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Dorsal-ventral axis formation in Xenopus laevis begins with a cytoplasmic rotation during the first cell cycle and culminates in a series of cell interactions and movements during gastrulation and neurulation that lead to the formation of dorsal-anterior structures. Evidence reported here indicates that mitochondria are differentially redistributed along the prospective dorsal-ventral axis as a consequence of the cortical-cytoplasmic rotation during the first cell cycle. This finding reinvigorates a possibility that has been considered for many years: asymmetries in cytoplasmic components and metabolic activities contribute to the development of morphological asymmetries.
FIG. 1. Mitochondrial RNA is more abundant in dorsal cells than in
ventral cells. RNAs were extracted from dorsal cells (lane D) and ven·
tral cells (lane V) of early gastrulae (stage 10), glyoxylated, and separated
on a 1% agarose gel for Northern blot analysis. Tracings from
densitometric analyses are presented on the right. (A) Northern blot
hybridized with radiolabeled probe for mt 1-rRNA. Band 1 is approximately
1.7 kb; the predicted size for mitochondrial 1-rRNA is 1631
bases. Band 2 is approximately 1.2 kb. (B) The Northern blot was
stripped and reprobed with 541- and 595-bp fragments from Hindiii
and Xbai double digests of pXlm-32 (Ramirez and Dawid, 1978), which
hybridized to mitochondrial mRNAs for cytochrome c oxidase subunit
1 (band 3, approximately 1.6 kb) and ATPase subunit 6 (band 4, approximately
700 bases). (C) As a loading control, the blot was repro bed
with the 3.4-kb EcoRI and BamHI fragment from pXlrl4 which hybridized
to cellular ribosomal RNA (Sollner-Webb and Reeder, 1979).
In this gel, cellular ribosomal RNA was slightly more abundant in the
ventral lane, indicating that the ventral lane was slightly overloaded
with respect to the dorsa] Jane. Densitometric comparison of mitochondrial
RNA signals (all bands were included in the analysis) with
loading controls indicated that approximately twice the amount of mitochondrial
RNA was present in dorsal cells than in ventral cells dissected
from blastula and gastrula stage embryos.
FIG. 2. Progressive redistribution of mitochondria during Xenopus early development. Paraffin-embedded seetions were hybridized with
sense (A) or antisense (B-F) mt l-rRNA dig-(U)-substituted probes, which were visualized via immunohistochemical means as described under
Materials and Methods. (A, B) Ovary from juvenile frog. (C) Stage V oocyte. (D) Fertilized egg fixed near the time of first cleavage (0.9 of the
cell cycle), sectioned in parallel to the prospective midsagittal plane, as defined by sperm entry point (SEP). The heavy accumulation of pigment
to the left of the animal pole denotes the region near the SEP. (E) Eight-cell embryo (stage 3), SEP on left. prospective dorsal on right. (F) Early
gastrula (stage 10). Dorsal lip of the blastopore is on the lower right marginal zone. Scale bars in A, B, 50 Jlm; C-F, 250 .urn. gv, germinal vesicle;
s. region of sperm entry point; be, blastocoel; bpi, blastopore lip.
FIG. 3. Mitochondrial distribution at the end of the first cell cycle.
Panels represent projections of nine successive confocal images taken
5 ~-Lm apart adjacent to the fracture plane. The animal pole is at the
top, and the prospective dorsal is to the right. (A) Control primary
antiserum. Light subcortical regions are due to autofluorescence resulting
from aldehyde-containing fixative. (B) BP128 antiserum. Note
small fluorescent patches throughout the cytoplasm, but particularly
in the region of the dorsal marginal zone. s, region of sperm entry
point.
FIG. 4. Effect of uv irradiation on the distribution of mitochondria
at the end of the first cell cycle. Eggs were fixed near the end of the
first cell cycle (0.9 NT) and sectioned parallel to the SEP meridian. In
situ hybridization with mt 1-rRNA was performed as in Fig. 2. (A)
Control. (B) uv-irradiated on vegetal surface to block cytoplasmic rotation
during the first cell cycle.
