252
JEAN BRÄCHET
3. Effects of Inhibitors of DNA Synthesis on Development
Studies on metabolic inhibitors of DNA synthesis (folic acid analogs,
fluoro-derivatives
of pyrimidines, etc.) all suggest that there must be a
change in the biochemical mechanism of DNA synthesis at the end of
cleavage. For example, Grant (1960) studied the effects on morphogenesis of a folic acid analog, amethopterin: he observed a block at the
late blastula stage, which can be partially lifted by the addition of
thymidine. He suggested that a change occurs in the metabolic pathways
for nucleic acid synthesis at gastrulation. As found, in a more direct way,
by Bieliavsky and Tencer (1960), the ribonucleotide pool would be
utilized preferentially for DNA synthesis during cleavage, and for RNA
synthesis during gastrulation and neurulation.
The findings of Tencer (1961b) who studied the effects of 5-fluorodeoxyuridine
on amphibian embryos are equally interesting. Like
amethopterin, this analog blocks cleavage at the young blastula stage,
the addition of thymidine allowing development to proceed until the
late blastula stage. Autoradiographic experiments showed that the
analog inhibits the incorporation of uridine into DNA, but not that of
thymidine into DNA (we shall soon find a similar situation in the case of
lethal hybrids). It has been suggested by Tencer (1961b) that the
availability of thymidylic acid might become a limiting factor in DNA
synthesis at the blastula stage and that an inhibition of thymidylate
kinase could stop development at this stage. Biochemical experiments
are of course required before this interesting hypothesis can be accepted.
It should be added, however, that fluoro derivatives of uracil, other than
fluoro-deoxyuridine,
i.e., fluoro-uracil and fluoro-uridine, also block
development of amphibian eggs at the blastula stage, without presumably inhibiting thymidylate kinase in a direct way (Bieliavsky and
Tencer, unpublished). All the fluoro derivatives tested inhibit protein
synthesis in dissociated morula cells, suggesting an effect on RNA as
well as on DNA. In the case of fluoro-uracil, it was demonstrated by
Bieliavsky and Tencer that this analog is, in fact, incorporated into
RNA only : addition of thymidine has no effect on the incorporation of
fluoro-uracil
into RNA, but it diminishes to a certain extent the
inhibition of DNA synthesis and cell division induced by this abnormal
pyrimidine. It appears that, at the end of cleavage, synthesis of new
enzymes required for DNA production (possibly thymidylate kinase) is
absolutely necessary for further development, and that RNA is involved
in this synthesis. Nothing more can be said in this regard until the key
enzymes for DNA synthesis have been studied by chemical means in
developing amphibian eggs.
The existence of two different metabolic pathways for DNA synthesis
in amphibian eggs can also be demonstrated when eggs are treated with
JEAN BRÄCHET
3. Effects of Inhibitors of DNA Synthesis on Development
Studies on metabolic inhibitors of DNA synthesis (folic acid analogs,
fluoro-derivatives
of pyrimidines, etc.) all suggest that there must be a
change in the biochemical mechanism of DNA synthesis at the end of
cleavage. For example, Grant (1960) studied the effects on morphogenesis of a folic acid analog, amethopterin: he observed a block at the
late blastula stage, which can be partially lifted by the addition of
thymidine. He suggested that a change occurs in the metabolic pathways
for nucleic acid synthesis at gastrulation. As found, in a more direct way,
by Bieliavsky and Tencer (1960), the ribonucleotide pool would be
utilized preferentially for DNA synthesis during cleavage, and for RNA
synthesis during gastrulation and neurulation.
The findings of Tencer (1961b) who studied the effects of 5-fluorodeoxyuridine
on amphibian embryos are equally interesting. Like
amethopterin, this analog blocks cleavage at the young blastula stage,
the addition of thymidine allowing development to proceed until the
late blastula stage. Autoradiographic experiments showed that the
analog inhibits the incorporation of uridine into DNA, but not that of
thymidine into DNA (we shall soon find a similar situation in the case of
lethal hybrids). It has been suggested by Tencer (1961b) that the
availability of thymidylic acid might become a limiting factor in DNA
synthesis at the blastula stage and that an inhibition of thymidylate
kinase could stop development at this stage. Biochemical experiments
are of course required before this interesting hypothesis can be accepted.
It should be added, however, that fluoro derivatives of uracil, other than
fluoro-deoxyuridine,
i.e., fluoro-uracil and fluoro-uridine, also block
development of amphibian eggs at the blastula stage, without presumably inhibiting thymidylate kinase in a direct way (Bieliavsky and
Tencer, unpublished). All the fluoro derivatives tested inhibit protein
synthesis in dissociated morula cells, suggesting an effect on RNA as
well as on DNA. In the case of fluoro-uracil, it was demonstrated by
Bieliavsky and Tencer that this analog is, in fact, incorporated into
RNA only : addition of thymidine has no effect on the incorporation of
fluoro-uracil
into RNA, but it diminishes to a certain extent the
inhibition of DNA synthesis and cell division induced by this abnormal
pyrimidine. It appears that, at the end of cleavage, synthesis of new
enzymes required for DNA production (possibly thymidylate kinase) is
absolutely necessary for further development, and that RNA is involved
in this synthesis. Nothing more can be said in this regard until the key
enzymes for DNA synthesis have been studied by chemical means in
developing amphibian eggs.
The existence of two different metabolic pathways for DNA synthesis
in amphibian eggs can also be demonstrated when eggs are treated with