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B3.0 > Developmental Mechanisms Of Morphological Change > Flashcards

Developmental Mechanisms Of Morphological Change Flashcards

1
Q

Developmental mechanisms of morphological change

A

Finding the pathways that change

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2
Q

When evolution changes development

A

Can we find the pathways/networks involved?

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3
Q

Two main strategies

A

1) candidate pathway approach
2) hypothesis-free approach

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4
Q

Transposition

A
  • “sliding” of homologous gene segments along a segmented body plan
  • e.g. no of ribs changes in different segments in vertebrate skeletons
  • change segment specification? Add segment?
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5
Q

Is transposition underpinned by

A

Hox genes?

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6
Q

Hox gene expression has changed between species

A
  • to modify the end-product of development
  • e.g. Hoxc6 in mice, chick and goose by in situ localisation
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7
Q

Modification of the Hox pathway correlated with

A

Development change

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8
Q

Crustacea

A
  • transposition
  • head appendages = feeding
  • thorax appendages = locomotion
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9
Q

Artemia

A
  • brine shrimp
  • Ubx-Ab stains thorax and posterior (swimming limbs /=/ feeding)
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10
Q

Ubx

A

A hox protein

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11
Q

Triops

A
  • Ubx-Ab stains thorax and posterior (swimming limbs /=/ feeding)
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12
Q

Mysidium

A
  • T1 modified for feeding, not locomotion
  • maxilliped
  • Ubx not expressed; slid
  • faint in T2
  • expressed in T3
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13
Q

Lobster (Homanes)

A
  • T1 & 2 modified for feeding
  • Ubx not rxot eeed
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14
Q

Crustacean evidence

A
  • correlational
  • we need interventional
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15
Q

Parhyale

A
  • amphipod
  • T1: Ubx not expressed (maxilliped)
  • T2: Ubx expressed (gnathopod)
  • RNAi @ embryo stage (siRNA injection) induces partial transformation: T2->T1-like
  • hatchling SEM
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16
Q

Oligodactyly

A
  • mice = ancestral mammal
  • cow/pig/camel digits: stands on 2, 2 highly reduced (more symmetrical)
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17
Q

Oligodactyly H

A
  • Shh
  • expressed in limb bud posterior
  • manipulating concs changes digits no
  • e.g. in chicle
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18
Q

Shh

A

It is not where the RNA, but the protein is, that matters

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19
Q

Compare Shh protein patterns

A
  • travels further
  • downstream target genes more symmetrical
  • cellular cascade of initiation
20
Q

patched (ptc) and smoothened; Gli

A
  • sequester Shh
21
Q

Less ptc in Oligodactyly

A
  • Shh diffuses further back
22
Q

Oligodactyly mutation

A
  • ptc receptor has insertion mutations in cis-regulatory region of limb regulatory module in intrinsic regions A (2.4kb) and B (1.4kb)
  • suppressed expression and altered spatial distribution
23
Q

Observing Shh regulation

A
  • GFP-lacZ transgenics
  • in mice: expression lower and shifter distribution
  • mutation expands Shh influence
  • not necessarily causative; could be many mutations in other genes
24
Q

Tunicates

A
  • Molgula oculata
  • tadpole larva
  • oral/atrial siphon
  • brachial basket
  • tunic
  • digestive system
  • tail-less form is derived; lost
25
Q

Hypothesis

A

In tail-less tunicates, notochord cells don’t stack and extend; was it changes to structural genes?

26
Q

M. oculata

A
  • 40 notochord cells that intercalate
27
Q

M. occulta

A

20 notochord cells in that do not intercalate

28
Q

Tunicate hybrid

A

20 notochord cells that intercalate

29
Q

Notochord

A
  • secrete collagen and stack as discs; causes tail growth
30
Q

Methodology :

A

1) extract RNA from different species @ right developmental stage
2) RNA-Seq
3) map
4) DEG analysis
- examine known notochord genes

31
Q

Tunicates results

A
  • 32 known notochord genes ^ in tailed but not tailless
  • not pseudogenes, so must be regulatory change
  • 2 collagen, 2 laminin, 2 collagen-processing enzymes
32
Q

Does changing collagen expression affect the tail?

A
  • intervention yet
  • Ciona robusta: CRISPR collagen KO
  • tail formation affected
  • mild: wonky
  • severe: lost
33
Q

What else to do re Tunicates?

A
  • Sequence before and after tail emergence ; what has changed?
  • are collagens or laminins represented
34
Q

Darwin’s finches

A
  • adaptive radiation
  • one of the first studies adopting a hypothesis-free approach
35
Q

Short, stumpy beaks

A
  • medium ground finch (G. fortis)
  • large ground finch (G. magnirostris)
36
Q

Long, pointy beaks

A
  • cactus finch (G. scadens)
  • large cactus finch (G. conirostris)
37
Q

Beak shape

A
  • shape of mandible (upper jaw) via skull
  • crucial
  • RNA-Seq
38
Q

How to study gene regulation in embryonic mandible

A

1) RNA-Seq
2) microarray

39
Q

RNA Seq

A
  • newer
  • more expensive
  • more sensitive
  • need a large amount of tissue
  • Darwin’s finches are protected
40
Q

Microarray

A
  • traditional
  • extract RNA from different species and compare expression to outgroup/ reference species
41
Q

Microarray process

A

1) extract from maxilla (embryo beak region)
2) different species different label (Cy3/Cy5)
3) hybridise
4) convert to cDNA
5) spot cDNA into glass slide; each spot represents an RNA
6) scan
7) set results: look for genes affecting beak length

42
Q

Microarray results

A
  • c100 show consistent difference correlating w beak shape but not overall size
  • e.g. calmodulin (Ca2+-binding protein) affects calcium envrt; signalling
43
Q

Test: is calmodulin capable of changing beak length? If

A
  • chickens: experimentally tractable
  • expression in frontonasal mandible
  • use RCAS delivery to constitutive express downstream CaMKII
  • beak length increases 10%
  • sufficient
44
Q

RCAS

A

Retroviral

45
Q

CaMKII

A

CaM effector

46
Q

Evolution has tweaked a calcium signalling pathway during radiation of Darwin’s finches

A
  • one developmental perspective
  • probably other pathways too
47
Q

Other potential factors

A
  • IGFBP (insulin)
  • β-catenin (Wnt pathway)
  • Kruppel Factor TF
  • incomplete understanding!

B3.0 (23 decks)

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