Primordial Germ Cells: From Genital Ridge to Gonads PDF

# Primordial Germ Cells: From Genital Ridge to Gonads - **In mammals**, PGCs first enter the hindgut and eventually migrate forward into the bipotential gonads. - They multiply as they migrate. - PGCs are surrounded by cells secreting **stem cell factor (SCF)** which is needed for PGC motility a...

# Primordial Germ Cells: From Genital Ridge to Gonads - **In mammals**, PGCs first enter the hindgut and eventually migrate forward into the bipotential gonads. - They multiply as they migrate. - PGCs are surrounded by cells secreting **stem cell factor (SCF)** which is needed for PGC motility and survival. - A cluster of SCF-secreting cells appears to migrate with the PGCs, forming a "traveling niche" that supports the persistence, division, and movement of PGCs. - **Once the PGCs are in the gonad**, they are sustained by BMPs that create a niche for them in the genital ridge. - Subsequently, the gonad determines whether the PGCs will initiate oogenesis (the formation of eggs) or spermatogenesis (the formation of sperm). - A fundamental difference between mammalian males and females involves the timing of meiosis. - In females, meiosis begins in the embryonic gonads. - In males, meiosis is not initiated until puberty. - The **"gatekeeper" for meiosis appears to be the Stra8 transcription factor**, which promotes a new round of DNA synthesis and meiotic initiation in the germ cells. - In the developing ovaries, Stra8 is upregulated by two factors - Wnt4 and retinoic acid (RA) - coming from the adjacent kidney. - In the developing testes, however, Stra8 is downregulated by Fgf9, and the retinoic acid produced by the developing kidney is degraded by the testes' secretion of the RA-degrading enzyme Cyp26b1. - During male puberty, retinoic acid is synthesized in the Sertoli cells and induces Stra8 in sperm stem cells. - Once Stra8 is present, the sperm progenitor cells become committed to meiosis. ## The Role of Stra8 - Stra8 promotes a new round of DNA synthesis and meiotic initiation in the germ cells - In the developing ovaries, Stra8 is upregulated by **Wnt4 and retinoic acid (RA)** - In the developing testes, Stra8 is down regulated by **Fgf9** - During male puberty, retinoic acid is synthesized in the **Sertoli cells and induces Stra8 in sperm stem cells.** ## Oogenesis vs. Spermatogenesis | Characteristic | Oogenesis | Spermatogenesis | |------------------------------|----------------------|-----------------------| | Meiosis initiated once in a finite population of cells | ✅ | ❌ | | One gamete produced per meiosis | ✅ | ❌ | | Completion of meiosis delayed for months or years | ✅ | ❌ | | Meiosis arrested at first meiotic prophase and reinitiated in a smaller population of cells | ✅ | ❌ | | Differentiation of gamete occurs while diploid, in first meiotic prophase | ✅ | ❌ | | All chromosomes exhibit equivalent transcription and recombination during meiotic prophase | ✅ | ❌ | | Meiosis initiated continuously in a mitotically dividing stem cell population | ❌ | ✅ | | Four gametes produced per meiosis | ❌ | ✅ | | Meiosis completed in days or weeks | ❌ | ✅ | | Meiosis and differentiation proceed continuously without cell cycle arrest | ❌ | ✅ | | Differentiation of gamete occurs while haploid, after meiosis ends | ❌ | ✅ | | Sex chromosomes excluded from recombination and transcription during first meiotic prophase | ❌ | ✅ | ## Spermatogenesis in Mammals - Spermatogenesis is the developmental pathway from germ cell to mature sperm. - Spermatogenesis begins at puberty and occurs in the recesses between the Sertoli cells. - Spermatogenesis is divided into three major phases: - **Proliferative phase:** Sperm stem cells (spermatogonia) increase by mitosis. - **Meiotic phase:** Involves the two divisions that create the haploid state. - **Postmeiotic "shaping" phase (Spermiogenesis):** Round cells (spermatids) eject most of their cytoplasm and become streamlined sperm. ## The Proliferative Phase of Spermatogenesis - The proliferative phase begins when the mammalian PGCs arrive at the genital ridge of a male embryo. - They are called gonocytes and become incorporated into the sex cords that will become the seminiferous tubules. - These gonocytes become undifferentiated spermatogonia residing near the basal end of the tubular cells. - These cells are **true stem cells** in that they can reestablish spermatogenesis when transferred into mice whose sperm production has been eliminated by toxic chemicals. - Spermatogonia appear to take up residence in stem cell niches at the junction of the Sertoli cells, Leydig cells, and the testicular blood vessels. - Adhesion molecules join the spermatogonia directly to the Sertoli cells, which will nourish the developing sperm. ## The Meiotic Phase: Getting To Haploid Spermatids - Type B spermatogonia are the precursors of the spermatocytes and contain high levels of Stra8. - These cells are the last of that lineage to undergo mitosis and they divide once to generate the primary spermatocytes -- the cells that enter meiosis. - Each primary spermatocyte undergoes the first meiotic division to yield a pair of haploid secondary spermatocytes, which complete the second division of meiosis. - The cells thus formed are called spermatids, and they are still connected to one another through their cytoplasmic bridges. - The spermatids that are connected in this manner have haploid nuclei, but are functionally diploid since a gene product made in one cell can readily diffuse into the cytoplasm of its neighbors. ## The Postmeiotic Phase: Spermiogenesis - In humans, the progression from spermatogonial stem cell to mature spermatozoa takes 65 days, and the last third of it (about 21 days) is taken up by spermiogenesis. - The mammalian haploid spermatid is a round, unflagellated cell that looks nothing like the mature vertebrate sperm. - For fertilization to occur, the sperm has to meet and bind with an egg, and spermiogenesis prepares the sperm for these functions of motility and interaction. # Oogenesis in Mammals - Oogenesis (egg production) differs greatly from spermatogenesis. - It is dependent on **hormones, paracrine factors, enzymes, chromatin structures, and tissue anatomy.** - Egg maturation can be seen as having four stages: - **Proliferation:** In the human embryo, the thousand or so PGCs reaching the developing ovary divide rapidly from the second to the seventh month of gestation. - **Primary Oocyte:** The surviving population, under the influence of retinoic acid, enters the next step and initiates the first meiotic division. - **Diplotene Stage:** The first meiotic division does not proceed very far, and the primary oocytes remain in the diplotene stage of first meiotic prophase. - This prolonged diplotene stage may last from 12 to 40 years. - **Secondary Oocyte and Maturation:** With the onset of puberty, groups of oocytes periodically resume meiosis. - At that time, LH from the pituitary gland releases this block and permits these oocytes to resume meiotic division. - They complete first meiotic division, and the resulting secondary oocytes proceed to second meiotic metaphase and undergo maturation steps. - This maturation involves the crosstalk of paracrine factors between the oocyte and its follicle cells, both of which are maturing during this phase. - The follicle cells activate the translation of stored oocyte mRNA encoding proteins such as the sperm-binding proteins that will be used for fertilization and the cyclins that control embryonic cell division. ## Oogenic Meiosis - The oocyte is a remarkable cell with two roles in reproduction: - Correct segregation of chromosomes during two successive rounds of meiosis - Sustaining viability of the early embryo until transcriptional activation (meiosis minimizes the loss of cytoplasm during this process). ## Oocytes and Age - The retention of the oocyte in the ovary for decades has profound medical implications - Most human embryos do not survive to birth, and a large proportion (perhaps even a majority) of fertilized human eggs have too many or too few chromosomes to survive. - Genetic analysis has shown that such **aneuploidy (incorrect number of chromosomes)** is usually due to errors in oocyte meiosis. - Only a few aneuploidies survive to be born (such as those of the sex chromosomes and chromosome 21), and the percentage of babies born with such aneuploidies increases greatly with maternal age. - Women in their twenties have only a 2-3% chance of bearing a fetus whose cells contain an extra chromosome. - This risk goes to 35% in women who become pregnant in their forties. - This increase appears to be due to the gradual loss of cohesin proteins from the chromosomes as the cell ages, causing a less stable linkage between the kinetochore and spindle during meiotic metaphase. # Sperm - Sperm were discovered in the 1670s, but their role in fertilization was not discovered until the mid-1800s. - It was only in the 1840s, after Albert von Kölliker described the formation of sperm from cells in the adult testes, that fertilization research could really begin. - Von Kölliker denied that there was any physical contact between sperm and egg, believing that the sperm excited the egg to develop in much the same way a magnet communicates its presence to iron - The first description of fertilization was published in 1847 by Karl Ernst von Baer, who showed the union of sperm and egg in sea urchins and tunicates. ## Sperm Anatomy - Each sperm cell consists of a haploid nucleus, a propulsion system to move the nucleus, and a sac of enzymes that enable the nucleus to enter the egg. - In most species, almost all of the cell's cytoplasm is eliminated during sperm maturation, leaving only certain organelles that are modified for spermatic function. - During the course of maturation, the sperm's haploid nucleus becomes very streamlined and its DNA becomes tightly compressed. - In front or to the side of this compressed haploid nucleus lies the acrosomal vesicle, or acrosome. - The acrosome is derived from the cell's Golgi apparatus and contains enzymes that digest proteins and complex sugars. - Enzymes stored in the acrosome can digest a path through the outer coverings of the egg. - In many species, a region of actin proteins lies between the sperm nucleus and the acrosomal vesicle. - These proteins are used to extend a fingerlike acrosomal process from the sperm during the early stages of fertilization. - In sea urchins and numerous other species, recognition between sperm and egg involves molecules on the membrane of the acrosomal process. - Together, the acrosome and the nucleus constitute the sperm head. ## Sperm Motility - The means by which sperm are propelled vary according to how the species has adapted to environmental conditions. - In most species, an individual sperm is able to travel by whipping its flagellum. - The major motor portion of the flagellum is the axoneme. - The axoneme is formed by microtubules emanating from one of the two centrioles at the base of the sperm nucleus. - The core of the axoneme consists of two central microtubules surrounded by a row of nine doublet microtubules. - These microtubules are made exclusively of the dimeric protein tubulin. - The other centriole is also important, as it will enter the egg to establish the mitotic spindle of first cleavage. # Egg - The egg, or ovum, is a single cell that stores all the material necessary to begin growth and development. - The egg eliminates most of its cytoplasm as it matures. - The oocyte conserves the material it has, and actively accumulates more. - The meiotic divisions that form the oocyte conserve its cytoplasm rather than giving half of it away. - The oocyte either synthesizes or absorbs proteins such as yolk that serve as food reservoirs for the developing embryo. - Even eggs with relatively sparse yolk are large compared with sperm. ## Sperm and Egg: Key Differences - While the sperm has eliminated most of its cytoplasm, the egg has maintained its cytoplasm and grown even larger. - The sperm is essentially a haploid nucleus with a propulsion system and an egg-recognizing membrane. - The egg has equipped its haploid nucleus with a cytoplasm full of ribosomes, mitochondria, and enzymes critical for development. # Recognition of Egg and Sperm - The interaction of sperm and egg generally proceeds according to five steps: 1. Chemoattraction of the sperm to the egg by soluble molecules secreted by the egg 2. Binding of the sperm to the extracellular matrix (jelly or zona pellucida) of the egg 3. Exocytosis of the sperm acrosomal vesicle and release of its enzymes 4. Passage of the sperm through the extracellular matrix to the egg cell membrane 5. Fusion of the egg and sperm cell membranes # External Fertilization in Sea Urchins - The events of sperm-egg meeting and fusing are outlined in the document. - Sea urchins release their gametes into the environment. - Sea urchins are faced with two problems: - How can sperm and eggs meet in such a dilute concentration? - How can sperm be prevented from attempting to fertilize eggs of another species? - To solve these problems, two major mechanisms have evolved: - Species-specific sperm attraction - Species-specific sperm activation ## Species-Specific Sperm Attraction - Species-specific sperm attraction has been documented in numerous species, including cnidarians, mollusks, echinoderms, amphibians, and urochordates. - Sperm are attracted toward eggs of their species by chemotaxis – that is, by following a gradient of a chemical secreted by the egg. - In sea urchins, the chemotactic agents are small peptides called sperm-activating peptides (SAPs). - One such SAP is resact, a 14-amino acid peptide that has been isolated from the egg jelly of the sea urchin Arbacia punctulata. - It has a profound effect at very low concentrations, such that the binding of even a single resact molecule may be enough to provide direction for the sperm, which swim up a concentration gradient of this compound until they reach the egg. ## The Acrosome Reaction - As the sperm reaches the egg surface, the acrosomal process adheres to the vitelline envelope and tethers the sperm to the egg. - It is possible that large complexes of acrosomal protein-digesting enzymes coat the acrosomal process, allowing it to digest the vitelline envelope at the point of attachment and proceed toward the egg cell membrane. ## Internal Fertilization in Mammals - It is very difficult to study any interactions between the mammalian sperm and egg that take place prior to these gametes making contact. - This is because mammalian fertilization occurs inside the oviducts of the female. - Although it is relatively easy to mimic the conditions surrounding sea urchin fertilization using natural or artificial seawater, we do not yet know the components of the various natural environments that mammalian sperm encounter as they travel to the egg. # Getting the Gametes into the Oviduct: Translocation and Capacitation - The female reproductive tract is not a passive conduit through which sperm race, but a highly specialized set of tissues that actively regulate the transport and maturity of both gametes. - Both male and female gametes use a combination of small-scale biochemical interactions and large-scale physical propulsion to get to the ampulla at the upper end of the oviduct, where fertilization takes place. ## Translocation - The meeting of sperm and egg must be facilitated by the female reproductive tract. - Different mechanisms are used to position the gametes at the right place at the right time. - A mammalian oocyte just released from the ovary is surrounded by a matrix containing cumulus cells. - The oocyte-cumulus complex is transported to the appropriate position for its fertilization in the oviduct by a combination of ciliary beating and muscle contractions. - Sperm must travel a longer path, and approximately 200-300 million sperm are ejaculated into the vagina. - Only about 200 sperm probably reach the vicinity of the egg. - The translocation of sperm from the vagina to the oviduct involves several processes that work at different times and places: - **Sperm motility:** Motility (flagellar action) is probably important in getting sperm through the cervical mucus in the vicinity of the oocyte. - **Hyperactivation:** As sperm enter the ampulla, they become **hyperactivated**, which involves the opening of the sperm-specific calcium channels, the CatSper proteins, in the sperm tail. - Hyperactivated sperm swim at higher velocities and generate greater force. - They detach their binding with the oviduct epithelial cells and continue their journey to the egg. - **Thermal and Chemical Gradients:** Sperm can sense thermal differences as small as 0.014 degrees Celsius over a millimeter and tend to migrate toward the higher temperature (thermotaxis). - **Progesterone Detection:** Sperm can also detect and respond to picomolar amounts of the hormone **progesterone**, which is secreted by the cumulus cells surrounding the egg. ## The Acrosome Reaction - As sperm enter the ampulla of the oviduct, **capacitated** sperm undergo the acrosome reaction. - "Successful" sperm (i.e., those that actually fertilize an egg) have usually undergone the acrosome reaction by the time they are seen in the cumulus. - Progesterone appears to trigger the release of Protein kinase A (PKA) from the sperm cell membrane, thereby allowing it to activate the sperm-specific cation channels. - These CatSper channels facilitate the transfer of calcium ions into the sperm, which causes exocytosis of the acrosome. ## Recognition at the Zona Pellucida - Once within the cumulus, the sperm can make contact with the zona pellucida, the extracellular matrix of the egg. - The zona pellucida is a far thicker and denser structure than the vitelline envelope in invertebrates. - The mouse zona pellucida is made of three major glycoproteins - ZP1, ZP2, and ZP3 (zona proteins 1, 2, and 3) - along with accessory proteins that bind to the zona's integral structure. - The human zona pellucida has four major glycoproteins - ZPI, ZP2, ZP3, and ZP4. - The binding of sperm to the zona is relatively, but not absolutely, species-specific, and a species may use multiple mechanisms to achieve this binding. - The egg may have pathways for accepting both types of capacitated sperm: - Sperm that have undergone the acrosome reaction bind directly to ZP2. - Sperm with intact acrosomes become bound to ZP3, which causes the acrosome reaction, and the sperm binding is then transferred to ZP2. - The mechanism for this is not known, but two important proteins may be the acrosomal proteins acrosin and MMP2 (matrix metalloproteinase-2). # Gamete Fusion and The Prevention of Polyspermy ## Sperm-Egg Fusion - The sperm and the egg now finally meet. - It is not the tip of the sperm head that makes contact with the egg (as happens in the perpendicular entry of sea urchin sperm) but the side of the sperm head. - The acrosome reaction, in addition to expelling the enzymatic contents of the acrosome, also exposes the inner acrosomal membrane to the outside. - The junction between the inner acrosomal membrane and the sperm cell membrane is called the equatorial region, and this is where membrane fusion between sperm and egg begins. - This fusion involves several proteins, two of which are Izumo, originally in the acrosomal membrane, and Juno, an oocyte membrane protein. - Mutations in either Juno or Izumo block fertilization. - As in sea urchin gamete fusion, the sperm is bound to regions of the egg where actin polymerizes to extend microvilli to the sperm. ## Blocks to Polyspermy - Polyspermy is a problem for mammals, just as it is for sea urchins. - No electrical fast block to polyspermy has yet been detected; it may not be needed given the limited number of sperm that reach the ovulated egg. - A **slow block to polyspermy** does occur, and involves the cortical granule reaction. - When the **cortical granules fuse with the egg cell membrane**, they release protein-digesting enzymes that modify the zona pellucida proteins such that they can no longer bind sperm. - One of these cortical granule proteases is **ovastacin**. - When ZP2 is cleaved by ovastacin, it loses its ability to bind sperm. - Polyspermy occurs more frequently in mouse eggs bearing mutant ZP2 that cannot be cleaved by ovastacin. - Another slow block to polyspermy comes from the **zinc spark**, the release of billions of zinc ions that is induced by the entry of the first sperm. - These released zinc ions are seen to bind to the zona pellucida. - Because **two acrosomal enzymes, acrosin and MMP2 are inhibited by zinc**, the accumulation of this heavy metal on the zona pellucida and in the surrounding cumulus may form a "zinc shield" that prevents further sperm entry. ## Activation of the Mammalian Egg - As in every other animal studied, a transient rise in cytoplasmic **Ca2+** is necessary for egg activation in mammals. - Fertilization triggers this release through the production of **IP3** by the enzyme **phospholipase C (PLC)**. - Unlike in sea urchins, in mammals, the PLC responsible for egg activation and pronucleus formation may come from the sperm rather than from the egg. - The PLC responsible for egg activation in mammals turns out to be a soluble sperm PLC enzyme, **PLCC (PLC-zeta)**, which is delivered to the egg by gamete fusion. ## Fusion of Genetic Material - In sea urchins, the single mammalian sperm that finally enters the egg carries its genetic contribution in a haploid pronucleus. - In mammals, the process of pronuclear migration takes about **12 hours**, compared with less than 1 hour in sea urchins. - The mammalian sperm enters the oocyte while the oocyte nucleus is arrested in **metaphase of its second meiotic division**. - Unlike in the sea urchin egg, which has already completed meiosis, the chromosomes of the mammalian oocyte are still in the middle of their second meiotic metaphase. - Oscillations in the level of Ca2+ activate another kinase that leads to the proteolysis of cyclin (thus allowing the cell cycle to continue) and securin (the protein that holds the metaphase chromosomes together), thereby allowing the completion of meiosis and the establishment of a mature female pronucleus. # Additional Notes - The document was written in the same language as the original text, which is assumed to be English, based on the spelling and word choice. - The document did not include any images, so I did not include them in the markdown output. - I converted mathematical formulas into LaTeX format where applicable. - I formatted the text using markdown headings, lists, and a table for easier readability.