Vertebrate Embryonic Patterning 6
Limb Formation
Tetrapod Limb
anterior
ventral
distal
proximal
dorsal
posterior
Limb Bud Formation
Establishment of Limb Fields
Hox Gene Expression in Trunk During
Limb Bud Development
Fig 23.21 Analysis of Biological Development 2nd ed, Kalthoff
FGF10 Induces FGF8
supernumerary limbs from
implanting FGF10 beads
Positioning of the Limb Field
• Forelimb position lies near anterior expression
boundary of hox b8, b9, c6, c9, & d9
• FGF8 expression in intermediate meso + hox
genes induces FGF10 in the lateral plate meso
• Wnts in the lateral plate maintain FGF expression
in regions of hoxd9 expression
• Between the fore & hindlimb buds, the
developing kidney (mesonephros) inhibits FGF
expression
• Exogenous FGF can induce supernumerary limbs
Forelimb vs Hindlimb
Tbx5
Tbx4
Forelimb vs
Hindlimb
Correlates with
Tbx5 & Tbx4
Expression
Respectively
Tbx4 Gain of Function Study
Tbx4
expressing
retrovirus
infected
embryo
Proximal Distal Limb Axis
• AER
– overlying ectoderm
– responsible for maintaining outgrowth of limb
• Progress zone
– underlying mesoderm
– region of cell division
– maintained in active mitotic state by AER
AER – Apical Ectodermal Ridge
AER Necessary for Limb Bud Outgrowth
Removal of AER at various times prematurely truncates limb
AER Necessary for Limb Bud Outgrowth
Reimplantation of younger
AER into older bud causes
duplication of distal structures
Reimplantation of older AER
into younger bud causes loss
of proximal structures
Interactions of
AER and PZ
FGF8 in the AER
FGF in ectoderm
AER
3 day embryo
limb buds
Hox Code of the Limb
Proximal
Hoxa & d = forelimb; Hoxc & d = hindlimb
Distal
Mammalian Hox Genes
Hox Genes Required for Proper
Limb Development
wild-type
Hoxa-11/Hoxd-11
double knockout
human with synpolydactyly
resulting from homozygous
HOXD-13 null mutation
Proximal-Distal Structures
Anterior-Posterior Limb Axis
Zone of Polarizing Activity
ZPA
Classic Experiments: Retinoic Acid
Induced Limb Duplication
• Implanting RA soaked beads caused duplication
of zeugopod/autopod structures
• Depletion of RA reduces the normal structures
Shh is Expressed in ZPA
Demonstration that Sonic Hedgehog
is Primary ZPA Component
Dorsal-Ventral Gene Patterning in Limb Bud
Fig 23.23 Analysis of Biological Development 2 nd ed, Kalthoff
Dorsal-Ventral Limb Axis
1.
2.
3.
4.
Lmx
BMPs
Fgf-8 induces Shh
Shh & Fgf-8 induce Fgf-4
All these induce Lmx-1
Wnt-7a in the ectoderm
represses Lmx-1 and
induces Shh
Molecular Interactions in Limb
Bud
Dorsal-to-Ventral Transformations
WT mouse autopod
Wnt7a-/- mouse autopod
Homology of Process: A-P Axis
Homology of Process: D-V