|
|
|
Fig. 4. RT-PCR analysis of the expression of the Xa,tr mRNA in different
embryonic stages. With the gene specific primers indicated in
Fig. 1, cDNA fragments corresponding either to Xa, (A, u) or Xczstr
(A, tr) were amplified from reverse transcribed mRNA ofthe indicated
stages. For the amplification of the Xcu, cDNA fragment (position:
23&:3183 BP; length: 837), the PCR was performed with 30 cycles,
whereas for the Xastr cDNA fragment (position: 1081-1985; length:
904), 45 cycles were required. PCR products isolated from stage I and
23 were digested with the restriciton enzyme HaeIll and yielded the
predicted pattern (B, XolS-HaeIII fragments: 52, 141, 167, 180 and 297
bp; X+r-Ha&I fragments: 141, 280, 234 and 349 bp).
|
|
Fig. 5. Localization of integrin aS mRNA during embryogenesis. Whole mount in situ hybridization with digoxygenin-rUTP-labeled antisense
riboprobes was used to determine the localization of integrin q mRNAs throughout development. In each panel, arrows indicate expression in
neural crest cells and arrowheads point to staining within the somitic mesoderm. (A) Dorsal view, stage 1 I .5-12 gastrula, oriented with anterior
end at the top of the panel. (B) Optical section of (A). The approximate plane of section is indicated by the line in (A). (C) Frontal view, stage
15 neurula, dorsal to the top. Integrin q transcripts are detected in the neural crest (arrow) and paraxial mesoderm (arrowhead). (D) Side view,
stage 14 neuruia, anterior to the right. (E) Optical section, stage 17 neurula, dorsal is to the top. Arrow indicates trunk neural crest cells, arrowhead
marks the somitic mesoderm. (F) Dorsal view, stage 19 neurula, anterior to the left. Cranial neural crest cells (black arrow) are intensely stained
and located lateral to the unstained neural tube. Trunk neural crest forms a single line of cells along the dorsal midline of the embryo (white arrow).
(G) Side view, stage 21 neurula, anterior to the right and dorsal at the top. The somatic staining of the integrin os is concentrated around the nuclei
of the post-rotation somites (arrowhead). Intensely stained cranial neural crest cells are visible in the anterior part of the embryo (arrow). (H) Optical
section, stage 22 neurula, dorsal to the top. Trunk neural crest a single line of cells dorsal to the neural tube (arrow). Staining within the somites
is also visible (arrowhead). (I) Dorsal view, stage 23 neurula, anterior to the top. Staining is localized in discrete foci in the post-rotation somitic
mesoderm (black arrowhead) and is diffuse in the pre-rotation mesoderm (white arrow). (J) Side view, stage 32 tailbud embryo, anterior to the
right. Note weaker staining in anterior somites. (K) High magnification view of the tailbud of the embryo in (J). Integrin (r5 expression in the tail
remains restricted to the neural crest (arrow) and somites (arrowhead).
|
|
Fig. 6. Characterization of the q antiserum. Inununoprecipitations
were performed, either with the anti q antiserum (lanes B and C) or
with a 8, specific mAB 8C8 (lanes A, D and E). The precipitates were
analyzed by immunoblotting with a polyclonal antiserum directed
against the 8, subunit (lanes A and B), or with the anti q antiserum
(lanes C-E). Electrophoresis was carried out under reducing (lanes
A-D) or under non-reducing conditions (lane E). The signal in the
lower part of (B) and (C) indicate IgGs which leaked from the
precipitation beads. The positions (bars) and size (M, x 10m3) of relative
molecular weight markers are indicated.
|
|
Fig. 7. Temporal expression of the Xenopus integrin a5 protein during
development. Extracts from embryos of different stages (eight embryos
per lane) were electrophoresed under non-reducing conditions
and immunoblotted with the antiserum anti-q (A) or with mAB 8C8
(B). Developmental stages and the positions (bars) and size
(Mr x 10-3) of relative molecular weight markers are indicated.
|
|
Fig. 8. Distribution of the 0~s integrin subunit in the embryo during cleavage, blastula and gastrula stages. Embryos were whole mount stained
with the anti os antiserum, embedded in glycolmethacrylate and sectioned. (A) Detail of a stage 6 embryo stained with the anti cq antiserum preabsorbed
with the peptide used for the immunization. The speckled signal is also seen in all following pictures and is regarded as background. (B)
Phase contrast image of the region of contact between two blastomeres of a stage 5 embryo, animal view. (C) Same embryo as in (B) immunostained.
(D) Transverse section of a stage 7 embryo. (E) Dorsal blastopore lip region of a stage 10.5 embryo, with apically constricted bottle cells. (F)
Blastopore region of a stage I3 embryo. ae, archenteron; an, animal pole region; bl, blastocoel; dbl, dorsal blastopore lip region; me, mesoderm;
vg, vegatal pole region; yp, yolk plug. Bars: 50 um.
|
|
Fig. 9. Distribution of (Ye integrin in the embryo during neurula and tadpole stages. (A) Transverse section of a stage 20 embryo. (B) Transverse
section through a stage 30 embryo. Arrows indicate the ectoderm/mesoderm and mesodermlendoderm boundaries. ae, archenteron; en, endoderm;
no, notochord; nt, neural tube; pm, paraxial mesoderm; p, pronephros. Histological procedure as in Fig. 8. Bars: 50 um.
|
|
Itga5 Ab1 staining of a NF stage 30 Xenopus laevis embryo. Coronal section, Dorsal up.
|
|
Itga5 (integrin, alpha 5 (fibronectin receptor, alpha polypeptide)) gene expression in a Xenopus laevis embryo as assayed by in situ hybridization, NF stage 19, dorsal view, anterior left.
|
|
Itga5 (integrin, alpha 5 (fibronectin receptor, alpha polypeptide)) gene expression in a Xenopus laevis embryo as assayed by in situ hybridization, NF stage 32. Lateral view: Dorsal up, anterior left.
|
