Download March 22 – signals in frog embryos

Signals in frog embryos
How can we identify developmental
signaling pathways and test their roles?
How do other vertebrate embryos develop?
Developmental biology jargon
Cell fate – the normal future identity of a cell (muscle, skin,
etc.). A fate map represents future cell fates.
Specification – the process by which a cell acquires a
particular fate. Specified cells could still switch to some other
fate in response to perceived signals. A specification map
represents the identities that cells have acquired at a
particular stage.
Determination – the process by which cells fix their fate stably.
Determined cells do not change fate even if they receive new
signals.
Differentiation – the process by which cells actually change to
take on a particular identity (e.g. express muscle-specific
genes).
Mesoderm induction and patterning
~64-cell stage
late blastula
gastrulation
What experimental results demonstrate that a
particular signal induces a response?
What experimental results demonstrate that a
particular signal induces a response?
Location – The signal must be present in the right place and at the
right time.
Sufficiency – The signal must be able to induce the response
(perhaps outside normal context).
Necessity – When the signal is removed or blocked, the response is
also blocked.
Using these logical criteria to discover and
test signals that might regulate a response.
Location – Find molecules that are present in the right place and at the
right time.
Separate mRNA or proteins from different places, look for differences.
Sufficiency – Identify potential signaling molecules that can induce the
response (perhaps outside its normal context).
Inject mRNAs or proteins corresponding to putative signals.
Use gain-of-function mutations or transgenes.
Necessity – Remove or block the signal, test whether the response still
occurs or not.
Loss-of-function mutations
Inhibitors of particular pathways
RNA interference
Mesoderm induction and patterning
~64-cell stage
late blastula
gastrulation
Vegetally localized transcripts and proteins include
both signals and transcription factors
Vg1 mRNA
TGF- receptors activate gene regulatory
proteins at the membrane
The TGF-
family
includes
Vg1, nodal,
Xnr, activin,
BMPs.
Some of
these are
localized to
the vegetal
region.
Maternal VegT (transcription factor,
vegetally localized) activates
zygotic Xnr (Xenopus Nodal-related)
genes on vegetal side.
VegT also specifies endoderm.
The TGF-
family
includes
Vg1, nodal,
Xnr, activin,
BMPs.
Some of
these are
localized to
the vegetal
region.
Activin can induce different mesodermal cell
types at different concentrations
Activin can induce different genes at
different concentrations
General mesoderm
Dorsal mesoderm (organizer)
TGF- receptors dimerize after binding ligand
The TGF-
family
includes
Vg1, nodal,
Xnr, activin,
BMPs.
Some of
these are
localized to
the vegetal
region.
Dominant negative activin receptor poisons dimers
(might also block other TGF- signals)
Another important pathway in inducing dorsal mesoderm
-catenin immunolocalizations:
dorsal side
-catenin is a transcription factor
ventral side
Wnt signaling pathway
-
+
-
+
-
+
+
-
-
+
-
+
Inject dominant-negative GSK3 protein at 2-cell stage
Current model of frog mesoderm induction
(=Xnr)
Mesoderm induction and patterning
~64-cell stage
late blastula
gastrulation
noggin expression in blastula
Dorsal side
[noggin]
none
low
Noggin protein
injection rescues
UV-irradiated
embryos
medium
high
very
high
(behaves as a morphogen)
noggin expression in dorsal mesoderm
(also acts at later stages after gastrulation has begun)
Model for
mesoderm
patterning
Antagonize BMP-4
Antagonizes Xwnt-8
TGF- receptors activate gene regulatory
proteins at the membrane
The TGF-
family
includes
Vg1, nodal,
Xnr, activin,
BMPs.
BMP4 acts in
ventral
mesoderm.
Mesoderm identities
~64-cell stage
late blastula
gastrulation
Brachyury expression in blastula
Maternal
patterning
signals set up
zygotic gene
expression
Brachyury and
Goosecoid are
transcription factors
Activin can induce different genes at
different concentrations
General mesoderm
Dorsal mesoderm (organizer)
Injection of goosecoid mRNA into ventral
vegetal cells of 16-cell embryo
Control embryos (not injected)
How do other embryos develop?
Chick gastrulation:
the squashed frog model
form yolk sac
(extraembryonic)
Chick gastrulation
(equivalent to organizer in frogs)
Helpful web links
Tutorials on echinoderm and Xenopus gastrulation:
http://worms.zoology.wisc.edu/embryology_main.html
Human embryo development:
http://embryology.med.unsw.edu.au/
Various movies:
http://www.sdbonline.org/archive/dbcinema/