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SEA URCHINS as model organisms

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1

SEA URCHINS as model organisms

are echinoderm

easily obtainable

transparent embryos

used in mosaic model and regulative development experiment

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2

C.ELEGANS as model organisms

small

large embryo batches

short generation time

easy to read

fully sequenced genome

easy scoring of phenotype

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3

cell division of embryo/cleavage

cleavage is asymmetric, and can determine what each cell will give rise to

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4

PAR proteins

allow one part of the cell to become different to the other (allow for asymmetry)

131 cells undergo apoptosis (essential for proper development)

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5

improper regulation

can lead to disease`

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6

RNA interference (RNAi)

controls the flow of genetic info during development

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7

Thomas Hunt Morgan

established Drosophila as a genetic model

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8

Why is drosophila used

have similar genes that control development to humans

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9

Forward genetics

if a developmental gene is mutated this should lead to defects and illustrate their function

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10

how to forward genetics

starts with a mutant

function is known but the gene sequence needs to be determined

positional identification/cloning to find gene

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11

Reverse genetics

starts with a known gene sequence but function needs to be determined

gene knockout experiment

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12

Saturation screening

uses a chemical to randomly damage and mutate DNA

treatment is adjusted so 1 in 500 genes are destroyed

if 2000 lines are screened 98% chance to find mutant gene

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13

CMS in flies

chemical used to destroy and mutate gene

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14

Mutant screens use

allow for a basic understanding of how gene control

molecular identification of new genes and signalling pathways

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15

use of studying genes

  • pathways

  • homeostasis

  • cancer

  • regeneration

  • aging

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16

Drosophila life cycle

  1. fertilisation

  2. zygotic nuclei undergoes rapid divisions to create syncytium

  3. nuclei migrate to periphery of cytoplasm after 90 mins

  4. syncystal blastoderm formed after 2 hours (poles separate)

  5. after 3 hours membrane invaginates each individual nuclei

  6. gastrulation: mesoderm invagination

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17

Drosophila after hatching

  • instar larvae

  • malt to become second

    • malt again to become third

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18

Imaginal discs

part of larva that will become part of adult insect during pupal transformation

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19

Life cycle duration

9 days

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20

Life span duration

140 days

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21
  1. ANTERIOR

  2. POSTERIOR

  1. front end

  2. back end

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22

Syncytium

a single cell with cytoplasm with a large number of nuclei

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23

Drosophila development

after 1 day larva is formed that has clearly visible body segments

3 Thoracic and 8 abdominal

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24

Zebra fish (adv. and disadv.)

ADV

small

large number of embryos

transparent

DISADV

90 days to mature (slow)

slow life cycle

complex genome and gene duplication

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25

Mouse (adv. and disadv.)

ADV

small

mammal

rapid generation time

inbred stains

DISADV

poor accessibility

small embryo batches

expensive

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26

Frogs (adv. and disadv.)

ADV

external fertilisation

large embryos

robust

large embryo batches

DISADV

not very transparent embryos

long generation time

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27

Chick (adv. and disadv.)

ADV

large embryo

tetrapod

DISADV

not accessible early

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28

All vertebrates are _____

very similar in development similar embryonic stages

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29

defining structures of vertebrates

pharyngeal pouches and somite’s and pharyngula segmented backbone

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30

metameric structure

repeated structures

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31

somatic cells

not passed on makes up body

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32

homologue

chromosome from each parent; only one is packaged into gamete

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33

Blastomeres

1st cells produced after fertilisation every 30 mins the cells divide

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34

equation for final number of cells after division

Nstart x 2^tf = Nfinish N → number of cells tf → time x frequency of divison

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35

how is the gg activated

Ca2 release triggered by sperm entry wave of Ca2 travels across the egg egg completes meiosis meaning development can begin kinases that control cell cycle to initiate cleavage are activated by Ca2+

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36

Ca2+ use in egg

increase is necessary and sufficient for egg development oscillations in Co2+ synchronise cell division

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37

mouse embryo compaction

cadherin molecules stick to each other and the cytoskeleton; expression causes compaction

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38

Gastrulation

different for each vertebrate formation of 3 germ layers movement of cells to inside of embryo to form the endoderm and mesoderm cells that remain on surface form ectoderm establishes AP and DV axis

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39

what is part of the ectoderm

neurones glia neural crest placodes epidermis pigment cells

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40

what is part of the mesoderm

muscle cartilage bone dermis heart blood

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41

what is the endoderm

gut lungs

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42

epithelium and mesenchyme

the first cell types mesenchyme has no defined shape and move easily gives rise to mesoderm and endoderm epithelium: more structural/cuboidal and stay in sheet/cluster

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43

forces that drive cell and tissue rearrangements

cell shape changes cytoskeletal rearrangements changes in expression of cell surface proteins migration localised cell proliferation cell death morphogenesis (creation of shape)

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44

Zebra fish development

early cleavage→ gastrulation → somite-ogenesis somites form from A→ P vertebrate body is segmented at the end of gastrulation, mesenchymal cells gather dorsally

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45

what are somites

a transient structures that form following gastrulation in the mesoderm

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46

neural tube development

it is the brain and spinal cord arises from the ectoderm morphogenesis takes place by cell shape changes

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47

drosophila summary

3 mm 30 day lifespan hatches from egg as a larva 2 larval stages: pupa and metamorphosis

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48

step 1 of drosophila development

mitosis begins after fertilisation however cytokinesis does not occur in early drosophila embryo syncytial blastoderm

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49

step 2 of drosophila development

at 10th division, nuclei migrate to periphery

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50

step 3 of drosophila development

at 13th division, 6000 nuclei are partitioned into separate cells cellular blastoderm

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51

step 4 of drosophila development

single epithelial layer of cellular blastoderm gives rise to 3 germ layers ectoderm, mesoderm and endoderm

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52

step 5 of drosophila development

gastrulation occurs when future mesoderm in the ventral region invaginates, germ band extension occurs and Para-segments can be seen

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53

step 6 of drosophila development instar stage

larva malts shedding its cuticle twice

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54

step 7 of drosophila development

after the 3rd instar, larva becomes pupa and metamorphosis occurs

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55

how is the neurogenic region formed

in asymmetry ventrolateral-ly low concentration than nuclear dorsal

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56

how are neurones formed

mesoderm invaginates and neuroectoderm comes to lie ventrally to give rise to neurones (and ectoderm skin cells)

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57

what is DPP

helps to set up the D/V axis high levels in flies define dorsal

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58

what is BMP

helps to define D/V axis in vertebrates high levels define ventral

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59

pro-neural cluster

group of equivalent cells a single neural cell is selected from

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60

lateral inhibition

process used to select a single cell from a small group

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61

how is the selection process initiated

notch-delta pathway (cell-cell signalling) Achaete Scute protein promote delta expression which is a transmembrane ligand- can only influence neighbouring cells delta binds to activate notch receptors small differences in cells = different delta expression levels

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62

role of notch

downregulates Achaete Scute signal and therefore small amount of A.S will be amplified high and continuous A.S expression activates neural genes and the bottom/losing cell reverts to an epidermal fate

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63

cell dropping following lateral inhibition

neural and glial cells generated asymmetric cell division cell drops from epithelium into embryo all cells have apico-basal polarity one will differentiate like a stem cell while the other will become a gangion mother cell

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64

A/P axis (antero-posterior )

Head, tail, thorax and abdominal region thorax and abdomen are segmented

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65

D/V axis (dorso-ventral)

Amnioserosa dorsal ectoderm ventral/neuroectoderm mesoderm

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66

Nusslein - Volhard

discovered the genes that control the development of body axes discovered through screens for developmental mutants

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67

Antero-posterior patterning

genes can be grouped in a hierarchy axes are fully established in syncytium blastoderm stage

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68

breaking of initial symmetry

initial BICOID gradients result in expression of GAP genes that define different embryo regions

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69

maternal gene and BIOCID

mother provides information to set up initial 2 axes BIOCID is an example of a morphogen a transcription factor and morphogen (unusal as t.fs cannot cross membranes)

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70

Morphogen

any substance that triggers growth, proliferation and differentiation of cells forms a gradient across the A/P axis of the syncytial embryo from A end

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71

Postulated

suggest a fact as a basis of reasoning

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72

Zygotic GAP genes

lead to periodic expression of pair-rule genes to specify para-segments

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73

Para-segmentation

elaborated by pair rule genes foreshadow actual larva segments leads to segmentation gene activation

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74

cell cellularisation

no longer syncytium cell-cell communication needs to be established to set up patterning in each segments patterning occurs after segmentation gene is activated → cellularisation occurs → cell signalling can occur

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75

selector genes

at this point there are 14 segments (fairly identical) Homeotic selector genes gives segments precise characteristics

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76

How is symmetry broken

perpendicular axes by maternal genes provided by the mother genetic screens can detect these genes

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77

what are Nanos and Caudal

proteins required for proper formation of the posterior segments

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78

how does Nanos work

its RNA is tightly localised but it is stuck to posterior end of the oocyte under the influence of Oskar protein RNA is translated to protein to form a BICOID opposing gradient

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79

role of NANO

control the translation of maternal RNA coding for hunchback by preventing its translation in the posterior therefore posterior patterning (prevents posterior expression of hunchback)

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80

Caudal role

important for posterior patterning prevented from translating in the anterior by BICOID

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81

Torso signal

comes from outside the embryo and only activated at the anterior and posterior pole

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82

Torso receptor

expressed on the outside of the embryo and present everywhere on egg membrane into the vitelline

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83

Perivitelline space

between the egg membrane and the vitelline membrane where the torso receptor projects

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84

Torso

a ligand that torso binds to

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85

how is the receptor only activated at the poles?

ligand needs to be proteolytically cleaved to function and the cleaving protein is localised to the poles of the egg active trunk is produced at the pole and captured by the receptor at the nearest source creating a gradient

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86

Cell-cell signalling

to communicate, cells use signals that cannot pass through membranes and instead is received by transmembrane receptors

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87

Dorsoventral polarity (back and belly)

generated under the influence of a signal through the Toll receptor and takes the form of Spätzle ligand

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88

spatzle ligand and Toll receptors

both are everywhere localised enzyme pipe is on ventral side and activates both locally activation = nuclear localisation for dorsal t.f.

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89

oocyte

an immature egg that requires maturation before fertilisation

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90

egg and oocyte formation

one cyst cell will become an oocyte other 15 are nurse cells follicle cells provide important signals for the oocyte to form an egg chamber ovariole strings are polar structures in the A/P directions and polarity info is transferred to the oocyte

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91

what do nurse cells do?

produce protein, RNA and other material for the egg

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92

polarisation process

stalk communicates to the follicle cells if the signal coincides with GURKEN signal from oocyte, they will become posterior follicle cells no Gurken signal = anterior follicle cells

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93

posterior follicle cells and microtubules

microtubules rearrange with positive end towards the end of the oocyte and their negative end at the anterior

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94

function of microtubules

transport BICOID RNA to anterior end and Oskar RNA to posterior end kinesin to positive end and Dynein to negative end

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95

how does Gurken set up the D/V axis

orientated microtubules can push nucleus to one side nucleus produces localised RNA that encodes for Gurken protein to create localised signals signal makes dorsal follicles (differs from the ventral ones)

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96

D/V polarity

nuclear localisation of dorsal protein is high on ventral side and low on dorsal side

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97

Twist and snail gene promoters

they have a low affinity to dorsal binding sites only expressed when high level of nuclear dorsal is present makes mesoderm

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98

Rhomboid

high affinity to dorsal binding site expressed by twist and snail expressed laterally on both sides of mesoderm makes neuroectoderm

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99

low level of nuclear dorsal leads to

rhomboid expression and neuroectoderm

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100

high levels of nuclear dorsal

snail expressed → snail expressed → rhomboid expression blocked

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