Inter-and intraspecific differences in fatty acid composition of freshwater crustacean zooplankton

1. The composition of fatty acids (FA) from C14 to C18 was measured for edible seston and for individual Daphnia galeata , Diaphanosoma brachyurum and Eodiaptomus japonicus from Lake Biwa using a pyrolysis–gas chromatography (Py–GC) method. Based on the relative abundance of the FA, inter-and intraspecific differences in composition were examined.
2. Among the FA, C16 : 0 was the most abundant, both in the three zooplankton species and in edible seston smaller than 20 μm, suggesting that the composition of the zooplankton was roughly reflected by their diet. However, the abundance of C18 : 1 relative to C16 : 1 and C18 : 0 was much higher in each zooplankton species than in the diet.
3. Comparison of the relative abundance of FA among the three zooplankton species revealed that intraspecific differences in FA composition are greater than interspecific differences. These results indicate that variability in FA composition is not necessarily species-specific, and can be obscured by variation in composition between individuals.     

CARBOHYDRATE METABOLISM IN THE EGGS OF THE SILKWORM, BOMBYX MORI. I. ABSENCE OF PHOSPHOFRUCTOKINASE IN THE DIAPAUSING EGG

In the diapausing eggs of the silkworm, Bombyx mori , glycogen is rapidly converted to sorbitol and glycerol, and this conversion is reversed at termination of the diapause (C hino , 1958). To elucidate the pathway leading to this polyol formation and its regulatory mechanisms, enzymes concerning carbohydrate metabolism were surveyed in diapausing as well as in developing eggs of the silkworm.
Most of the enzyme activities concerning citric acid cycle are low at the beginning of the embryogenesis and during diapause, but increase at the later stages of the development. Making an exception, reduction rate of malate and fumarate was rather high from the onset of the embryonic development. Several glycolytic enzymes were also studied. Most remarkable fact is that phosphofructokinase activity could not be demonstrated in the diapausing and also in the early stages of the developing eggs. Other enzymes, viz. α-glycerophosphate dehydrogenase, aldolase, glyceraldehyde-3-phosphate dehydrogenase were detected from the beginning of the embryogenesis.
Absence of phosphofructokinase, together with the high activity in glucose-6-phosphate dehydrogenase, suggests that predominant pathway in carbohydrate metabolism in the early stages of embryogenesis and in the diapause period is by way of pentose phosphate pathway. This supposition is confirmed by the experiments using labeled glucose. Incorporation of the label into glycerol of the diapausing eggs was three to four fold when G-6-¹⁴C was injected into pupae as compared with the case of G-1-¹⁴C injection. The above experiments provide evidence supporting the theory that glycogen is converted into sorbitol and glycerol mostly by way of the pentose phosphate pathway in the diapausing eggs.     

Troponin in Cultured Chicken Breast Muscle Cells

The onset of troponin accumulation and the localization of troponin in cultured chick embryo skeletal muscle cells were studied by means of indrect immunofluorescent microscopy. At 31 hr after plating, troponin components were detected in 54–62% of total mononucleated myogenic cells and in all myotubes as longitudial fibrous structures. ³H-thymidine incorporation stduy coupled with the immunofluorescent microscopy showed that mononucleated myogenic cells at the mitotic stage did not contain troponin. As myotube maturation proceeds, the troponin-containing fibers were organized into cross-striated structures. At the myotube stage, muscle cells were labeled with ³⁵S-methionine and proteins synthesized were analyzed by two-dimensional gel electrophoresis. It was found that myotubes in culture synthesized both fast and slow types of troponin-I and-C. Our results suggest that fast and slow types of troponin components are synthesized in cultured skeletal muscle cells before or at the early phase of myofibrillogenesis.     

Direct Evidence for the Presence of Germ Cell Determinant in Vegetal Pole Cytoplasm of Xenopus laevis and in a Subcellular Fraction of It

To test for the presence of germ cell determinant in Xenopus embryos, vegetal pole cytoplasm containing the "germ plasm", or a subcellular fraction of it, was microinjected into single somatic blastomeres isolated from 32-cell embryos. Injected or non-injected (control) blastomeres were cultured in ³H-thymidine until normal control embryos reached the neurula stage. The labeled explants were then implanted into unlabeled host neurulae, which were allowed to develop to the tadpole stage. Labeled PGCs of explant origin in the genital ridges of the experimental tadpoles were examined by autoradiography.
Isolated blastomeres were injected with vegetal pole cytoplasm of 32-cell embryos or with a 20,000 g pellet made from vegetal pole cytoplasm of 2-cell embryos. Labeled PGCs were found in 7.6% and 2.3% of the experimental tadpoles, respectively. No labeled PGCs were found in the control tadpoles, except for one tadpole in the first experiment. These results strongly suggest that the vegetal pole cytoplasm and its subcellular fractions act as germ cell determinant.     

The Wave Pattern of Free Calcium Release Upon Fertilization in Medaka and Sand Dollar Eggs

A transient rise in the concentration of Ca²⁺ in the cortex upon fertilization was demonstrated in medaka eggs injected with aequorin. Detection of the aequorin luminescence with an ultra-high sensitivity photonic microscope system revealed a wave of increased Ca²⁺ concentration starting at the site of sperm entry (animal pole) and being propagated along the cortex of the egg toward the antipode. The wave traversed the entire egg surface within 2–3 min. The peak value of the aequorin luminescence, and therefore the peak value of the Ca²⁺ transient, was generally higher at the site of sperm entry than in other regions. The peak values of the luminescence (and therefore of the Ca²⁺ concentration in the cortex) remained fairly constant during propagation of the wave. Microinjection of Ca²⁺ into the cortex also induced a Ca²⁺ wave. When the egg was stimulated by microinjection of Ca²⁺ at the equatorial region, the Ca²⁺ wave was propagated at a fairly constant speed over the egg surface, except at the region near the vegetal pole where the wave was retarded. Simultaneous recording of the Ca²⁺ wave and the wave of cortical change (breakdown of cortical alveoli) in eggs during fertilization revealed that the Ca²⁺ wave preceded the wave of cortical change.
A Ca²⁺ wave was also demonstrated in sand dollar eggs, although due to their smaller size the phenomenon was not as clear as in medaka eggs.     

Implications of Differential Chemotaxis and Cohesiveness for Cell Sorting in the Development of Dictyostelium discoideum

Cell sorting behavior was observed during the development of Dictyostelium discoideum Ax-2, between cells grown with [G(+) cells] and without [G(−) cells] glucose. Development of the G(−) cells was about 2-3 hr faster, as reflected by differences in chemotactic sensitivity of the cells and cell cohesiveness. Among various mixing combinations of G(−) and G(+) cells, the most clear sorting occured when vegetative G(+) cells were mixed with G(−) cells which had been allowed to develop for 3 hr, the G(−) cells being located in the anterior prestalk region of a migrating slug. In contrast, vegetative G(−) cells moved to the posterior prespore region of a slug when mixed with G(+) cells which had developed for 6 hr. These findings indicate a close relationship of the cellular developmental stage to the sorting behavior. Possible implications of the differential chemotactic ability and cohesiveness for the sorting mechanism are disscussed.     

SPLITTING OF {varepsilon}-CAPROLACTAM AND OTHER LACTAMS BY BACTERIA

-Caprolactam-utilizing bacteria split -caprolactam, -valerolactamand -butyrolactam, and produce the -amino acids correspondingto them. This activity is lost when cells are grown on 6-amino-caproicacid or ammonium adipate, and reappears when cells are incubatedwith either -caprolactam or -valerolactam as the sole majororganic nutrient. Chloramphenicol inhibits this adaptation.The enzyme splitting those lactams is one and the same. It maybe called "lactam-splitting enzyme". But attempts to demonstratethe enzymic activity in a cell-free system has not yet beensuccessful. (Received September 9, 1965; )     

BACTERIAL BREAKDOWN OF {varepsilon}-CAPROLACTAM AND ITS CYCLIC OLIGOMERS

Eleven different types of bacteria were isolated which werecapable of growing on -caprolactam, the monomeric material fornylon 6 polyamide, as the sole source of both carbon and nitrogen. The optimal concentration of -caprolactam for the bacterialgrowth was about 0.6% in a synthetic liquid medium enrichedwith a small amount of yeast extract. The bacterial strains grew also on -butyrolactam, -valerolactamand the -amino acids corresponding to these lactams and -caprolactam.Ammonium adipate was a good substrate for the growth of allthe strains. One strain of Corynebacterium aurantiacum was found to be capableof utilizing cyclic and linear oligomers of 6-aminocaproic acidwith an exception of cyclic dimer. The strains of corynebacteria required vitamin B₁ for growth. Metabolism of -caprolactam and related compounds is discussedbriefly. (Received September 9, 1965; )     

Micromere Differentiation in the Sea Urchin Embryo: Two-Dimensional Gel Electrophoretic Analysis of Newly Synthesized Proteins

A method for large-scale culture of isolated blastomeres of sea urchin embryos in spinner flasks was developed. Micromeres and meso-, macromeres isolated from sea urchin embryos at the 16-cell stage were cultured by this method and the patterns of protein synthesis by their descendants were examined by two-dimensional gel electrophoresis of [³⁵S] methionine-labeled proteins. Six distinct proteins with molecular weights of 140–kDa, 105–kDa, 43–kDa, 32–kDa, and 28–kDa (two components) were specifically synthesized by differentiating micromeres. Quantitative analysis of the two-dimensional gel patterns demonstrated that all these proteins, except the 32–kDa protein, appeared at the time of ingression of primary mesenchyme cells (PMC's) in vivo , several hours earlier than the onset of spicule formation. The synthesis of 32–kDa protein was paralleled to active spicule formation and the uptake of Ca²⁺. Cell-free translation products directed by poly (A)⁺ RNAs isolated from descendant cells of micromeres and meso-, macromeres were compared by two-dimensional gel electrophoresis. Several spots specific to the micromere lineage were detected. However, none of them comigrated with the proteins synthesized specifically by the cultured micromeres. The results suggest that the expression of these proteins specific to differentiating micromeres may involve post-translational modification.