Factors controlling the rhythmic contraction of collagen gels by neonatal heart cells

Summary A floating collagen matrix culture of neonatal rat heart myocardial cells shows rhythmic contractions which are dependent on localization of cells, cell density, and collagen concentration. The rhythmic contractions of the collagen matrix can be registered by a device scanning the optical density at the edge of the gel and have been observed over a temperature range from 9° to 40° C. The results of the present study underline the usefulness of myocardial cell populated collagen matrixes for studies on coherent contractions of heart cell cultures.     

Regulation of amidase formation in mutants from Pseudomonas aeruginosa PAO lacking glutamine synthetase activity

The formation of amidase was studied in mutants from Pseudomonas aeruginosa PAO lacking glutamine synthetase activity. It appeared that catabolite repression of amidase synthesis by succinate was partially relieved when cellular growth was limited by glutamine. Under these conditions, a correlation between amidase and urease formation was observed. The results suggest that amidase formation in strain PAO is subject to nitrogen control and that glutamine or some compound derived from it mediates the nitrogen repression of amidase.     

Computer analysis of two-dimensional gels

New methods and computer programs are described which enable one to analyze autoradiograms produced by two-dimensional gel electrophoresis. These programs are completely automatic with respect to finding spots resolved by such gels and quantitating the radioactivity in them. Semiautomatic programs have also been developed to match the spot patterns of different autoradiograms, and to follow the synthesis of any individual polypeptide through a series of gels.     

Immobilization of protoplasts by anchoring to microcarriers

Datura innoxia cell suspension-derived protoplasts were anchored to Cytodex 1 microcarriers pre-swollen in buffered concanavalin A. As many as 34 protoplasts were estimated to attach per microcarrier, in comparison to a potential 47 as determined from a model based on random anchorage. Fluorescein diacetate was used as localizing agent as well as to assess viability. When included in the swelling medium fluorescence was observed almost instantaneously, first in the protoplast at its interface with the microcarrier, and later throughout the cytoplasm. However, the dye was not conjugated with the lectin, and leakage eventually resulted in fluorescence also of non-anchored protoplasts. Fluorescein-labelled concanavalin A on the other hand permitted detection of microcarriers but not of anchored protoplasts, suggesting the use of differentially fluorescing microcarriers, as an aid in identification of fusion partners.     

The enzymes of the ammonia assimilation in Pseudomonas aeruginosa

Glutamine synthetase from Pseudomonas aeruginosa is regulated by repression/derepression of enzyme synthesis and by adenylylation/deadenylylation control. High levels of deadenylylated biosynthetically active glutamine synthetase were observed in cultures growing with limiting amounts of nitrogen while synthesis of the enzyme was repressed and that present was adenylylated in cultures with excess nitrogen.NADP-and NAD-dependent glutamate dehydrogenase could be separated by column chromatography and showed molecular weights of 110,000 and 220,000, respectively. Synthesis of the NADP-dependent glutamate dehydrogenase is repressed under nitrogen limitation and by growth on glutamate. In contrast, NAD-dependent glutamate dehydrogenase is derepressed by glutamate. Glutamate synthase is repressed by glutamate but not by excess nitrogen.     

Location and orientation relative to the micelle surface for glucagon in mixed micelles with dodecylphosphocholine EPR and NMR studies

The spin labels, 5-doxylstearate, 12-doxylstearate, 16-doxylstearate and 1-oxyl-2,2,6,6-tetramethyl-4-dodecylphospiperidine, have been incorporated into dodecylphospocholine micelles and mixed dodecylphosphocholine/ glucagon micelles. The EPR spectral parameters for the different spin labels and the ¹H- and ¹³C-NMR relaxation rates for nuclei of the detergent molecules indicated that inclusion of up to one spin label molecule per micelle had little influence on the spatial organization of the micelles. Furthermore, the location and environment of the spin labels in the dodecylphosphocholine micelles were not noticeably affected by the addition of glucagon and the ¹H-NMR spectra observed for glucagon in mixed spin label/deuterated dodecylphosphocholine/glucagon micelles showed that the different spin labels had essentially no effect on the conformation of glucagon. Approximate spatial locations within the micelle for the nitroxide moieties of the different spin labels were determined from the NMR relaxation rates observed for different nuclei of dodecylphosphocholine. On this basis, the line broadening of individually assigned glucagon ¹H-NMR lines by the different spin labels was used to determine the approximate orientation of the polypeptide chain with respect to the micelle surface. Overall, the data indicate that the glucagon backbone runs roughly parallel to the micelle surface, with the depth of immersion adjusted so that polar and apolar side chains can be oriented towards the surface or interior of the micelle, respectively.     

On the study of two interacting populations one of which acts independently of the other

Many ecological and biological systems can be studied in terms of a bivariate stochastic branching process, {X ₁ (t), X ₂ (t)}, each of whose components (or populations) varies in magnitude according to the laws of a generalized birth-death process. Of particular interest is such a model in which the birth and death rates of the first population,X ₁, are constant while those of the second population,X ₂, exhibit a functional dependence upon the magnitude of the first. It is shown, first, that the existence of the stochastic mean of a birth death process implies the existence of all higher moments. The values of all the factorial moments of such a process are then determined. The moments of the dependent population of the bivariate process are given in terms of its expectation and the joint probability density function of the process is determined. It is possible, therefore, to use Bayesian techniques to infer conclusions about the independent population, given information about the variation of the dependent one.     

Free cytoplasmic messenger ribonucleoprotein complexes from rabbit reticulocytes

Free cytoplasmic globin mRNA containing mRNP-particles were isolated from rabbit reticulocytes by zonal sucrose gradient centrifugation and their properties were compared with mRNP particles isolated in the same way from EDTA-dissociated reticulocyte polyribosomes. The average poly(A)-length of 9S mRNA from free cytoplasmic mRNP was 17–20 nucleotides being about two times shorter than the average poly(A)-length of polysomal 9S mRNA. The protein composition of the free cytoplasmic mRNP particles disclosed the absence of the 76,000 dalton protein which is associated with the 3poly(A)-segment of polysomal globin mRNA. It was concluded that free cytoplasmic mRNP-particles from rabbit reticulocytes can be classified as old mRNP in a post-translational phase. Free cytoplasmic mRNPs were translated in heterologous cell-free systems as well as in Xenopus laevis oocytes. Addition of hemin stimulated the synthesis of -globin in all systems, while the presence of the cap analogue m⁷G(5)p inhibited translation of free cytoplasmic mRNA completely. The latter finding suggested that free cytoplasmic mRNA has a 5 terminal cap. Shortening of the poly(A)-segment with concomitant loss of the 76,000 dalton protein may lead to less efficient translation of free cytoplasmic mRNP.     

A model for ganglioside behaviour in cell membranes

Gangliosides from beef brain have been spin-labeled using two different attaching groups and employed to investigate the physical nature of ganglioside behaviour in membranes. Results obtained using EPR spectroscopy indicate that, in phosphatidylcholine bilayers at physiological pH, ganglioside oligosaccharide chains are quite mobile and show a measurable tendency towards cooperative interaction amongst themselves. We suggest that the source of this interaction is the formation of H-bonds between sugar residues in adjacent ganglioside molecules. We present evidence that physiological (extracellular fluid) levels of Ca²⁺ and Mg²⁺ lead to cross-linking and condensing of ganglioside headgroups by complexing sialic acid carboxyl residues. Ganglioside headgroup interactions are not very sensitive to changes in the buffer ionic strength, suggesting that ionic interactions are of minor importance. We have found no measurable tendency for headgroup carbohydrate to penetrate hydrophobic regions of lipid bilayers. EPR spectroscopy was also used to follow the interaction of spin-labeled gangliosides with the glycoprotein, glycophorin, and with intact BHK cells.We suggest that these carbohydrate-based interactions should contribute significantly to the properties of the eucaryotic cell glycocalyx. We predict that laterally mobile carbohydrate-bearing components of cell surfaces will show a tendency to cluster about complex glycoprotein arrays, especially if the species involved bear accessible carboxylic acid functions.