274
JOHN RUNNSTRÖM
but the nuclear divisions went on. Runnström and Kriszat (unpublished
data, 1964) varied the experiment in the following way. Unfertilized
eggs of Psammechinus
miliaris were pretreated in the manner described,
but after 14 min pretreatment the eggs were inseminated, and 1 min
later the trypsin was removed by thorough washing. Under these conditions the cortical changes were complete, the fertilization membrane
was well elevated, and the segmentation was even more regular than in
the untreated control eggs. This experiment points again to the importance of proteolytic enzymes in the primary steps of the fertilization
process.
There may be an additive effect of the trypsin present around the
egg and the proteolytic enzyme activated in the egg upon fertilization.
In this way the gelated state may be overcome. This is analogous to a
slower process that can be demonstrated in unfertilized eggs, treated,
for example, with 1 X 10
- 4
% trypsin for 15 min. A gelation of these eggs
occurs. If, however, the treatment is prolonged to 23-30 min or the concentration increased to 2.5 X 10"
4
%, the gelation is overcome. The experiment reported above shows that a rapid shift may occur from a
gelated state to one that is more sol-like, as was earlier inferred from the
behavior of ribonuclease-treated eggs (Runnström, 1962). The implications of these results will be further examined in Section IV,C.
Olsson (1960) showed that 5 min after insemination there is a 10%
decrease of the ribonucleic acid obtained by phenolic extraction from the
egg homogenates as compared with unfertilized eggs. A corresponding
increase in acid-soluble nucleotides was demonstrated. These results
indicate the activation of a ribonuclease that may release proteins, particularly enzymes attached to ribosomes or membranes present in the
cortical layer (see Hagström and Runnström, 1962; Runnström, 1962).
An enzyme of particular interest in the fertilization process is adenosinetriphosphatase (ATPase), the presence of which in egg homogenates
of Paracentrotus
was indicated by Runnström (1933), who also showed
that the egg is rich in acid-soluble phosphate; 25% of this proved to be
easily hydrolyzable (see örström and Lindberg, 1940). According to
T. Hultin (1957) the main content of this fraction is adenosine triphosphate (ATP).
Connors and Scheer (1947) studied the ATPase activity in homogenates of unfertilized and fertilized eggs of Strongylocentrotus
purpuratus; this activity was definitely higher in the latter. Mullins (1949)
reported also an ATPase activity that was higher in brei of fertilized
than of unfertilized Arbacia and Paracentrotus
eggs. The main activity
was found to be in the residue after treatment with ether and water and
a subsequent extraction with KCl. In addition, these "residual granules"
JOHN RUNNSTRÖM
but the nuclear divisions went on. Runnström and Kriszat (unpublished
data, 1964) varied the experiment in the following way. Unfertilized
eggs of Psammechinus
miliaris were pretreated in the manner described,
but after 14 min pretreatment the eggs were inseminated, and 1 min
later the trypsin was removed by thorough washing. Under these conditions the cortical changes were complete, the fertilization membrane
was well elevated, and the segmentation was even more regular than in
the untreated control eggs. This experiment points again to the importance of proteolytic enzymes in the primary steps of the fertilization
process.
There may be an additive effect of the trypsin present around the
egg and the proteolytic enzyme activated in the egg upon fertilization.
In this way the gelated state may be overcome. This is analogous to a
slower process that can be demonstrated in unfertilized eggs, treated,
for example, with 1 X 10
- 4
% trypsin for 15 min. A gelation of these eggs
occurs. If, however, the treatment is prolonged to 23-30 min or the concentration increased to 2.5 X 10"
4
%, the gelation is overcome. The experiment reported above shows that a rapid shift may occur from a
gelated state to one that is more sol-like, as was earlier inferred from the
behavior of ribonuclease-treated eggs (Runnström, 1962). The implications of these results will be further examined in Section IV,C.
Olsson (1960) showed that 5 min after insemination there is a 10%
decrease of the ribonucleic acid obtained by phenolic extraction from the
egg homogenates as compared with unfertilized eggs. A corresponding
increase in acid-soluble nucleotides was demonstrated. These results
indicate the activation of a ribonuclease that may release proteins, particularly enzymes attached to ribosomes or membranes present in the
cortical layer (see Hagström and Runnström, 1962; Runnström, 1962).
An enzyme of particular interest in the fertilization process is adenosinetriphosphatase (ATPase), the presence of which in egg homogenates
of Paracentrotus
was indicated by Runnström (1933), who also showed
that the egg is rich in acid-soluble phosphate; 25% of this proved to be
easily hydrolyzable (see örström and Lindberg, 1940). According to
T. Hultin (1957) the main content of this fraction is adenosine triphosphate (ATP).
Connors and Scheer (1947) studied the ATPase activity in homogenates of unfertilized and fertilized eggs of Strongylocentrotus
purpuratus; this activity was definitely higher in the latter. Mullins (1949)
reported also an ATPase activity that was higher in brei of fertilized
than of unfertilized Arbacia and Paracentrotus
eggs. The main activity
was found to be in the residue after treatment with ether and water and
a subsequent extraction with KCl. In addition, these "residual granules"