Two redundant transcription factor binding sites in a single enhancer are essential for mammalian sex determination
Meshi Ridnik , Elisheva Abberbock, Veronica Alipov, Shelly Ziv Lhermann, Shoham Kaufman, Maor Lubman, Francis Poulat, Nitzan Gonen
Development and Pathology of the gonad
Genetics, cell biology and development
Reproductive development requires the highly coordinated and regulated determination/differentiation of embryonic gonads, and maturation of the reproductive organs. Abnormalities in these processes during early embryo development, due to genetic or environmental factors, will induce a wide spectrum of syndromes named “disorders of sexual development” (DSD), going from complete gonadal dysgenesis (sex reversal), gonadoblastoma to mild phenotypes as cryptorchidy (undescended testis).
Embryonic gonads originate from a bipotential primordium (the genital ridge) that can differentiate into a testis or an ovary via the action of distinct transcription factors. In the mouse embryo, sex determination occurs between embryonic day (E) 10.5 and E12.5 within the genital ridge by differentiation of progenitors into a specific cell population, the supporting cells. These cells will then organize the embryonic gonads morphologically and induce sex determination in the germinal lineage.
In males, differentiation of supporting cells, also known as Sertoli cells, is driven by the expression of the Y chromosome-encoded gene Sry, that activates the autosomal gene Sox9, a SOX E group family member. In the absence of Sry in females, progenitors differentiate into granulosa cells, the female supporting cell lineage. While Sry is the upper switch of the male cascade, Sox9 is considered as the major effector downstream of Sry in the Sertoli cell differentiation process. After its activation by Sry, maintenance of expression of the Sox9 gene is a key event in the mammalian sexual differentiation program. In the mouse XY gonad, Sox9 upregulates Fgf9, which initiates a Sox9/Fgf9 feedforward loop (for review (1)).
Our team identified the prostaglandin D2 (PGD2) (Figure 1) which is produced through both L- and H- prostaglandin D synthases (Ptgds1 and 2 respectively) and that stimulates Sox9 nuclear translocation (2) and Sox9mRNA expression, acting as a second amplification loop to accelerate and maintain the male pathway (3,4). The PGD2 signalling pathway will also contribute to the differentiation of the primordial male germ cells (5) and then to the regulation of the spermatogonial stem cell pool in the postnatal testis.
Figure 1: PGD2 signalling is involved in the somatic and germ cell differentiation in the embryonic testis
Prostaglandins synthesis is inhibited by Nonsteroidal anti-inflammatory drugs (NSAIDs) and analgesic drugs, such as paracetamol (acetaminophen: ACE), that are widely used by pregnant women. On the basis of the results of epidemiological studies, a potential link between DSD and exposure to such molecules during the first and second trimesters of gestation has been hypothesized. We found that embryonic gonads are targeted after in utero exposure at the time of the sex determining period to ACE, aspirin and/or ibuprofen therapeutic doses similar to those administered to humans. The anticipated differentiation of the male germline is induced, affecting the spermatogonial stem cell pool in the postnatal testis and the sperm count of the adult testis of the F1 and F2 generations and suggesting a transgenerational impact of these drugs.
As Sox9 is the major autosomal sex-determining gene in mammals and in the goal of identifying the genetic networks directly regulated by Sox9, we combined analysis of Sox9-bound chromatin regions from murine and bovine foetal testes with sequencing of RNA samples from mouse testes lacking Sox9. We found that Sox9 controls a conserved genetic programme that involves most of the sex-determining genes. In foetal testes, Sox9 modulates both transcription and directly or indirectly sex-specific differential splicing of its target genes through binding to genomic regions with sequence motifs that are conserved among mammals and that we called ‘Sertoli Cell Signature’ (SCS). The SCS is characterized by a precise organization of binding motifs for the Sertoli cell reprogramming factors Sox9, Gata4 and Dmrt1. Starting from the hypothesis that nuclear factors that bind to genomic regions with SCS could functionally interact with SOX9, we identified Trim28 (Figure 2) as a new Sox9 partner in foetal testes (6).
Figure 2: Super-resolution OMX fluorescence microscopy (0.12 µm Z-sections) shows the co-localization of Sox9 (red) and Trim28 (green) in the nucleus of E13.5 Sertoli cells (orange/yellow spots indicated by white arrows in the insets). DNA (blue). Sc: Sertoli cells; Gc: germ cells. Bar: 2µm.
References
- Wainwright, E.N. and Wilhelm, D. (2010) The Game Plan Cellular and Molecular Mechanisms of Mammalian Testis Development. Curr Top Dev Biol, 90C, 231-262.
- Malki, S., Nef, S., Notarnicola, C., Thevenet, L., Gasca, S., Mejean, C., Berta, P., Poulat, F. and Boizet-Bonhoure, B. (2005) Prostaglandin D2 induces nuclear import of the sex-determining factor SOX9 via its cAMP-PKA phosphorylation. Embo J, 24, 1798-1809.
- Moniot, B., Declosmenil, F., Barrionuevo, F., Scherer, G., Aritake, K., Malki, S., Marzi, L., Cohen-Solal, A., Georg, I., Klattig, J., Englert, C., Kim, Y., Capel, B., Eguchi, N., Urade, Y., Boizet-Bonhoure, B. and Poulat, F. (2009) The PGD2 pathway, independently of FGF9, amplifies SOX9 activity in Sertoli cells during male sexual differentiation. Development, 136, 1813-1821.
- Moniot, B., Farhat, A., Aritake, K., Declosmenil, F., Nef, S., Eguchi, N., Urade, Y., Poulat, F. and Boizet-Bonhoure, B. (2011) Hematopoietic prostaglandin D synthase (H-Pgds) is expressed in the early embryonic gonad and participates to the initial nuclear translocation of the SOX9 protein. Developmental dynamics : an official publication of the American Association of Anatomists, 240, 2335-2343.
- B. Moniot, S. Ujjan, J. Champagne, H. Hirai, K. Aritake, K. Nagata, E. Dubois, S. Nidelet, M. Nakamura, Y. Urade, F. Poulat and B. Boizet-Bonhoure. (2014). Prostaglandin D2 acts through the Dp2 receptor to influence male germ cell differentiation in the fetal mouse testis. Development, 141(18):3561-71. doi: 10.1242/dev.103408. Epub 2014 Aug 19.
- Rahmoun, M., Lavery, R., Laurent-Chaballier, S., Bellora, N., Philip, G.K., Rossitto, M., Symon, A., Pailhoux, E., Cammas, F., Chung, J., Bagheri-Fam, S., Murphy, M., Bardwell, V., Zarkower, D., Boizet-Bonhoure, B., Clair, P., Harley, V.R. and Poulat, F. (2017) In mammalian foetal testes, SOX9 regulates expression of its target genes by binding to genomic regions with conserved signatures. Nucleic Acids Res, 45, 7191-7211.
Publications of the team
Transposable elements acquire time-and sex-specific transcriptional and epigenetic signatures along mouse fetal gonad development
Isabelle Stevant Nitzan Gonen Francis Poulat
Cocktails of NSAIDs and 17α Ethinylestradiol at Environmentally Relevant Doses in Drinking Water Alter Puberty Onset in Mice Intergenerationally
Pascal Philibert, Stéphanie Déjardin, Mélissa Girard, Quentin Durix, Anne-Alicia Gonzalez, Xavier Mialhe, Mathieu Tardat, Francis Poulat, Brigitte Boizet-Bonhoure
TRIM28-dependent SUMOylation protects the adult ovary from activation of the testicular pathway
Moïra Rossitto , Stephanie Déjardin , Chris M Rands , Stephanie Le Gras , Roberta Migale , Mahmoud-Reza Rafiee , Yasmine Neirijnck , Alain Pruvost , Anvi Laetitia Nguyen , Guillaume Bossis , Florence Cammas , Lionel Le Gallic , Dagmar Wilhelm , Robin Lovell-Badge , Brigitte Boizet-Bonhoure , Serge Nef , Francis Poulat
Using Experimental Models to Decipher the Effects of Acetaminophen and NSAIDs on Reproductive Development and Health
B. Boizet-Bonhoure, S. Dejardin, M. Rossitto, F. Poulat and P. Philibert
In utero exposure to acetaminophen and ibuprofen leads to intergenerational accelerated reproductive aging in female mice
Rossitto M, Ollivier M, Déjardin S, Pruvost A, Brun C, Marchive C, Nguyen AL, Ghettas A, Keime C, de Massy B, Poulat F, Philibert P, Boizet-Bonhoure B
The Mouse Microbiome Is Required for Sex-Specific Diurnal Rhythms of Gene Expression and Metabolism.
Weger BD, Gobet C, Yeung J, Martin E, Jimenez S, Betrisey B, Foata F, Berger B, Balvay A, Foussier A, Charpagne A, Boizet-Bonhoure B, Chou CJ, Naef F, Gachon F
Microtubule polyglutamylation and acetylation drive microtubule dynamics critical for platelet formation.
van Dijk J, Bompard G, Cau J, Kunishima S, Rabeharivelo G, Mateos-Langerak J, Cazevieille C, Cavelier P, Boizet-Bonhoure B, Delsert C, Morin N
Intergenerational effects on mouse sperm quality after in utero exposure to acetaminophen and ibuprofen.
Rossitto M, Marchive C, Pruvost A, Sellem E, Ghettas A, Badiou S, Sutra T, Poulat F, Philibert P, Boizet-Bonhoure B
Sex reversal following deletion of a single distal enhancer of Sox9.
Gonen N, Futtner CR, Wood S, Garcia-Moreno SA, Salamone IM, Samson SC, Sekido R, Poulat F, Maatouk DM, Lovell-Badge R
A role for SOX9 in post-transcriptional processes: insights from the amphibian oocyte.
Penrad-Mobayed M, Perrin C, L'Hôte D, Contremoulins V, Lepesant JA, Boizet-Bonhoure B, Poulat F, Baudin X, Veitia RA
In mammalian foetal testes, SOX9 regulates expression of its target genes by binding to genomic regions with conserved signatures
Rahmoun M, Lavery R, Laurent-Chaballier S, Bellora N, Philip GK, Rossitto M, Symon A, Pailhoux E, Cammas F, Chung J, Bagheri-Fam S, Murphy M, Bardwell V, Zarkower D, Boizet-Bonhoure B, Clair P, Harley VR, Poulat F.
Development and pathology of the gonad
Boizet-Bonhoure, B.
Molecular events and signalling pathways of male germ cell differentiation in mouse
Rossitto M, Philibert P, Poulat F, Boizet-Bonhoure B.
The multiples roles of the Prostaglandin D2 signaling pathway in reproduction
Rossitto M, Ujjan S, Poulat F, Boizet-Bonhoure B
Prostaglandin D2 synthase/GPR44: a signaling axis in PNS myelination
Trimarco A, Forese MG, Alfieri V, Lucente A, Brambilla P, Dina G, Pieragostino D, Sacchetta P, Urade Y, Boizet-Bonhoure B, Boneschi FM, Quattrini A, Taveggia C.
Anatomical and Molecular Analyses of XY Ovaries from the African Pygmy Mouse Mus minutoides
Rahmoun M, Perez J, Saunders PA, Boizet-Bonhoure B, Wilhelm D, Poulat F, Veyrunes F.
Prostaglandin D2 acts through the Dp2 receptor to influence male germ cell differentiation in the foetal mouse testis
Moniot B, Ujjan S, Champagne J, Hirai H, Aritake K, Nagata K, Dubois E, Nidelet S, Nakamura M, Urade Y, Poulat F, Boizet-Bonhoure B
Unilateral Cryptorchidism in Mice Mutant for Ptgds
Philibert P, Boizet-Bonhoure B, Bashamboo A, Paris F, Aritake K, Urade Y, Leger J, Sultan C, Poulat F.
The Impressionistic Landscape of Meiotic Recombination
Lichten, M., de Massy, B.
Hematopoietic prostaglandin D synthase (H-Pgds) is expressed in the early embryonic gonad and participates to the initial nuclear translocation of the SOX9 protein
Moniot B, Farhat A, Aritake K, Declosmenil F, Nef S, Eguchi N, Urade Y, Poulat F, Boizet-Bonhoure B.
Inhibition of SRY-Calmodulin Complex Formation Induces Ectopic Expression of Ovarian Cell Markers in Developing XY Gonads.
Sim H, Argentaro A, Czech DP, Bagheri-Fam S, Sinclair AH, Koopman P, Boizet-Bonhoure B, Poulat F, Harley VR
Temporal and spatial SOX9 expression patterns in the course of gonad development of the caudate amphibian Pleurodeles waltl.
Dumond, H., Al-Asaad, I., Chesnel, A., Chardard, D., Boizet-Bonhoure, B., Flament, S., Kuntz, S.
Hematopoietic-Prostaglandin D2 synthase through PGD2 production is involved in the adult ovarian physiology
Farhat A, Philibert P, Sultan C, Poulat F, Boizet-Bonhoure B.
MiniSOX9, a dominant-negative variant in colon cancer cells
Abdel-Samad R, Zalzali H, Rammah C, Giraud J, Naudin C, Dupasquier S, Poulat F, Boizet-Bonhoure B, Lumbroso S, Mouzat K, Bonnans C, Pignodel C, Raynaud P, Fort P, Quittau-Prévostel C, Blache P
Relationship between glutathione S-transferase P1 polymorphisms and chronic obstructive pulmonary disease in a Tunisian population
Lakhdar R, Denden S, Knani J, Leban N, Daimi H, Hassine M, Lefranc G, Ben Chibani J, Haj Khelil A.
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IMGT® - the international ImMunoGeneTics information system®
Shuttling of SOX proteins
Malki S, Boizet-Bonhoure B, Poulat F
Unusual Virilization in Girls with Juvenile Granulosa Cell Tumors of the Ovary Is Related to Intratumoral Aromatase Deficiency.
Kalfa N, Méduri G, Philibert P, Patte C, Boizet-Bonhoure B, Thibaut E, Pienkowski C, Jaubert F, Misrahi M, Sultan C.
Shuttling of SOX proteins
Malki S, Boizet-Bonhoure B, Poulat F
The new molecular biology of granulosa cell tumors of the ovary.
Kalfa, N., Veitia, RA., Benayoun, BA., Boizet-Bonhoure, B., Sultan, C.
Is hypospadias a genetic, endocrine or environmental disease, or still an unexplained malformation?
Kalfa, N., Philibert, P., Sultan, C
Hypospadias and genetics
Kalfa, N., Philibert, P., Sultan, C.
The PGD2 pathway, independently of FGF9, amplifies SOX9activity in Sertoli cells during male sexual differentiation
Moniot, B., Declosmenil, F, Barrionuevo, F., Scherer, G., Aritake, K., Malki, S., Marzi, M., Cohen-Solal, A., Georg, I., Klattig, J., Englert, C., Kim, Y., Capel, B., Eguchi, N., Urade, Y., Boizet-Bonhoure, B., Poulat, F.
Juvenile granulosa-cell tumor: Clinical and molecular expression.
Kalfa N, Philibert P, Patte C, Thibaud E, Pienkowski C, Ecochard A, Boizet-Bonhoure B, Fellous M, Sultan C.
Male specific expression of Lipocalin-type Prostaglandin D Synthase (cPTGDS) during chicken gonadal differentiation: relationship with cSOX9
Moniot, B., Boizet-Bonhoure, B., Poulat, F.
La Prostaglandine D2: nouveaux rôles dans la gonade embryonnaire et pathologique.
Malki, S., Declosmenil, F., Farhat, A., Moniot, B., Poulat, F., Boizet-Bonhoure, B.
Granulosa cell tumor of the testis in children : evidence of localized intratesticular sex reversal through aberrant expression of FOXL2, an ovary-determining gene.
Kalfa, N., M. Fellous, B. Boizet-Bonhoure, C. Patte, P. Duvillard, C. Pienkowski, F. Jaubert, A. Ecochard and C. Sultan.
Sex-specific expression of SOX9 during gonadogenesis in the amphibian Xenopus tropicalis.
El Jamil, A., Kanhoush, R., Magre, S., Boizet-Bonhoure, B., Penrad-Mobayed, M.
The regulation of Sox9 gene expression by the GATA4/FOG2 transcriptional complex in dominant XX sex reversal mouse models
Manuylov, NL., Fujiwara, Y., Adameyko, II., Poulat, F., Tevosian, SG.
Expression and biological role of the prostaglandin D synthase / SOX9 pathway in Human Ovarian Cancer Cell
Malki, S., Bibeau, F., Notarnicola, C., Roques, S., Berta, P., Poulat, F., Boizet-Bonhoure, B.
Transcription factors involved in pancreas development are expressed in pediatric solid pseudopapillary tumors
Galmiche, L., Sarnacki, S., Verkarre, V., Boizet, B. Duvillie, B., Fabre, M., Jaubert, F
Coexpression of Wilms' tumor suppressor 1 (WT1) and androgen receptor (AR)in the genital tract of human male embryos and regulation of AR promoter activity by WT1
Köhler, B., Delezoide, AL., Boizet-Bonhoure, B., McPhaul, MJ., Sultan, C., Lumbroso, S
Fgf9 and Wnt4 act as antagonistic signals to regulate mammalian sex determination.
Kim, Y., Kobayashi, A., Sekido, R., DiNapoli, L., Brennan, J., Chaboissier, M.C., Poulat, F., Behringer, R.R., Lovell-Badge, R. and Capel, B.
Transient expression of SOX9 protein during follicular development in the adult mouse ovary.
Notarnicola C, Malki S, Berta P, Poulat F, Boizet-Bonhoure B.
Transient expression of SOX9 protein during follicular development in the adult mouse ovary.
Notarnicola C, Malki S, Berta P, Poulat F, Boizet-Bonhoure B.
Cytoplasmic retention of the sex-determining factor SOX9 via the microtubule network
Malki, S., Berta, P., Poulat, F., Boizet-Bonhoure, B.
Prostaglandin D2 induces nuclear import of the sex-determining factor SOX9 via its cAMP-PKA phosphorylation
Malki, S., Nef, S., Notarnicola, C., Thevenet, L., Gasca, S., Méjean, C., Berta, P., Poulat, F., Boizet-Bonhoure, B.
Gene expression during sex determination reveals a robust female genetic program at the onset of ovarian development.
Nef, S., Schaad, O., Stallings, N.R., Cederroth, C.R., Pitetti, J.L., Schaer, G., Malki, S., Dubois-Dauphin, M., Boizet-Bonhoure, B., Descombes, P., Parker, K.L. and Vassalli, J.D.
NHERF2/SIP-1 interacts with mouse SRY via a different mechanism than human SRY
Thevenet, L., Albrecht, K.H., Malki, S., Berta, P., Boizet-Bonhoure, B. and Poulat, F.
Regulation of human SRY subcellular distribution by its acetylation/deacetylation.
Thevenet, L., Mejean, C., Moniot, B., Bonneaud, N., Galeotti, N., Aldrian-Herrada, G., Poulat, F., Berta, P., Benkirane, M., Boizet-Bonhoure, B.
SOX9 specifies the pyloric sphincter epithelium through mesenchymal-epithelial signals.
Moniot, B., Biau, S., Faure, S., Nielsen, C.M., Berta, P., Roberts, D.J. and de Santa Barbara, P.
SOX9 interacts with a component of the human thyroid hormone receptor associated protein complex
Zhou, R., Bonneaud, N., Yuan, C., de Santa Barbara, P., Boizet, B., Schomber, T., Scherer, G., Roeder, G., Poulat, F., Berta, P.
Anosmin-1 is a regionally restricted component of basement membranes and interstitial matrices during organogenesis : implications for the developmental anomalies of X chromosomlinkes Kallmann syndrome.
Hardelin, J.P., Julliard, A.K., Moniot, B., Soussi-Yanicostas, N., Verney, C., Schwanzel, M., Ayer-Le-Lièvre, C. and Petit, C.
Thèses et hdr
Impact of in utero exposure to non-steroidal analgesics and anti-inflammatory drugs on early development and maturation of male reproductive organs in mice 09/12/2016
Defended by Moïra Rossitto on 09-12-2016
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