Mammalian Gametogenesis
The gametes are the product of a germ line that is separate from the somatic cell lineages that divide mitotically to generate the differentiated somatic cells of the developing individual. Cells in the germ line undergo meiosis, a remarkable process of cell division by which the chromosomal content of a cell is halved so that the union of two gametes in fertilization restores the full chromosomal complement of the new organism. Sexual reproduction means that each new organism receives genetic material from two distinct parents, and the mechanisms of meiosis provide an incredible amount of genomic variation upon which evolution can work. Gametogenesis and fertilization are both the end and the beginning of the circle of life. This chapter describes how the sex of an individual organism is determined, which in turn will determine whether that individual’s gametes will become sperm or eggs.
Chromosomal Sex Determination
There there are several ways chromosomes can determine the sex of an embryo. In mammals, the presence of either a second X chromosome or a Y chromosome determines whether the embryo will be female (XX) or male (XY). In birds, the situation is reversed: the male has the two similar sex chromosomes (ZZ) and the female has the unmatched pair (ZW). In flies, the Y chromosome plays no role in sex determination, but the number of X chromosomes appears to determine the sexual phenotype. In other insects (especially hymenopterans such as bees, wasps, and ants), fertilized, diploid eggs develop into females, while unfertilized, haploid eggs become males.
The Mammalian Pattern of Sex Determination
Mammalian sex determination is governed by the gonad-forming genes and by the hormones elaborated by the gonads. Primary sex determination is the determination of the gonads—the egg-forming ovaries or sperm-forming testes. Secondary sex determination is the determination of the male or female phenotype by the hormones produced by the gonads. The formation both of ovaries and of testes is an active, gene-directed process. Both the male and female gonads diverge from a common precursor, the bipotential gonad e (sometimes called the indifferent gonad)
Development of gonads and their ducts in mammals.
Originally, a bipotential (indifferent) gonad develops, with undifferentiated Müllerian ducts (female) and Wolffian ducts (male) ducts both present. If XY, the gonads becomes testes and the Wolffian duct persists. If XX, the gonads become ovaries and the Müllerian duct persists. Hormones from the gonads will cause the external genitalia to develop either in the male direction (penis, scrotum) or the female direction (clitoris, labia majora).
In mammals, primary sex determination is dictated by whether an organism has an XX or an XY karyotype c. In most cases, the female’s karyotype is XX and the male’s is XY. Every individual must carry at least one X chromosome. Since the diploid female is XX, each of her haploid eggs has a single X chromosome. The male, being XY, generates two populations of haploid sperm: half will bear an X chromosome, half a Y. If at fertilization the egg receives a second X chromosome from the sperm, the resulting individual is XX, forms ovaries, and is female; if the egg receives a Y chromosome from the sperm, the individual is XY, forms testes, and is male ( Figure A, below )
Sex determination in placental mammals.
(A) Mammalian chromosomal sex determination results in approximately equal numbers of male and female offspring.
(B) Postulated cascades leading to male and female phenotypes in mammals. The conversion of the genital ridge into the bipotential gonad requires, among others, the Sf1, Wt1, and Lhx9 genes; mice lacking any of these genes lack gonads. The bipotential gonad appears to be moved into the female pathway (ovary development) by the Foxl2, Wnt4, and Rspo1 genes and into the male pathway (testis development) by the Sry gene (on the Y chromosome), which triggers the activity of Sox9. (Lower levels of Wnt4 are also present in the male gonad.) The ovary makes thecal cells and granulosa cells, which together are capable of synthesizing estrogen. Under the influence ofestrogen (first from the mother, then from the fetal gonads), the Müllerian duct differentiates into the female reproductive tract, the internal and external genitalia develop, and the offspring develops the secondary sex characteristics of a female. The testis makes two major hormones involved in sex determination. The first, anti-Müllerian hormone (AMH), causes the Müllerian duct to regress. The second, testosterone, causes differentiation of the Wolffian duct into the male internal genitalia. In the urogenital region, testosterone is converted into dihydrotestosterone (DHT), which causes the morphogenesis of the penis and prostate gland.
The Y chromosome carries a gene that encodes a testis-determining factor that organizes the bipotential gonad into a testis. This was demonstrated in 1959 when karyotyping showed that XXY individuals (a condition known as Klinefelter syndrome) are male (despite having two X chromosomes), and that individuals having only one X chromosome (XO, sometimes called Turner syndrome) are female. XXY men have functioning testes. Women with a single X chromosome begin making ovaries, but the ovarian follicles cannot be maintained without the second X chromosome. Thus, a second X chromosome completes the ovaries, whereas the presence of a Y chromosome (even when multiple X chromosomes are present) initiates the development of testes.
If the fetus is XY, the mesodermal cells continue to proliferate through week 8, when a subset of these cells initiate their differentiation into Sertoli cells. During embryonic development, the developing Sertoli cells secrete the anti-Müllerian hormone that blocks development of the female ducts. These same Sertoli epithelial cells will also form the seminiferous tubules that will support the development of sperm throughout the lifetime of the male mammal.
Once primary (chromosomal) determination has established the gonads, the gonads begin to produce the hormones and paracrine factors that govern secondary sex determination—development of the sexual phenotype outside the gonads. This includes the male or female duct systems and the external genitalia. A male mammal has a penis, scrotum (testicle sac), seminal vesicles, and prostate gland. A female mammal has a uterus, oviducts, cervix, vagina, clitoris, labia, and mammary glands. In many species, each sex also has a sex-specific body size, vocal cartilage, and musculature. Secondary sex characteristics are usually determined by hormones and paracrine factors secreted from the gonads. In the absence of gonads, it appears the female phenotype is generated. When Jost (1947, 1953) removed fetal rabbit gonads before they had differentiated, the resulting rabbits had a female phenotype, regardless of whether their genotype was XX or XY. The general scheme of primary sex determination is shown in Figure B above. If the embryonic cells have two X chromosomes and no Y chromosome, the gonadal primordia develop into ovaries. The ovaries produce estrogen, a hormone that enables the development of the Müllerian duct into the uterus, oviducts, cervix, and upper portion of the vagina . If embryonic cells contain both an X and a Y chromosome, testes form and secrete two major factors. The first is a TGF-b family paracrine factor called anti-Müllerian hormone (AMH ; sometimes called Müllerian-inhibiting factor, MIF ). AMH destroys the Müllerian duct, thus preventing formation of the uterus and oviducts. The second factor is the steroid hormone testosterone. Testosterone masculinizes the fetus, stimulating formation of the penis, male duct system, scrotum, and other portions of the male anatomy, as well as inhibiting development of the breast primordia.
Primary Sex Determination in Mammals
Mammalian gonads embody a unique embryological situation. All other organ rudiments normally can differentiate into only one type of organ—a lung rudiment can only become a lung, a liver rudiment can develop only into a liver. The gonadal rudiment, however, has two options: it can develop into either an ovary or a testis, two organs with very different tissue architectures. The path of differentiation taken by the gonadal rudiment is dictated by the genotype and determines the future sexual development of the organism. But before this decision is made, the mammalian gonad first develops through a bipotential, or indifferent, stage during which it has neither female
nor male characteristics
The developing gonads
In humans, two gonadal rudiments appear during week 4 and remain sexually indifferent until week 7. These gonadal precursors are paired regions of the mesoderm adjacent to the developing kidneys , see Figure A, below:
Differentiation of human gonads shown in transverse section.
(A) Genital ridge of a 4-week embryo.
(B) Genital ridge of a 6-week indifferent gonad showing expanded epithelium.
(C) Testis development in week 8. The sex cords lose contact with the cortical epithelium and develop the rete testis.
(D) By week 16 of development, the testis cords are continuous with the rete testis and connect with the Wolffian duct through the efferent ducts remodeled from the mesonephric duct.
(E) Ovary development in an 8-week embryo.
(F) In the 20-week embryo, the ovary does not connect to the Wolffian duct, and new cortical sex cords surround the germ cells that have migrated into the genital ridge.
The germ cells—the precursors of either sperm or eggs—migrate into the gonads during week 6 and are surrounded by the mesodermal cells. If the fetus is XY, the mesodermal cells continue to proliferate through week 8, when a subset of these cells initiate their differentiation into Sertoli cells. During embryonic development, the developing Sertoli cells secrete the anti-Müllerian hormone that blocks development of the female ducts. These same Sertoli epithelial cells will also form the seminiferous tubules that will support the development of sperm throughout the lifetime of the male mammal. During week 8, the developing Sertoli cells surround the incoming germ cells and organize themselves into the testis cords. These cords form loops in the central region of the developing testis and are connected to a network of thin canals, called the rete testis, located near the developing kidney duct (Figure C,D). Thus, when germ cells enter the male gonads, they will develop within the testis cords, inside the organ. Later in development (at puberty in humans; shortly after birth in mice, which procreate much faster), the testis cords mature to form the seminiferous tubules. The germ cells migrate to the periphery of these tubules, where they establish the spermatogonial stem cell population that produces sperm throughout the lifetime of the male. In the mature seminiferous tubule, sperm are transported from the inside of the testis through the rete testis, which joins the efferent ducts. The efferent ducts are the remodeled tubules of the developing kidney. During male development, the Wolffian duct differentiates to become the epididymis (adjacent to the testis) and the vas deferens (the tube through which sperm pass into the urethra and out of the body). Note that both sperm and urine will use the urethra to exit the body. Meanwhile, the other group of mesoderm cells (those that did not form the Sertoli epithelium) differentiate into a mesenchymal cell type, the testosterone-secreting Leydig cells. Thus, the fully developed testis will have epithelial tubes made of Sertoli cells that enclose the germ cells, as well as a mesenchymal cell population, the Leydig cells, that secrete testosterone. Each incipient testis is surrounded by a thick extracellular matrix, the tunica albuginea, which helps protect it.
What is anti-Müllerian hormone?
About eight weeks after conception the human fetus has two sets of ducts, one of which can develop into the male reproductive tract and the other into the female reproductive tract. If the fetus is genetically male (XY chromosomes) then the embryonic testes will produce anti-Müllerian hormone. This causes the Müllerian (female) ducts to disappear – hence the term anti-Müllerian hormone, whilst testosterone produced by the testes causes the male (Wollfian) ducts to survive. The Wollfian ducts go on to develop into the different parts of the male reproductive system: the epididymis, the vas deferens, the seminal vesicles and the prostate gland. In a female fetus (XX chromosomes) the Wollfian ducts disappear (because of the lack of testosterone) and the Müllerian ducts develop into the fallopian tubes, uterus (womb), cervix and the upper part of the vagina.
Anti-Müllerian hormone may also have a role in regulating sex steroid production in puberty and in the adult ovaries and testes. In the ovaries, anti-Müllerian hormone appears to be important in the early stages of development of the follicles, which contain and support the eggs prior to fertilisation. The more ovarian follicles a woman has, the more anti-Müllerian hormone her ovaries can produce, and so AMH can be measured in the bloodstream to assess how many follicles a woman has left in her ovaries: her ‘ovarian reserve’. 3
If the fetus is XX, the sex cords in the center of the developing gonad degenerate, leaving sex cords at the surface (cortex) of the gonad. Each germ cell gets enveloped by a separate cluster of sex cord epithelial cells (Figure E,F). The germ cells will become ova (eggs), and the surrounding cortical epithelial cells will differentiate into granulosa cells. The remaining mesenchyme cells of the developing ovary differentiate into thecal cells. Together, the thecal and granulosa cells form follicles that envelop the germ cells and secrete steroid hormones such as estrogens and (when pregnant) progesterone. Each follicle contains a single germ cell—an oogonium (egg precursor)— which will enter meiosis at this time.
There is a reciprocal relationship between the germ cells and the somatic cells of the gonads. The germ cells are originally bipotential and can become either sperm or eggs. Once in the male or female sex cords, however, they are instructed to either (1) begin meiosis and become eggs, or (2) remain meiotically dormant and become spermatogonia. In XX gonads, germ cells are essential for the maintenance of ovarian follicles. Without germ cells, the follicles degenerate into cordlike structures and express male-specific markers. In XY gonads, the germ cells help support the differentiation of Sertoli cells, although testis cords will form even without the germ cells, albeit a bit later . When an ovary is being formed, the Müllerian duct remains intact (there is no AMH to destroy it), and it differentiates into the oviducts, uterus, cervix, and upper vagina. In the absence of adequate testosterone, the Wolffian duct degenerates.
Genetic mechanisms of primary sex determination: Making decisions
Several human genes have been identified whose function is necessary for normal sexual differentiation. Because the phenotype of mutations in sex-determining genes is often sterility, clinical infertility studies have been useful in identifying those genes that are active in determining whether humans become male or female. The story starts in the bipotential gonad that has not yet been committed to the male or female direction. The genes for transcription factors Wt1, Lhx9, GATA4, and Sf1 are expressed, and the loss of function of any one of them will prevent the normal development of either male or female gonads. Then the decision is made:
If no Y chromosome is present, these transcription and paracrine factors d are thought to activate further expression of Wnt4 protein (already expressed at low levels in the genital epithelium) and of a small soluble protein called
R-spondin1 (Rspo1). Rspo1 binds to its cell membrane receptor and further stimulates the Disheveled protein of the Wnt pathway, making the Wnt pathway more efficient at producing the transcriptional regulator β-catenin. One
of the several functions of β-catenin in gonadal cells is to further activate the genes for Rspo1 and Wnt4, creating a positive feedback loop between these two proteins. A second role of β-catenin is to initiate the ovarian pathway of
development by activating those genes involved in granulosa cell differentiation. Its third role is to prevent the production of Sox9, a protein crucial for testis determination .
If a Y chromosome is present, the same set of factors in the bipotential gonad activates the Sry gene on the Y chromosome. Sry protein binds to the enhancer of the Sox9 gene and elevates expression of this key gene in the testis-determining pathway. Sox9 and Sry also act to block the ovary-forming pathway, possibly by blocking β-catenin
Following figure below shows one possible model of how primary sex determination can be initiated. Here we see an important rule of animal development: a pathway for cell specification often has two components, with one branch that says “Make A” and another branch that says “… and don’t make B.” In the case of the gonads, the male pathway says “Make testes and don’t make ovaries,” while the female pathway says “Make ovaries and don’t make testes.”
Possible mechanism for the initiation of primary sex determination in mammals.
While we do not know the specific interactions involved, this model attempts to organize the data into a coherent sequence. If Sry is not present (pink region), the interactions between paracrine and transcription factors in the developing genital ridge activate Wnt4 and Rspo1. Wnt4 activates the canonical Wnt pathway, which is made more efficient by Rspo1. The Wnt pathway causes the accumulation of β-catenin, and large accumulation of β-catenin stimulates further Wnt4 activity. This continual production of β-catenin both induces the transcription of ovaryproducing genes and blocks the testis-determining pathway by interfering with Sox9 activity. If Sry is present (blue region), it may block β-catenin signaling (thus halting ovary generation) and, along with Sf1, activate the Sox9 gene. Sox9 activates Fgf9 synthesis, which stimulates testis development and promotes further Sox9 synthesis. Sox9 also prevents β-catenin’s activation of ovary-producing genes. Sry may also activate other genes (such as TCF21 and NT3) that help generate Sertoli cells. In summary, then, a Wnt4/β-catenin loop specifies the ovaries, whereas a Sox9/Fgf9 loop specifies the testes. One of the targets of the Wnt pathway is the follistatin gene, whose product organizes the granulosa cells of the ovary. Transcription factor Foxl2, which is activated (in a still unknown way) in the ovary, is also involved in inducing follistatin synthesis. The XY pathway appears to have an earlier initiation; if it does not function, the XX pathway takes over.
One of the most important events in sex determination is the determination of the germ cells to undergo gametogenesis, the formation of gametes (sperm and egg). As in the case of the genital ridges, the mammalian primordial germ cells (PGCs) are bipotential and can become either sperm or eggs; if they reside in the ovaries they become eggs, and if they reside in the testes they become sperm. All of these decisions are coordinated by factors produced by the developing gonads a. First and importantly, the cells that generate the sperm or eggs do not originally form inside the gonads. Rather, they form in the posterior portion of the embryo and migrate into the gonads . This pattern is common throughout the animal kingdom: the germ cells are “set aside” from the rest of the embryo and the cells’ transcription and translation are shut down while they migrate from peripheral sites into the embryo and to the gonad. It is as if the germ cells were a separate entity, reserved for the next generation, and repressing gene expression makes them insensitive to the intercellular commerce going on all around them. Although the mechanisms used to specify the germ cells vary enormously across the animal kingdom, the proteins expressed by germ cells to suppress gene expression are remarkably conserved. These proteins, which include the Vasa, Nanos, Tudor, and Piwi family proteins, can be seen in the germ cells of cnidarians, flies, and mammals. Vasa proteins are required for germ cells in nearly all animals studied. They are involved in binding RNA and most likely activate germ-cell-specific messages. In chickens, experimentally induced Vasa can direct embryonic stem cells toward a germ cell fate. Nanos proteins bind to their partner, Pumilio, to form a very potent repressive dimer. Nanos can block RNA translation, and Pumilio binds to the 3′ UTRs of specific mRNAs. In Drosophila, Nanos and Pumilio repress the translation of numerous mRNAs, and in so doing they
(1) prevent the cell from becoming part of any germ layer;
(2) prevent the cell cycle from continuing; and
(3) prevent apoptosis
Tudor proteins were discovered in Drosophila, in which females carrying these genes are sterile and do not form pole cells. It appears that Tudor proteins interact with those Piwi proteins that are involved in transcriptionally silencing portions of the genome, especially active transposons.
The decision of germ cells to differentiate as spermatocytes or oocytes is dramatically different from other decisions made during development. First, the magnitude of the response is far greater than in most cell-fate decisions. For example, microarray analyses identified at least 250 oocyte-enriched genes and 650 spermatocyte-enriched genes in Caenorhabditis elegans a In C. elegans, for example, primordial germ cells begin spermatogenesis or oogenesis as part of a syncytium. b Developing oocytes contain a variety of messenger RNAs and proteins that are needed for embryonic development, and some of these molecules must be prevented from influencing the sperm/oocyte decision itself. Thus, this regulatory decision is unique. In most animals, primordial germ cells differentiate into spermatocytes in males or oocytes in females.
a Caenorhabditis elegans is a free-living, transparent nematode ( roundworm), about 1 mm in length, that lives in temperate soil environments. It is the type species of its genus. 1 It is a species in which females have the ability to produce their own self-fertilizing sperm, thereby allowing these "hermaphrodites" the strategic choice to self-reproduce or outcross with males.
b A cytoplasm that contains many nuclei is called a syncytium,4 and the specification of presumptive cells within such a syncytium is called syncytial specification. A notable example of an embryo that goes through a syncytial stage
is found in insects, as illustrated by the fruit fly Drosophila melanogaster.
c A karyotype is the number and appearance of chromosomes in the nucleus of a eukaryotic cell. The term is also used for the complete set of chromosomes in a species or in an individual organism[1][2][3] and for a test that detects this complement or measures the number.
d Paracrine signaling is a form of cell-to-cell communication in which a cell produces a signal to induce changes in nearby cells, altering the behavior of those cells.
e A gonad, sex gland, or reproductive gland is a mixed gland that produces the gametes (sex cells) and sex hormones of an organism. In the female of the species the reproductive cells are the egg cells, and in the male the reproductive cells are the sperm. The male gonad, the testicle, produces sperm in the form of spermatozoa. The female gonad, the ovary, produces egg cells. Both of these gametes, are haploid cells. 2
1. https://en.wikipedia.org/wiki/Caenorhabditis_elegans
2. https://en.wikipedia.org/wiki/Gonad
3. http://www.yourhormones.info/hormones/anti-muellerian-hormone/
The gametes are the product of a germ line that is separate from the somatic cell lineages that divide mitotically to generate the differentiated somatic cells of the developing individual. Cells in the germ line undergo meiosis, a remarkable process of cell division by which the chromosomal content of a cell is halved so that the union of two gametes in fertilization restores the full chromosomal complement of the new organism. Sexual reproduction means that each new organism receives genetic material from two distinct parents, and the mechanisms of meiosis provide an incredible amount of genomic variation upon which evolution can work. Gametogenesis and fertilization are both the end and the beginning of the circle of life. This chapter describes how the sex of an individual organism is determined, which in turn will determine whether that individual’s gametes will become sperm or eggs.
Chromosomal Sex Determination
There there are several ways chromosomes can determine the sex of an embryo. In mammals, the presence of either a second X chromosome or a Y chromosome determines whether the embryo will be female (XX) or male (XY). In birds, the situation is reversed: the male has the two similar sex chromosomes (ZZ) and the female has the unmatched pair (ZW). In flies, the Y chromosome plays no role in sex determination, but the number of X chromosomes appears to determine the sexual phenotype. In other insects (especially hymenopterans such as bees, wasps, and ants), fertilized, diploid eggs develop into females, while unfertilized, haploid eggs become males.
The Mammalian Pattern of Sex Determination
Mammalian sex determination is governed by the gonad-forming genes and by the hormones elaborated by the gonads. Primary sex determination is the determination of the gonads—the egg-forming ovaries or sperm-forming testes. Secondary sex determination is the determination of the male or female phenotype by the hormones produced by the gonads. The formation both of ovaries and of testes is an active, gene-directed process. Both the male and female gonads diverge from a common precursor, the bipotential gonad e (sometimes called the indifferent gonad)
Development of gonads and their ducts in mammals.
Originally, a bipotential (indifferent) gonad develops, with undifferentiated Müllerian ducts (female) and Wolffian ducts (male) ducts both present. If XY, the gonads becomes testes and the Wolffian duct persists. If XX, the gonads become ovaries and the Müllerian duct persists. Hormones from the gonads will cause the external genitalia to develop either in the male direction (penis, scrotum) or the female direction (clitoris, labia majora).
In mammals, primary sex determination is dictated by whether an organism has an XX or an XY karyotype c. In most cases, the female’s karyotype is XX and the male’s is XY. Every individual must carry at least one X chromosome. Since the diploid female is XX, each of her haploid eggs has a single X chromosome. The male, being XY, generates two populations of haploid sperm: half will bear an X chromosome, half a Y. If at fertilization the egg receives a second X chromosome from the sperm, the resulting individual is XX, forms ovaries, and is female; if the egg receives a Y chromosome from the sperm, the individual is XY, forms testes, and is male ( Figure A, below )
Sex determination in placental mammals.
(A) Mammalian chromosomal sex determination results in approximately equal numbers of male and female offspring.
(B) Postulated cascades leading to male and female phenotypes in mammals. The conversion of the genital ridge into the bipotential gonad requires, among others, the Sf1, Wt1, and Lhx9 genes; mice lacking any of these genes lack gonads. The bipotential gonad appears to be moved into the female pathway (ovary development) by the Foxl2, Wnt4, and Rspo1 genes and into the male pathway (testis development) by the Sry gene (on the Y chromosome), which triggers the activity of Sox9. (Lower levels of Wnt4 are also present in the male gonad.) The ovary makes thecal cells and granulosa cells, which together are capable of synthesizing estrogen. Under the influence ofestrogen (first from the mother, then from the fetal gonads), the Müllerian duct differentiates into the female reproductive tract, the internal and external genitalia develop, and the offspring develops the secondary sex characteristics of a female. The testis makes two major hormones involved in sex determination. The first, anti-Müllerian hormone (AMH), causes the Müllerian duct to regress. The second, testosterone, causes differentiation of the Wolffian duct into the male internal genitalia. In the urogenital region, testosterone is converted into dihydrotestosterone (DHT), which causes the morphogenesis of the penis and prostate gland.
The Y chromosome carries a gene that encodes a testis-determining factor that organizes the bipotential gonad into a testis. This was demonstrated in 1959 when karyotyping showed that XXY individuals (a condition known as Klinefelter syndrome) are male (despite having two X chromosomes), and that individuals having only one X chromosome (XO, sometimes called Turner syndrome) are female. XXY men have functioning testes. Women with a single X chromosome begin making ovaries, but the ovarian follicles cannot be maintained without the second X chromosome. Thus, a second X chromosome completes the ovaries, whereas the presence of a Y chromosome (even when multiple X chromosomes are present) initiates the development of testes.
If the fetus is XY, the mesodermal cells continue to proliferate through week 8, when a subset of these cells initiate their differentiation into Sertoli cells. During embryonic development, the developing Sertoli cells secrete the anti-Müllerian hormone that blocks development of the female ducts. These same Sertoli epithelial cells will also form the seminiferous tubules that will support the development of sperm throughout the lifetime of the male mammal.
Once primary (chromosomal) determination has established the gonads, the gonads begin to produce the hormones and paracrine factors that govern secondary sex determination—development of the sexual phenotype outside the gonads. This includes the male or female duct systems and the external genitalia. A male mammal has a penis, scrotum (testicle sac), seminal vesicles, and prostate gland. A female mammal has a uterus, oviducts, cervix, vagina, clitoris, labia, and mammary glands. In many species, each sex also has a sex-specific body size, vocal cartilage, and musculature. Secondary sex characteristics are usually determined by hormones and paracrine factors secreted from the gonads. In the absence of gonads, it appears the female phenotype is generated. When Jost (1947, 1953) removed fetal rabbit gonads before they had differentiated, the resulting rabbits had a female phenotype, regardless of whether their genotype was XX or XY. The general scheme of primary sex determination is shown in Figure B above. If the embryonic cells have two X chromosomes and no Y chromosome, the gonadal primordia develop into ovaries. The ovaries produce estrogen, a hormone that enables the development of the Müllerian duct into the uterus, oviducts, cervix, and upper portion of the vagina . If embryonic cells contain both an X and a Y chromosome, testes form and secrete two major factors. The first is a TGF-b family paracrine factor called anti-Müllerian hormone (AMH ; sometimes called Müllerian-inhibiting factor, MIF ). AMH destroys the Müllerian duct, thus preventing formation of the uterus and oviducts. The second factor is the steroid hormone testosterone. Testosterone masculinizes the fetus, stimulating formation of the penis, male duct system, scrotum, and other portions of the male anatomy, as well as inhibiting development of the breast primordia.
Primary Sex Determination in Mammals
Mammalian gonads embody a unique embryological situation. All other organ rudiments normally can differentiate into only one type of organ—a lung rudiment can only become a lung, a liver rudiment can develop only into a liver. The gonadal rudiment, however, has two options: it can develop into either an ovary or a testis, two organs with very different tissue architectures. The path of differentiation taken by the gonadal rudiment is dictated by the genotype and determines the future sexual development of the organism. But before this decision is made, the mammalian gonad first develops through a bipotential, or indifferent, stage during which it has neither female
nor male characteristics
The developing gonads
In humans, two gonadal rudiments appear during week 4 and remain sexually indifferent until week 7. These gonadal precursors are paired regions of the mesoderm adjacent to the developing kidneys , see Figure A, below:
Differentiation of human gonads shown in transverse section.
(A) Genital ridge of a 4-week embryo.
(B) Genital ridge of a 6-week indifferent gonad showing expanded epithelium.
(C) Testis development in week 8. The sex cords lose contact with the cortical epithelium and develop the rete testis.
(D) By week 16 of development, the testis cords are continuous with the rete testis and connect with the Wolffian duct through the efferent ducts remodeled from the mesonephric duct.
(E) Ovary development in an 8-week embryo.
(F) In the 20-week embryo, the ovary does not connect to the Wolffian duct, and new cortical sex cords surround the germ cells that have migrated into the genital ridge.
The germ cells—the precursors of either sperm or eggs—migrate into the gonads during week 6 and are surrounded by the mesodermal cells. If the fetus is XY, the mesodermal cells continue to proliferate through week 8, when a subset of these cells initiate their differentiation into Sertoli cells. During embryonic development, the developing Sertoli cells secrete the anti-Müllerian hormone that blocks development of the female ducts. These same Sertoli epithelial cells will also form the seminiferous tubules that will support the development of sperm throughout the lifetime of the male mammal. During week 8, the developing Sertoli cells surround the incoming germ cells and organize themselves into the testis cords. These cords form loops in the central region of the developing testis and are connected to a network of thin canals, called the rete testis, located near the developing kidney duct (Figure C,D). Thus, when germ cells enter the male gonads, they will develop within the testis cords, inside the organ. Later in development (at puberty in humans; shortly after birth in mice, which procreate much faster), the testis cords mature to form the seminiferous tubules. The germ cells migrate to the periphery of these tubules, where they establish the spermatogonial stem cell population that produces sperm throughout the lifetime of the male. In the mature seminiferous tubule, sperm are transported from the inside of the testis through the rete testis, which joins the efferent ducts. The efferent ducts are the remodeled tubules of the developing kidney. During male development, the Wolffian duct differentiates to become the epididymis (adjacent to the testis) and the vas deferens (the tube through which sperm pass into the urethra and out of the body). Note that both sperm and urine will use the urethra to exit the body. Meanwhile, the other group of mesoderm cells (those that did not form the Sertoli epithelium) differentiate into a mesenchymal cell type, the testosterone-secreting Leydig cells. Thus, the fully developed testis will have epithelial tubes made of Sertoli cells that enclose the germ cells, as well as a mesenchymal cell population, the Leydig cells, that secrete testosterone. Each incipient testis is surrounded by a thick extracellular matrix, the tunica albuginea, which helps protect it.
What is anti-Müllerian hormone?
About eight weeks after conception the human fetus has two sets of ducts, one of which can develop into the male reproductive tract and the other into the female reproductive tract. If the fetus is genetically male (XY chromosomes) then the embryonic testes will produce anti-Müllerian hormone. This causes the Müllerian (female) ducts to disappear – hence the term anti-Müllerian hormone, whilst testosterone produced by the testes causes the male (Wollfian) ducts to survive. The Wollfian ducts go on to develop into the different parts of the male reproductive system: the epididymis, the vas deferens, the seminal vesicles and the prostate gland. In a female fetus (XX chromosomes) the Wollfian ducts disappear (because of the lack of testosterone) and the Müllerian ducts develop into the fallopian tubes, uterus (womb), cervix and the upper part of the vagina.
Anti-Müllerian hormone may also have a role in regulating sex steroid production in puberty and in the adult ovaries and testes. In the ovaries, anti-Müllerian hormone appears to be important in the early stages of development of the follicles, which contain and support the eggs prior to fertilisation. The more ovarian follicles a woman has, the more anti-Müllerian hormone her ovaries can produce, and so AMH can be measured in the bloodstream to assess how many follicles a woman has left in her ovaries: her ‘ovarian reserve’. 3
If the fetus is XX, the sex cords in the center of the developing gonad degenerate, leaving sex cords at the surface (cortex) of the gonad. Each germ cell gets enveloped by a separate cluster of sex cord epithelial cells (Figure E,F). The germ cells will become ova (eggs), and the surrounding cortical epithelial cells will differentiate into granulosa cells. The remaining mesenchyme cells of the developing ovary differentiate into thecal cells. Together, the thecal and granulosa cells form follicles that envelop the germ cells and secrete steroid hormones such as estrogens and (when pregnant) progesterone. Each follicle contains a single germ cell—an oogonium (egg precursor)— which will enter meiosis at this time.
There is a reciprocal relationship between the germ cells and the somatic cells of the gonads. The germ cells are originally bipotential and can become either sperm or eggs. Once in the male or female sex cords, however, they are instructed to either (1) begin meiosis and become eggs, or (2) remain meiotically dormant and become spermatogonia. In XX gonads, germ cells are essential for the maintenance of ovarian follicles. Without germ cells, the follicles degenerate into cordlike structures and express male-specific markers. In XY gonads, the germ cells help support the differentiation of Sertoli cells, although testis cords will form even without the germ cells, albeit a bit later . When an ovary is being formed, the Müllerian duct remains intact (there is no AMH to destroy it), and it differentiates into the oviducts, uterus, cervix, and upper vagina. In the absence of adequate testosterone, the Wolffian duct degenerates.
Genetic mechanisms of primary sex determination: Making decisions
Several human genes have been identified whose function is necessary for normal sexual differentiation. Because the phenotype of mutations in sex-determining genes is often sterility, clinical infertility studies have been useful in identifying those genes that are active in determining whether humans become male or female. The story starts in the bipotential gonad that has not yet been committed to the male or female direction. The genes for transcription factors Wt1, Lhx9, GATA4, and Sf1 are expressed, and the loss of function of any one of them will prevent the normal development of either male or female gonads. Then the decision is made:
If no Y chromosome is present, these transcription and paracrine factors d are thought to activate further expression of Wnt4 protein (already expressed at low levels in the genital epithelium) and of a small soluble protein called
R-spondin1 (Rspo1). Rspo1 binds to its cell membrane receptor and further stimulates the Disheveled protein of the Wnt pathway, making the Wnt pathway more efficient at producing the transcriptional regulator β-catenin. One
of the several functions of β-catenin in gonadal cells is to further activate the genes for Rspo1 and Wnt4, creating a positive feedback loop between these two proteins. A second role of β-catenin is to initiate the ovarian pathway of
development by activating those genes involved in granulosa cell differentiation. Its third role is to prevent the production of Sox9, a protein crucial for testis determination .
If a Y chromosome is present, the same set of factors in the bipotential gonad activates the Sry gene on the Y chromosome. Sry protein binds to the enhancer of the Sox9 gene and elevates expression of this key gene in the testis-determining pathway. Sox9 and Sry also act to block the ovary-forming pathway, possibly by blocking β-catenin
Following figure below shows one possible model of how primary sex determination can be initiated. Here we see an important rule of animal development: a pathway for cell specification often has two components, with one branch that says “Make A” and another branch that says “… and don’t make B.” In the case of the gonads, the male pathway says “Make testes and don’t make ovaries,” while the female pathway says “Make ovaries and don’t make testes.”
Possible mechanism for the initiation of primary sex determination in mammals.
While we do not know the specific interactions involved, this model attempts to organize the data into a coherent sequence. If Sry is not present (pink region), the interactions between paracrine and transcription factors in the developing genital ridge activate Wnt4 and Rspo1. Wnt4 activates the canonical Wnt pathway, which is made more efficient by Rspo1. The Wnt pathway causes the accumulation of β-catenin, and large accumulation of β-catenin stimulates further Wnt4 activity. This continual production of β-catenin both induces the transcription of ovaryproducing genes and blocks the testis-determining pathway by interfering with Sox9 activity. If Sry is present (blue region), it may block β-catenin signaling (thus halting ovary generation) and, along with Sf1, activate the Sox9 gene. Sox9 activates Fgf9 synthesis, which stimulates testis development and promotes further Sox9 synthesis. Sox9 also prevents β-catenin’s activation of ovary-producing genes. Sry may also activate other genes (such as TCF21 and NT3) that help generate Sertoli cells. In summary, then, a Wnt4/β-catenin loop specifies the ovaries, whereas a Sox9/Fgf9 loop specifies the testes. One of the targets of the Wnt pathway is the follistatin gene, whose product organizes the granulosa cells of the ovary. Transcription factor Foxl2, which is activated (in a still unknown way) in the ovary, is also involved in inducing follistatin synthesis. The XY pathway appears to have an earlier initiation; if it does not function, the XX pathway takes over.
One of the most important events in sex determination is the determination of the germ cells to undergo gametogenesis, the formation of gametes (sperm and egg). As in the case of the genital ridges, the mammalian primordial germ cells (PGCs) are bipotential and can become either sperm or eggs; if they reside in the ovaries they become eggs, and if they reside in the testes they become sperm. All of these decisions are coordinated by factors produced by the developing gonads a. First and importantly, the cells that generate the sperm or eggs do not originally form inside the gonads. Rather, they form in the posterior portion of the embryo and migrate into the gonads . This pattern is common throughout the animal kingdom: the germ cells are “set aside” from the rest of the embryo and the cells’ transcription and translation are shut down while they migrate from peripheral sites into the embryo and to the gonad. It is as if the germ cells were a separate entity, reserved for the next generation, and repressing gene expression makes them insensitive to the intercellular commerce going on all around them. Although the mechanisms used to specify the germ cells vary enormously across the animal kingdom, the proteins expressed by germ cells to suppress gene expression are remarkably conserved. These proteins, which include the Vasa, Nanos, Tudor, and Piwi family proteins, can be seen in the germ cells of cnidarians, flies, and mammals. Vasa proteins are required for germ cells in nearly all animals studied. They are involved in binding RNA and most likely activate germ-cell-specific messages. In chickens, experimentally induced Vasa can direct embryonic stem cells toward a germ cell fate. Nanos proteins bind to their partner, Pumilio, to form a very potent repressive dimer. Nanos can block RNA translation, and Pumilio binds to the 3′ UTRs of specific mRNAs. In Drosophila, Nanos and Pumilio repress the translation of numerous mRNAs, and in so doing they
(1) prevent the cell from becoming part of any germ layer;
(2) prevent the cell cycle from continuing; and
(3) prevent apoptosis
Tudor proteins were discovered in Drosophila, in which females carrying these genes are sterile and do not form pole cells. It appears that Tudor proteins interact with those Piwi proteins that are involved in transcriptionally silencing portions of the genome, especially active transposons.
The decision of germ cells to differentiate as spermatocytes or oocytes is dramatically different from other decisions made during development. First, the magnitude of the response is far greater than in most cell-fate decisions. For example, microarray analyses identified at least 250 oocyte-enriched genes and 650 spermatocyte-enriched genes in Caenorhabditis elegans a In C. elegans, for example, primordial germ cells begin spermatogenesis or oogenesis as part of a syncytium. b Developing oocytes contain a variety of messenger RNAs and proteins that are needed for embryonic development, and some of these molecules must be prevented from influencing the sperm/oocyte decision itself. Thus, this regulatory decision is unique. In most animals, primordial germ cells differentiate into spermatocytes in males or oocytes in females.
a Caenorhabditis elegans is a free-living, transparent nematode ( roundworm), about 1 mm in length, that lives in temperate soil environments. It is the type species of its genus. 1 It is a species in which females have the ability to produce their own self-fertilizing sperm, thereby allowing these "hermaphrodites" the strategic choice to self-reproduce or outcross with males.
b A cytoplasm that contains many nuclei is called a syncytium,4 and the specification of presumptive cells within such a syncytium is called syncytial specification. A notable example of an embryo that goes through a syncytial stage
is found in insects, as illustrated by the fruit fly Drosophila melanogaster.
c A karyotype is the number and appearance of chromosomes in the nucleus of a eukaryotic cell. The term is also used for the complete set of chromosomes in a species or in an individual organism[1][2][3] and for a test that detects this complement or measures the number.
d Paracrine signaling is a form of cell-to-cell communication in which a cell produces a signal to induce changes in nearby cells, altering the behavior of those cells.
e A gonad, sex gland, or reproductive gland is a mixed gland that produces the gametes (sex cells) and sex hormones of an organism. In the female of the species the reproductive cells are the egg cells, and in the male the reproductive cells are the sperm. The male gonad, the testicle, produces sperm in the form of spermatozoa. The female gonad, the ovary, produces egg cells. Both of these gametes, are haploid cells. 2
1. https://en.wikipedia.org/wiki/Caenorhabditis_elegans
2. https://en.wikipedia.org/wiki/Gonad
3. http://www.yourhormones.info/hormones/anti-muellerian-hormone/
