Chromatin reconstitution on small DNA rings. II. DNA supercoiling on the nucleosome

DNA supercoiling on the nucleosome was investigated by relaxing with topoisomerase I mono- and dinucleosomes reconstituted on small DNA rings. Besides 359 base-pair (bp) rings whose linking differences were integers, two additional series of rings with fractional differences, 341 and 354 bp in size, were used. Mononucleosomes reconstituted on 359 bp rings were found to relax into a single mononucleosome form. In contrast, 341 and 354 bp mononucleosomes relaxed into a mixture of two forms, corresponding to two adjacent topoisomers. The observation that the ratio between these two forms was, within each ring series, virtually independent of the initial linking number of the topoisomer used for the reconstitution suggested that each partition reflected an equilibrium. Comparison with the equilibria observed for the same rings in the absence of histones showed that the formation of a single nucleosome is associated with a linking number change of -1.1(+/-0.1) turn. Dinucleosomes, in contrast, were not relaxed to completion and do not reach equilibria. The corresponding linking number change per nucleosome was, however, estimated to be similar to the above figure, in agreement with previous data from the literature obtained with circular chromatins containing larger numbers of nucleosomes. DNA structure in mononucleosomes was subsequently investigated by means of high-resolution electron microscopy and gel electrophoresis. It was found that the above linking number reduction could be ascribed to a particle with a large open extranucleosomal DNA loop and with no more than 1.5 turns of a superhelix around the histone core. A theoretical model of a nucleosome on a small ring was constructed in which one part of the DNA was wrapped around a cylinder and the other part was free to vary both in torsion and flexion. The linking number reduction predicted was found to be most consistent with experimental data when the twist of the DNA in the superhelix was between 10.5 and 10.65 pb per turn, suggesting that wrapping on the nucleosome does not alter the twist of the DNA significantly. A lower estimate of the linking number reduction associated with a two-turn nucleosome was also derived, based on an analysis of recent data obtained upon treatment of reconstituted minichromosomes with gyrase. The value, 1.6 turns, set a lower limit of 10.44 bp per turn for the twist of nucleosomal DNA, in agreement with the above estimate.(ABSTRACT TRUNCATED AT 400 WORDS)     

Stimulation of the chemiluminescence of human polymorphonuclear leukocytes in various stages of the intraerythrocyte development of Plasmodium falciparum

The synchronized cultures of Plasmodium falciparum were used to stimulate in vitro the chemiluminescence of human polymorphonuclear leukocytes in the presence of immune serum. The schizonts were concentrated by Percoll gradient centrifugation method (density 1.085 and osmolarity 285 mOsmol), and placed in culture, treated 6 hours later by sorbitol. Under incubation at constant temperature and pressure, the rate of synchronization reached 85% for schizonts during 5 replicative cycles. Every asexual stages of Plasmodium falciparum were used separately to stimulate polymorphonuclear leukocytes: merozoites were the most effective, followed by schizonts, trophozoites, and lastly supernatants of cultures containing degradation products of parasites.     

Brain metabolism of sarcoma 45-bearing rats undergoing radiation and the effect of hyperthermia on the tumor

Activity of hexokinase and acetylcholinesterase and pyridoxal co-enzyme content of brain subcellular fractions were studied in rats, bearing sarcoma 45, after local exposure of the tumor to 20 Gy X-radiation and microwave hyperthermia. The carbohydrate metabolism was sharply inhibited while the pyridoxal coenzyme content and acetylcholinesterase activity increased.     

Accelerated tissue expansion with the "overfilling" technique

Temporary overexpansion of the tissues at each filling session increased the actual inflation volume by about 59 percent. This shortened the process of tissue expansion considerably. "Overexpansion" means that the pressure in the expander is increased to a point where the dermal capillary flow is zero or the patient experiences so much discomfort that inflation cannot be carried out any further. The pressure in the expander is thereafter gradually decreased by taking out saline until both the capillary refill and all other clinical criteria are sufficient for safe dermal circulation. Comparison of the expander volume at the final pressure to the initial filling volume at the same pressure before the "overfilling" confirmed an average gain of about 59 percent in 14 patients.