Embryology - Biology 104, Spring 2006 - Albert Harris and Corey Johnson   OUTLINE OF EIGHTH LECTURE: Feb 1, 2006, by Corey Johnson Fertilization Gamete Structure: Sperm Anton van Leeuwenhoek first observed them (1678). Spalanzini demonstrated that filtered semen would not fertilize, that something fairly large must be responsible for fertilization. However, both thought of sperm/spermatozoa as invading organisms! Head : Haploid nucleus, acrosomal vesicle (acrosome), acrosomal process (sea urchins, not vertebrates) Neck : centriole Midpiece : mitochondria, axoneme Tail : Flagellum/axoneme- the axoneme is made of a core of 2 microtubules surrounded by 9 pairs. The microtubules are made of tubulin subunits. Propulsion is achieved by the hydrolysis of ATP by dynein Gamete Structure: Egg Cell is much larger than sperm. Sea urchin egg has 10,000x the volume of a sperm. 1. Large nucleus . The nucleus, depending on the species, may be haploid or diploid. Most mammals (including humans and mice) undergo a 'meiotic arrest' at Metaphase II. Dogs are diploid / sea urchins are haploid at the time of fertilization 2. Cytoplasm: contains Yolk, RNAs, and regulatory proteins 3. Cortex: outermost cytoplasm, rich in actin that polymerizes to produce microvilli. Also contains cortical granules that are released upon fertilization (contain proteolytic enzymes, and proteins that prevent other sperm from entering the egg and physically support the cleaving blastomeres) 4. Vitelline envelope. Called the zona pellucida in mammals: covers the cell membrane, has a protective function, and prevents other species' sperm from entering the egg. 5. Cumulus or follicle cells (mammals): surrounding the zona pellucida    OR, Egg Jelly (Sea Urchins): gelatinous covering that attracts and/or activates sperm Illustrations of these two types of egg Egg and Sperm Interactions 1. Chemoattraction of sperm to egg 2. release of the acrosomal vesicle 3. Sperm binds vitelline layer/zona pellucida 4. Fusion of sperm and egg cell membranes Notice: there are three membrane fusions in the entire process: -Fusion of acrosomal membrane with sperm plasma membrane -Fusion of sperm plasma membrane with oocyte plasma membrane -Fusion of oocyte plasma membrane with cortical granule membranes Blocks to polyspermy Monospermy-a single sperm enters the egg, the usual and desired outcome Polyspermy-more than one sperm enters the egg 1. fast block - membrane depolarization prevents sperm from binding the egg membrane. Normal resting potential of -70mV turns positive as Na+ channels allow an influx of Na+. Sperm cannot bind a membrane with a positive membrane potential. Occurs in Sea urchins and frogs, but not most mammals, 2. slow block - Calcium-induced cortical granule release. Granules empty contents between cell membrane and vitelline envelope - contents of the granules: 1. proteases - cleave sperm binding proteins 2. mucopolysaccharides - cause osmotic gradient which in turn causes water to rush in; this expands the vitelline membrane forming the fertilization membrane - here's a little movie showing the fertilization of a sea urchin embryo 3. other proteins that harden the fertilization membrane (hyaline) Egg activation The egg is in a relatively slow metabolic state before fertilization Upon fertilization (sperm binding): -Ca++ levels rise. The increase in Ca++ comes mostly from the endoplasmic reticulum Watch a quick movie: A calcium reactive dye is shown on the right panel. -pH rises (sea urchin) -Those eggs in meiotic arrest complete meiosis -sperm pronucleus migrates to egg center -protein synthesis and metabolism rise dramatically -DNA synthesis and mitosis -cleavage -The sperm loses its flagellum and mitochondria as it enters the egg. All (approximately) of the zygote's mitochondria come from the mom. -Fusion of the pronuclei -Centrioles come from the sperm - in the case of 2 sperms fertilizing, there are then 4 centrioles and 3 haploid nuclei (a very bad and awkward situation for everyone). Mammals: The role of the maternal and paternal genetic material in development is unequal: Male-directed development emphasizes placenta and a large size Female-directed development has opposite affect: smaller embryo and extraembryonic investment Amphibians: Cytoskeletal rearrangement follows sperm entry. The cortical cytoskeleton rotates 30 degrees towards the sperm entry point. This forms the grey crescent on the opposite side. The dorsal lip of the blastopore will form where the grey crescent forms (emerges). At this point, 3 axes have been established. Tunicates (ascidians) also have cytoplasmic rearrangements following sperm entry: the homogenous yellow cytoplasm (the yellow stuff is actually associated with cytoskeleton) is moved/moves toward the sperm pronucleus, eventually resulting in crescent formation.

 

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