Interactions between glucocorticoid receptor-bearing chromatin and antireceptor antibody preparations

Recent evidence indicates that the control of gene expression by steroid hormones is mediated by hormone-receptor complexes bound at specific chromosomal locations. The isolation of these in vivo sites of binding would be useful in an analysis of the mechanism of this control. We have therefore examined the interaction between two different antiglucocorticoid receptor antibody preparations and a defined chromatin fraction containing bound glucocorticoid receptors in order to test the feasibility of this approach. Both antibody preparations, when attached to sepharose, removed nucleosome-bound and nucleosome free receptor from solution, indicating that chromatin-bound receptor was exposed and available for reaction with antibody. The bulk of the chromatin, not containing receptor, was mainly unaffected during these reactions, showing that the antibodies exhibited significant specificity. To determine whether the nucleosome-bound receptor remained attached to the nucleosome during reaction with antibody, studies using soluble antibody were performed. One of the antibodies caused a shift in the sedimentation rate of the nucleosome-bound receptor from 11S to 11.5S, suggesting that an intact ternary complex of antibody-receptor-nucleosome had formed. The other antibody produced various-sized aggregates of the free and bound receptors. Surprisingly, we found that one of the antibodies reacted strongly with free and nucleosome-bound estrogen receptors as well as glucocorticoid receptors. These studies suggest that an antibody preparation with appropriate characteristics should permit isolation of chromosomal receptor binding sites.     

Propagation of cortical differences in tetrahymena

Progeny clones were derived from crosses arranged so that the number of ciliary meridians (corticotype) was unusually high in one partner, and normal in the other. An analysis of the propagation of corticotypes during maintenance of these clones for up to 1,000 fissions indicated that corticotypes above 21 undergo a rapid downward shift, while corticotypes in the range of 18 to 21 change slowly. Although these observations are consistent with Nanney's earlier deduction of a "stability center" at corticotype 19, there appears to be little if any difference in the stability of perpetuation of corticotypes 18, 19 and 20. Within this "stability range," the inertia of maintenance of pre-existing corticotypes is sufficiently strong that sister clones derived from an exconjugant pair can remain different for 1,000 fissions. These findings are consistent with observations made earlier, and those in the present study, indicating that cells in stock cultures express a substantial range of corticotypes even when maintained with frequent transfer. The results suggest that mechanisms of spatially ordered structural assembly within the cell can show sufficient fidelity to allow long-term vegetative perpetuation of phenotypic differences without artificial selection.     

Development of the ciliature of Tetrahymena thermophila: I. Temporal coordination with oral development

The number of basal bodies and cilia along pole-to-pole ciliary rows was enumerated in Tetrahymena thermophila cells sampled during the rapid-exponential phase of culture growth in three different media that supported generation times ranging from 2 to 4 hr. The time required for oral development was nearly constant in the three media, and thus most of the differences in generation time were accounted for by differences in the interval prior to the onset of oral development (stage 0), which ranged from 50% of the generation time in the “poorest” medium to 20% in the “richest.” There was very little increase in number of basal bodies and of cilia along ciliary rows during stage 0, irrespective of the duration of this stage. The bulk of the increase took place during oral development, following a time course suggestive of coordination wth oral development. The same temporal pattern of increase was found in several ciliary rows, although the proportion of basal bodies that were ciliated differed among rows. There is no simple relationship between the number of basal bodies along ciliary rows and cell length, surface area, or volume. However, a large and constant proportion of the total division-to-division cell growth took place during the interval prior to the onset of oral development, suggesting that an ensemble of developmental events, including oral development and an associated activation of the remainder of the cell surface, may be triggered by attainment of a threshold cell size.     

Hormone release during computer-monitored sexual behavior in mature and aged male rats

Sexual behavior and the increase in plasma hormone levels of LH, prolactin, and testosterone associated with sexual behavior were examined in three age groups of sexually naive male rats. The two younger groups (5- and 11-month-old) mated normally and their behavioral latencies decreased significantly following sexual experience. Both plasma testosterone and LH concentrations increased significantly following entrance of a receptive female into the mating arena. Plasma prolactin levels rose but not significantly. However, the 27-month-old rats neither mated nor showed an increase in the three plasma hormone concentrations during exposure to a receptive female. Only basal testosterone levels were significantly lower than those of the younger animals. Low testosterone levels possibly contributed to deficiencies in both behavior and its associated hormone release. The monitoring of sexual behavior was facilitated by a computer, programmed to record, store, and analyze behavioral events.     

STUDIES ON THE MAINTENANCE OF ORAL DEVELOPMENT IN TETRAHYMENA PYRIFORMIS GL-C : II. The Relationship of Protein Synthesis to Cell Division and Oral Organelle Development

The effects of puromycin on synchronized Tetrahymena pyriformis were investigated at two different concentrations, 43 µg per ml and 430 µg per ml. The rate of incorporation of histidine-¹⁴C into hot TCA-insoluble material was reduced by 30% at the low concentration and by 80–90% at the high concentration. The rate of oxygen uptake was lowered by only 10–20% at both concentrations. Cell division was prevented at both concentrations, if the drug was added prior to a "transition point" at about 45 min after the end of the synchronizing treatment. Development of "anarchic field" oral primordia was arrested, while primordia in early stages of membranelle differentiation were resorbed. Resorption began shortly after addition of the drug, and proceeded most rapidly at the lower concentration. If the drug was added after the "transition point," cell division and oral primordium formation were completed with only slight delay at the low concentration, and with considerable delay (in some cases complete arrest) at the high concentration. The results thus indicate that protein synthesis is involved in the later as well as the earlier stages of development; what specially characterizes the earlier stages, prior to the "transition point," is a dramatic response to partial inhibition of protein synthesis. It is suggested that this response involves the activation or release of a latent intracellular degradative system which is specific for developing structures.