BIOL 205: GRQs Test 3

Where else have you seen Catenin in the class? Compare and contrast their role in these 2 very diverse biological events. What accounts for this difference?

1) A-catenin and B-catenin link cadherin molecules in the cell membrane to actin microfilaments. 2) B-catenin works within the cell to act as TF • B-catenin is freed from the complex to alter transcription in the nucleus

Two Important Mechanisms of epiboly in Xenopus gastrulation

1) Animal cap and non-involuting marginal zone (NIMZ) expand by epiboly to cover entire embryo and form surface ectoderm • occurs through increase in cell number via division & integration of several deep layers 2) assembly of fibronectin into fibrils by blastocoel roof • allows vegetal rotation of animal cap and enclosure of embryo • fibronectin is essential for cell migration since fibronectin-containing ECM provides substrate for adhesion and cues for direction of cell migration

In Situ Hybridization

1) Dissect Tissue 2) Fix 3) Add proteinase K (permeabilize, remove protein) 4) Add probe and hybridize 5) Add RNAse (remove RNA and excess probe) 6) Block 7) Add antibody to detect probe 8) Add 2° antibody 9) Add detection reagent 10) Observe using microscope

3 primary germ layers of all embryos and their derivatives

1) Ectoderm: outer layer of embryo produces surface layer of skin (epidermis), brain, and nervous system 2) Endoderm: innermost layer of embryo produces epithelium of digestive tube and its associated organs, as well as the lungs 3) Mesoderm: middle layer of embryo forms blood, heart, kidney, gonads, bones, muscles, and connective tissue

Explain how one ligand can illicit more than one transcriptional response across a sheet of cells.

1) In order for induction to occur, the tissue must be competent 2) Another factor that affects cell fate specification is the gradient of paracrine factors known as morphogens • diffusible biochemical molecules that determine cell's fate by its concentration • differing levels/thresholds of morphogen gradients activate different genes

Give at least 2 experiments that demonstrates there is a difference between the Nieuwkoop Center verses the Spemann and Mangold Organizer.

1) Nieuwkoop Center dorsal vegetal cells can induce animal cells to become dorsal mesodermal tissue 2) Organizer can induce host and donor tissue into secondary embryo with A-P and D-V axes

Draw a table that compares and contrasts the molecular signaling properties of the Nieuwkoop Center verses the Spemann and Mangold Organizer.

1) Nieuwkoop Center signals to induce the organizer in the dorsal mesoderm 2) Dorsal mesoderm signals to the rest of the mesoderm

Why might a cell's response to a signal be slow?

1) Rapid cellular response happens when the signal affects the activity of proteins already present in the target cell. 2) For slower cell response (cell growth and division), the response to extracellular signals requires changes in gene expression and the synthesis of new proteins.

Why can one sperm not fertilize cross species?

1) Species-specific sperm attraction and species-specific sperm activation prevent cross species fertilization. • Each species has its own chemotaxis or SAPs. 2) Polysaccharides and egg jelly factors from one species fail to activate the acrosome reaction 3) Bindin (acrosomal protein mediating recognition of egg surface) is species-specific; bindin from one species binds only to its own bindin receptor of the dejellied egg

Draw the schematic for the signal transduction pathway that functions as the chemotactic pathway for sperm.

1) Sperm are attracted towards chemotaxis 2) Sperm mobility is activated by outside factor 3) Direction is provided by sperm-activating peptides (SAPs) • Ex: Resact 4) Resact binds to receptor's guanylyl cyclase (RGC) 5) Forms cGMP from GTP 6) cGMP opens Ca2+ channels in cell membrane 7) Ca2+ influx activates sperm motility 8) Sperm begins to swim up the resact gradient

Draw out the steps of vegetal rotation and describe what initiates it. (Fig. C-E)

1) bottle cells change shape so main body of each cell is displaced toward the inside of the embryo from slitlike blastopore 2) gastrulation begins in the marginal zone (region surrounding the equator of blastula)

The same signaling molecule can elicit a different response in different cells. Explain how this works.

1) certain cells do not have a receptor for signaling molecules so they will be "deaf" to these signals. 2) signal is conveyed to the inside of the cell via intracellular signaling molecules. These molecules work to alter the activity of effector proteins, which directly affect cellular behavior. • the intracellular relay system and effectors differ from cell to cell so different cell types respond to the same signal in different ways 3) Morphogen gradients

How does the organizer form?

1) first signal tells the cells that they are dorsal 2) second signal tells the cell that they are medoderm

Differentiation

1) follows determination 2) the cell elaborates a cell-specific developmental program.

Two Components of Acrosome Reaction

1) fusion of acrosomal vesicle with the sperm cell membrane • exocytosis that results in release of contents of acrosomal vesicle 2) extension of the acrosomal process

Upon fertilization intercellular Ca2+ is released. Where from? What is the signal transduction pathway that leads to its release?

1) sperm binds to receptors on egg and alters their conformation, thus initiating intracellular cascades 2) membrane phospholipid phosphatidylinositol 4,5-biphosphate (PIP2) is split by phospholipase C (PLC) 3) The split results in two molecules: IP3 and diacylglycerol (DAG) 4) IP3 is the molecules responsible for releasing Ca2+ from the ER by binding to and opening calcium channels 5) DAG activates PKC activates Na+-H+ pump (requires Ca2+) • results in alkalinization and pH increase 6) Ca2+ from ER is responsible for cortical granule reaction and reactivation of development

Detailed Steps of Acrosome Reaction

1) sulfate-containing polysaccharides in egg jelly bind to specific receptors 2) Receptor activates 3 membrane proteins • Ca2+ transport channel that allows Ca2+ to enter sperm head • sodium-hydrogen exchanger that pumps Na+ into the sperm as it pumps H+ out • phospholipase enzyme that makes IP3 3) IP3 releases Ca2+ from inside the sperm (probably within the acrosome itself) 4) Elevated Ca2+ levels triggers acrosome fusion with sperm cell membrane

the 2 major breakthroughs of modern biology

1. all organisms share similar cellular machinery 2. all animals use this machinery in similar ways to direct embryonic development bc common ancestor

describe the different steps of fertilization, comparing mechanisms and molecules

1. binding of sperm to zona pellucida 2. sperm-eggshell contact triggers acrosomal rxn and bindin presentation 3. penetration of sperm through ZP 4. fusion of pm's 5. sperm nucleus enters egg cytoplasm, mediated by fertilin and ADAM2, also izumo and juno in mammals

Sperm's view of fertilization

1. find egg and swim twd it- resact in sea urchins 2. bind to and burrow through egg shell 3. bind to and fuse PM with eggs PM

Calculating Splice Variants

2n-1 (where n is the number gene sites)

Mid-blastula transition for Drosophila

After the nuclei reach the periphery: 1) Slowdown of nuclear division 2) cellularization 3) simultaneous increase in new RNA transcription • maternally provided mRNAs are degraded and control of development is handed over to the zygote's own genome

desmosomes

Anchoring junctions that prevents cells subjected to mechanical stress from being pulled apart; button like thickenings of adjacent plasma membranes connected by fine protein filaments

Does a blastomere cleave or divide? Explain the difference using the terms nucleus and cytoplasm.

Blastomeres cleave since they effectively half the size of the cytoplasm as the cytoplasm of the egg is cleaved into progressively smaller blastomere cytoplasms

Acrosome Reaction

Ca2+-dependent fusion of the acrosome with the sperm cell membrane, resulting in exocytosis and release of proteolytic enzymes that allow the sperm to penetrate the egg's ECM and fertilize the egg

What molecules are critical to cell sorting?

Cell adhesion molecules are critical to cell sorting. • Ex: Cadherins

contributions of father to developing embryo

DNA

Epiboly

Epiboly: movement of epithelial sheets (usually of ectodermal cells) that spread as a unit to enclose the deeper layers of the embryo • can occur by the cells dividing, cells changing their shape, or by several layers of cells intercalating into fewer layers • often all 3 mechanisms are used

Intercalation

Expansion process whereby cells from different layers lose contact with their neighbors and rearrange into a single layer, which consequently spreads laterally, owing to an increase in surface area • use of cells and sheets of cells that are already present

difference between focal adhesion and hemidesmosomes

FC- links to IC actin and EC fibronectin, HD- links to IC keratin and EC laminin

Describe one experiment that demonstrates that Bicoid is necessary to induce anterior portion of the Drosophila embryo.

Remove anterior cytoplasm (and bicoid) • CANNOT form a head.

List a few molecules that serve as signaling molecules in multi-cellular organisms.

Signal molecules can be proteins, peptides, amino acids, nucleotides, steroids, fatty acid derivatives, or even dissolved gasses.

Hormones

Stem from endocrine cells and function as a "public" style of cellular communication • involves broadcasting signal throughout the whole body by secreting it into the bloodstream [or plant's sap] Ex: Insulin from the pancreas

Why in A-P axis formation are all the critical/essential genes transcription factors?

That way new proteins do not need to be synthesized. Instead, signals can merely alter the activity of already existing proteins to bring about changes in transcription levels and gene regulation. • allows for differential gene activity in spite of all cells sharing the same genome

Explain how Maternal Effect Mutations differ from a zygotic mutation

The mutant individual appears phenotypically normal • mutation appears in the progeny of the mutant

Describe one experiment that demonstrates that Bicoid is sufficient to induce anterior portion of the Drosophila embryo.

When bicoid mRNA was added to Bicoid-deficient embryos in different places, the place where bicoid mRNA was injected became the head.

Draw out the Wnt/β-Catenin pathway using any portions of the text necessary (Hint: go back to previous lectures and look at Chapter 4). Compare this pathway to that of BMP.

Wnt/B-catenin pathway works by inhibiting an inhibitor: 1) Wnt interacts with a pair of transmembrane proteins (Frizzled & LRP5/6) to form a multimeric complex 2) enables LRP5/6 to bind both Axin and GSK3 and Frizzled to bind Disheveled 3) Disheveled keeps Axin and GSK3 bound which prevents B-catenin from being phosphorylated by GSK3 4) B-catenin accumulates and binds to LEF/TCF TF and converts the transcription repressor into an activator

potency

a cell can become any type of cell; over time has fewer and fewer possibilities until it is a specific type of cell

cell-cell matrix anchoring junctions

adherens, desmosomes, hemidesmosomes, actin-linked

compare the components, localization, and function of adherens junctions and desmosomes

adherens- on apical end of lateral surface, cadherin protein, hold cells together to make sheets/barriers, anchor a belt of actin filaments; desmosomes- on lateral sides of epi cells, different- cadherin, IC anchors, keratin filaments; give tissues additional strength under mechanical stress, link to intermediate filament cytoskeleton

explain how the cytoskeleton and adhesion machinery could cooperate to allow cells to change shape

adhesion machinery connected to the cytoskeleton moves cells as a sheet (ex. invagination)

explain the evolutionary utility and mechanisms of hormonal control of human oogenesis

all organisms share conserved developmental mechanisms; oogenesis is a big investment, so evolution made it so it only happens in animals at the right time through cyclic changes in peptide and steroid hormone signals (FSH and LH); oocytes arrested in prophase I until puberty, cortical granules in developing follicle, antral follicle, LH surge during puberty causes completion of meiosis 1 to make a secondary oocyte, oocyte detaches from follicle with first polar body and leaves corpus luteum

describe the organization of simple epithelial tissues

apico-basal polarity, tight/gap/desmosome/hemidesmosome junctions, 1 single-layer sheet of cells, most common tissue architecture

Cortical Granule Reaction

basis of the slow block to polyspermy in many animal species (sea urchins and most mammals); mechanical block to polyspermy, in which enzymes from the egg's cortical granules contribute to the formation of a fertilization envelope that blocks additional sperm entry

which model explains identical twins?

both (mosaic and regulatory)

contrast cadherin and integrin-based cell junctions (components, functions, cytoskeletal connections)

cadherin- tight cell-cell junctions, transmembrane, homophilic interactions; integrin- transmembrane, allow cells to interact w/ EC matrix, heterophilic interactions, cell receptors for the ECM, dimers of alpha and beta chains

adherens junctions

cadherins (tm proteins) are linked to actin filaments by caterins; different tissues express distinct but related cadherins

Undifferentiated

cell that has not yet acquired a special structure and function; pertaining to an immature cell or a primitive cell • undifferentiated cells go through a process of maturation that begins when they become committed to a specific cell lineage, progresses through a stage when cell fate is determined to become that of a specific cell type, and ends in differentiation

mosaic model of development + exp

cells do not influence each other; cells already know what they are from the start; killed half cells made half embryo

describe cell specialization in terms of gene expression

cells have different transcription factors that turn on different genes at different times to make them different

regulative model of development +exp

cells influence each other; cells adjust fates in response to their environment; separated cells from each other, made 4 smaller embryos

Fast Block to Polyspermy

change in electric potential of egg cell membrane due to influx of Na+ ions through sodium channels 1) resting membrane potential goes from -70 mV to +20 mV 2) sperm cannot fuse with eggs that have positive membrane potentials 3) does NOT occur in mammals (happens in sea urchins and frogs) 4) only a temporary fix

gap junctions

channel-forming; gated channels of transmembrane connexon subunits

Blastomere

cleavage-stage cell resulting from mitosis

what do mutations in ECM matrix components cause

defects in skin, connective tissue, and bones

contrast different patterns of early division

depends on the amount of yolk in eggs, asymmetrical (except in the simplest cases), plane changes from meridonial to equatorial at the 4 cell stage, does not go to completion in yolk-rich eggs

Determination

determined state, assumed irreversible, is when a cell or tissue is capable of differentiating autonomously—even when placed into a non-neutral environment (other region of embryo or with differently specified cells) • stage of commitment following specification • still "look" like their undetermined neighbors. • fate of determined cells does NOT change

compare the protein and carbohydrate components of the ECM

different tissues secrete different types of ECM for their different roles ex. bones vs. eyeballs; carbs- hyaluron HUGE, glycogen, proteins- spectrin, collagen

Cell division

division of a cell into two daughter cells with the same genetic material. • creation of new cells from old ones

Nieuwkoop Center

dorsalmost vegetal blastomeres of the amphibian blastula, formed as a consequence of the cortical rotation initiated by sperm entry; important signaling center on the dorsal side of the embryo • one of its main functions is to induce the Spemann and Mangold Organizer

explain which cells give rise to the 3 germ layers in the sea urchin embryo

ectoderm on outside, then endoderm, middle is mesoderm

contributions of mother to developing embryo

eggs are huge cells and have nutrients and everything else the embryo needs until it becomes its own organism (yolk, macromolecules- mitochondria, RNA, proteins); shapes pattern formation of embryo

Specification

first stage of commitment of cell or tissue fate during which the cell or tissue is capable of differentiating autonomously, when placed in an environment that is neutral with respect to developmental pathway • cell commitment is still capable of being reversed

explain how cell junctions help mediate cell change

focal contacts at the leading edge of migratory cells; coordination of adhesion and cytoskeleton

Cell Lineages

following individual embryonic cells to see what those cells become • tracks cells in vivo by labeling embryonic cells and seeing what that area becomes in adult organisms

Fertilization Envelope

forms from the vitelline envelope of the sea urchin egg following cortical granule release • allows water to expand space between cell membrane and fertilization envelope

compare the origins of germline and somatic cells

germline- pass genetic info to the next gen, the cells are set aside early as a separate lineage, segregation of determination (P granules) and undergo meiosis to become haploid gametes somatic- all body cells, no passing of genetic info, diploid

Define the zona pellucida. How many cells make up the zona pellucida?

glycoprotein coat (ECM) around the mammalian egg, synthesized and secreted by the growing oocyte • it is made of NO cells since it is the ECM

cell-matrix junctions

hemidesmosomes; link to intermediate cytoskeleton; has analagous components to cell-cell junctions except have integrins instead of adherens for the transmembrane linker protein; integrins that localize to focal contacts (rapidly assemble at leading edge of migrating cells) anchor actin filaments; many different types of alpha and beta subunits creates diversity

explain how tight junctions make epithelial barriers

hold cells together so nothing can get through the sheet; attached to both cell's cytoskeletons; transmembrane claudins make cell-cell connections; allow epithelial cells to maintain different concentrations of molecules on different sides of the sheet

Spemann and Mangold Organizer

in amphibians, dorsal lip cells of blastopore and their derivative (notochord and head endomesoderm) • progeny of dorsal lip cells induces dorsal axis and neural tube, aka primary embryonic induction • transform flanking mesoderm into the A-P body axis

Extracellular signaling molecules

large and hydrophilic (most common) • bind to receptors on cell surface to relay their message inwards

Syncytium

many nuclei residing in a common cytoplasm, resulting either from karyokinesis without cytokinesis or from cellular fusion • interaction between nuclei and differing amounts of determination factors specifies cell fate

compare functions of mitotic and meiotic sisters of the oocyte

meiotic- polar bodies, create poles of embryo; mitotic- nurse cells, 15 for each egg, smaller, attached to egg by cytoplasmic bridges, synthesize macromolecules and pump them into the oocyte thru bridges, provide oocyte with key molecular machinery in the form of mRNA's and proteins that drive all events of early development

Vegetal Rotation

morphogenetic movement that drives mesoderm internalization during gastrulation in amphibian embryos

define the mechanism by which the fruit fly anterior-posterior axis is established

nurse cells synthesize molecules that localize specifically in oocyte to differentiate A/P axis; anterior- bicoid TF, posterior= nanos mRNA transcription regulator; mRNAs are transported along mts by motor proteins, makes localized mRNA's and localized protein, vertebrates- VEG1 localizes to vegetal pole and encodes TGFB family signaling molecule

compaction

occurs at the 8-cell stage in mammals; sudden change in cell-cell adhesion; requires E-cadherin to group cells together

Signal Transduction

pathways of response whereby paracrine factors bind to a receptor that initiates a series of enzymatic reactions within the cell that in turn have often several response as their end point: 1) regulation of TFs (alter gene activity) 2) regulation of cytoskeleton (alter shape or enable migration)

contrast the cell cycle just after fertilization with that of later in cell devo + explain why they're different

post-fert- rapid increase in # of cells without growth bc no gap phase; cytokinesis is driven by F-actin and myosin contractile rings

describe in cell biological terms the common changes in cell shape/arrangement that underlie gastrulation in all animals

powered by the cytoskeleton; changes in individual cell shapes drive gastrulation as a whole contraction of actin belt; apical constriction and elongation/shortening of cell sheet

Differentiated

process by which an unspecialized cell becomes specialized into one of the many cell types that make up the body • cell ceases to divide and develops specialized structural elements with distinct functional properties • cell acquires gene expression pattern characteristic of a specific cell type

identify the 2 main sets of molecular machinery cells use when choosing fate

protein machines carry out the work; cells choose fates and cells change shape and move; Cell signals from neighbors regulate gene transcription and alter cell behavior and Transcription factors tell RNA polymerase which genes to transcribe and thus determine which protein machines are made

mid-blastula transition (MBT)

rapid cell divisions end; key events- cell cycle lengthens, zygotic transcription starts- the embryo makes its own mRNA; triggered by N/C ratio nearing 1

apical constriction

sheet of epithelial cells -> invagination by organized tightening along adhesion belts in selected regions -> epithelial tube pinches off from overlying sheet of cells

explain the roles of different egg components in protecting and equipping the embryo

shell protects it; fast and slow blocks prevent polyspermy, yolk has nutrients

Local Mediators

signal molecules diffuse locally through the extracellular fluid, remaining near the cell that secreted it • involved in paracrine signaling

Intracellular signaling molecules

small and hydrophobic enough to pass thru membrane • either activate enzymes or bind to intracellular receptor proteins that regulate gene expression ϖ Ex: steroid & thyroid hormones

Maternal Effect Mutation

special class of genes that have their effect in the reproductive organs of the mutant • mutant organism may appear phenotypically normal • the progeny express detectable differences • they do so whether the progeny have inherited the mutant gene or not

Morphogen

substance that differentially specifies the fates of cells by different concentrations

size of things (sugar, AA's, NTPs, globular proteins, DNA, ribosomes)

sugar/AA's/NTPs- ~.5-1 nm, globular protein= ~2-10 nm, DNA= ~ 2nm wide, but v long, ribosomes= ~30nm

Mid-Blastula Transition (MBT) for Xenopus

transition from early rapid biphasic (only M and S phases) mitoses of embryo to stage characterized by: 1) mitoses that include "gap" stages (G1 & G2) of cell cycle 2) loss of synchronicity of cell division 3) transcription of new (zygotic) mRNAs needed for gastrulation and cell specification • blastomeres acquire capacity to become motile • chromatin modification

Slow Block to Polyspermy or Cortical Granule Reaction

upon sperm entry, cortical granules fuse with the egg cell membrane and release their contents into the space between fibrous mat of vitelline envelope proteins (or zona pellucida receptors) 1) granule serine protease cleaves protein posts that connect the vitelline envelope and clips off bindin receptors 2) components bind to vitelline envelope to form fertilization envelope 3) In mammals, another slow block involves the release of the egg surface protein Juno, which is responsible for binding to Izumo proteins on sperm's surface

distinguish between molecules that can/cannot pass through gap junctions

~1.5 nm wide; can- inorganic ions, AA's, vitamins, NTPs; cannot- proteins, nucleic acids