principles of animal development embryonic...

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Chapter 52Animal Development

n Principles of Embryonic Developmentn General Events of Embryonic Developmentn Control of Cell Differentiation and

Morphogenesis During Animal Developmentn Impact on Public Health

Key Concepts:n Biological information that controls embryonic

development resides in bothq Organism’s DNAq Cytoplasm of the egg

n Morphology − refers to organism’s organization and structure

n Morphogenesis − process that creates morphology

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Principles of Embryonic Development

n Embryonic developmentq Fertilized egg becomes an organism with

physiological systems and body parts

n Development and growth are differentq Growth produces more or larger cellsq Development produces organisms with a

defined set of characteristics

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n Body planq Dorsoventral axisq Anteroposterior axisq Right-left axis

n Cellular differentiationq Process by which different cells within a developing

organism acquire specialized forms and functions, due to the expression of cell-specific genes

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n Most animals are triploblastsq Develop from embryos with 3 cell layersq Vertebrates, arthropods, echinoderms, and mollusks

n Five general events of embryogenesis:q Fertilization, cleavage, gastrulation, neurulation,

and organogenesis

n May also include metamorphosis of larva to adult

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General Events of Embryonic Development

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Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

Event 1: FertilizationInitiates the process ofembryonic development.

Development and growthcontinue as the embryomatures through larvalstage to adulthood.

Event 5: Organogenesis results in tissues becoming organizedinto functional organs composed of differentiated cells.

Event 4: Neurulation producesthe future nervous system;during this time, a segmentedbody plan also develops.

Event 2: Cleavage produces a mass of smaller cells in the embryo.

Event 3: Gastrulation produces3 cell layers, called ectoderm,mesoderm, and endoderm.

Future bodysegment

Future nervous system

Stages of Development

Tadpole

Adult

Sperm

Egg

Endoderm

Mesoderm

Ectoderm

Event 1: Fertilization

n Initial contact of egg and sperm followed by acrosomal reactionsq Enzymes dissolve jelly-like layer allowing sperm

to contact the plasma membrane of the egg

n Sperm head fuses with egg membrane and penetrates

n Fast block to polyspermy – fusion depolarizes eggs so that no more sperm may enter

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n Cortical reactionq Ca2+ released from ER into cytosolq Results in calcium wave in which

n Sperm-binding proteins are inactivatedn Outer coating hardens and separates

q Slow block to polyspermyn First cell cycle is activatedn Increased protein synthesis and metabolism

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Spermhead

Spermnucleus

Acrosome

Sperm-bindingproteins

Jellycoat

(a) Acrosomal reaction

Egg plasmamembrane

Cortical granules

Hydrolyticenzymes

Egg cellcytoplasm

Vitellinelayer

21 When a sperm cellcontacts an egg, theacrosome releaseshydrolytic enzymesthat dissolve the jellycoat.

3 The sperm and eggplasma membranesfuse. The spermnucleus will thenenter the egg.

This exposes sperm-binding proteins on the egg cell plasmamembrane that bindto the sperm.


Egg cellcytoplasm

EndoplasmicreticulumEndoplasmicreticulum

IP3 is released from the plasma membrane nearthe site of sperm fusionwith the egg.

1 The contents of the corticalgranules destroy the sperm-binding proteins and causethe vitelline layer and plasmamembrane to separate. Thevitelline layer of the egghardens. This preventspolyspermy.

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2 IP3 stimulatesCa2+ releasefrom theendoplasmicreticulum.

IP3

Ca2+ 3 Ca2+ stimulates

exocytosis ofcortical granules.

(b) Cortical reaction


© Courtesy Dr. M. Whitaker, University of Newcastle upon Tyne

Site of sperm entry

Time after sperm entry15 seconds 25 seconds 31 seconds 36 seconds

(c) The Ca2+ wave of the cortical reaction in a sea urchin egg

Event 2: Cleavage

n Repeated cell divisions without cell growth

n “Biphasic” – alternates between mitotic (M) phase and DNA synthesis (S) phase (no G1 or G2)

n Blastomeres – half-size daughter cell produced at each division

n Blastula – hollow ball of cells

n Blastocoel – space inside ball12

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n Incomplete cleavage – birds and fishesq Eggs have large amount of yolk

q Vegetal pole – where yolk is more concentrated in the egg

q Animal pole – where there is less yolk and more cytoplasm in the eggn Meroblastic cleavage – only this area undergoes cell divisionn Forms flattened disc called blastoderm

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Animal pole

Cleavage-stageembryo

Vegetal pole

Cells in the vegetal hemispherecontain much yolk and littlecytoplasm.

Cells in the animal hemispherecontain little yolk and muchcytoplasm.

n Complete cleavage – amphibians and mammalsq Smaller amount of yolk

q Holoblastic cleavagen Cleavage during the first cell division is complete and bisects

the entire zygote into two equal-sized blastomeresn Mammals undergo compaction creating a morula and forming

a blastocyst (mammalian blastula)

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Amphibians

Mammals Blastocyst

Blastula

2.2 mm 2.5 mm 2.3 mm 2.7 mm 3.1 mm

3 mm 7.7 mm 4 mm

2.3 mm

Meroblastic (incomplete)cleavage

Holoblastic (complete)cleavage

Birdsand fishes

Animalpole

Vegetalpole

Morula

Blastoderm

4.4 mm 4 mm

(bottom): © Tom Fleming

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n In mammals, fertilization and cleavage occur in the oviductq Blastocyst has a different morphological appearance

than the blastula or blastoderm embryos in nonmammalian species

q Blastocyst hasn Outer epithelial layer – trophectoderm – will give rise to

placentan Inner cell mass develops into embryo

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Ovulation

Oviduct

Blastocystbefore hatching

Implantation

Blastocystafter hatching 2-cell

stageZonapellucida

First cleavage

Fertilization

Secondary oocyte

Ovary

Morula

Uterus

Inner cell mass

BlastocystTrophectoderm

n Toward the end of cleavage, cell cycles become less synchronous

n Embryo begins to express its own genes

n Embryo shifts from existing exclusively on maternal factors to developing in response to products derived from its own genome

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Event 3: Gastrulation

n Major cell movement occurs

n Blastula develops into gastrula

n 3 germ layersq Ectoderm, endoderm, and mesoderm

n First time that anteroposterior and dorsoventralbody axes are clearly evident in the embryo

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BlastulaBlastomereslabeledwith dye

Larval structures arisingfrom labeled blastomeres

(bottom): Courtesy Hiroki Nishida, Biological Sciences, Osaka University

n Three distinct cell layers:q Endoderm – forms epithelial lining of gut, liver,

pancreas, thyroid, lungs, and bladder

q Mesoderm – forms heart, limbs, muscles, kidneys, blood, connective tissues, and notochord

q Ectoderm – forms epidermis and nervous system

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Ectoderm (outer layer)

Neurons of peripheralnervous system

Cells ofepidermis

Neuronsof brain

Red bloodcells

Cells of thenotochord

Kidneytubule cells

Skeletalmuscle cells

Endoderm (inner layer)

Pancreaticacinar cells

Lung alveolarcells

Thyroidfollicular cells

Mesoderm (middle layer)

Ectoderm

Mesoderm

Endodermn Invagination

q Gastrulation begins when band of tissue pinches in creating opening – blastoporen Defines the anteroposterior axis

q Initiating site of invagination becomes dorsal lip of blastopore

n Involutionq Cells of animal pole spread out and move downq Enter blastopore and migrate along roof of blastocoel

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Animal pole

Formation of the blastopore byinvagination of bottle cells.Gastrulation is initiated by theinvagination of bottle cells, whichforms a blastopore. Invaginationof bottle cells forces cells behindthem to involute toward the futureanterior end of the embryo. Thecurved arrows indicate directionsof cell movements.

Bottlecells

Dorsallip

Involutingmigratorycells

BlastocoelVegetal pole

Blastula

Dorsal lip ofblastopore

EndodermMesodermEctoderm

KEY


2 Formation of the archenteron byinvagination and involution.The cavity that begins at the blastoporeexpands to form the archenteron (futuredigestive tract). Ectoderm spreads overthe embryo.

Blastopore

InvolutionSpreading

Blastocoelbecomingdisplaced

Archenteron

Invaginationandinvolution


KEY


3 Completion of gastrulation with thebeginning of notochord formation.By the end of gastrulation, thearchenteron has displaced the blastocoeland becomes closed by a yolk plug.Involution continues; some of theinvoluting cells become the mesodermlayer of the gastrula. The dorsal surfaceof the gastrula begins to thicken, and thedorsal mesoderm begins to form thenotochord.

Yolk plug

EctodermArchenteron(futuredigestive tract)Notochord

Endoderm


KEY n Archenteron displaces blastocoel to become organism’s digestive tract

n In chordates and echinoderms, blastopore becomes the anus

n During involution, surface cells spread from the animal hemisphere to surround the entire vegetal hemisphere to become the future ectoderm

n Result of these cellular rearrangements is an embryo with three distinct germ layers

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n Changes in cell shape and position

q Apical constrictionn Bottle cellsn Actin rings constrict and cells

elongate

q Convergent extensionn Spreading of ectodermn 2 rows of cells merge to from

a single elongated layer

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Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.Basal end

Apical end

Contractileactin network

(a) Apical constriction (b) Convergent extension

n Notochord – mesodermal structure providing structural rigidity along anteroposterior axis on dorsal side of embryoq Presence of notochord defines phylum Chordata

q Notochord persists in the trunk and tail of fishes and amphibians; in birds and mammals, the notochord disappears by the time vertebrae have formed

q By the time the notochord forms, the dorsal ectoderm overlying the notochord thickens and next stage begins – neurulation

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n Primordial germ cells (PGCs)q Specialized group of cells migrates to future site of

gonads

q Often arise independently of the three cell layers

q Two functions:n Protect and propagate genetic contentn Undergo meiosis to produce gametes

q Stem cells can undergo mitosis to replicate themselves

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(a) Germ plasm

Germ plasm around the vegetalpole of an amphibian embryo

Pole cells at the posteriorend of an early fly embryo

a: © Courtesy Dr. Laurence D. Etkin, University of Texas M.D. Anderson Cancer Center. With permission of Malgorzata Kloc. This article was published in Mechanisms of DevelopmentVol. 75, No. 1-2, Malgorzata Kloca, Carolyn Larabellb, Agnes Pui-Yee Chana and Laurence D. Etkin “Contribution of METRO pathway localized molecules to the organization of the

germ cell lineage,” pages 81-93. Elsevier Science Ireland Ltd. July 1998; b: © F. R.Turner/Indiana University

(b) Pole cells

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Event 4: Neurulation

n Formation of neural tube from ectoderm located dorsal to notochord

n All neurons and their supporting cells in the CNS originate from neural precursor cells derived from neural tube

n During neurulation, the embryo also develops segmented structures

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n Steps of neural tube formationq Ectoderm overlaying the notochord thickens by the

elongation of cells in the dorsal region to form the neural platen Will form neural crest and epidermis

q Neural plate folds into the neural tube through a series of apical constrictions

q Two sides of neural groove converge to form a tubelike structure

q Fusion – neural tube closes

q Neural crest cells migrate away

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Future neural crest Future epidermis

Neuralplate Thickening and elongation:

Ectoderm over the notochord thickens to form the neural plate,which elongates. Adjacent regions will form a neural crest andepidermis.

Folding:Cells along the medial hinge point undergo apical constriction,causing the formation of a neural groove.

Convergence:Two dorsal lateral hinge points undergo apical constriction toproduce a tubelike structure.

Fusion:The dorsal-most cells on each side of the neural tube begin tofuse. Epidermis also fuses dorsal to the neural tube.

Neural crest cells migrate away as fusion is completed.

Migrating neural crest cells

Notochord

Neuralgroove

MedialHinge point Unsealed

dorsalsurface

Dorsal lateralhinge points

Neuralcrest

Neural tube

Epidermis

Courtesy Kathryn Tosney

n Cells overlying dorsal side of neural tube

n Unique to vertebrates

n Migrate to other regions of embryo to form all neurons and supporting cells of peripheral nervous system

n Also migrate to form q Melanocytesq Facial skeleton and cartilage q Medulla of adrenal gland 36

Neural crest

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n Segmentation of the body plan along the anteroposterior axis becomes apparent during neurulationq Allows individual body segments to have more

specialized functions

q In vertebrates, the segmentation process helps define repeated structures such as vertebrae and ribs, which form later during development

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n Somitesq Mesoderm becomes segmented at the anterior end

first, giving rise to blocklike structures

q Continues toward the posterior end of the animal and into the tail

q Number of somite pairs is constant in a given species but can vary widely among species

q Stages of development often standardized according to number of somites formed

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a: Courtesy Kathryn Tosney; b: © Ed Reschke/Getty Images

Neural tube

(b) Segmentation in an early chick embryo as seen with alight microscope

(a) Segmentation in an early chick embryo asseen with a scanning electron microscope

Toward posterior end

Epithelial somite

Unsegmentedmesoderm

Toward anterior end

AnteriorSomitesPosterior

5 μm

Event 5: Organogenesis

n Organs have two or more tissue types

n Each germ layer gives rise to particular types of cells found within different organs

n Many organs form during or just after neurulation

n Organs can become functional at different times during development (lungs do not function until after birth)

n Development controlled by Hox genes

n Proteins or growth factors induce cells in local vicinity to differentiate along a specific pathway

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n Two primary mechanisms to convey positional informationq Autonomous specification – differential acquisition

of various cytoplasmic factors during cell divisionq Conditional specification – acquisition of properties

through cell-to-cell signaling mechanisms

n Together these provide cells a continuously changing internal and external environment –in which cells fulfill their unique spatial and functional fates 41

Control of Cell Differentiation and Morphogenesis in Animal Development n Two main mechanisms mediate communication

q Morphogensq Cell-to-cell contacts

n Morphogens – substances eliciting cellular responses based on concentrationq Autonomous specification − morphogen gradient

results in daughter cells that have unequal amounts of the morphogen in their cytoplasm - determines how the cell differentiates

q Conditional specification – morphogenic gradient can be established in the embryo by secretion into extracellular fluids

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Cytoplasmic factors, including morphogens,are asymmetrically distributed in the egg.

Following fertilization and cell division, the factorsare asymmetrically distributed to daughter cells.

(a) Autonomous specification

(b) Conditional specification

Cell-to-cell signaling by exocytosisof stored morphogens

Binding of membrane proteinsto each other

Cytoplasmicfactors

Signal

Signal n Cell-to-cell contactsq In conditional specification involves proteins that are

present in the cell membrane and that can interact with other proteins in the cell membranes of other cells

q Interaction generates intracellular signals

q Play a major role in determining the final positioning of individual cells within different regions of an embryo

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n Conditional specification often involves cellular inductionq Process by which a cell or group of cells induces

a response in a neighboring group of cells in the embryo

q Animal cap assay – has been used extensively to identify morphogens secreted by embryonic cells that induce cells in the animal hemisphere to differentiate into mesoderm

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Clumps ofectodermresemblingskin

Bloodlikecells

Musclecells

Notochord-like cells

Heart cellsActivin(very highconcentration)

Activin (highconcentration)

Activin(moderateconcentration)

Activin (lowconcentration)

Control(no activin)Animal

pole

Vegetalpole

Animalcap

Culture medium

Late-stageblastula

n Different classes of cells are held together by cell-to-cell contact

n Cells within ectoderm, mesoderm, and endoderm express different genes that encode distinct cadherin proteins

n Only cadherins of the same type can bind to one another

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Ectodermalcell

Cytoskeletalelements

Cadherin

Canbind

Cannotbind

Cadherin-cadherinbinding

Mesodermalcells

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Groups of Embryonic Cells Can Produce Specific Body Structures Even When Transplanted into Different Animals

n Morphogenetic fieldq Group of embryonic cells that ultimately produce a

specific body structureq Removing limb field produces an embryo lacking that

limb and transplanting the field to other areas produces the limb in that area

q Cells within fields are uniquely specified to become particular embryonic structures before any physical evidence of that structure can be observed

n Spemann’s organizerq Hans Spemann discovered this field in the early gastrulaq “Organizer” secretes morphogens responsible for

inducing formation of embryonic axisq First Nobel Prize in developmental biology


1 2 Completion of embryonic developmentTransplantation

Pigmented gastrula of donorembryo (Triton taeniatus)

Dorsal lip of blastopore

Nonpigmented gastrula ofhost embryo (Triton cristatus)

Secondary (induced)nonpigmented embryo

Primary embryo

Primary structures(nonpigmented cells)

Neural tube

Notochord

Neural tube

NotochordSecondary structures(mostly nonpigmentedcells)

Richard Harlan and Coworkers Identified Genes Expressed Specifically in the Organizer

n Hypothesized that morphogens in the organizer would promote formation of anterior structures (the head)

n Used expression cloningq Purified mRNA from organizer, constructed cDNA

library and transcribed each back into mRNA

n Injected mRNAs into UV treated eggs that would not have formed dorsal mesoderm

n Looked for evidence of “rescue” of UV treated embryos

n Noggin protein later revealed to act by inhibiting two other morphogens known to induce ventral mesoderm

n Antagonistically acting proteins specify structures in a concentration dependent fashion

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Conceptual level

UV light inactivates a morphogen and prevents dorsalmesoderm formation; the embryos usually die.

Most fertilized eggs formed only ventral mesoderm anddied. One, however, developed into a tadpole (rescue).

Isolate mRNA from dorsallip tissue, which containsthe organizer.

Expose unfertilized Xenopuslaevis eggs to UV light, theninject with an mRNA. Fertilizethe eggs to determine which,if any, develop. Note: Eachegg was injected with asingle type of mRNA.

Inject with UV-treated eggsincreasing amounts of themRNA that rescued theembryo in step 3.

Rescue mRNA from step 3

Protein product of mRNA

Low number ofproteins formed.

Moderatenumber ofproteins formed.

Many proteinsformed.

Many copiesof mRNA

Moderatenumber ofcopies of mRNA

Few copiesof mRNA

Egg

Fertilize

mRNA injected into egg

UV

mRNAFertilized egg

GastrulaDorsal lip

Blastopore

Extract andpurify mRNA.

Experimental level

Surgical scissors

HYPOTHESIS Cells within Spemann’s organizer express specific genes that encode proteins that regulate the development of dorsal structuresduring gastrulation.

KEY MATERIALS Xenopus laevis gastrulas and eggs.

mRNA

+2 DNA is isolatedand transcribedinto mRNAin vitro.

Clones of bacteria,each containing adifferent cDNA

Create cDNA library. UsecDNA library to make15,000 different mRNAs.(See Chapter 20 for adescription of cDNAlibraries.) Each bacterium

takes up 1 plasmid.

Plasmid

cDNA ligatedinto vectorBacterium

Contains manydifferent mRNAsexpressed in theorganizer (~15,000different mRNAs)

Xenopus laeviseggs

Vegetalpole


(5): © Richard Harland, U.C. Berkeley

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THE DATA

Embryo fromUV-treated egg

Partialrescue

Normaltadpole

Abnormal tadpoles with excessdorsal and anterior development

CONCLUSION Cells in Spemann’s organizer secretea morphogen—termed noggin—thatdirects normal dorsal development inXenopus laevis embryos.

SOURCE Smith, W.C., and Harland, R.M. 1992.Expression cloning of noggin, a newdorsalizing factor localized to theSpemann organizer in Xenopus embryos.Cell 70:829–840.

Increasing amounts of dorsal mesoderm-promoting mRNA and its protein product

n Spina bifida – failure of neural tube to closeq 7 in 10,000 births in USq Up to 75% may be prevented if women take folic acid

n Tobacco, alcohol, and other drugs can also have severe and devastating consequences on a developing fetus

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Impact on Public Health n Fetal alcohol syndrome (FAS) – the leading cause of mental retardationq Morphological features caused by effects of ethanol

on cell divisionq Cognitive deficits result from death of developing

CNS neurons

n Down syndrome (trisomy 21) – results in multiple physical and neurological disorders

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n Cleft lip or palate (or both combined)q 1:1,000 births in the US)q Appear to share both genetic and environmental basisq Treatment is surgical repair

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