptosis, reactive oxygen species detoxification and cell proliferation. Interestingly, one of the main roles of FOXL2 is also to preserve the identity of ovarian granulosa cells even at the adult stage and to prevent their transdifferentiation into Sertoli-like cells. All these recent advances indicate that FOXL2 is central to ovarian development and maintenance. The elucidation of the impact of FOXL2 germinal and somatic mutations will allow a better understanding of the pathogenesis of BPES and of OGCTs

GnRH regulates the expression of its receptor accessory protein SET in pituitary gonadotropes.

Reproductive function is under the control of the neurohormone GnRH, which activates a G-protein-coupled receptor (GnRHR) expressed in pituitary gonadotrope cells. GnRHR activates a complex signaling network to regulate synthesis and secretion of the two gonadotropin hormones, luteinizing hormone and follicle-stimulating hormone, both regulating gametogenesis and steroidogenesis in gonads. Recently, in an attempt to identify the mechanisms underlying GnRHR signaling plasticity, we identified the first interacting partner of GnRHR, the proto-oncogene SET. We showed that SET binds to intracellular domains of GnRHR to enhance its coupling to cAMP pathway in αT3-1 gonadotrope cells. Here, we demonstrate that SET protein is rapidly regulated by GnRH, which increases SET phosphorylation state and decreases dose-dependently SET protein level. Our results highlight a post-translational regulation of SET protein involving the proteasome pathway. We determined that SET phosphorylation upon GnRH stimulation is mediated by PKC and that PKC mediates GnRH-induced SET down-regulation. Phosphorylation on serine 9 targets SET for degradation into the proteasome. Furthermore, a non-phosphorylatable SET mutant on serine 9 is resistant to GnRH-induced down-regulation. Altogether, these data suggest that GnRH-induced SET phosphorylation on serine 9 mediates SET protein down-regulation through the proteasome pathway. Noteworthy, SET down-regulation was also observed in response to pulsatile GnRH stimulation in LβT2 gonadotrope cells as well as in vivo in prepubertal female mice supporting its physiological relevance. In conclusion, this study highlights a regulation of SET protein by the neurohormone GnRH and identifies some of the mechanisms involved