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mice initially suggested that Wnt signaling is not active in the blastocyst prior to
implantation (Mohamed et al. 2004 ; Na et al. 2007 ). Recent studies with more sensitive
reporter constructs have detected some beta-catenin-dependent activity in the ICM/
epiblast of peri-implantation blastocysts (E3.0–4.5; Granier et al. 2011 ; ten Berge
et al. 2011 ). Similarly, transient nuclear beta-catenin has been observed by immuno-
localization at the same stage (Chazaud and Rossant 2006 ). However, the extent of
activity is unclear, since overexpression of stabilized beta- catenin does not ectopi-
cally induce Wnt target genes before gastrulation (Kemler et al. 2004 ). Thus,
although there are indications of early but non-polarized beta- catenin activity in the
preimplantation mammalian embryo, the role of this signaling appears to be dis-
pensable or easily compensated for, at least with regard to axis formation.
Correspondingly, mutants for the major Wnts, Wnt/beta-catenin coreceptors, as well
as regulators of Wnt secretion and activity all undergo normal preimplantation devel-
opment, including anteroposterior patterning in the AVE. These mutants also fail to
form mesoderm and do not gastrulate. These Wnt/beta-catenin pathway mutants
include (but are not limited to) Wnt3 (Liu et al. 1999b), Lrp5/6 (Kelly et al. 2004 ),
Mesdc1 (Hsieh et al. 2003 ), Wntless (Fu et al. 2009 ), and Porcn (Biechele et al. 2011 ,
2013 ; Barrott et al. 2011 ). Since beta-catenin mutants exhibit these same mesoderm
defects but additionally fail to form the AVE, a Wnt ligand-independent role for beta-
catenin in anteroposterior patterning has been proposed (Morkel et al. 2003 ), possibly
mediated by activation of Tdgf1 expression and subsequent effects on Nodal activity
(see Sect. 6.5.2). These results suggest possible similarities to the potential role of Wnt
ligand independent signaling in Xenopus axis formation (see Sect. 6.3.3), but too little
is known about either mechanism to draw meaningful comparisons. There are other
agonist ligands of Wnt/beta-catenin signaling, such as Norrin and R-spondin (Cruciat
and Niehrs 2013 ), but these also rely on Lrp5/6. Other potential mechanisms of regulat-
ing beta-catenin activity may exist, such as regulation through Hippo signaling (Varelas
et al. 2010 ) or through Seven in abstentia homologs (Siah) (Topol et al. 2003 ), although
these have not been extensively investigated in the context of axis formation.
Rather than relying on polarized Wnt activity, the earliest asymmetries in the
mouse blastocyst are the expression of Nodal antagonists Cerl and Lefty1 in the
primitive endoderm of the peri-implantation blastocyst (~E4.0; Torres-Padilla et al.
2007b). Expression of these genes likely depends on FoxH1-mediated Nodal signal-
ing (Takaoka et al. 2006 ; Torres-Padilla et al. 2007b), initiating a cascade of both
positive and negative feedback regulation of Nodal signaling in the embryo. Cerl/
Lefty1 asymmetry is likely to arise stochastically at this early stage, as the blastocyst
contains a rather small number of cells and it is unlikely that the few Cerl/Lefty1
expressing cells would arise precisely in the center. These cells are then thought to
lead and propagate AVE migration toward the future anterior side, i.e., towards the
side of initial asymmetry (Takaoka et al. 2011 ; Morris et al. 2012a), restricting
Nodal activity to the prospective posterior. This aspect will be discussed further in
Sect. 6.5. Additionally, Nodal is expressed at low levels in the ICM prior to implan-
tation, but there is no evidence that Wnt/beta-catenin signaling activates this expres-
sion. Later in development, prior to and during gastrulation, Nodal expression is
maintained by Wnt3 signals in the posterior epiblast and in the forming primitive
D.W. Houston