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Fig. 3. (A-P) Phenotypic analysis of embryos stained with phalloidin-rhodamine
after injection of CsDelta1, CsDelta2 and CsNotch dsRNA.
(A-J) Confocal micrographs of flat preparations of prosomal segments of
180-hour-old embryos; anterior is towards the top. (K-P) Transverse sections
through the fourth prosomal segments; medial is towards the left. Prosomal
regions of embryos injected with GFP dsRNA as a control (A,F,K), with
CsDelta1 dsRNA (B,G,L), CsDelta2 dsRNA (C,H,M) and
CsNotch dsRNA (D-P), respectively. (F-J) Higher magnifications of the
fourth prosomal hemisegments. (A,F) After injection of GFP dsRNA, the
ventral neuroectoderm shows the normal number of invagination sites (about 30
per hemisegment; dots of high phalloidin-rhodamine staining, arrows). (B,G)
After injection of CsDelta1 dsRNA, the number of invagination sites
is reduced in individual segments (arrows). (C,H) A more severe reduction of
invagination sites can be detected after injection of CsDelta2 dsRNA;
invagination sites are absent in the whole ventral neuroectoderm. (D,I,) After
injection of CsNotch dsRNA, dots of high phalloidin-rhodamine
staining can be detected in the positions that correspond to invagination
sites in control injected embryos (arrows), although they are much smaller.
(E,J) In a more severely affected embryo there is only diffuse
phalloidin-rhodamine staining visible in positions that correspond to
invagination sites in control injected embryos (arrow). (K) Confocal
micrograph of a transverse optical section through an invagination site
(arrowheads) of an embryo injected with GFP dsRNA. The cell processes
of the basally enlarged cells extend to the apical surface. (L) Transverse
optical section through the fourth prosomal hemisegment of an embryo injected
with CsDelta1 dsRNA, showing that in a region where invagination
sites are missing the neuroectoermal cells form a bulge (arrowhead). (M)
Transverse section through the fourth prosomal hemisegment of an embryo
injected with CsDelta2 dsRNA. Two bulges of neuroectermal cells are
visible (arrowheads). (O,P) The transverse sections through the fourth
prosomal hemisegments of embryos injected with CsNotch dsRNA reveal
that, although dots of higher phalloidin-rhodamine staining are visible on the
apical surface (arrowheads), there are no bottle-like cells visible underneath
these dots. The presence of several cell layers suggests that cells that
normally invaginate occupy space in the apical layer so that newly formed
cells were pushed basally. l1, l2, walking legs 1 to 2 (corresponding
to prosomal segments 3 and 4). Scale bars: in A, 150 µm for A-E; in F, 50
µm for F-J; in K, 20 µm for K-P.
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