WNT4 and RSPO1 together are required for cell proliferation in the early mouse gonad

Abstract

SUMMARYThe gonad arises from the thickening of the coelomic epithelium and then commits into the sex determination process. Testis differentiation is activated by the expression of the Y-linked gene Sry, which promotes cell proliferation and differentiation of Sertoli cells, the supporting cells of the testis. In absence of Sry (XX individuals), activation of WNT/CTNNB1 signalling, via the upregulation of Rspo1 and Wnt4, promotes ovarian differentiation. However, Rspo1 and Wnt4 are expressed in the early undifferentiated gonad of both sexes, and Axin2-lacZ, a reporter of canonical WNT/CTNNB1 signalling, is expressed in the coelomic region of the E11.5 gonadal primordium, suggesting a role of these factors in early gonadal development. Here, we show that simultaneous ablation of Rspo1 and Wnt4 impairs proliferation of the cells of the coelomic epithelium, reducing the number of progenitors of Sertoli cells in XY mutant gonads. As a consequence, in XY Wnt4 -/-; Rspo1 -/-foetuses, this leads to the differentiation of a reduced number of Sertoli cells and the formation of a hypoplastic testis exhibiting few seminiferous tubules. Hence, this study identifies Rspo1 and Wnt4 as two new regulators of cell proliferation in the early gonad regardless of its sex, in addition to the specific role of these genes in ovarian differentiation. Jeays-Ward et al., 2004;Kim et al., 2006;Vainio et al., 1999). Similarly, Rspo1 is specifically upregulated in XX foetal gonads at E11.5 Parma et al., 2006). Loss of function of either Rspo1 or Wnt4 in XX gonads promotes: (1) ectopic steroidogenic precursors, endothelial cell migration and the formation of a coelomic vessel (Chassot et al., 2008b;Jeays-Ward et al., 2003); and (2) sex reversal of the supporting cell lineages with expression of the Sertoli cell markers, SOX9 or Dhh, and the development of ovotestes around birth (Chassot et al., 2008b;Tomizuka et al., 2008;Vainio et al., 1999). WNTs and R-spondins act synergistically by interaction of their respective receptors LRP5/6 and LGR4/5 (Carmon et al., 2011;de Lau et al., 2011;Glinka et al., 2011). R-spondin binding activates WNT/CTNNB1 signalling and can disrupt WNT/planar cell polarity signalling (Hao et al., 2012). How RSPO1 and WNT4 interact in the gonad remains to be elucidated; however, in absence of Rspo1 and its effector CTNNB1, Wnt4 upregulation is impaired, indicating that Rspo1 is required for Wnt4 upregulation in the ovary after 11.5 dpc (Chassot et al., 2008b;Liu et al., 2009;Manuylov et al., 2008;Tomizuka et al., 2008). WNT/CTNNB1 signalling is required for Foxl2 upregulation (Manuylov et al., 2008), which in turn promotes follicular differentiation (Ottolenghi et al., 2005;Schmidt et al., 2004) and homeostasis in the ovaries (Uhlenhaut et al., 2009). In addition to its role in ovarian development, Wnt4 is also involved in testis differentiation. Indeed, in XY gonads, loss of function of Wnt4 induces a delay in sex cord formation that is compensated at birth (Jeays-Ward et al., 2004). By contrast, Rspo1-/-males are nor...

R-spondin1 is a novel hormone mediator for mammary stem cell self-renewal

“…Indeed, it has been proposed that Wnt4 can act on luminal cells (Hiremath et al 2007;Joshi et al 2010). Recent studies have demonstrated that Rspo1 and Wnt4 cooperate during early gonad proliferation in ovary differentiation (Chassot et al 2008(Chassot et al , 2012Tomizuka et al 2008). It will be interesting to understand whether such cooperation mediates the influence of hormones in these processes.…”

From Sex Determination to Initial Folliculogenesis in Mammalian Ovaries: Morphogenetic Waves along the Anteroposterior and Dorsoventral Axes

“…LGR5-positive cells are restricted to the cortical region of the ovaries throughout the fetal and perinatal stages, and are fated to differentiate into granulosa cells of several growing follicles even in the first folliculogenesis [Rastetter et al, 2014; review by Hummitzsch et al, 2015]. As described above, the proliferation and expansion of coelomic epithelium is induced by RSPO1/WNT4/ CTNNB1 signals in the initial stages of gonadogenesis [Chassot et al, 2012]. This appears to occur continuously in the later stages (i.e.…”

“…Two key factors secreted by the ovary, wingless-type MMTV integration site family member 4 (WNT4) and R-spondin homolog RSPO1, a ligand of the LGR5 receptor that regulates the WNT/ CTNNB1 (β-catenin) signaling pathway [Carmon et al, 2011;de Lau et al, 2011], synergistically regulate the thickening of the coelomic epithelium and its subsequent expansion toward the subepithelial region in both XY and XX gonads [Chassot et al, 2012]. RSPO1/WNT4/ CTNNB1 signals have been shown to antagonize the masculinizing factors SOX9 and FGF9 downstream of Sry [Kim et al, 2006;Hiramatsu et al, 2009;Jameson et al, 2012;Nicol and Yao, 2015].…”

“…This is because, after E12.5, somatic cells in the XY gonads are separated from the coelomic epithelium by the testis-specific formation of the tunica albuginea with vasculatures [Karl and Capel, 1998;Mork et al, 2012]. Also considering the high-level activation of canonical WNT/ CTNNB1 signaling in the E11.5-12.5 coelomic epithelia in both XY and XX gonads [Chassot et al, 2011[Chassot et al, , 2012, these data suggest that RSPO1/WNT4/CTNNB1 signals play crucial roles in the expansion of proliferative coelomic epithelial cells in both XY and XX gonads until E12.5. Thereafter, the signals continuously appear to induce expansion of the cortical region only in XX gonads in a sexually dimorphic manner.…”

Primordial germ cell migration and the Wnt signaling pathway

“…Several Wnt ligands have been implicated in PGC development in the mouse: Wnt3 and Wnt3a in specification (Ohinata et al, 2009;Bialecka et al, 2012;Aramaki et al, 2013;Tanaka et al, 2013), Wnt5a, and its receptor Ror2, in migration (Laird et al, 2011;Chawengsaksophak et al, 2012), and Wnt4 in female sex-differentiation (Vainio et al, 1999;Chassot et al, 2012). Both the specification and sex-differentiation of PGCs utilize the β-catenin-dependent, canonical arm of the Wnt pathway (Chassot et al, 2008(Chassot et al, , 2011Aramaki et al, 2013), while migration is regulated by the noncanonical Wnt pathway (Laird et al, 2011;Chawengsaksophak et al, 2012).…”