Cytogenetic replication studies with short thymidine pulses in bromodeoxyuridine-substituted chromosomes of different mouse cell lines

Summary In normal diploid fibroblasts of the mouse, 3T3-, SV-3T3-, and Meth A-cells, the chromosome replication patterns were studied by a bromodeoxyuridine (BrdU)-labelling technique. SV-3T3 is a subline of 3T3 transformed by SV 40 and Meth A is a permanent cell line from Balb c transformed by methylcholanthrene. The use of 1 h thymidine pulses permits high resolution of the S-phase after partial synchronization of the cells at G₁/S in an otherwise BrdU-substituted S-phase. It could be shown that the autosomal heterochromatin of the mouse (Mus musculus) starts replication during the early S-phase (R-band replication), continues while R-band chromatin finishes, and still replicates when G-band chromatin starts. The heterochromatin finishes before the majority of G-bands have been replicated. There is no fundamental difference in the course of chromosome replication between the different cell lines studied here. It is concluded that there are no obligate changes in the course of the S-phase linked to the process of transformation.     

Effects of potassium,lanthanum and black widow spider venom on miniature synaptic potentials in theTorpedo electroplax

Summary Using a method of focal drug application it is demonstrated that high potassium concentration, lanthanum, and black widow spider venom accelerate spontaneous transmitter release inTorpedo electric tissue.This investigation was supported by the Deutsche Forschungsgemeinschaft (Sonderforschungsbereich 138). Thanks are due to Dr. R. Martin and the staff of the Stazione Zoologica, Naples, for supplyingTorpedo, and Dr. N. Frontali, Rome, for a gift of frozen black widow spiders.     

The Isolation of Colicine V and a Study of Its Immunological Properties

Colicine V has been obtained from the culture medium in which the colicinogenic bacillus E. coli K357 L_T is grown. The material is electrophoretically homogeneous and proves to be a lipocarbohydrate protein complex identical with the type-specific O antigen of the parent bacillus. Colicine V is toxic both for mice and for rabbits and readily stimulates the elaboration of precipitins and bacterial agglutinins, as well as antibodies which neutralize the antibacterial activity of the colicine itself. The colicine is also toxic for certain strains of Enterobacteriaceae. Although colicine V and colicine K, previously described in this laboratory, have many properties in common, they exhibit no cross-serological relationship whatsoever.     

An Electron Microscope Study of Myelin Figures

In the electron microscope, thin sections of OsO₄-fixed myelin figures from the phospholipide fraction of human brain show a pattern of parallel dark lines with a repeating period of about 40 A. It is shown that the dark lines probably represent the reaction product of OsO₄ with double bonds in the fatty acid chains, thereby marking the central portion of one bimolecular lamella. The addition of globin results in dense lines 25 to 50 A wide that cover the surface of the myelin figures. When such a figure consists of only two bimolecular leaflets of lipide covered with globin, the structure shows striking similarity to the image of cell membranes in fixed tissue sections. A hypothetical schema is given of the molecular structure of the figure, and the distribution of OsO₄ in it.     

How does inactivation change timing of replication in the human X chromosome?

Summary The kinetics of replication of the inactive (late replicating) X chromosome (LRX) were studied in karyotypically normal lymphocytes and human amniotic fluid cells. Both cell types were successively pulse labeled with 1-h or 1/2-h thymidine pulses in an otherwise BrdU-substituted S phase after partial synchronization of the cultures at G₁/S. For the first time with this technique, the entire sequence of replication was analyzed for the LRX from the beginning to the end of the S phase, with special reference to mid S (R-band to G-band transition replication). The inactive X is the last chromosome of the metaphase to start replication, with a delay of 1 or 2h, after which time a thymidine pulse results in R-type patterns. In mid S, the inactive X is the first chromosome to switch to G-type replication (without overlapping of both types and without any detectable replication pause). Until the end of S, a thymidine pulse results in G-type patterns. To rule out artifacts that might arise by the synchronization of cultures in these experiments, controls were carried out with BrdU pulses and the BrdU antibody technique without synchronization. In the course of replication, no fundamental difference was seen between the two different cell types examined. In contrast to studies using continuos labeling, this study did not reveal an interindividual difference of replication kinetics in the LRXs of the seven individuals studied; thus it is concluded that the inactive X chromosome shows only one characteristic course of replication.