DEVELOPMENTAL BIOLOGY
CHAPTER 1
Developmental biology: The anatomical tradition
- without DNA , without cell biology
Development
1. slow progressive change
2. a zygote proceeds to organism by mitosis
3. embryology: fertilization to birth
4. development biology: fertilization onwards (puberty, aging, etc.)
Questions:
differentiation
-the descendants of the zygote are of hundreds (200) of kinds of cells; How is that possible?
morphogenesis
- How are cells organized into tissues, organs, and limbs
growth
- How can it be rapid, yet tightly controlled?
reproduction
- how are gametes produced?
evolution
- changes in mature organisms are dependent on changes in developmental priorities; How do they occur?
environmental integration
- how does the environment affect development?
Approaches to Developmental Biology
The Anatomical Approach
Comparative embryology – looks at how differences between organisms develop
Aristotle, 352 BC:
Oviparity- egg birth
Viviparity- live birth
Ovoviviparty- eggs retained internally (some sharks)
Holoblastic cleavage
Meroblastic cleavage
Harvey, 1651 AD:
Ex ovo omnia
Blastoderm of chicken
Epigensis and preformation
Epigenesis- it meant: a “force” , “form”, or “soul” directed the formation of organs from formless “goo,” associated with upward social mobility : now it refers to the control of developmental processes independent of genetic control
Preformation- the egg, or sperm, contains the embryo in miniature before fertilization, the gamete is like a Russian doll, containing all future generations, the form of all future generations has been decided and can’t be improved, associated with stratified class systems
Now we know: In a sense, the “preformed” DNA contains the information that allows the epigenesis from the organized (preformed) material of the egg. Epigenesis is more correct than preformation.
1820’s: Naming the parts: The primary germ layers and early organs
ectoderm
endoderm
mesoderm
triploblastic
diploblastic- 2 layers; sponges and jelly fish
induction- one “determined” tissue tells another what it should become
pharyngeal arches: discovered by Rathke, become gill supports in fish and other structures in mammals
The four principles of Karl Ernst von Baer (discoverer of the notochord)
1. The general features of a large group of animals appear earlier in development
than do the specialized features of a smaller group.
2. Less general characters develop from the more general, until finally the most specialized appear.
3. The embryo of a given species, instead of passing through the adult stages of lower animals, departs more and more from them
4. Therefore, the early embryo of a higher animal is never like a lower animal, but only like its early embryo.
Fate mapping the embryo: what part of the embryo becomes what
cell lineages: who are the ancestors of the adult’s differentiated cells?
Observing Living Embryos: tunicates; different cells, different pigments
Vital Dye Marking: fig. 1.8
Radioactive Labeling and Fluorescent Dyes: fig. 1.9
Genetic Marking: chimeric embryos
Cell Migration of melanocytes from neural crest: fig. 1.11
Evolutionary Embryology
Embryonic homologies
Homologous
Analogous (homoplastic)
Medical Embryology and Teratology
Malformations (lack of coordination)
syndromes
animal models
disruptions
teratology
Mathematical Modeling of Development
The mathematics of organismal growth
isometric growth
allometric growth (or allometry)
The mathematics of patterning: reaction-diffusion model
CHAPTER 2
Life cycles and the evolution of developmental patterns
The Frog Life Cycle- compare to sea urchins, etc.
vegetal hemisphere vs. animal hemisphere in the egg
fertilization
egg pronucleus and fusion of the pronuclei
Cleavage makes blastomeres that form a blastula
gastrulation: forms gastro-intestinal tract
- the gastrula has the 3 germ layers
blastocoel
blastopore- opposite the site sperm entry
dorsal vs. ventral
blastopore lip
organogenesis- forming the organs, begins with the neural tube
neurula
neural tube
neural crest cells
somites- segmented precursors to bone and muscle
larvae are free-living embryos
gametes come from germ cells ( not somatic cells) by gametogenesis, the first differentiation in development
metamorphosis- radical changes
The Evolution of Developmental Patterns in Unicellular Protists
morphogensis: The role of the nucleus; Acetabularia (web: 2.2)
Unicellular protests and the origins of sexual reproduction
reproduction
conjugation: paramecium
sex: Chlamydomonas
- sexual reproduction
- mating types (not male and female)
isogamous
heterogamy
oogamy
Multicellularity: The Evolution of Differentiation
The volvocaceans (colony or multicellular?)
4 cells: flat plate
16 cells: flat plate
16
32
64 cells in a regular sphere (pattern formation)
2000 cells: Volvox: germ cells-> specialization and death
-> death becomes inevitable as the result of multicellualrity and sex
inversion = proto-gastrulation?
Differentiation and morphogenesis in Dictyostelium: Cell adhesion
The Life Cycle of Dictyostelium
slug -like pseudoplasmodium or grex
prestalk
prespore
Aggregation of Dictyostelium Cells
cyclic adenosine 3’5’ –monophosphate (cAMP)
Cell Adhension Molecules in Dictyostelium
Differentiation in Dictyostelium
Bias (change their minds when isolated)-> Labile Specification (only change their minds when transplanted) -> Commitment and Differentiation (will not change their minds)
Developmental Patterns among the Metazoa
Metazoans
- sponges: no digestive tract, or tissue, Archeocyte
Diploblastic
Triploblastic
Protostomes and deuterostomes
Protostomes
Coelom
Ecdysozoa
Lophotrochozoa
Deuterostome
Schizocoelous vs. Enterocoelous
amniote egg
yolk sac
amnion
allantois
chorion
Chapter 3
Principles of Experimental Embryology
Environmental Developmental Biology
Environmental sex determination
Sex Determination In An Echiuroid Worm: Bonellia
Sex Determination In A Vertebrate: Alligator
Adaptation of embryos and larvae to their environment
Phenotypic Plasticity
- map butterfly
- oaks and caterpillars
The Abnormal Influence of UV Light
a) sea urchin
b) frogs
-interaction of UV, species and frog
The Developmental Dynamics of Cell Specification
differentiation
-the zygote/Embryonic stem cell proliferates quickly, has no specific function yet can give rise to any of the 200 cell types as its descendants (totipotent)
-as cells differentiate in to different types, they lose proliferation capacity and potency while gaining specific functions
-pluripotent ("lots of potential") stem cells are found in umbilical chords and bone marrow etc
-cancer cells typically lose specific function and gain proliferation capacity, most cancers look like stem cells
commitment
specification- they can change their mind
determination- they can't change their mind
Autonomous specification: leads to mosaic development because of morphogenetic determinants (chemicals like mRNA or proteins in the cytoplasm that control cell fate)
-common in invertebrates
Conditional specification: leads to regulative development because pluripotent cells respond to their environment, position and each other
syncitial specification: interactions happen within one cytoplasm not between cells
Morphogen Gradients
-morphogen: a hormone that controls morphogenesis
-concentration gradient: a difference in concentration over a distance
germ plasm theory- incorrect, chromosomes are broken up into the appropriate cell
-the opposite of genomic equivalence (Weisman)
Tests of germ plasm theory:
1. defect experiment- incorrect interpretation
2. isolation experiment
3. The recombination experiment
4. transplantation experiment
The Influence of Neighboring Cells
-a pair of "animal" cells from a 16 cell sea urchin embryo will produce ectoderm and mesoderm, although they usually only make ectoderm
-several pairs together can't make mesoderm; probably every cell wants to become mesoderm but they send out signal that inhibits mesoderm formation by each other
- in another experiment, the exact same number of presumptive gut (endoderm) cells will be converted into skeletal mesoderm (the closest to the animal pole) as the number of presumptive skeletal mesoderm cells that are removed; probably every gut cell has tendency to become skeletal mesoderm but is inhibited by skeletal mesoderm signals
Morphogen Gradients
-morphogen: a hormone that controls morphogenesis
-concentration gradient: a difference in concentration over a distance
-activin is a morphogen in Xenopus toad's vegetal hemisphere; higher concentrations induce presumptive ectoderm to become more dorsal/posterior mesoderm
- each animal cap cell has 500 activin receptors on their cell surface
- when no activin receptors are occupied, the cells become ectoderm
- at higher concentrations, 100 are occupied, the brachyury gene is expressed, this is only found in mesoderm , they become ventrolateral mesoderm
- at still higher concentrations, 300 are occupied, and goosecoid is also expressed, they become notochord cells, a dorsal mesoderm
Regeneration: parts of the flatworm know what they are supposed to reproduce because of their position in a concentration gradient; it seems the head produces a morphogen, the tail destroys it, this maintains a gradient, the lack of cell to cell contact tells the planaria that something is missing; the gradient tells it what direction the missing part lies in and at what position in the body the cut lies.
Morphogenetic Fields: the region within the field commits to making an organ but the cells within the region can still regulate themselves
example: a presumptive Drosophila "knee" transplanted from the leg imaginal disc to the antenna imaginal disc tip becomes a claw (a tip of a leg); the knee cells were committed towards making a leg but not which part
analogy: flags
limb field: salamander forelimb; frogs with extra limbs because of damage from parasites
Stem Cells and Commitment
Stem cells
-totipotent zygote and embryonic stem cells
-pluripotent stem cells
-committed stem cells
Progenitor cells, Precursor cells:
Morphogenesis and Cell Adhesion
1. How are tissues formed from populations of cells
2. How are organs constructed from tissues?
3. How do organs form in particular locations, and how do migrating cells reach their destinations?
4. How do organs and their cells grow, and how is their growth coordinated throughout development?
5. How do organs achieve polarity?
mesenchymal cells vs. epithelial cells
Differential Cell Affinity
selective affinity
- cells stick to some other cell types better than others
histotypic aggregation
- is when cells of the same tissue are attracted to each other
The thermodynamic model of cell interactions
-WHATEVER cells have the greater homotypic affinity will end central, etc.
Cadherins and cell adhesion
cadherins
catenins
E-cadherin
P-cadherin
N-cadherin
EP-cadherin
Protocadherins
homophilic binding
trophoblast- cadherins to the uterine wall
inner cell mass- cadherins to each other, not the uterine wall
Chapter 4: The Genetic Core of Development
The Embryological Origins of the Gene theory
Nucleus or cytoplasm: Which controls heredity?
-Acetabularia
- morphogenetic determinants are genes: Morgan (originally)
- chromosomes contain genes: Wilson
-Boveri:
fertilized sea urchins by polyspermy (2 sperm)
the first cleavage results in 4 cells (not 2)
the resulting cells have abnormal distribution of chromosomes and an abnormal phenotypes
-Nettie Stevens:
fruit fly:
XO or XY = male
XX= female
- Morgan:
-sex-linked traits
- becomes a geneticist rather than an embryologist
- genetics = transmission of traits vs. embryology
The split between embryology and genetics
geneticists, please explain:
1. genomic equivalence and differentiation
2. genetics of development
3. environmentally controlled sex determination
Early attempts at developmental genetics ( a fusion of the 2 fields)
1. brachyury mutation: aberrant notochord, disrupts dorsal-ventral axis in the posterior of the mouse
2. Waddington: discovered fly wing mutations and then showed how mutants abnormally develop
Evidence for Genomic Equivalence
Amphibian cloning: The restriction of nuclear potency
- somatic nuclear transfer
- restriction of efficiency during development
Amphibian cloning: The totipotecy of somatic cell nuclei
- serial transplantation
1. take nuclei from foot webbing
2. transfer to enucleated egg
3. take nuclei from that blastula
4. transfer to a second enucleated egg
5. those embryos develop
Cloning mammals
1. take udder cell
2. grow in a dish
3. serum starve (food, but no growth hormones)
4. take egg cell in metaphase 2 of meisosis
5. remove spindle and nucleus
6. fuse egg and de' uddder cell with electrical current
7. implant blastocyst
Differential Gene Expression, evidence:
- cytokinesis - the division of the contents of the cytoplasm
- mitosis- duplication and division of the nucleus
- cells can go through mitosis without cytokinesis (like the syncytial embryo)
-polytene chromosome: DNA duplication without mitosis or cytokinesis
RNA Localization Techniques
-Northern blotting
-electrophoresis
-Reverse transcriptase-polymerase chain reaction
-polymerase chain reaction (PCR)
-reverse transcriptase (RT)= a viral enzyme that produces a DNA (cDNA) version of a RNA molecule
-Microarrays and macroarrays
- In situ (at the site)hybridization
-Antisense mRNA; inhibits translation by producing a complementary RNA to your gene of interest
Interfering RNA - iRNA
- when you produce a double stranded RNA corresponding to your gene of interest, most cells will think it is a virus and destroy RNA with that sequence, shut down expression of that gene, and may even prevent its expression in the next generation
-whole-mount in situ hybridization
Determining the Function of Genes During Development
microinjection
transfection
electroporation
transposable element or retroviral vector
P elements
Transgenic Mice
- produces a mouse with an extra gene, it is useful if you are looking at the effects of mutated genes that would not be obscured by the normal genes still being there (like mutated oncogenes)
-embryonic stem cells (ES cells)
-chimeric mouse
Gene Targeting (Knockout) Experiments
-eliminate normal genes from zygotes, so you can see the results of their loss
Antisense RNA
Morpholino Antisense Oligomers
The Paradigm of Differential Gene Expression
Developmental genetics- transforming genotype into phenotype
Anatomy of the gene:
chromatin- DNA and protein
histones- major protein of chromatin
nucleosome- histones and a loop of DNA
exons- coding sequences that exit the nucleus as mRNA
introns- sequences of DNA that do not end up in mRNA
promoter- a DNA sequence that directs mRNA sequences
transcription initiation site
cap sequence- 5' sequence
translation initiation site- in an exon
5’ UTR/leader sequence
translation termination codon
3’ untranslated region
polyadenylation- poly (A) tail
Anatomy of the gene: Promoters and enhancers
TATA box- most promoters, 35 bp upstream
basal transcription factors
TFIID; [TATA-binding protein (TBP)]
TBP-associated actors (TAFs)
upstream promoter elements
Cell-specific transcription factors (Pax 6, Myf)
enhancer- a sequence of DNA that binds proteins that
- control the efficiency of promoters,
- determines in what cells and when genes will be transcribed
reporter genes:
β-galactosidase gene
green fluorescent protein (GFP)
silencers
Transcription factors
DNA-binding domain
trans-activating domain
protein-protein interaction domain
MITF- transcription factor that recruits histone acetyltransferase ; www.devbio.com
Pax6
Transcription factor cascades: no transcription factor need be tissue specific, combinatorial control is at work
Silencers- DNA sequences that actively repress transcription
- neural restrictive silencer element (NRSE)
-histone deacetylase: repress transcription
Methylation Patterns and the Control of Transcription
DNA methylation and gene activity
- Chromatin modification
Acetylation vs. Methylation!!!! (stop him!)
Insulators- insulate enhancer activity
Transcriptional Regulation of an entire Chromosome Dosage Compensation
dosage compensation
X chromosome inactivation
heterochromatin
euchromatin
Barr body
Differential RNA Processing
Control of early development by nuclear RNA selection
nuclear RNA (nRNA)
Creating families of proteins through differential nRNA splicing
Alternative nRNA splicing
spliceosomes
splicing isoforms
proteome-all the proteins an organism produces
Control of Gene Expression at the Level of Translation
Differential mRNA longevity
Selective inhibition of mRNA translation
small regulatory RNAs (micro-RNAs)
Control of RNA expression by cytoplasmic localization
Posttranslational regulation of gene expression
Chapter 6
Cell-Cell Communication in Development
induction- one group of cells changes the behavior of another set of cells
- optic vesicle induces ectoderm to become lens
competence- the ability to respond to induction
- head ectoderm is competent because it expresses Pax6
Cascades of induction: Reciprocal and sequential inductive events
Instructive interactions- induction of new gene expression
permissive interaction- the environment allows gene expression
Reciprocal Inductions- induced structures in turn induce the inducer (lens and neural retina)
Regional Specification- inducers from different regions of the embryo can induce region appropriate structures (wing, thigh, foot epidermis induced from prospective wing epidermis by wing mesenchyme), the specificity is an instructive interaction, the formation of any cutaneous structure is permissive
Genetic specification- a cell is limited by genes and epigenetic factors which control what it can become
Epithelial-mesenchymal interactions
epithelial-mesenchymal
Regional Specificity of Induction
Genetic Specificity of Induction
Paracrine Factors
juxtacrine interactions
paracrine interaction
paracrine factors
growth and differentiation factors
endocrine factors
The fibroblast growth factors
fibroblast growth factor (FGF)
fibroblast growth factor receptors
The Hedgehog family
The Wnt family
The TGF-β superfamily
bone morphongenetic proteins
Other paracrine factors
Cell Surface Receptors and Their Signal Transduction Pathways
signal transduction pathways
The receptor tyrosine kinase (RTK) pathway
receptor tyrosine kinase (RTK)
G protein
Ras
Guanine nucleotide releasing factor (GNRP)
GTPase-activating protein (GAP)
sevenless
bride of sevenless
Vulval induction in Caenorhabditis elegans
vulval precursor cells VPCs)
anchor cell
equivalence group
stem cell factor
Kit
The Smad pathway
Smad
The JAK-STAT pathway
STAT
thanatophoric dysplasia
chondrocytes
The Wnt pathway
Frizzled
The Hedgehog pathway
Cell Death Pathways
Programmed cell death/apoptosis
Bcl-2 family
Apaf1
caspase-9
caspase-3
Juxtacrine Signaling
The Notch pathway: Juxtaposed ligands and receptors
Delta, Jagged, or Serrate
Notch
The extracellular matrix as a source of critical developmental signals
Proteins and Functions of The Extracellular Matrix
extracellular matrix
Fibronectin
Laminin and type IV collagen
basal lamina
Integrins, The Receptors for Extracellular matrix moleclules
integrins
Direct transmission of signals through gap junctions
gap junctions
connexin
Cross-Talk between Pathways
cross-talk
Maintenance of the Differentiated State
Coda
Chapter 7
Fertilization: Beginning a new organism
Structure of the Gametes
Sperm
acrosomal vesicle/acrosome
acrosomal process
head
flagellum
axoneme
tubulin
dynein
capacitation- is the final maturation of the sperm that take place in response to chemical signal from the egg (sea urchin) or the reproductive tract (mammals)
The egg
ovum
oocyte
Proteins
Ribosomes and tRNA
Messeger RNA
Morphogenetic factors
Protective chemicals
cell membrane
vitelline envelope- invertebrate glycoprotein membrane becomes the fertilization envelope, sperm binds here in a species specific way
zona pellucida- the mammalian vitelline envelope
cumulus- "cloud" of ovarian follicular cells around ovum
corona radiata- inermost layer of cumulus
cortex- outermost layer of cytoplasm, contains globular actin (will become microfilaments which also support microvilli)
cortical granules- female equivalent of the acrosomal vessicle, there are 15,000 in a sea urchin, contains proteolytic enzymes and mucopolysaccharides
egg jelly- glycoprotein that attracts sperm
Recognition of Egg and Sperm
I) Sperm attraction: Action at a distance
resact- chemotaxis, sea urchin, also a sperm-activating peptide that causes increase in metabolism and motility
II) The acrosome reaction in sea urchins
acrosome reaction- caused by specific proteins in egg jelly,
- exocytosis of the acrosomal vessicle,
- extension of the acrosomal process,
-exposure of bindin
III) Gamete binding and recognition
Species-specific recognition in sea urchins
bindin- only 1500 sperm can bind to sea urchin egg at one time, therefore their seems to be a bindin receptor on the cell surface
Gamete binding and recognition in mammals
ZP3: The Sperm-Binding Protein of the Mouse Zona Pellucida
Induction of the Mammalian Acrosome Reaction by ZP3: binding, then acrosomal reaction (no process)
galactosyltransferase-I - binds ZP3
Traversing the Zona Pellucida- digest through, bind ZP2 (ZP3 binding is lost)
Gamete Fusion and the Prevention of Polyspermy
IV) Fusion of the egg and sperm cell membranes
fertilization cone- of the egg grabs sperm, formed by actin, membranes fuse
The prevention of polyspermy
monospermy
polyspermy- disaster
The fast block to polyspermy in sea urchins:
- resting membrane keeps out Na+, keeps K+ in
-Na+ comes in, membrane potential changes
- membranes can no longer fuse
The slow block to polyspermy in sea urchins:
cortical granule reaction
cortical granule serine protease - dissolves vitelline posts
fertilization envelope- mucopolysaccharides absorb water, cause membranes expansion
peroxidase- crosslinks adjacent proteins, hardens membrane,
hyaline- forms a layer right over cell membrane
hyaline layer
In mammals:
ZP3 is disabled by n-acetylglucosaminidase
Calcium as The Initiator of the Cortical Granule Reaction
The Activation of Egg Metabolism
Early responses
IP3: Releaser of Calcium Ions
inositol 1,4,5-trisphosphate (IP3)
phospholipids phosphatidylinositol 4,5-bisphosphate (PIP2)
phospholipase C (PLC)
diacylglycerol (DAG)
protein kinase C
Phospholipase C: Generator of IP3
Late responses: preparation for cell growth, increase in pH, protein synthesis, eventually mitosis, movement of morphogentic determinants
V) Fusion of the Genetic Material
Fusion of genetic material in sea urchins
female pronucleus
male pronucleus
mitochondria usually come from mom, while the centrosome usually comes from dad
zygote nucleus
Rearrangement of the Egg Cytoplasm
gray crescent
parthenogenesis
CHAPTER 8
Early Development In Selected Invertebrates
Cleavage- rapid cell division (fig. 8.1)
-no increase in embryo size,
-completely dependent on maternal factors (except in mammals)
- G1 and G2 abolished
blastomeres
mitosis-promoting factor (MPF): makes mitosis possible by phosphorylating histones, nuclear envelope proteins, etc.
- composed of:
-cyclinB: is destroyed after every mitosis; the time it takes to accumulate is the time till the next mitosis
-cyclin-dependent kinase: adds phosphate groups to proteins, activated by cyclins
mid-blastula transition- when the blastomeres take over the cell cycle from the maternal cytoplasmic factors; G1 and G2 appears again
The cytoskeletal mechanisms of mitosis
karyokinesis- mitosis, can happen without cell division
mitotic spindle- microtubules, etc., that moves the chromosomes
cytokinesis- division of the cytoplasmic contents
contractile ring- actin filaments that pinch the cell membrane, perpendicular to the mitotic spindle
cleavage furrow- bisects the plane of mitosis
Patterns of embryonic cleavage
vegetal pole- yolk slows down cleavage furrows
animal pole- divides faster
isolecithal- sparce, evenly distributed yolk
- must have a hungry larval form or help from a placenta
holoblastic- cleavage furrow extends through the entire egg
meroblastic- a portion of the egg never divides
- yolk inhibits membrane formation
centrolecithal- yolk in the center forces superficial cleavage
telolecithal- lots of dense yolk
-both merblastic and discoidal
- only a small portion of the animal pole divides: called discoidal cleavage
Gastrulation
- the formation of the three germ layers
Invagination- infolding of sheet of cells into the embryo
Involution- one layer folds in and under another
Ingression- individual cells move into the embryo
Delamination- one sheet splits into two
Epiboly- one sheet moves over another
Cleavage in Sea urchins
radial holoblastic cleavage- meridional (meridian) and perpendicular to each other
mesomeres- medium-sized cells of the animal hemisphere produced by the third cell division
macromeres- big cells of the vegetal hemisphere produced by the cell division
micromeres- very small cells of the vegetal hemisphere, induce the blastopore
Blastula formation
blastocoel- every cell is contact with this
vegetal plate- flat, thick cells at the vegetal pole where the blastopore will form
hatched blastula- swimming embryo
Fate maps and the determination of sea urchin blastomeres
Cell Fate Determination- potential is greater than fate except for large micromeres which will always be skeletal and induce the blastopore
β-catenin- is high in all vegetal hemisphere cells
- induces endo/mesoderm
- with otx in micromeres it turns on:
Pmar1- which represses a represser of mesenchymal genes
Specification of the Vegetal Cells- micromeres induce adjacent cells to be secondary mesenchyme rather than endoderm
Sea Urchin Gastrulation
- forms the pluteus larva
Ingression of primary mesenchyme- skeletogenic mesenchyme
filopodia- skinny psuedopods
Importance of Extracellular Lamina Inside the Blastocoel
- primary mesenchyme cells lose their affinity for the hyaline layer by 10 fold and increase their affinity for the extracellular matrix inside the blastocoel by 100 fold
- primary mesenchyme also "reads" the extracellular matrix to find their proper, equatorial location
First stage of archenteron invagination
1.the blastopore is caused by bottle cells that swell on the blastocoel side and the swelling inner lamina of the hyaline layer caused by the presumptive secondary mesenchyme that secrete a chondroitin proteoglycan
Second and third stages of archenteron invagination
2. convergent extension: cells intercalcate
- extends and narrows gut
3. secondary mesenchyme cells grab the ventral surface of the balstocoel with filopodia and pull the archenteron up
filopodia- they are like skinny pseudopods
Chapter 9
The Genetics of Axis Specification in Drosophila
Cleavage- the egg is centrolecithal, therefore it produces a syncytial blastoderm
Superficial cleavage- nuclei migrate to the periphery;
energids- after cylce 10, the area under the control of one nucleus
after cycle 13, cells form the cellular blastodernm, undergoes the the mid-blastula transition (maternal cytoplasmic determinants lose control) cell division becomes asynchronous
pole cells-germ cells at the posterior
Gastrulation
ventral furrow- pinches off to become a tube of mesoderm within the embryo, it will wrap around the periphery, and cover endoderm that pinches off from the anterior and posterior
cephalic furrow- a bend that cross sections the anterior the embryo
germ band- the ectoderm wraps around the posterior of the embryo
The Anterior-Posterior Axis
maternal effect genes- cytoplasmic determinants, Mom's nucleus is in control
Bicoid - mRNA is sequestered at the anterior
Hunchback- protein is only translated at the anterior
Nanos- mRNA is sequestered at the posterior
Caudal- protein is only translated at the posterior
Segmentation Genes
gap genes- whole portions of the embryo (gap regions) are missing when they are mutated
pair-rule genes- divide the gap regions into parasegments
segment polarity genes- distinguish between the anterior and posterior of each segment
homeotic selector genes- are turned on in response
The gap genes
Giant, Kruppel, Knirps, tailless all are expressed in specific regions of the embryo in response to maternal effect genes
Giant- activated by high levels of Hunchback and Caudal
Kruppel- inhibited by hunchback, knirps and tailess
Knirps- acitvated by bicoid inhibited by hunchback, activated by Caudal
tailless- activated by torso, hunchback(?)
Pair-rule Genes
These genes are expressed in stripes.
Each stripe has its an enhancer that drives the expression of the gene in one or a couple of stripes.
Theses enhancers are controlled by the complex interaction of maternal effect genes and gap genes.
even-skipped is a good example of this.
fushi tarazu expression is repressed by even-skipped and enhanced by itself. This creates alternating bands of eve and ftz separated by narrow gaps, each band and gap corresponds to a parasegment
Segment Polarity Genes
wingless- is expressed in these gaps, it is repressed by eve and ftz, this will be the middle of the segment
engrailed- drives the expression of hedgehog, engrailed is expressed wherever there are high levels of eve or ftz
wingless and ftz cells give each other positive feedback
engrailed expression marks the posterior of each segement
The Homeotic Selector Genes
- determine the identity of each segment
Patterns of homeotic gene expression
homeotic selector genes
Antennapedia complex
bithorax complex
homoetic complex (Hom-C)
homeotic mutants
halteres
Initiating the patterns of homeotic gene expression
Maintaining the patterns of homeotic gene expression
Realisator genes
Dorsal: The Morphogenetic Agent for Dorsal-Ventral Polarity
Dorsal
Translocation of Dorsal to the nucleus
The signal cascade
Signal from the oocte nucleus to the follicle cells
lgurken
torpedo
Signal from the follicle cells to the oocyte cytoplasm
pipe
Establishing the dorsal patterning gradient
separation of the dorsal and cactus proteins
Effects of the dorsal protein gradient
Axes and Organ Primordial: The Cartesian Coordinate Model
Chapter 10
Early Development and Axis Formation in Amphibians
Cleavage in Amphilbians
- radial and holoblastic but the egg is mesolecithal
- cleavage is extemely slow in the vegetal hemisphere
- the 2nd division starts before the 1 st finishes
- no G stages until after the 12 th division
-cd2 activity inhibited by lack of cyclin B and phosphorylation
morula- 16-64 cells; "mulberry"
Amphibian Gastrulation
The Xenopus fate map
- both the ectoderm and the endoderm are on the outside of the the blastula
- in Xenopus, the mesoderm is inside around the equator
prechordal plate- head mesoderm enters, first mesoderm to enter the blastopore
chordamesoderm- notochord, the second mesoderm to enter the blastocoel, important for nervous system induction
The mid-blastula transition: Preparing for gastrulation
mid-blastula transition (MBT) - specific embryonic genes' promoters are de-methylated and support gastrulation
Positioning the blastopore
- opposite the site of sperm entry, becomes the anus and the end of the dorsal surface
- the egg already has an anterior/posterior axis
-the blastopore determines the dorsal/ ventral axis and bilateral symmetry
Invagination and involution
Cell movements during amphibian gastrulation
bottle cells
- swell into the blastocoel under the dorsal lip of the blastopore, below the equator (the marginal zone) and on the opposite side of sperm entry,
- important, but not critical
-the endoderm (IMZ-S) folds in and under the ectoderm, through the blastopore, dragging the mesoderm along with it, called vegetal rotation
involuting marginal zone (IMZ)- those cells that move into the blastocoel
The convergent extension of the dorsal mesoderm: cells move forward by intercalating (two layers fusing to form a longer one)
cause intercalating cells to preferentially adhere to each other:
- the expression of these cell adhesion molecules: paraxial protocadherin and axial protocadherin:
- calcium causes actin contraction which allows cells to migrate towards each other
noninvoluting marginal zone (NIMZ) and animal cap cells move by epiboly to cover the entire embryo surface (collectively, ectoderm)
Epiboly of the ectoderm
-cells increase in number, deep and superficial cells also intercalate
The Progressive Determination of the Amphibian Axes
Hans Spemann:
gray crescent- future blastopore, only embryos with grey crescent develop normally
regulative (conditional or dependent) development- possible in early gastrulas
autonomous (independent or mosaic) development- likely in late gastrulas
all this indicates that the blastopore is causing embryonic cells to commit, acts as the embryonic organizer
and Hilde Mangols:
- in the amphibian, only the BPL is determined by maternal morphogenetic determinants
primary embryonic induction:
- the blastopore causes the first induction in a chain reaction of inductions
Mechanisms of Axis Determination in Amphilbians
Nieuwkoop center
- at the vegetal pole, sequesters Dishevelled , which then rotates to the grey crescent and inhibits GSK-3, which usually degrades β-catenin which helps cadherins and acts as a nuclear factor
- so β-catenin accumulates on the dorsal side
Tgf-β like proteins- are expressed throughout the marginal zone, induced by vegetal endoderm
together, these drive goosecoid expression which is found in cells on the dorsal side of the embryo
The Functions of the Organizer
1. self-differentiate
2. dorsalize mesoderm into paraxial mesoderm
3. induce the neural tube
4. initiate the movements of gastrulation
The diffusible proteins of the organizer : The BMP inhibitors
bone morphogenesis protein 4 (BMP4)- induces epidermis formation
Noggin - induces neural ectoderm, by blocking epidermis formation
Chordin and Nodal-related protein 3- prechordal and chordal mesoserm
Follistatin - inhibits epidermis formation
Chapter 11
The Early Development of Vertebrates: Fish, Birds, and Mammals
Cleavage in Fish Eggs
telolecithal yolk, causes meroblastic,discoidal cleavage in the blastodisc
yolk cell - birds don't have this
blastoderm- cap of cells
yolk synchytial layer (YSL)- multiple nuclei in the yolk cell
internal YSL - deep
external YSL- in the cortex
enveloping layer - superficial blastoderm that is pulled down over the embryo, becomes the transient periderm
deep cells- will form the almost all of the embryp, see fate map
Gastrulation in Fish Embryos
The formation of germ layers:
germ ring- equatorial swelling of the embryo
epiblast - upper layer of cells
hypoblast- lower layer of cells, presumptive mesoderm and ectoderm, probably produced by involution from the epiblast
embryonic sheild- a thickening of the two layers that extends forward
chordamesoderm and notochord forms from the sheild hypoblast
paraxial mesoderm- the muscle and bone that will be associated with the spinal column
neural keel- the presumptive nervous system
Cleavage in Bird Eggs
telolecithal yolk, causes meroblastic,discoidal cleavage in the blastodisc
subgerminal cavity- under hypoblast
area pellucida-t he circle of cells one cell thick
area opaca- thick layers of cells on the periphery of the blastodisc
marginal zone (or marginal belt)- the border between the AP and AO
Gastrulation of the Avian Embryo
The hypoblast forms from a bridge that starts from Koller’s sickle to the polyinvagination islands (primary hypoblast)
The primitive streak is a thickening of the epiblast that forms anterior - posterior axis the primitive groove is a crevice in it that acts as the blastopore
primitive knot or Hensen’s node- the anterior end of primitive streak
primitive pit- the anterior end of the primitive groove
Migration Through The Primitive Streak: Formation of Endoderm and mesoderm
germinal crescent- where the germ cells live, formed by the hypoblast
Regression of the Primitive Streak
Leaves behind the notochord, the anterior embryo is more mature
Axis Formation in the Chick Embryo
The role of pH in forming the dorsal-ventral axis- the subgerminal cavity is acidic, this controls the D-V axis
-if you make the area above the epiblast acidic, you get an upside down embryo
The role of gravity in forming the anterior-posterior axis
The egg rotates, lighter components accumulate under one side of the blastodisc, which is pushed up, this will be the posterior end of the chick where Koller's sickle will form