2616.5.2 Molecular Regulation of Anteroposterior Patterning
6.5.2.1 Nodal and Wnt Antagonists as Regulators of Anterior Patterning
Clues to the molecular regulation of anteroposterior patterning first came from the
characterization of conserved Nodal and Wnt growth factor antagonists expressed
in the amphibian organizer endoderm region and in the amniote hypoblast/AVE (see
below). Xenopus cerberus (cer) was identified through differential screening of dor-
sal lip-specific molecules (Bouwmeester et al. 1996 ). Cer overexpression on the
ventral side could induce multiple second axes consisting only of heads (Kerberos
was the multi-headed guard dog of the underworld in Greek mythology; Hesiod and
Evelyn-White 1914 ). This head-inducing activity was attributed to the ability of Cer
to antagonize multiple developmental signaling molecules extracellularly, including
BMP, Wnt and Nodal proteins (Piccolo et al. 1999 ), although Nodal antagonism is
likely paramount. The activity of Cer could largely be recapitulated by co- expression
of the Cerberus Nodal antagonist domain (Cerberus-short, CerS) with Chrd.
Additionally, overexpression of nodal1 (nodal homolog 1) after the mid-blastula
transition (i.e., from a plasmid vector) resulted in anterior truncations (Piccolo et al.
1999 ). Additional studies with amniote Cerl suggested the anti-Wnt activity is mini-
mal in mouse and chick and that Cerl represents mainly a Nodal and BMP antago-
nist (Marvin et al. 2001 ). These and studies in zebrafish using Lefty, another Nodal
antagonist, established that inhibition of Nodal activity is critical for proper pattern-
ing of anterior fate in the axial mesendoderm (Thisse et al. 2000 ).
In addition to anti-Nodal signals, Wnt inhibitors are expressed within the orga-
nizer and in the hypoblast/AVE and Wnt signaling is antagonistic to anterior develop-
ment in a stage-dependent manner. Although Wnt signaling is critical for axis
induction, other experiments show that activating Wnt/beta-catenin signaling during
gastrulation causes anterior truncations (Christian and Moon 1993 ; Kelly et al. 1995b;
Hoppler et al. 1996 ) whereas inhibiting late Wnt signaling results in hyperanterioriza-
tion (Hoppler et al. 1996 ; Leyns et al. 1997 ; Wang et al. 1997 ; Glinka et al. 1997 ,
1998 ). Additionally, inhibition of BMP and Wnt signaling together is needed to
induce secondary axes with head structures, whereas induction with BMP antagonists
alone generates only trunk structures (i.e., notochord; Glinka et al. 1997 ). Genetic
studies in zebrafish tcf3 (headless, hdl) mutants show a reduction in anterior struc-
tures and expanded ventrolateral derivatives (Kim et al. 2000 ), whereas the opposite
occurs in wnt8a mutants (Kim et al. 2000 ; Lekven et al. 2001 ). Wnt antagonists are
also required in vivo for anterior patterning. Interference with Dkk1 function in fish
and frogs leads to anterior truncations (Hashimoto et al. 2000 ; Shinya et al. 2000 ),
mimicking Wnt overexpression. Similarly, genetic deletion of Dkk1 in the mouse
epiblast led to loss of forebrain neural markers (Mukhopadhyay et al. 2001 ).
Additionally, Dkk1 is a direct target of Otx2, a key anterior specifying gene, in both
the anterior mesendoderm (Ip et al. 2014 ) and anterior visceral endoderm (Kimura-
Yoshida et al. 2005 ). And, either overexpression of Dkk1 or additional deletion of one
Ctnnb1 allele is sufficient to partially rescue loss of Otx2 (Kimura-Yoshida et al.
2005 ), suggesting that controlling Wnt levels is a critical role of Otx2.
6 Vertebrate Axial Patterning: From Egg to Asymmetry