Supplemental Figure 1
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Fig. S1.
Immunohistochemistry for H+-V-ATPase subunits in early Xenopus embryos. (A) Cartoon showing the arrangement of subunits in the H+-V-ATPase
(after Nishi 2002). (B) Subunit A
in an AV section (parallel to the animal-vegetal axis) of an unfertilized egg;
subunits are found localized in the vegetal cortex. (C) A just-fertilized egg stained with anti-subunit F. A
small area of staining can be seen next to the brown curve of pigment granules
marking the path of sperm entry. (D)
In post-rotation zygotes, staining for subunit A has moved up into the vegetal
cytoplasm (AV section). (E-H)
Two-celled embryos stained for subunits F and A. E illustrates a commonly seen
pattern: subunit F staining appears to form fingers that reach up from the
vegetal to the animal cytoplasm. In flat sections (perpendicular to the AV
axis), staining is usually asymmetric and heavy near the cell membrane: (F)
subunit F; (G) subunit A; (H) subunit A revealed with a different primary
antibody. (I,J) Four-cell embryos, flat sections: (I) subunit A; (J)
oriented section, subunit B. Staining is on the right hand side. (K) Flat section, oriented eight-cell embryo stained for
subunit B. Staining is on the right hand side. (L) AV section through gastrula-stage embryo stained for
subunit A. By this stage, staining is mostly uniformly distributed with respect
to the LR axis. (M) Embryo treated
with cytochalasin B to disrupt actin filaments, fixed when controls reached the
two cell stage (no cleavage furrows form in actin-disrupted embryos). Staining
for subunit B shows that the normal vegetal-plus-fingers localization of H+-V-ATPase
subunits (compare to E) has become entirely animal. (N) Embryo treated with latrunculin to disrupt actin
filaments, imaged when controls reached the four-cell stage. Subunit A staining
reveals that localization has been completely disrupted (compare with I or J).
Although the patterns presented in this figure match those found with the
anti-subunit A (Fig. 3), the antibodies used for this figure, although highly
specific in mammals, did not, in our hands, work for westerns.
