31P-NMR analysis of sea urchin sperm activation: Reversible formation of high energy phosphate compounds by changes in intracellular pH

³¹P-NMR has been used to estimate the internal pH (pH_i) of sperm from the sea urchin Strongylocentrotus purpuratus. The values for pH_i obtained from the chemical shift of inorganic phosphate agree well with those obtained from amine accumulation. At low pH_i, when sperm are quiescent (immotile and non-respiring), they accumulate phosphocreatine (PCr), but have a low level of inorganic phosphate (P_i). Conversely, when the pH_i is elevated, sperm respiration and motility are activated, PCr is decreased and P_i is increased. This change is reversible upon decrease of the pH_i, whereupon respiration and motility are arrested, P_i disappears and PCr increases. We conclude that the overall balance of energy metabolism, and thus the phosphate potential, of sea urchin sperm are under the control of the pH_i.     

Genetic nature of one of the traits of malignant cell transformation in vitro

The genetic events controlling the ability of transformed cells to grow in a medium with a low serum content (ser+) were studied. A hypodiploid clone of Chinese hamster cells with normal serum requirements (49a5ser-) was used as starting material. The results of the fluctuation tests have shown that serum-independence is a random spontaneous event. Its rate of occurrence is 1-2 . 10(-5). The concomitant study of a gene mutation (resistance to 6-mercaptopurine) revealed similar characteristics with respect to the distribution of the number of mutants in replicate cultures and the mutation rate. N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and the oncogenic SV40 virus significantly increased the frequency of ser+ colonies. In the majority of clones isolated in a medium with 1% serum (11 spontaneous and 7 induced by MNNG), the ser+ character proved to be stable after different periods of cultivation without selective pressure. The degree of serum-independence varied in different clones. The results suggest that the ability to grow in a medium with a low serum content originates, in most cases, from a mutation event.     

Distinguishing behavioral interactions from visual cues as causes of adult sex change in a coral reef fish

Two experiments tested the hypothesis that adult female-to-male sex reversal in protogynous fish is induced by the loss of close-contact behavioral interactions between males and females, and not by the loss of simple visual cues from the male. Twenty-six laboratory groups of Anthias squamipinnis were manipulated so that females within each group (1) retained chemical and acoustic access to a male, (2) were denied behavioral access to a male, and (3) were either allowed or denied visual access to a male alone or to a male interacting with another female. At least one female subsequently changed sex in each of 22 groups. While acoustic and chemical cues were not completely eliminated as possible causes, sex change is apparently induced by loss of male-female behavioral interaction in combination with continued interaction between females.     

The effects of bisulfite on growth and macromolecular synthesis in Escherichia coli

Bisulfite reversibly inhibits the growth of a variety of microorganisms and has been used as a preservative in foods and beverages for that reason. We have now measured macromolecule synthesis in Escherichia coli K12 after bisulfite treatment. RNA synthesis, the synthesis of total protein, and of an inducible enzyme, beta-galactosidase, stopped almost immediately upon addition of 2 mM (or higher concentrations) of bisulfite. These functions resumed after a lag whose duration depended on the concentration of bisulfite added. The synthesis of DNA was slowed upon bisulfite addition, but did not stop entirely. The inhibition of RNA synthesis by bisulfite took place in both stringent and relaxed strains of E. coli and was not relieved upon addition of chloramphenicol. Stringent control was therefore not involved in this effect. No effect on protein synthesis was observed in the cell-free system of E. coli (using poly(U) or MS2 RNA as messenger) at bisulfite concentrations up to 10 mM. Protein synthesis inhibition in vivo was apparently not due to a reaction of bisulfite with a component of this system. In additional experiments, RNA polymerase was not impaired by bisulfite, and the growth inhibition effect was shown to proceed in the presence of inhibitors of free radical chain reactions.     

Immunological and Biophysical Separation of Dengue-2 Antigens

Antigenic compositions of slowly sedimenting dengue-2 hemagglutinin (SHA) and soluble complement-fixing antigen (SCF) were compared with the virion (rapidly sedimenting hemagglutinin, RHA) by radioimmune precipitation (RIP), RIP inhibition, kinetic neutralization, and neutralization blocking tests with the use of hyperimmune mouse ascitic fluids. RHA and SHA were unable to inhibit completely the RIP of each other by anti-RHA, and neutralization by anti-RHA was not blocked by SHA. This indicated that SHA is serologically related, but not identical, to RHA. SHA differed from RHA in that SHA lacked the “core” polypeptide but contained the two envelope polypeptides. In addition, SHA contained a polypeptide with a molecular weight of 16,500 daltons and a suggestion of several other proteins. These data, when considered with other evidence, suggest that SHA is a special form of “incomplete virus.” SCF was unable to inhibit the RIP of SHA or RHA or to block neutralizing antibodies. Further, anti-SCF did not neutralize RHA or precipitate significant levels of SHA or RHA. Polyacrylamide gel electrophoresis separated SCF from structural polypeptides by molecular size. This evidence suggests that SCF is a nonstructural antigen.     

Chronology of chromosome reduplication in Chinese hamster cells incubated at low temperature

Experiments have been carried out on Chinese hamster fibroblasts. Cultures at the log-stage of growth were incubated at 15 or 25° C for 24 hrs. In two groups of experiments the cells were labeled with H³-TdR for 6 hrs at the respective temperature, washed and further incubated at 37° C. In each group of experiments cultures labeled with H³-TdR at 37° C for 20 min were used as a control. It was found: 1. the delay in the onset of cell passage through the mitotic cycle at 37° in cultures exposed to 15 or 25° C was about equal to 1,5-1 hr resp. and cells proceeded through the life cycle without blockages at any phase of the cycle; 2. the patterns of chromosome reproduction during the second half of the S-phase were the same after labeling at 15, 25 and 37° C. — In the third group the cells were labelled with HP-TdR for 10–60 min and 6 hrs resp. at 25° C. The patterns of reproduction of chromosome pairs 1–4 and small metacentrics were found to be the same in cells labeled briefly and those labeled for 6 hrs. After brief labeling asynchronous reduplication of different segments in many chromosomes became evident. It was masked because of heavy labeling after 6 hrs treatment.