351primitive streak begins when the hypoblast is displaced by invasion of the definitive
endodermal cells from the KS (Lawson and Schoenwolf 2003 ). These movements
displace the Cerberus expressing cells in the hypoblast and relieves the inhibition of
Nodal in the overlying epiblast, thereby activating the positive feedback loop of
cNR-1 expression.
7.7.2.3 Xenopus
The xnr genes are expressed in the margin, in the presumptive mesoderm (Jones
et al. 1996 ; Joseph and Melton 1997 ; Takahashi et al. 2000 ). An Activin/Nodal
response element from the first intron of xnr1 is highly active in the marginal blas-
tomeres in Xenopus, where xnr1 is normally expressed, and can drive expression of
a reporter gene in the mouse primitive streak (Osada et al. 2000 ). Expression of
xnr1, -2, and -4 is induced in response to Nodal signals, and lost when Nodal-
signaling is inhibited by a Nodal antagonist (Agius et al. 2000 ; Cheng et al. 2000 ;
Takahashi et al. 2000 ). xnr1, -2, -4, -5, -6 expression is expanded in embryos lack-
ing Antivin/Lefty, and reduced when Antivin/Lefty is overexpressed (Cha et al.
2006 ). Thus, nodal-related gene expression is controlled by an autoregulatory loop
in frogs as it is in mammals and birds. Maternal TGF-β signals act to reinforce
zygotic expression of the xnr genes. In overexpression assays, the xnr genes are
upregulated in response to Activin, are required for Activin to induce mesoderm in
animal caps and are reduced when Activin B is depleted (Osada and Wright 1999 ;
Agius et al. 2000 ; Piepenburg et al. 2004 ). This provides strong evidence
that Activin is required to induce the expression of the xnr genes (Fig. 7.9a).
Although Activin B is required for xnr expression, some Activin target genes are
induced independently of Nodal (Ramis et al. 2007 ). xnr gene expression is induced
in response to Vg1, but their expression has not been fully characterized in embryos
lacking Vg1 (Joseph and Melton 1998 ; Osada and Wright 1999 ; Agius et al. 2000 ;
Birsoy et al. 2006 ). The phenotype of Vg1 depleted embryos is consistent with a
reduction in Nodal signaling (Joseph and Melton 1998 ; Birsoy et al. 2006 ).
Therefore, it is likely that Vg1 acts in parallel to Activin to induce xnr expression
(Fig. 7.9a). The T-box transcription factor, VegT, is also required to induce expres-
sion of xnr genes (Clements et al. 1999 ; Kofron et al. 1999 ; Xanthos et al. 2001 ).
Thus, in frogs, maternally provided TGF-β signals act in parallel with the VegT
transcription factor to induce xnr gene expression, which is subsequently main-
tained by an autoregulatory loop mediated in part by an Activin/Nodal Response
element in the first intron of xnr1 (Fig. 7.9a).
7.7.2.4 Zebrafish
ndr1/sqt and ndr2/cyc are expressed in the marginal zone and in the YSL in the pre-
gastrula stages (Erter et al. 1998 ; Feldman et al. 1998 ; Rebagliati et al. 1998a; Fan
et al. 2007 ). ndr2/cyc is expressed a bit later than ndr1/sqt, and its expression largely
depends on ndr1/sqt function (Dougan et al. 2003 ). The maintenance of ndr1/sqt
7 Establishment of the Vertebrate Germ Layers