Opposing Pathways in Mammalian Sex Determination

描述(申请人提供)：性发育的病理在人类中很常见，反映了性别决定和性别分化的不稳定过程。性腺形成为一个双潜能器官，由可分化为睾丸或卵巢细胞类型的细胞组成。性别决定的主要步骤是发生在体细胞支持细胞谱系中的命运决定。我们之前已经证明，支持细胞最初是通过对立和拮抗的信号网络在支持(睾丸)和卵泡(卵巢)命运之间平衡的。在XY胚胎中，Y连锁基因Sry在支持细胞前体中的瞬时表达打破了这种平衡。我们对性腺转录组的全球分析显示，在XY双潜在性腺中有两个同时存在的网络，一男一女。这一数据表明，在男性命运决定过程中，男性性别决定既需要男性途径基因的激活，也需要潜在的女性途径的稳定抑制。这一观点得到了多梳蛋白M33/CBX2在小鼠或人类中功能丧失导致男性到女性性别逆转的发现的支持。在这一资助期间，我们发现Notch信号在快速分裂的细胞中产生支持细胞前体，而Numb定位于这些细胞的基底部，但对称分布于性腺内部的细胞中，这些细胞建立了SRY及其下游靶SOX9或女性支持细胞前体的最早标记FOXL2的表达。我们发现，无论男女，数量表达的支持细胞前体都是p27/p21阳性的，并在分化开始时滞留在细胞周期的G0/G1期。我们假设Numb通过多梳复合体(PRC1)的活性来调节细胞周期停滞，这是染色质重塑所必需的，以建立和/或维持支持细胞和颗粒细胞的命运。在这个提案中，我们将通过以下目标来确定性腺细胞如何接近和执行决定性别的细胞命运的决定：(1)确定Notch/Numb信号和细胞周期停滞在指定支持细胞谱系中的作用；(2)确定M33/PRC1是否是建立支持细胞谱系和维持支持或颗粒细胞命运所必需的。将细胞周期与细胞命运指定和表观遗传重编程相结合，将在器官发生和干细胞生物学领域具有广泛的相关性，并将为研究性发育和不孕症的发育基础提供洞察。

DESCRIPTION (provided by applicant): Pathologies of sexual development are common in humans, reflecting the precarious processes of sex determination and sexual differentiation. The gonad forms as a bipotential organ, composed of cells that can differentiate as testis or ovarian cell types. The primary sex-determining step is the fate decision that occurs in the somatic supporting cell lineage. We previously showed that supporting cells are initially balanced between Sertoli (testis) and follicle (ovary) fates by opposing and antagonistic signaling networks. In XY embryos, this balance is disrupted by the transient expression of the Y-linked gene Sry in supporting cell precursors. Our global analysis of the gonad transcriptome revealed two concurrent networks in the XY bipotential gonad, one male and one female. This data suggests that male sex determination requires both the activation of male pathway genes and the stable repression of the underlying female pathway during male fate determination. This idea is supported by the finding that loss of function of the polycomb protein M33/CBX2 in mice or humans results in male to female sex reversal. During this funding period, we found that Notch signals in rapidly dividing cells that give rise to the supporting cell precursors, whereas NUMB localizes basolaterally in these cells, but is symmetrically distributed in cells in the interior of the gonad that establish expression of either SRY and its downstream target SOX9, or the earliest marker of female supporting cell precursors, FOXL2. We found that NUMB-expressing supporting cell precursors of both sexes are p27/p21-positive and arrested in G0/G1 of the cell cycle at the stage when differentiation begins. We hypothesize that Numb mediates cell cycle arrest, which is required for chromatin remodeling through the activity of the polycomb complex (PRC1) to establish and/or maintain Sertoli and granulosa cell fate. In this proposal, we will determine how gonadal cells approach and execute the sex-determining cell fate decision through the following aims: (1) Determine the role of Notch/Numb signaling and cell cycle arrest in specification of the supporting cell lineage; (2) Determine whether M33/PRC1 is required to establish the supporting cell lineage and to maintain Sertoli or granulosa cell fate. Integration of the cell cycle with cell fate specification and epigenetic reprogramming will have broad relevance to the fields of organogenesis and stem cell biology and will provide insight into the developmental basis for disorders of sexual development and infertility.