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181 Cards in this Set
- Front
- Back
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What are the general events of fertilization?
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Contact recognition b/t sperm/egg
Regulation of sperm entry into egg and prevention of polyspermy Fusion of genetic material Activation of egg metabolism |
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How does sea urchin sperm become motile following spawning?
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There is a pH change from the testes to the sea water (7.2-7.6)
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What are directional cues with relation to sea urchin sperm finding egg after spawning?
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Chemotactic response to attractant synthesized in egg jelly of urchins.
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What is the specific chemotactic attractant made by sea urchin egg jelly?
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Resact
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How does resact lead sea urchin sperm to egg?
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14 AA peptide that diffuses with potent effect
Resact binds to receptors on sperm Activates guanylyl cyclase: increases cGMP Ca2+ channels open and calcium enters from sea water Species-specific |
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What are the events that trigger acrosomal reaction in sea urchins?
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Interaction b/t sperm and egg jelly
Specific complex carbos. in egg jelly bind to specific receptors in sperm PM Binding activates a Ca 2+ channel, a Na+/H exchanger, and an enzyme that catalyzes the formation of IP3 Acrosome fuses with PM of sperm |
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What happens after the acrosome fuses with the PM of sperm in sea urchins?
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Digestive enzymes are released that digest through egg jelly
Formation of the acrosomal process |
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Why is bindin needed in sea urchins?
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Once through egg jelly, bindin is needed for species-specific binding of sperm to vitelline envelope.
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What is the fast block in sea urchins?
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Change in electrical potential of egg caused by influx of Na+ into the egg.
Sperm can bind at -70 but not at +20mv. (Transient-only stays like this for about one min.) |
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When does the fast block occur?
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1-3 seconds after the binding of the first sperm.
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What is the slow block in sea urchins?
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A wave of Ca2+ ions from point of sperm entry causes cortical granules to fuse with egg cell membrane. The release of contents of granules cause the vitelline envelope to rise and harden into the fertilization envelope.
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What are the contents of the cortical granules in the slow block of sea urchins?
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Digestive enzymes that cut extracellular regions of bindin receptors and attached sperm.
Proteins that fuse with vitelline envelope to make hyaline layer and fertilization envelope. |
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What are some details about the formation of fertilization envelope in sea urchins?
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Fertilization envelope is elevated from the cell membrane by mucopolysaccharides released by granules.
These compounds make an osmotic gradient the forces water to go into space between cell membrane and fertilization envelope, causing envelope to expand from egg. |
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What hardens the fertilization envelope in sea urchins?
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A perioxidase enzyme released from granules that crosslinks with tyrosine residues on adjacent proteins.
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Where does the fertilization envelope start from?
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It begins to form at the site of sperm entry.
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What does hyaline layer do in sea urchins?
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It forms a coating around the egg during slow block. The egg extends microvilli that attach to hyaline layer. Provides support for blastomeres during cleavage.
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Where does the Ca2+ influx come from after fertilization in sea urchins?
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A wave of Ca2+ from the point of sperm entry to other side of egg triggers slow block. Ca2+ comes from internal stores in egg, complete release in 30 sec. and resequestered after that. Ca2+ stored in ER as well.
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What are the first events of egg metabolism in sea urchins?
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Ca2+ activates NAD+ kinase which converts NAD+ to NADP+, which leads to lipid biosynthesis(needed for new cell membranes). NADP+ also boosts Ca2+ which effects oxygen consumption.
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What is the oxygen reduction during egg metabolism in sea urchins used for?
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Oxygen reduction to hydrogen peroxide is used to crosslink fertilization envelope.
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What are the late responses of fertilization in sea urchins involved in egg metabolism?
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Activation of DNA and protein synthesis. The fusion of egg/sperm lead to increase of pH intracellularly.
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What causes the rise in pH after fertilization in sea urchins during beginning of egg metabolism?
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A second influx of Na+ ions from the seawater and a loss of hydrogen ions from the egg. The pH increase and Ca2+ elevation act together to stimulate DNA/protein synthesis.
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What do the elevated Ca2+ ions do in sea urchin egg metabolism?
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Inactivate enzyme MAP kinase, converting it from phosphorylated to unphosphorylated, which removes inhibition on DNA synthesis.
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How does protein synthesis occur several minutes after sperm entry in sea urchins?
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Uses mRNAs already present in oocyte.
An inhibitor binds translation intitiation factor eIF4E at the 5' end of several mRNAs. After fertilization, this inhibitor becomes phosphorylated and degraded, allowing translation. |
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What are some of the mRNAs released after the inhibitor is degraded after fertilization in sea urchins?
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Cyclin B
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What is a complex formed after cyclin B is released after fertilization in sea urchins?
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Cyclin B combines with CDK1 cyclin to create mitosis promoting factor (MPF)
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Why is MPF required after fertilization in sea urchins?
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Mitosis promoting factor (MPF) is required for initiating cell division.
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What are some factors that contribute to mammalian sperm movement toward an ovulated egg?
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Uterine muscle contractions
Sperm capacitation Specialized regions of uterus control sperm movement. Flagellar beating Temperature/Chemical gradients |
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What is the region where fertilization in mammals occurs?
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Ampulla
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What is a mammalian oocyte just released from the ovary surrounded with?
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Cumulus cells
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What are cumulus cells?
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Cells of the ovarian follicle that the oocyte is attached to. Cumulus-oocyte complex required.
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How does the cumulus- oocyte complex get the the appropriate position for fertilization in mammals?
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Ciliary beating along with muscle contractions bring oocyte-cumulus complex into oviduct.
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What is the set of physiological changes that lead to a competent sperm called?
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Capacitation
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What is the first molecular change for sperm capacitation in mammals?
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Sperm cell membrane loses cholesterol b/c of sterol-binding proteins in female oviduct. Changes location of lipid rafts Lipid rafts cluster at anterior of sperm head. Increase membrane fluidity and lead to acrosome reaction.
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What is second molecular change for sperm capacitation in mammals?
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Loss of proteins and carbohydrates from plasma membrane surface that exposes binding sites for receptors on zona pellucida
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What is third molecular change for sperm capacitation in mammals?
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Loss of K+ ions through K+ channels: leads to influx of Ca2+ and HCo3-. Loss of K+ makes sperm cell more negative, which opens Ca2+ channels. Ca2+ and HCO3- activate cAMP which activates membrane fusion events of the acrosome reaction.
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What is the fourth molecular change for sperm capacitation in mammals?
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Protein phosphorylation occurs; two chaperone proteins migrate to the surface of the sperm head when they are phosphorylated-they have a role in forming the receptor that binds to zona pellucida.
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What is the fifth molecular change for sperm capacitation in mammals?
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Initiation of acrosomal reaction by changes in acrosomal membrane.
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What are lipid rafts?
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Contain saturated fatty acids
Moves/floats through non-raft parts Enriched for certain lipids/proteins: -sphingolipids and hydrocarbons -certain proteins like GPI-linked proteins and tyrosine kinases(receptors) |
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What are three distinct events in oviduct that help activate sperm and direct their movement in mammals?
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Hyperactivation
Thermotaxis Chemotaxis |
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What is hyperactivation in mammalian sperm?
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Occurs at beginning of oviduct:
Ca2+ channels in sperm tail are opened, leads to more beating/greater force. |
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What is thermotaxis?
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Occurs in isthmus region prior to ampulla:
capacitated sperm can sense a 2*C difference between isthmus(cooler) and ampulla(warmer). |
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What is chemotaxis?
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Occurs in ampulla:
Oocyte and surrounding cumulus cells secrete chemoattractants. |
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What does a sperm head consist of?
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Haploid nucleus and an acrosome
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Where is the acrosome derived from?
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Golgi apparatus
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What does the acrosome contain?
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Digestive enzymes needed to get through coats surrounding egg.
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What do the midpiece and neck of sperm contain?
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mitochondria and the centriole
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What does the centriole generate?
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microtubules of the flagellum
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Where does energy for flagellar motion come from?
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mitochondrial ATP and a dynein ATPase in the flagellum.
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What can the female gamete be?
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egg(sea urchins) and oocyte(mammals)
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What surrounds the egg?
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vitelline envelope(sea urchins)
zona pellucida(mammals) |
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Where do cortical granules reside?
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Just beneath egg cell membrane
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In sea urchins, the acrosomal reaction is activated by?
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compounds in the egg jelly
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What polymerizes to extend the acrosomal process?
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globular actin
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What recognizes bindin on the acrosomal process in sea urchins?
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a protein complex on the egg surface
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What is IP3 responsible for?
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releasing Ca2+ from storage in endoplasmic reticulum.
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What is DAG responsible for?
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Diacylglycerol is thought to initiate rise in egg pH.
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How is IP3 generated?
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By phospholipases.
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What must capacitated sperm penetrate to bind to the zona pellucida?
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cumulus layer
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What initiates acrosomal reaction in mammals?
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ZP3
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What is the zona pellucida comprised of?
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Three glycoproteins ZP1, ZP2, and ZP3.
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What are the events that trigger acrosome reaction in mammals?
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Binding of sperm receptor (GaIT) to ZP3 triggers
Increase in intracellular Ca2+ and phospholipase C products activate TRPC2 channels Dramatic rise in Ca2+ leads to exocytosis of acrosome. |
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After the acrosome reaction how does mammalian sperm move through ZP?
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Following the fusion of acrosomal vesicle, sperm possesses bindin proteins on surface.
Sperm bindin proteins interact with ZP2 Loss of anterior portion of sperm PM |
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What are some details involved in sperm/egg fusion in mammals?
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cell-cell fusion rare
acrosome reaction required for sperm/egg fusion sperm only bind to microvilli rich region of egg PM |
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What is zona pellucida analogous to?
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Vitelline envelope
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What are the events of sperm and zona pellucida?
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Sperm binds weakly to ligand protein coating ZP.
The sperm surface SED1 binds to ZP complex A protein on sperm form strong link with ZP3. Acrosome reaction |
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What is first event of ZP3 and acrosome reaction?
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Induced when ZP3 crosslinks the receptors on sperm cell membrane.
The sperm cell protein galactosyltransferase active site faces outward an binds to carbos on ZP3 |
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Why is the acrosome reaction necessary?
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It makes it possible for the sperm to concentrate it's proteolytic enzymes directly at the point of attachment and digest hole through ZP.
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What is second event of ZP3 and acrosome reaction?"
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The crosslinked ZP3 and galactosyltransferase activates specific G proteins in sperm cell membrane.
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What is third event in ZP3 and acrosome reaction?
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The G proteins initiate a cascade that opens membranes Ca2+ channels and causes calcium-mediated exocytosis of acrosomal vesicle.
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What is the fourth event in ZP3 and acrosome reaction?
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The exocytosis of acrosomal vesicle releases a variety of proteases that lyse the zona pellucida.
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What is the fifth event in ZP3 and acrosome reaction?
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During the acrosome rxn. the anterior portion of the sperm cell membrane is shed.(area bound to ZP3), secondary binding to ZP is accomplished via proteins in inner acrosomal membrane that bind to ZP2.
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Where does sperm contact egg in mammals?
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On the side of the sperm head.
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What is the junction between the inner acrosomal membrane and the sperm cell membrane called?
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The equatorial region
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Where does the fusion between sperm and egg begin?
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equatorial region
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Mammalian gamete fusion may depend on an interaction between...?
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sperm protein and integrin-associated CD9 protein on egg.
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What protein on the sperm side is needed in mammalian fusion process?
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Izumo
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What does the cortical granule reaction do in mammals?
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Releases enzymes that modify ZP sperm receptors so they can no longer bind sperm.
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What do cortical granules contain in mammals?
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N-acetylglucosamine enzymes that cleave N-acetylglucosamine from ZP3 carbohydrate which bind sperm.
Clip binding to ZP2 via protease. |
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How long does pronuclear migration take in mammals?
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12 hours
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How long does pronuclear migration take in sea urchins?
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1 hour
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How does the mammalian sperm enter the egg?
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Tangentially to the surface of the egg
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What does the sperm fuse to?
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Numerous microvilli
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What is cleavage?
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rapid mitotic divisions that divide the cytoplasm of an egg.
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What is gastrulation?
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dramatic cellular movements throughout embryo to create the three germ layers.
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What is MPF?
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mitosis promoting factor-heterodimer
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What is cyclin B?
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a regulatory subunit, non-catalytic
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What happens to the levels of cyclin B during the cell cycle?
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The levels fluctuate due to protein synthesis and degradation.
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When does the mRNA for cyclin B become active?
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After fertilization.
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what is Cdk?
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cyclin-dependent kinase
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What does Cdk require to be active?
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Must be bound to cyclin B
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What is different in the cell cycle during cleavage compared to other times?
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No G1 and G2 phase
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What is isolecithal?
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sparse, even distribution of yolk
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What is mesolecithal?
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moderate vegetal yolk distribution.
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What is the vegetal pole?
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the yolk-rich pole
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What is the animal pole?
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low yolk concentrated pole
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What is holoblastic cleavage?
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In isolecithal species the cleavage furrow extends through entire egg.
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What is meroblastic cleavage?
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Only a portion of the cytoplasm is cleaved, the cleavage furrow does not penetrate the yolky portion of the cytoplasm.
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What is centrolecithal?
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When the yolk is in the center
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What is telolecithal?
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Only one small area of the egg is free of yolk. Bilateral cleavage, incomplete cell divisions on the periphery.
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What is mesolecithal?
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displaced radial cleavage- cells bigger on vegetal side-smaller on animal.
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What is invagination?(cell movement during gastrulation)
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Single layer of cells made during cleavage is punched in a specific spot. No disruption of cell-cell contacts
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What is involution?(cell movement during gastrulation)
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Cells along vegetal side migrate inside making a second layer.
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What is ingression?(cell movement during gastrulation)
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Epithelial to mesenchymal switch. Migrate as individuals.
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What is delamination?(cell movement during gastrulation)
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Single epithelial sheet that gives rise to two epithelial sheets.
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What is epiboly?(cell movement during gastrulation)
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Cells that are in the outermost layer, spread until they encompass entire embryo.
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What cleavage pattern is exhibited in early development of sea urchins?
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Radial holoblastic cleavage-spindle axes are parallel or at right angles to axis of egg.
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What occurs in the first seven divisions in early development of sea urchins?
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128 blastomeres generated
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What happens at the 128-cell stage of sea urchins in early development?
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Blastula is formed-cells arranged in single epithelial sheet connected by tight junctions.
Blastocoel in center. |
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What happens to the vegetal pole during early development of sea urchins?
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Vegetal pole thickens forming the vegetal plate.
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What happens to the fertilization envelope during early development of sea urchins?
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It is digested releasing a free swimming 'hatched blastula'
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Where is the fertilization envelope derived from in sea urchins?
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It came from the vitelline envelope, but has different composition and is further from the plasma membrane.
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In a fate map of 60 cell urchin embryo what comprises Veg 1?
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Some ectoderm and endoderm
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In the 60 cell fate map of sea urchin what comprises Veg 2?
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endoderm/secondary mesenchyme/coelom
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What does autonomous mean?
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Inherited some protein that allows them to become something-unconditionally. Other ones require combination of cell-cell communication and inheritance.
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What is B-catenin required for?
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Specification of micromeres
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How does B-catenin mediate cell-cell communication?
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Cadherins
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What signaling pathway is B-catenin an important player in?
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WNT signaling
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Where does B-catenin accumulate?
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In the nucleus of micromeres.
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What needs to be present for B-catenin to move into the nucleus?
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Dissheveled
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What happened when LiCl allowed B-catenin to be in all cell nuclei?
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Embryo only had Veg 1 and Veg 2 cells.
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What happened when B-catenin was eliminated?
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No Veg 1 or Veg 2 cells
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What else are micromeres called?
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Primary mesenchyme cells-specified autonomously
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What does B-catenin do in the nucleus of micromeres?
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Transcriptionally activates Pmar 1.
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What is Pmar 1?
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A transcription factor that represses a repressor.
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What happens after Pmar 1 represses the repressor?
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Activation of DELTA which is a ligand for notch signaling. A secondary signal for veg 2.
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Other than repressing a repressor, what else does Pmar 1 do?
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Leads to the production of an early signal for Veg 2.
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What is required for the specification of veg 2 cells?
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Veg 2 cells closest to micromeres effected by notch ligand imbedded in cytoplasm of micromeres.
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How does Nemo-like kinase (NLK) function to specify mesoderm?
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By down regulation of B-catenin
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What do the animal-vegetal axis in sea urchins lead to?
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anterior and posterior axis
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What is oral-aboral axis (ventral-dorsal) axis specified by?
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TGF-B signaling
Nodal promotes oral fates BMP2/4 promote aboral fates |
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What does sea urchin gastrulation begin with?
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ingression of primary mesenchyme
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What are the steps to ingression of primary mesenchyme in gastrulation of sea urchins?
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Group of micromere-derived cells undergo epithelial to mesenchymal transition.
Exhibit altered cytoskeleton altered cell morphology changes in adhesion patterns enter blastocoel |
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What are the features of Filopodia for movement of mesenchyme cells?
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Actin filament polymerization
Thin extensions Dynamic-extension and retraction |
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What are the features of lamellipodia for movement of mesenchyme cells?
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actin filament polymeriztion
broad, short, highly branched move forward and adhere anchoring cell |
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What does primary mesenchyme migrate a specific distance to do?
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initiate skeleton formation
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What are the steps of primary mesenchyme action followed by archenteron formation?
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cells thicken to form vegetal plate
invagination occurs about 1/2 length of blastocoel. Driven by dramatic cell shape changes Let by secondary mesenchyme cells Archenteron forms primitive gut. |
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After gurken signals to torpedo in follicle cells posteriorly, and oocyte signals back to cause MT rearrangement, how are the plus and minus ends directed?
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Minus end anterior
Positive end posterior |
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What does oocyte do after MT elongate?
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oocyte size increases relative to nurse cells
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What does bicoid get moved by?
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dynein motor-minus end directed-anterior
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What does oscar get moved by?
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kinesin motor-plus end directed-posterior
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Where does the nucleus in drosophila get moved to?
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dorsal/anterior
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After A-P axis formation, what is the second role of Gurken?
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After nucleus has moved while tethered to Gurken, Gurken signals again in Dorsal/anterior end.
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What does Gurken do to form the dorsal side of Drosophila?
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Gurken protein binds to torpedo on dorsal side. Inhibits formation of Pipe-which forces cells to take on dorsal fate.
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What occurs on the ventral side of drosophila during Gurken's second signaling?
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No gurken on ventral side. The ventral follicle cells have pipe made in them. A signaling pathway takes place eventually phosphorylating and degrading cactus.
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Why is the degradation of cactus important where gurken does not inhibit pipe?
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cactus is bound to Dorsal protein which is now free to move into the nucleus.
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What happens when dorsal moves into the nucleus?
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It activates twist and snail, they lead to inpocketing and gastrulation.
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What leads to rearrangement of microtubules during A-P axis formation in drosophila?
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Reciprocal signaling between oocyte and follicle cells. + and - ends of MTs gets flipped by gurken/torpedo.
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D-V axis occurs over time, prefertilization and post fertilization, what are some details?
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Nurse cells disappear
multiple nuclei are effected by dorsal cells are already specified prefertilization |
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More examples of pipe in follicle cells...
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Pipe in follicle cells activates proteins in extracellular space that leads to a receptor in embryo that leads to localized accumulation of dorsal in nuclei close to follicle cells where pipe was made.
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What does twist activate after dorsal activates it?
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gastrulation movements-ventral furrow-then pinching off to make mesodermal tube
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Why is nanos localized posteriorly?
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Oscar being translated their creates trap for nanos.
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What do the bicoid and nanos protein gradients lead to?
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hunchback gradients
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When there is lots of nanos there is no...?
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no hunchback
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Bicoid is a transcription factor that activates..
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Zygotic transcription of hunchback in anterior; reinforces maternal gradient already established.
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Bicoid represses translation of...
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Caudal
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Hunchback is a.../
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Gap gene
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What are the sequences of genes?
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Bicoid/nanos cytoplasmic polarity
HB protein gradient sets up gap gene Gap genes set up pair rule genes Pair rule genes set up segment polarity genes(homeotic genes) |
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What are bicoid, nanos, and hunchback?
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morphogens
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What does staufen allow for oskar?
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Staufen is tethered to oskar and allows it to be translated.
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What is an example of gap gene expression?
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Setting up kruppel domain; bicoid activates kruppel while HB represses kruppel at high concentrations, but activates it at low concentrations. Knirps represses kruppel
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What maternal proteins, and gap proteins affect expression of pair-rule genes like even-skipped(Eve)?
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Multiple repressor sites and multiple activation sites. Giant and kruppel repress. Bicoid and hunchback activate.
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What does Hox gene expression along A-P axis direct?
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Development of different structures
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What happens at fourth division in animal tier in sea urchins?
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Four cells from animal tier divide into eight blastomeres, each with same volume, mesomeres.
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What happens in fourth division in vegetal tier in sea urchins?
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Undergoes unequal equatorial cleavage to make four large cells, macromeres. And four small cells, micromeres.
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Where does B-catenin accumulate?
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Nuclei of cells fated to become endoderm/mesoderm
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What other genes does Pmar 1 activate after B-catenin activates it?
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Delta and Tbr which signal veg 2 cells to become endoderm and secondary mesenchyme.
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In normal veg 2 cells B-catenin biases them to be what?
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endomesoderm
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What does Delta protein on the micromeres activate?
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Notch pathway in adjacent veg 2 cells. Causing them to be secondary mesenchyme.
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WNT 8 is made by micromeres and endoderm cells, in what manner does WNT 8 behave?
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In an autocrine manner, to make veg 2 cells endoderm.
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What is convergent/extension with regards to archenteron?(primitive gut)
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narrowing tissue at the same time it is moving forward.
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After primary mesenchyme leave vegetal region what happens?
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Cells left behind thicken-vegetal plate
remain bound to one another and hyaline layer Move to fill gap caused by ingression of primary mesenchyme |
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How far does vegetal plate invaginate?
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about 1/2 way into blastocoel
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After invagination ceases, what is the secondary elongation?
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Cells rearrange by migrating over one another and flattening.
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After secondary elongation, what is the third elongation event in formation of archenteron?
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Tension from secondary mesenchyme cells extend filopodia through blastocoel fluid till contact inner surface of the blastocoel wall. The filopodia attach to blastomeres then shorten pulling archenteron up.
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Delta signal produced by micromeres, imbedded in micromeres, veg 2 cells adjacent will bind via notch receptor, and activate NLK-1, what does this activation do?
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Down regulates B-catenin to form mesoderm
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What is basal lamina?
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extracellular matrix on blastocoel side.
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Par 3, Par 6, and aPKC are located where?
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anteriorly
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Par 1 and Par 2 are located where?`
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posteriorly
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aPKC phosphorylates?
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Par-1
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What is NMY-2?
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Non-muscle myosin 2, it creates a static end where the sperm enters, also the posterior.Par 2 and NMY-2 form a complex.
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Where is Par 3 originally located?
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The cortical cytoplasm
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