Effects of nucleotides and divalent cations on phospholipase activity in Saccharomyces cerevisiae

Divalent cations activate the lysophospholipase and transacylase reactions catalyzed by the same enzymes in the yeast Saccharomyces cerevisiae. The activation was observed at neutral pH, but not at the pH optimum of lysophospholipase/transacylase, near 3.5. Adenine nucleotides, especially AMP and ADP, are strong inhibitors of the same group of enzymes. Half maximal inhibition by AMP was found at a concentration of about 20 M. The inhibition by nucleotides in low concentrations is enhanced by divalent cations.     

Temperature jump study of charge translocation during the bacteriorhodopsin photocycle

Temperature jump experiments were carried out on purple membranes oriented and fixed in polyacrylamide gel. With green background illumination a relaxation of the photocurrent after an infrared laser pulse could be observed. To simulate the temperature jump signals different models of the bacteriorhodopsin photocycle were tested. The parameters of these models were obtained by measuring absorbance changes and photocurrent after excitation with a 575-nm laser flash.

A model with a temperature-dependent branching before the M state turned out to be satisfying. Other models, especially those with a late branching or without branching, could not reproduce the temperature jump measurements.

     

Protein kinase C inhibition by calmodulin and its fragments

Inhibition of protein kinase C (PKC) by calmodulin is investigated and we describe the localization of inhibitory sequences within the calmodulin molecule. We present evidence that calmodulin inhibits PKC through an inhibition of the activation of PKC associated with lipid membranes: Binding of PKC to lipid vesicles is not affected, but activation is abolished. The potent calmodulin antagonist R24571 (calmidazol) did not affect the inhibition of PKC by calmodulin at concentrations up to 10^–5 M. Two tryptic fragments of calmodulin were isolated which inhibited PKC. They were only slightly less potent than intact calmodulin with an IC₅₀ of 6 µ M compared to 1 µ M of intact calmodulin. They were identified as Ser³⁸-Arg⁷⁴ and His¹⁰⁷-Lys¹⁴⁸. Each of the inhibiting fragments contains an intact Ca²⁺-binding domain with complete helix-loop-helix structure (EF hand). Other calmodulin peptides showed only weak inhibitory activity. Both fragments did not stimulate cAMP phosphodiesterase even at concentrations 100-fold higher than the calmodulin concentration needed for maximal stimulation. None of the fragments acted as a calmodulin antagonist.     

Differentiation of embryonic chick sympathetic neurons in vivo: ultrastructure,and quantitative determinations of catecholamines and somatostatin

Summary The ultrastructural and transmitter development of lumbar sympathetic ganglia was studied in embryonic day-6 through-18 chick embryos. At embryonic day 6, ganglia are populated by two morphologically distinct types of neuronal cells and Schwann cell precursors. The neuronal populations basically comprise a granule-containing cell and a developing principal neuron. Granule-containing cells have, an irregularly shaped or oval nucleus with small clumps of chromatin attached to the inner nuclear membrane and numerous large (up to 300 nm) membrane-limited granules. Developing principal neurons display a more rounded vesicular nucleus with evenly distributed chromatin, prominent nucleoli, more developed areas of Golgi complexes, and rough endoplasmic reticulum and large dense-core vesicles up to 120 nm in diameter. There are granule-containing cells with fewer and smaller granules which still display the nucleus typical for granule-containing cells. These granule-containing cells may develop toward developing principal neurons or the resting state of granule-containing cells found in older ganglia. Both granule-containing cells and developing principal neurons proliferate and can undergo degeneration. At embryonic day 9 there are far more developing principal neurons than granule-containing cells. Most granule-containing cells have very few granules. Mitotic figures and signs of cell degeneration are still apparent. Synapse-like terminals are found on both developing principal neurons and granule-containing cells. Ganglionic development from embryonic day 11 through 18 comprises extensive maturation of developing principal neurons and a numerical decline of granule-containing cells. Some granule-containing cells with very few and small granules still persist at embryonic day 18. The mean catecholamine content per neuron increases from 0.044 femtomol at embryonic day 7 to 0.22 femtomol at embryonic day 15. Concomitantly, there is a more than 6-fold increase in tyrosine hydroxylase activity. Adrenaline has a 14% share in total catecholamines at embryonic day 15. Somatostatin levels are relatively high at embryonic day 7 (1.82 attomol per neuron) and are 10-fold reduced by embryonic day 15. Our results suggest the presence of two morphologically distinct sympathetic neuronal precursors at embryonic day 6: one with a binary choice to become a principal neuron or to die, the other one, a granule-containing cell, which alternatively may develop into a principal neuron, acquire a resting state or die.     

Changes in the auditory system of locusts (Locusta migratoria and Schistocerca gregaria) after deafferentation

Deafferentation experiments during postembryonic development show morphological and/or physiological changes of receptor fibers and of identified auditory interneurons in the CNS of the locusts Locusta migratoria and Schistocerca gregaria after unilateral ablation of one tympanic organ either in the larva or the adult animal.

Characterization of the state of differentiation of six newly established human non-small-cell lung cancer cell lines

Six new non-small-cell lung cancer (NSCLC) cell lines were established directly from human tissue or indirectly via nude mouse xenografts in serum-supplemented media with success rates of 8% and 13%, respectively. They comprised one adenocarcinoma (ADLC-5M2), two squamous cell carcinomas (EPLC-32M1, EPLC-65H), two large cell carcinomas (LCLC-97TM1, LCLC-103H), and one malignant biphasic mesothelioma (MSTO-211H). All cell lines grew adherent to culture vessels with population doubling times (PDT) of 16-40 h, formed colonies in soft agarose with efficiencies of 0.1%-5.1%, and all grew in athymic nude mice. Xenograft histologies appeared as follows: (a) undifferentiated carcinomas with feeble resemblance to the original tumors in the case of adenocarcinomas and squamous cell carcinomas; (b) large cell carcinoma with high resemblance to the original tumor; (c) an undifferentiated tumor with predominance of large epithelial cells and few fibrous cells in the case of mesothelioma. Human chorionic gonadotropin (HCG) was found by radioimmunoassay and high-affinity binding sites for epidermal growth factor (EGF) by radio-receptor assay in 4/4 cell lines. A very low activity of L-DOPA decarboxylase (DDC) was detectable only in the adenocarcinoma cell line. All cell lines overexpressed the c-myc protooncogene, and no gene rearrangement or amplification was observed. Chromosome analysis revealed modal chromosome numbers of 70-73 in ADLC-5M2, EPLC-32M1, EPLC-65H, and MSTO-211H. Cell lines derived from large cell carcinoma had modal values of 65 and 170 and a wider chromosome distribution than all other cell lines. A NSCLC specific chromosomal aberration has been undetectable until now. These cell lines may aid in elucidating the biology of NSCLC and its interrelationship to other lung tumors.     

Acoustic orientation in adult, female crickets (Gryllus bimaculatus de Geer) after unilateral foreleg amputation in the larva

Summary InGryllus bimaculatus females one foreleg was amputated at the coxa-trochanter joint in the 2nd, 4th or 8th/9th larval instar. A leg of up to normal length is regenerated (Fig. 1) but it lacks a functional ear. In spite of the, usually shorter, regenerated foreleg, the adult one-eared crickets show no impairments in walking when tested on a locomotion compensator. Without sound they walk erratically and most of them weakly circle towards the intact side (Fig. 2).With calling song presentation three response types can be distinguished:tracking (Fig. 3A), hanging on (Fig. 3B) or continuouscircling towards the intact side (Fig. 3C, D). Turning tendencies in monaurals increase with song intensity and exceed those of intact and bilaterally operated animals (Fig. 4). Course deviations towards the intact side also slightly increase with intensity (Fig. 5). Course stability is reduced compared to that of intact animals but exceeds that of bilaterally operated crickets (Figs. 5, 6). It is best at 60 dB and deteriorates at higher sound intensities (Fig. 6). The percentage of monaurals tracking or hanging on decreases with increasing intensity (Fig. 7B). Tracking is established in most animals but it is limited to a narrow intensity range (Fig. 7A, C). Apart from an increased percentage of tracking after early operations (Fig. 7D), there are no prominent changes in orientational parameters with the date of foreleg amputation.Reamputation of the regenerated leg in the adult monaurals does not significantly impair acoustic orientation (Figs. 8, 9), but occlusion of the ipsilateral prothoracic spiracle does (Figs. 10, 11).An attempt is made to correlate the behavioral performance with the activity of auditory interneurons which have undergone morphological and physiological changes (Fig. 12).     

Oxygen binding properties,capillary densities and heart weights in high altitude camelids

Summary The oxygen binding properties of the blood of the camelid species vicuna, llama, alpaca and dromedary camel were measured and evaluated with respect to interspecific differences. The highest blood oxygen affinity, not only among camelids but of all mammals investigated so far, was found in the vicuna (P₅₀=17.6 Torr compared to 20.3–21.6 Torr in the other species). Low hematocrits (23–34%) and small red blood cells (21–30 m³) are common features of all camelids, but the lowest values are found in theLama species. Capillary densities were determined in heart and soleus muscle of vicuna and llama. Again, the vicuna shows exceptional values (3720 cap/mm² on average in the heart) for a mammal of this body size. Finally, heart weight as percent of body weight is higher in the vicuna (0.7–0.9%) than in the other camelids studied (0.5–0.7%). The possibility that these parameters, measured in New World tylopodes at sea level, are not likely to change considerably with transfer to high altitude, is discussed.In the vicuna, a unique combination of the following features seems to be responsible for an out-standing physical capability at high altitude: saturation of blood with oxygen in the lung is favored by a high blood oxygen affinity, oxygen supply being facilitated by low diffusion distances in the muscle tissue. Loading, as well as unloading, of oxygen is improved by a relatively high oxygen transfer conductance of the red blood cells, which is due to their small size and which compensates the negative effect of a low hematocrit on the oxygen conductance of blood. Blood oxygen transport is presumably favored by two factors: a relatively large heart mass and, as a result of low hematocrit, a low blood viscosity. Both are advantageous for achieving a high maximal cardiac output.     

Detection of a single-copy gene on plant chromosomes by in situ hybridization

Summary DNA recombinant technology, combined with improvements in hybridization efficiency and quality of chromosome spreads, has made the method of in situ hybridization a reliable tool for gene mapping used by mammalian cytogeneticists to complement other methods. By appropriate alterations of the method, we demonstrate that detection of unique genes can be achieved along plant chromosomes despite some inherent disadvantages of the plant material. Using genomic subclones homologous to 6.6 kb of the single-copy chalcone synthase gene in parsley, we report the first example of chromosomal detection and localization of a unique endogenous gene in plants.