347of the germ layers. In addition, the orthologues of Activin-B and Vg1 are expressed
maternally in both frogs and fish, and can induce mesoderm and endoderm when
overexpressed. Maternal Activin is required for mesoderm formation in frogs and
Medaka fish, and is involved in oocyte maturation in zebrafish. Vg1 is required for
axis formation in both frogs and zebrafish, although the defects in zebrafish DVR1
morphants have not been well characterized.
Fgf signals have a dual role during mesoderm formation. Several members of the
Fgf family can induce mesoderm when overexpressed in vertebrate embryos, and
are expressed in the presumptive mesoderm. bFgf/Fgf2 is expressed maternally in
frogs, and bFgf/Fgf2 and Fgf4 are expressed maternally in zebrafish. Fgf3, Fgf4,
and Fgf8 are expressed zygotically in the blastula and gastrula stages in frogs and
fish. These expression patterns suggest that Fgf ligands act redundantly in meso-
derm formation. Experiments in frogs, fish, chicken, and mice indicate that both
Fgf4 and Fgf8 are required to specify mesoderm. Fgf signals also act independently
of its role inducing mesoderm to direct cell movements during gastrulation in frogs,
fish, chicken, and mice. Overexpression of the Fgf antagonist Sprouty2 blocks
convergence- extension movements in frogs without affecting mesoderm formation.
In the chicken epiblast, Fgf4 acts as a chemoattractant, while Fgf8 act as a chemore-
pellant. Fgf4 may play a similar role attracting cells to the primitive streak in mice,
but this has not been tested due to the early lethality of fgf4−/− mutants. fgf8 and/or
fgf4 are required in the mouse for ingression at the primitive streak.
7.7 Hierarchy of Mesoderm Inducing Signals
7.7.1 TGF-β and Fgf
The identification of such a large number of secreted factors presented researchers
with the challenge of understanding how these pathways interact with each other to
induce and pattern the germ layers. A series of experiments in Xenopus, zebrafish,
chicken, and mice indicated that Fgf signals acts in parallel to the TGF-β signals to
induce mesoderm, and are interdependent on each other for mesoderm formation. In
the chicken, Fgf signaling is required for formation of the endogenous streak, and
for the induction of ectopic primitive streaks by misexpressed Vg1 or Nodal
(Lawson et al. 2001 ; Skromne and Stern 2002 ; Bertocchini et al. 2004 ). In the nor-
mal embryo, cNR-1 expression precedes that of Fgf8 in the primitive streak. Finally,
Fgf8b and Nodal are more potent inducers of a secondary axis when expressed
together than when either protein is expressed alone (Bertocchini et al. 2004 ).
The synergistic action of Fgf and TGF-β is conserved in Xenopus, since animal
cap cells exposed to Fgf respond to lower doses of purified Activin or TGF-β than
cells not exposed to Fgf, and adopt mesodermal fates at lower concentration thresh-
olds (Kimelman and Kirschner 1987 ; Green et al. 1992 ). Animal cap cells expressing
a dominant negative Fgf receptor are unable to respond to Activin and other meso-
derm inducing signals (Cornell and Kimelman 1994 ; LaBonne and Whitman 1994 ;
7 Establishment of the Vertebrate Germ Layers