Bio 120 2005 lecture 11

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Bio 120 2005 lecture 11

• Drosophila
• Dorsoventral axis formation
• segmentation

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summary of A-P pattern

• oocyte microtubules become polarized in egg
  chamber (ultimate cause polarity of ovariole)
• bicoid mRNA transported to anterior
• specifies acron, head, thorax by activating
  hunchback, repressing caudal
• nanos (etc) mRNA ltransported to posterior
• allows posterior development by repressing
  maternal hunchback
• signals from follicle cells specify termini

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are genes conserved?

• caudal--yes
• functions in posterior development in worms and
  vertebrates
• nanos--sort of
• function in germ cell development, not polarity
• bicoid--only found in some flies
• ancestral anterior morphogen may be hunchback
• as in vertebrates, earliest polarities are crude
  and flexible in evolution

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The dorsoventral axis

• like AP polarity, involves reciprocal signals
  between oocyte and follicle cells
• coordination of oocyte and eggshell polarity
• 1. oocyte to follicle cells, dorsalizing
  gurken ? torpedo
• mutants have ventralized eggshell and oocyte (13
  genes)
• 2. follicle cells to oocyte, ventralizing
  spätzle ? Toll
• mutants have normal eggshell, dorsalized oocyte
  (12 genes)
• 3. zygotic dorsalizing factors (dpp/sog)
• mutants ventralized (12 genes)

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1. symmetry breaking oocyte signals to specify
dorsal follicle cells

• oocyte nucleus moves to one side along
  MTs--stochastic?
• signals to nearby follicle cells (gurken again!)
• gurken transcribed by oocyte and locally
  translated receptor ubiquitous

Fig 5.12
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2. follicle cells signal back to ventralize
follicle cell epithelium

• mutants dorsalized--little tubes of naked cuticle
  (hence snake, nudel, spätzle)
• all follicle cells can send this signal, but
  gurken inhibits those on dorsal side
• protease cascade in perivitelline space, cleaves
  spätzle precursor into active ligand on ventral
  side
• Toll receptor on oocyte plasma membrane,
  ubiquitous

oocyte
perivitelline space
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Toll pathway in embryo
Fig 5.8
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The dorsoventral pattern of fates
dorsal

• 1. amnioserosa (extra-embryonic)
• 2. dorsal ectoderm (epidermis)
• 3. ventral ectoderm (neurectoderm)
• 4. mesoderm (will invaginate)

ventral
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3. zygotic readout of DV polarity

• gradient of Dorsal protein in ventral half
  turns on various genes, represses dpp
• in dorsal half, dpp expressed
• lateral cells express sog

Fig 5.14
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protein gradients along DV axis
Fig 5.15

• countergradients of DPP and SOG (not simple LSDS)
• DPP BMP-like, SOG chordin-like

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Do insects and vertebrates use the same mechanism
to pattern DV axes?

• Vertebrate (Xenopus)
• BMP ventralizes
• noggin/chordin dorsalize by inhibiting BMP
• Insect (Drosophila)
• DPP dorsalizes
• SOG dorsalizes by inhibiting DPP
• Xenopus noggin can dorsalize fly embryo
• convergent or divergent evolution? if divergent,
  how come the gradients are inverted?

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are there similarities in body plans?

• Cuvier animals belong to four divisions
  (vertebrates, articulates, molluscs, radiates)
  that are fundamentally different

Geoffroy St. Hilaire note the similarity
between body plan of lobster and vertebrate if
one is inverted
See section 15.7
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Geoffroy may have been right (after 170 years)
DPP

• similarity BMP inhibition on side that forms
  CNS
• difference is in relative position of mouth
• from blastopore in insects (protostome)
• forms secondarily on other side in vertebrates
  (deuterostome)

insect
blue CNS red CV system
Inhibitor (SOG)
vertebrate
chordin
sog
inverted insect
BMP-4
DPP
Figure 15.18
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Development of AP axis

• Segmentation
• division into regular repeating units (metameres)
• Segment diversification
• making segments different--next lecture
• two processes are independent but must be
  coordinated

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Models for periodic patterns in space

• Clock and wavefront
• progressive segmentation
• somitogenesis
• Reaction-diffusion models
• self-organizing standing waves
• stripes of pigmentation (zebra etc)

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segmentation genes

• Nusslein-Volhard and Wieschaus, 1980
• screens for zygotic pattern mutants
• 30 genes, 3 classes, forming spatial hierarchy
• GAP
• PAIR-RULE
• SEGMENT POLARITY

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phenotypic hierarchy

• gap mutants
• lack multiple adjacent segments
• other segments form OK
• pair-rule mutants
• lack alternating segments
• have 7 double-wide stripes (instead of 14)
• segment polarity mutants
• every segment missing same part of pattern
• since 1980s genes cloned and regulation studied
  at molecular level

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The gap genes

• 7 genes hunchback, Kruppel, giant
• transcriptionally activated by maternal proteins
• themselves encode txn factors
• proteins unstable, form transient concentration
  gradients

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the gap proteins are local morphogens

• hunchback protein in gradient (thanks to bcd,
  nos)
• high HB represses Kr
• medium HB activates Kr
• low HB has no effect
• result Kr txn turns on only in single thin
  stripe!
• how did they figure it out? by altering HB gene
  dosage and looking at Kr expression

Fig 5.18
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cross-inhibition sharpens gap domains

• gradients overlap at first
• each gap gene activates itself and represses
  others
• result each level of AP axis has unique
  combination of gap proteins at blastoderm stage
• each gap domain spans several segments--how are
  specific segments defined?

an example of lateral inhibition
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the pair-rule genes

• gt8 genes even-skipped (eve), fushi tarazu
  (ftz), etc
• txn turned on by gap proteins before
  cellularization
• encode transcription factors
• expressed in alternating parasegments (PS), not
  segments

Fig 5.21
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Parasegments

• first overt signs of segmentation (at extended
  germ band) are pits in ectoderm
• originally thought to correspond to segment
  boundaries
• in fact out of register--segment boundaries form
  later, between pits
• embryonic repeating unit named the parasegment

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Animations from scanning EMs by Thom Kaufman
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pair-rule gene expression is dynamic

• example even-skipped
• stage 10 expression low, uniform
• stage 14 (cellularization) 7 distinct stripes
• stripes initially fuzzy, then sharpen anterior
  borders (refinement)--involves autoactivation

one parasegment
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how do we get from non-periodic gap domains to
periodic pair-rule patterns?

• eve stripe 2 (PS3)
• combinatorial control by gap proteins
• BCD and HB activate
• GT, KR repress
• equivalent mechanisms for other stripes

Fig 5.22
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combinatorial control of transcription involves
binding to enhancer regions
eve

• the eve stripe 2 enhancer
• 600 bp regulatory element in DNA of gene
• many binding sites for gap proteins
• note e.g. GT binding site overlaps HB site

activators
repressors
Fig 5.23
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evidence that stripes are made piecemeal
make flies expressing these transgenes
see the LacZ or GFP turn on in these patterns
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reporter (LacZ, GFP)
1. eve regulatory DNA
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