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Universal mechanism of animal development
Gene expression controls 4 essential process
Homologous proteins=functionally interchangeable
Eyeless =Pax-6
Share basic anatomical features
Epidermal cellGut cellsMuscle cellsNeuron & sensory cells
GastrulationEctoderm-epidermis, nervousEndoderm-gut, lung, liverMesoderm-muscle, connective
gastrulation
Multicellular animals are enriched in proteins mediating cell-cell interaction and gene regulation
Genome sequencingC. Elegans 19,000 genes Drosophila 14,000Homo sapiens 30,000
50% homologsNon-conserved---1) minor importance---free to mutation2) Gene duplication
Two classes of genes (same molecules for body construction) 1. TM (cell-cell interaction) cell adhesion and cell signaling2. Gene regulatory proteins (differential gene expression)
Regulatory DNA define the program of developmentDifferent arrangement of regulatory modules
Same cell types, different body structureAssembling the components in different combination—instruction in non-coding region
Descriptive embryology—track the individual cells (cell lineage)
Xenopus : cell division, growth, movement
Experimental embryology—remove, rearrange, transplantcell and tissue interaction
Chick & Xenopus
Developmental Genetics—action of genes
Developmental Genetics—action of genes
1.Isolation of mutant animals—genetic screen Mutations in their germ cells2.Interesting abnormality3.Discover genes4.Cloning and sequencing5.How does gene work6.Regulatory DNA that controls its expression
Fruit flyC. ElegansZebrafishMouseHuman—medical care system (abnormalities compatible with life)
Cell fate determinationtransplanting test—alter environments
Specified or committed—strong tendency
Positional values—reflect their location in the bodyregionally determined—switch on and maintain expression
Signaling system that controls the differencesbetween the parts of the limb is the same—tip to be toesdetermined as leg already
Markers of position
Gene regulatory proteins-T-box
Same genome to different cell fates
(Neuronblast)
Different environmentsadjacent similar cells –exchange signals
Cell-cell contact
Notch pathway—gain an advantage—stronger inhibitory signalSelf-reinforcing
Inductive interaction—signal is limited in time and space
Short-rang-----transmitted via cell-cell contactLong range----molecule diffuses through extracellular medium
Similar cells—equivalence group or morphogenetic field
Morphogen—finely graded
Localized sourceDifferent cell fatesHigh conc.Medium conc.low. Conc.
Sonic hedgehog protein (shh) –thumb to little finger axis
Mirror duplication of the pattern of digits
2,3,4 according to their distances from the source of shh
Extracellular inhibitors of signal molecules shape the response to the inducer
Chordin—neuron tissueInhibitor of BMP/TGFb—induces epidermal
Refined by sequential induction
Morphogen—1mmCell proliferation increase sizeLocal induction-more cell fates
C.elegans--anatomically simple
1000 somatic cells, 1000-2000 germ cells
Hermaphrodite –female + limited number of sperms—self-fertilized (homozygote progeny)Male-cross fertilized
Single fertilized egg-558 cells (egg shell)Growth (further division) and sex maturationThrough 4 larval stages and molts to adults in three days
Small and transparent—follow individual cells by direct observationGenome is small
Hard to do transplantationNo similar body structure as human
Lineage analysis
Clone-one founder cell-germ line and intestineCell-cell interaction
Maternal-effect genes in asymmetric division
Sperm entry point—posterior poleMother’s mRNA—to proteins (organized in relation to this point)Par-partitioning defectiveP granules—ribonucleoprotein particles to the posterior poleVasa homologs—RNA binding protein (germ cell determining material)
Complex pattern by cell-cell interaction
What point: decisive internal changes signals from other cellsMethod:1. Microsurgery a. laser microbeam microsurgery b. early embryo, cell pushed around and rearranged c. remove egg shell—in culture
2. Genetic screen (gene cloning, sequencing) p2-EMS interaction Screen for a. no gut cells –mom mutant : more mesoderm (Wnt, Frizzled) b. extra gut-pop mutant: plenty of pharynx (LEF-1/TCF)—E cells
Developmental Biology by Scott F. GilbertSinauer Associates, publishersSix edition, 2000
Cell change over time in their responsiveness to developmental signals
Age and past history
4 cell stage ABp –Notch signalABa
12 cell stageGrand daughter ABp (no response) Aba-Notch signal--Pharynx
Heterochronic genes
Loss of lin-14—prematureGain of lin-14—stay at 1st larva stage
Division and differentiation
Less cells Cell death
Green with lin-14 which disappears at larva feeding
Lin 4, Let 7 upstream of lin 14
short untranslated RNA (21-22nts)–complementary sequences in non-coding regionControl rate of translation or degradation
Lin 4 RNA increase—in stage 1 but like in stage 3Let 7 RNA increase– late larva to adult stages
Let7 homologs , and its target homologs in human, fly and zebrafishmicroRNA84 family-down regulate Ras (Let60)Low expression in lung cancer
Programmed cell death
Control cell numbers131 dieGenetic screen-Cell death abnormalced-3—caspaseced-4—Apaf-1ced-9—Bcl-2egl-1--Bad
Cell movementXenopus
Three daysBig-transplantation
The polarity of embryo depends on the polarity of the egg
VegT-T box familyVg1-TGF-Wnt-DishevelledVegetal pole—inner tissueAnimal pole—outer tissueD/V rotate—cortex Dishevelled
cleavage
Blastomeres—smaller cells 12 cleavages—synchronously without transcriptionAsymmetric—vegetal (fewer cells, but larger)
Three germ layers
Determinants distributed asymmetrically—different cell fatesEctodermMesodermEndoderm
Blastula—epithelial sheet
Na+ pump in water in
Only outer most cells
After gastrulation, the arrangement of three germ layers
Endoderm-digestive tractMesoderm-connective tissue, muscle (vascular system)Ectoderm-epidermisNervous system
Gastrulation
Gastrulation
Mesoderm: somite, notochordSegregate from the epithelium
Chemical signals trigger the mechanical processes
Organizer –gastrulation & pattern of specialization of tissues
Chick & Xenopus
Changes of cell packing provide force
Bottle cells-narrow necks-anchor them to the surface of epithelium
Convergent extension (main force)
Frizzled/Dishevelled polarity signaling pathway
Selective cell-cell adhesion
Sorting outReconstructionCadherins (Ca2+ dependent)Differentially expressed in the various tissuesInvolved in gastrulation, neurulation, somite formation
Red-epidermalGreen-mesodermBlue-neural plate
neurulation
Mesoderm-notochord-convergent extensionNotochord expresses Brachyury (T box family)
Neural tube formation
Gene expression oscillationmesoderm to somite segmentation
C-hairy-1: pair-rule genePeak-one set of genesTrough-another set of genes
Cadherin familyParaxial protocadeherinMark out the somite
somite- to muscle-cell precursors
Muscle precursor-myoblast (MyoD)
nucleoli
The main pathways of neural crest cell migration
From epidermis
Fibronectin-provide adhesive siteChondroitin sulfate proteoglycan-repel
Effect of mutations in the kit gene
Pigment cells depend on the kit product as a receptor for a survival factor
Albino-no pigment, megacolon-lack Endothelin-3
Drosophila melanogaster
From egg to adult
Body segments
Syncytial blastoderm
Extended germ bandGastrulation
Clear segment
Segments in different stages
Development of Drosophila egg
Fate map
Egg polarity genes
Maternal effect genes
Oocyte in its follicle
Egg polarity gradient system
Dorsal gradientNF-kB
Cactus-I-kB
Patterning the dorsoventral axis
Patterning the dorsoventral axis
Mesoderm from cells expressing twist
Ventral furrow formation--invagination
Dorsoventral inversion
Dpp=BMP4 (TGF-)Sog=Chordin (high-neurogenic tissue)
Three types of segmentation genes
The regulatory hierarchy
Modular organization of the regulatory DNA of the eve gene.
The formation of ftz and eve stripes in the Drosophila blastoderm
Interactions between Gap and pair-rule genesMaintained by segment polarity and homeotic selector genes
The expression of engrailed, a segment-polarity gene
Throughout the life!
A homeotic mutation
Antenaapedia mutantRegulatory region of the gene(expressed in the head)
The effect of deleting most of the genes of the bithorax complex
Molecular address labels(each segment)-segment identityDNA binding homeodomain(60 amino acids)DNA contains homeobox
All like ps5
The spatial pattern of expression of
genes of the bithorax complex
Bithorax—Ultrabithorax –5-12 Abdominal-A—7-13 Abdominal-B—10-13
Bithorax mutant –PS 4 default state+Ubx—5,6+Abd-A—7,8,9+Abd-B—10Combinatorial mannerLack Ubx—5,6 to 4 also 7-14 thorax structure in the abdomen
Hox—gap, pair-rule for the first 4 hours, then polycomb (repression), and Trithorax (activation)
The patterns of expression compared to the chromosomal locations of the genes of
the Hox complex
Dark-high levelMedium-lowerLight (not affect phenotype)
14 parasegment-at extended germ band stage
Action of genes of the polycomb group
Permanent record of positional information1. autoactivation-homeotic selector genes2. Polycomb-represses Trithorax-on
Affect histone H4 acetylationHyperacetylated H4-Hox exposed to activator
polycomb on chromosome
The Hox complex of an insect and the Hox complexes o
f a mammal
Expression domains of Hox genes in a mouse
Hox BTransgenic mouse withLacZ reporter
Segmentation and Hox gene expression in the hindbrain
Rhombomeres=segemnt
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