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Fig. 1. Smed-βcatenin1 silencing induces a gradual
anteriorization of regenerating planarians. (A-E) Stereomicroscope
views of regenerating trunk pieces showing the different phenotypes.
(F-W,A'-H') Several markers were used at
different regeneration stages to characterize the phenotype. Analysis of
opsin (F-I), cintillo (J-M) and H.10.2f (N-Q)
revealed the differentiation of ectopic anterior structures in the posterior
blastema, which subsequently expanded throughout the periphery of the
planarian body. Anti-arrestin immunostaining (R,S) showed that, in radial-like
hypercephalized planarians, all photoreceptors were connected through ectopic
visual axons. Synapsin immunostaining (T,U) and in situ hybridization for
GluR (V,W) and H.10.2f (P,Q) demonstrated that the cephalic
ganglia (cg) and ventral nerve cords (vnc) are transformed into a thick ring
from which cephalic branches ectopically differentiate all around the body of
radial-like hypercephalized planarians. (A'-H') Defects in the
pattering of gut and secretory cells were visualized with anti-P-Tyr (green)
and anti-arrestin (pink pseudocolour) antibodies. A control planarian viewed
at the level of the gut has one anterior and two posterior gut branches (gb;
B',F'), along with more dorsal secretory cells (sc) in the
pre-pharyngeal but not the post-pharyngeal region (D',H'). The two
posterior gut branches converge into one in Smed-bcatenin1-silenced
planarians (E'), and secretory cells ectopically differentiate in the
post-pharyngeal region (G'). (I') The dose dependence of
the phenotype and a quantification of the different degrees of anteriorization
(1, 2 or 3d x 3inj/d indicates that planarians were injected three times
during 1, 2 or 3 consecutive days, respectively). Anterior is shown to the
left in those cases where an anteroposterior axis is present.
(R-U,A'-H') Confocal z-projections. Yellow asterisk
indicates the pharynx. Yellow/black arrows and red arrows indicate the
differentiation of anterior structures in normal and ectopic positions,
respectively. d, days after wounding; a, anterior; p, posterior. Scale bar in
A: 270 µm in A-C,E,F,N; 180 µm in D; 250 µm in H,L,P,R,T,W; 395 µm
in J; 437 µm in K; 300 µm in all other images.
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