I began this series of posts as a background with the goal of discussing Polycystic Ovary Syndrome in trans men. I thought that in order to understand what happens when things go wrong in ovarian function, we first need to talk about what happens when things go right.
So first, we had a discussion about where ovaries come from, and we learned that the gonads arise from the genital ridges, indifferent embryonic structures that are bi-potential. The genital ridges can become ovaries or testes depending on the genetic signals that push their development.
In addition, we discussed how the nascent gonads are populated by primordial germ cells (PGCs) that migrate from the yolk sac. PGCs can become either oogonia or spermatogonia, again, depending on the genes that drive their differentiation.
In support of the young germ cells are undifferentiated somatic (i.e. non-germ) cells that eventually become supportive Sertoli cells in the testis and granulosa cells in the ovary. In the embryonic testis, the early Sertoli cells encase the germ cells in sex cords which eventually develop into seminiferous tubules, which is where the sperm cells develop in the adult. In the ovary, the pre-granulosa cells form follicles by joining in a single layer to encapsulate the germ cells, which then enter meiosis, subsequently becoming arrested part way through the process.
All in all, gonadal development is a relatively intriguing phenomenon when you think about it. The primordial gonads are bi-potential and can develop into either ovaries or testes. And the germ cells don’t even arise from within the nascent gonads! They migrate in from outside the embryo, and are also bi-potential — they can become immature eggs or sperm. These steps require genetic orchestration and direction, where some signaling pathways are turned on while others are inhibited — a genetic balancing act, so to speak.
So how does this all work? And what happens when things don’t go according to plan?
In this post, we will learn about the genes and signaling pathways responsible for sex determination of the gonads. In a subsequent post, we will talk about what happens when the genes and signals that drive gonadal development and differentiation deviate from the “master plan.” In those cases, the results are intersex conditions, also known as disorders of sexual development (DSDs).
This series of posts is about the ovary, with the goal of talking about Polycystic Ovary Syndrome in trans men. But in order to understand ovarian development, it’s easier to first talk about development of the testes, because it’s the absence or inhibition of signals that occur in testicular differentiation that contribute, in part, to ovarian development.
Differentiation of the Embryonic Testis
Male sex determination, as we currently know it, begins when the somatic cells differentiate into the supportive Sertoli cells by expressing the gene SRY, which, in humans, begins at 40-50 days of gestation. The young Sertoli cells proliferate and form the sex cords around the primordial germ cells (PGCs). The germ cells differentiate into spermatogonia through contact with the Sertoli cells.
The gene SRY, called sex-determining region Y, was discovered in humans and mice in 1990 (Gubbay et al., 1990; Sinclair et al., 1990). Located on the Y-chromosome, the SRY gene encodes a protein that contains sequences that both target it to the nucleus and allow it to bind to a particular DNA sequence. This type of protein is a transcription factor in that it can turn genes on by binding to the DNA of the promoter sequences of other genes and initiate transcription of the genes’ DNA into RNA.
In essence, SRY is a master switch — it’s expression sets off a series of events that includes turning on a host of other genes and induces differentiation of cells in the genital ridge to go down the male path, ultimately resulting in the production of hormones that direct male sexual differentiation and development, including regression of the female Müllerian ducts, production of testosterone, development of the male Wolfian ducts and development of secondary sex glands and sex characteristics (reviewed in Ostrer, 2003 and Wainright & Wilhelm, 2010). I have discussed some of these steps in another post.
[We know that there are genes and signaling factors upstream of the SRY gene that regulate its expression and turn it on when it's time for the bipotential gonad to commit to differentiation, but I will not go into those details here. Readers are invited to read the brief review in Wainwright & Wilhelm (2010) for that information.]
So there is SRY, the master switch for male sex determination — its expression in the early Sertoli cells of the human embryonic testis reaches a peak at the 44th day of gestation and then declines to low levels up until at least 18 weeks of gestation (Hanley et al., 2000). Its expression turns on the gene SOX9 (SRY-related HMG BOX gene 9; Sekido & Lovell-Badge, 2008) in Sertoli cells.
The expression timing and pattern of SOX9 in the developing testis is similar to that of SRY, beginning between Days 41 to 44 of gestation and continuing until at least 18 weeks (Hanley et al., 2000). Also similar to SRY, the protein encoded by the gene SOX9 is a transcription factor that binds to DNA and turns on other genes. Two important products of SOX9 up-regulation during male sex determination are FGF9 (fibroblast growth factor 9) and PGD2 (prostaglandin D2).
FGF9 and PGD2 feedback to the early Sertoli cells and reinforce their development, and also signal to adjacent somatic cells in the nascent testis and induce their proliferation and development to Sertoli cells (reviewed in Nef & Vasselli, 2009 and DiNapoli & Capel, 2008). These signals result in expansion of the supporting cells and growth of the developing testis. In addition, these two factors inhibit “female” signals that would induce ovarian differentiation and development.
[FGF9 belongs to a 22-member family of proteins that have quite a list of effects on processes in both the developing embryo/fetus and adult. PGD2 is a member of a family of lipid signaling molecules that are formed through the action of cyclooxygenase enzymes. Both factors play important roles in normal physiology and disease. Interested readers can learn about them in free reviews by Krejci et al., 2009 and Smyth et al., 2009.]
Differentiation of the Embryonic Ovary
When I was a baby scientist, I was taught that there were two sexes and ‘female’ was that default state. Now we scientists know better. We know that there are signals that drive ovarian development just as there are signals that drive testicular development. There is no true “default” when it comes to biological sex.
In the XX embryonic gonads, SRY is absent and, unlike the testis where one pathway determines male development (i.e. SRY), differentiation of the ovary involves multiple signaling pathways (reviewed in Schlessinger et al., 2010). One of the first steps occurs when retinoic acid from the mesonephros turns on the STRA8 gene (Stimulated by Retinoic Acid Gene 8) in the primordial germ cells, inducing them to become oogonia and enter meiosis. This is a step that is obligatory for formation of ovarian structure (reviewed in Nef & Vassalli, 2009; Kocer et al., 2009) — without the oogonia, follicles will not form.
Retinoic acid cannot induce differentiation of germ cells in the XY embryonic gonad because the up-regulation of SOX9 in nascent Sertoli cells induces expression of CYP26B1, a gene that encodes an enzyme that breaks down retinoic acid, thereby stopping the germ cells from entering meiosis. In the XX embryonic ovary that does not have SOX9, there is also no CYP26B1 and so the retinoic acid is not degraded and can push the germ cells into meiosis, down the female pathway (reviewed in Piprek, 2010).
One of the main differentiation genes for the ovary is R-spondin1 (RSPO1; Roof plate-specific Spondin 1). The gene is turned on in the somatic (pre-granulosa) cells of the nascent ovary, and the protein encoded by this gene is secreted by the cells to modulate other genes, such as WNT4 and the β-catenin pathways. The RSPO1 pathway acts to both promote ovarian differentiation and also to suppress testis formation by silencing the SOX9 and FGF9 genes (reviewed in Nef & Vasselli, 2009).
[The Wnt family of signaling proteins were initially characterized by developmental biologists due to their importance during embryogenesis, but they also play roles in adult processes and during cancer. β-catenin is one of a group of proteins that is important in cell growth and adhesion. Wikipedia, the most accurate source for information in the world, has information about Wnt and β-catenin family members and signaling pathways. RSPO1 is a member of the R-spondin family of genes which are important during development, particularly in the Wnt/β-catenin signaling pathway. In addition to sex determination, RSPO1 is involved in skin differentiation and the regulation of growth of epithelial cells of intestinal crypts.]
Another ovary-promoting differentiation gene in the embryonic gonad is FOXL2 (Forkhead box L2), which encodes a winged-helix/forkhead transcription factor that is expressed in the nascent granulosa cells (reviewed in Schlessinger et al., 2010). FOXL2 is believed to act in concert to RSPO1 to push the XX bipotential gonad in the diretion of the ovary and to block genes that promote differentiation in the direction of the testis (reviewed in Nef & Vasselli, 2009).
[FOXL2 belongs to the family of Forkhead Box ("FOX") transcription factors that are involved in the regulation of genes important during development and other processes where there is cellular growth, proliferation and differentiation.]
So in essence, with these competing pathways, there is a push and pull between male and female, testis and ovary. Regarding differentiation of the gonad, Nef and Vassalli (2009) refer to these signaling processes as “complementary pathways” but DiNapoli and Capel (2008) categorize them as more of a balancing act:
“This precarious balance of the gonad between these two developmental pathways is likely what confers its dual potential. Normally, the system employs complex cell signaling loops that reinforce a single fate decision in the supporting cell lineage and recruit all gonadal cells behind the testicular or ovarian pathways. Defects in these reinforcing signaling loops may explain many disorders of incomplete sexual development that manifest as gonadal dysgenesis, ovotestis formation, ambiguous ductal or genitalia development, or a combination of these features.”
In the next post of this series, we will look at what happens when this balance is lost and things don’t go according to the genetic plan of sex determination.
PS – I apologize for the delay in making this post. I came down with a cold, got buried by work and am now traveling again. You should see the next couple of posts come more quickly in this series. Thank you for your patience and for your interest in this blog!
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