Limb: Anterior-Posterior
Gilbert - Chapter 16
Today’s Goals
• Become familiar with several aspects of limb
formation in the tetrapod
– Limb initiation
• Forelimb vs. hindlimb
• Where to make a limb?
– Limb patterning
• Dorsal/ventral
• Anterior/Posterior
• Proximal/distal
Anterior-Posterior Limb Axis
• Surgical experiments (Saunders again)
shows a portion of limb tissue is
important for A-P axis of limb
– ZPA tissue (Zone of Polarizing Activity)
– Located at posterior, medial portion of limb
– When transplanted to anterior, medial area
• Limb develops with 2 sets of digits, mirrorimaged to each other
The ZPA molecule!
• After lots of hunting, it was found that
the vertebrate homolog of Drosophila
hedgehog gene was expressed in this
region (in situ hybridization)
– SHH (sonic hedgehog!)
• How do we know SHH is involved?
– SHH is expressed in the posterior portion
of the limb! (ZPA)
• If ectopic SHH is placed on the anterior
margin of the wing, the same type of
mirror image 2nd set of digits forms
Ectopic SHH in Mouse
How does SHH work?
• How is SHH expression localized to the
ZPA?
• What does SHH do once it is expressed
to give the A-P identity to the limb bud?
SHH and FGF
• It appears that SHH is regulated by FGF
expression in the AER
– FGF8 activates SHH expression in cells
– THEN - why not all cells express SHH?
• Only ZPA cells must be competent to receive
SHH signal! HOW??
– Cells destined to become ZPA express Hox b-8,
dHAND
– Both are transcription factors
– Appear to allow competency to receive FGF8
signal and turn on SHH expression
• FGF and SHH then continue to promote
expression of each other in feedback loop
• SEE FIGURE 16.20 in book
SHH and Digit Identity
• Not fully understood, but. . .
– SHH appears to work in a gradient - longer
exposures to the gradient changes digit identity
– SHH appears to specify digits (1,2,3,4,5) by
determining the identity of the webbing between
the digits
– This webbing eventually undergoes apotosis and
degenerates
• SHH regulates expression of the final wave of
Hox expression in the digital tissue
– Ectopic SHH bead reverses Hox expression in a
mirror image pattern
Dorsal-Ventral Axis of Limb
• Appears to be controlled by Wnt
molecules
• Signals from the overlying ectoderm
– Wnt 7a expressed in dorsal limb ectoderm
but not ventral limb ectoderm
Coordinating the Limb Axes
• All 3 axes appear to be coordinated via
a variety of feedback interactions
– Both positive and negative feedback occur
• This is important!
– To form properly proportioned limb along
all axes
Apoptosis and Digit Formation
• Programmed cell death is important for
shaping certain areas of the limb
• Focus: Digits
– Interdigital necrotic zone
– Here, it appears BMP signaling induces
cellular apoptosis
– Noggin expression inhibits this
Limb Patterning Conclusions
• Although the formation and patterning of the
limbs is one of the best studied models in
vertebrate embryology. . .
• “There is a very large gap in our understanding of
how the activity of SHH, BMP, FGF and Wnt genes
influences, for example, where the cartilaginous
condensation will form, how the elements are
sculpted, how the number of phalangeal elements
are specified, and where the tendon/muscle will
insert” (Niswander 2002)
Introduction to Article
Assignment
• Increase our awareness of the changing
field of limb patterning.