Molecular mechanisms governing the meiosis entry in mammals – molmechmeiosis

For this study, the acquisition of XX and XY GC sexual identity driven by the surrounding somatic tissues, the induction or prevention of meiosis entry, as well as the post-natal recovery of GC proliferation are dissected in multiple mouse lines bearing either germinal or somatic gene ablations of key components relaying these pathways. These developmental processes are investigated by means of morphological approaches such as histology, immuno-histochemistry, in situ hybridization, but also through more sophisticated, holistic, molecular approaches such as global micro-RNA profiling, transcriptome analysis using DNA microarrays and high throughput parallel sequencing of RNA (RNA-Seq).

Partner 1 has shown that PGD2 signalling, acting through DP2 receptor, has an essential role in the control of proliferation and differentiation of male germ cells (GC) via an activation of the mitotic block and the inhibition meiosis entry (manuscript in preparation).

Partner 2 has shown that the RSPO1/ß-catenin pathway is involved in the control of germ cell differentiation XX (Chassot et al., 2011). Moreover, partner 2 results show that RSPO1 regulates cell proliferation of precursor of supporting cells (Chassot et al., Revision) and clarify the role of RSPO1 in sex reversal (Lavery et al., In revision).

Partner 3 showed that the homeobox genes Msx are required for the development of fetal female germ cells (Le Bouffant et al. Development). Mice deficient for the gene Riken described in the project are sterile, because of a block in the early stages of meiosis. Finally, the partner 3 has identified Nodal as an autocrine factor able to block the meiosis entry in the fetal testis (Souquet et al. Endocrinology 2012).

Partner 4 showed that (i) MAFB is a direct target of RARA in Sertoli cell, and that the retinoic acid (RA) from Sertoli cells is required for the differentiation of spermatogonia responsible of the first wave of spermatogenesis (Raverdeau et al. , 2012 Proc. Natl. Acad. Sci. USA in press), (ii) initiation of meiosis requires the RA produced by spermatocytes (Raverdeau et al., 2012 Proc. Natl. Acad. Sci. U.S. in press), (iii) RARG in spermatogonia controls the transition between undifferentiated spermatogonia and differentiated spermatogonia (Gély-Pernot et al., 2012 Endocrinology 153 (1) :438-49) (iv) RARG operates as a heterodimer with RXR to control genes responsible of spermatogonia differentiation, including SALL4 (manuscript in preparation).

The purpose of the Molmechmeiosis project is to identify signals produced by GC or somatic cells surrounding the GC able to induce either meiosis entry in the foetal ovary and post-natal testis, either preventing meiosis in the foetal testis. Our goal is to integrate the different signalling pathways studied in this project to build a scheme describing the mechanisms controlling the meiosis entry.

1. Chassot et al. (2011). PLoS ONE 6 : e25641. (Partner 2)
2. Chassot et al. (2012). Development, en révision (Partner 2).
3. Gely-Pernot et al. (2012) Endocrinology 153(1):438-49 (Partner 4)
4. Raverdeau et al. (2012) Proc. Natl. Acad. Sci. USA in press (Partner 4)
5. Le Bouffant et al (2011) Development 138(24):5393-402 (Partner 3)
6. Souquet et al (2012) Endocrinology 153(5):2466-73 (Partner 3)

Central to the process of germ cell (GC) differentiation, meiosis allows the mixing of genetic material between male (XY) and female (XX) individuals. In mammals, XX GC meiosis is initiated during foetal life, at a time when XY GC become quiescent in the G0/G1 phase of their cell-cycle. Then in females, the number of gametes (oocytes) is determined at birth, whereas in males, XY GC proliferation resumes post-natally, yielding spermatogonia stem cells (SSC), which subsequently insure the continuous production of gametes during all the life-time of the individual. The molecular mechanisms driving these gender differences in GC proliferation and commitment to meiosis are not fully understood, but the somatic environment proves to play a critical role in this process, independently of the chromosomal content of GC. In Molmechmeiosis project, we propose to identify yet uncovered cascades and to decipher the genetic and molecular interactions set up between several signalling pathways, which all appear required for somatic sexual determination and GC commitment to meiosis, namely the prostaglandin D2 (PGD2), fibroblast growth-factor 9 (FGF9), MSX homeogene, NODAL, R-spondin (RSPO1), beta-catenin (CTNNB1) and retinoic acid (RA) dependent pathways. To reach this goal, the acquisition of XX and XY GC sexual identity driven by the surrounding somatic tissues, the induction or prevention of meiosis entry, as well as the post-natal recovery of GC proliferation are being dissected in multiple mouse lines bearing either germinal or somatic gene ablations of key components relaying these pathways. These developmental processes are being investigated by means of morphological approaches such as histology, immuno-histochemistry, in situ hybridization, but also through more sophisticated, holistic, molecular approaches such as global micro-RNA profiling, transcriptome analysis using DNA microarrays and high throughput parallel sequencing of RNA (RNA-Seq), as well as through an innovative in vivo tandem-affinity purification tag-facilitated immuno-precipitation of nuclear complexes coupled to mass-spectrometry aimed at identifying RA-receptor interacting proteins. Collectively, the broad combination of both state-of the art and cutting-edge methodologies will allow us (i) elucidating the role played by PDG2, FGF9 and NODAL-dependent signalling pathways in the mitotic arrest and meiosis inhibition occurring in foetal XY GC, (ii) identifying the molecular factors downstream of MSX and RSPO1/CTNNB1 allowing XX GC to become competent for meiosis, and (iii) uncovering the genetic networks and characterizing the molecular mechanism through which CTNNB1 and RA control XY GC and SSC proliferation, as well as entry in meiosis in the post-natal testis. Overall, the combination of our respective expertises in the Molmechmeiosis project offers a unique opportunity to unravel fundamental genetic networks and molecular cascades governing the sexual fate of GC.