Mechanisms of cell differentiation: Affinity of a morphogenetic factor to DNA

Summary A morphogenetic factor which induces inTriturus gastrula ectoderm tissues which are derived from mesoderm and endoderm has been extracted from chicken and amphibian embryos. The factor which is protein in nature has been obtained from chicken embryos in a highly purified state.The biological activity of the chicken factor is partially inhibited when the factor is combined with chicken DNA or sonicated chicken DNA.When the 3H-labelled factor is combined with sonicated DNA and then centrifuged on a sucrose gradient the factor migrates in part with the DNA. This indicates that the factor is bound to DNA.The inferences from these results are discussed with regard to the possible mechanism of action of the factor and the molecular mechanism of differentiation.     

Stimulation by 6-N,2′-O-dibutyryladenosine 3′:5′-cyclic monophosphate and glucocorticoids of phosphoenolpyruvate carboxykinase in the isolated perfused rat liver (Short Communication)

Dibutyryl cyclic AMP stimulated the activity of phosphoenolpyruvate carboxykinase in perfused livers of rats, fed on a low-protein diet, linearly over a 6h period. The enzyme activity was also significantly elevated by dexamethasone, the effect being considerably lower than that of the cyclic nucleotide. Since the time-course of phosphoenolpyruvate carboxykinase activity in response to dibutyryl cyclic AMP resembled that observed after dibutyryl cyclic AMP injection into intact animals, it is suggested that induction of the enzyme in vivo is due to a direct action of the cyclic nucleotide on the liver. Combined administration of dibutyryl cyclic AMP and glucocorticoids did not lead to an additive increase of liver phosphoenolpyruvate carboxykinase activity, either in vivo or in the perfused organ.     

Über den Einfluß der Vorbelichtung auf die anschließende Phosphorylierung im Dunkeln bei einzelligen Grünalgen (Ankistrodesmus braunii)

Zusammenfassung Wird einzelligen Algen (Ankistrodesmus braunii) nach Vorbelichtung in anschließender Dunkelheit ³²P-markiertes Phosphat geboten, so tritt gegenüber Dauerdunkel eine erhebliche Förderung der ³²P-Einlagerung auf. Die nach Vorbelichtung bestimmte Markierung der aufgetrennten Phosphatfraktionen ähnelt sehr derjenigen im Dauerlicht. Die erhöhte Dunkelphosphorylierung nach Vorbelichtung hängt von der CO₂-Konzentration, von der Lichtintensität und der Zeit der Vorbelichtung ab. Unter den vorliegenden Bedingungen waren 7 min Vorbelichtung zur maximalen Förderung nötig. Die Halbwertzeit des Abklingens betrug etwa 4 min.Aus den Experimenten geht hervor, daß durch die Belichtung der Algen auch in vivo ein Zustand gebildet wird, der noch nach Belichtung eine Zeitlang im Dumkeln eine Erhöhung der ³²P-Einlagerung erlaubt.Es wird diskutiert, ob es sich einerseits um die Bildung einer im Licht reduzierten Substanz R handeln könnte, die für eine begrenzte Zeit in Dunkelheit noch einen cyclischen, mit Phosphorylierung gekoppelten Elektronentransport aufrechterhalten kann. Andererseits könnte durch die Vorbelichtung ein energiereiches Zwischenprodukt X _E — oder auch ein Protonenpool — gebildet werden, das bei dem Energietransfer vom Elektronentransportsystem zur ATP aufgebaut wird. Schließlich muß berücksichtigt werden, daß durch die Vorbelichtung an den Chloroplastenmembranen ein verstärkter ATP-Pi-Austausch zustande kommen könnte, der nach Belichtung nur langsam abklingt.

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Influence of preillumination on subsequent phosphorylation in the darkness of unicellular green algae (Ankistrodesmus braunii)

Summary Preilluminated unicellular green algae (Ankistrodesmus braunii) were treated in the subsequent darkness with ³²PO₄. The post-illumination dark incorporation was considerably increased compared with the control in continuous dark. The labeling of the separated phosphate-fractions was similar to that of continuous light. The light-induced dark incorporation depended from the light intensity as well as from the time of preillumination. A preillumination of 7 min was required for a maximal enhancement of this preillumination effect. On the other hand the effect diminished in darkness with a half life of approximately 4 min. Finally the enhancement was found to be greater in the absence of CO₂ than in the presence of CO₂.The experiments demonstrate the light-induced formation of a state in the algae, which permits the enhancement of ³²P-incorporation into several phosphate-fractions for a limited time during subsequent darkness.It is discussed, that this may be performed through the formation of a light-reduced substance R maintaining for a limited time a cyclic electron transport in darkness, coupled with phosphorylation. On the other hand it seems possible, that preillumination induces a high energy intermediate X _E—this could also be a pool of protons—formed in the course of energy-transfer from electron transport to ATP-formation. But we must consider also the possibility that light accellerates the ATP-Pi exchange on chloroplast-membranes for a time after preillumination.

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Abkürzungen ATP Adenosintriphosphat - ADP Adenosindiphosphat - Pi Orthophosphat - Poly-P anorganisches Polyphosphat - RNS Ribonucleinsäure - TCE Trichloressigsäure - 2,4-DNP 2,4-Dinitrophenol Stipendiat der Nishina-Gedächtnis-Stiftung (Japan) für 1963.     

Kinetics of insulin internalization and processing in adipocytes: effects of insulin concentration

We have studied the effect of insulin concentration on the kinetics of insulin internalization and efflux in isolated rat adipocytes. To determine internalization rates adipocytes were incubated with 125I-insulin at 37 degrees C; and at frequent, early time points surface-bound and intracellular insulin were quantitated. Surface-bound and intracellular insulin were discriminated by the sensitivity of the former to rapid dissociation by a pH 3.0 buffer at 4 degrees C. From this data the endocytotic (internalization) rate constant (ke) was calculated for six insulin concentrations ranging from 0.3 to 100 ng/ml. Ke was found to decrease in an insulin concentration-dependent manner (P less than .001). Thus, values for ke were 0.121 +/- 0.006 min-1 versus 0.074 +/- 0.011 min-1 at 0.3 ng/ml and 100 ng/ml, respectively. The decrease in ke did not parallel insulin concentration-dependent changes in insulin receptor affinity indicating it was not the result of an inability of low affinity receptors to be internalized. The kinetics of insulin efflux were determined by loading various concentrations of 125I-insulin into the adipocyte interior, washing away surface-bound and extracellular insulin, and then monitoring the subsequent efflux of pre-loaded insulin into medium that contained the same concentration of insulin used in the loading step. The overall rate of efflux was independent of insulin concentration. In summary, these results show that at high insulin concentrations the efficiency of insulin internalization is impaired. In contrast, the rate of insulin efflux is unaffected.     

Control of Mn status and infection rate by genotype of both host and pathogen in the wheat take-all interaction

Take-all is a world-wide root-rotting disease of cereals. The causal organism of take-all of wheat is the soil-borne fungus Gaeumannomyces graminis var tritici (Ggt). No resistance to take-all, worthy of inclusion in a plant breeding programme, has been discovered in wheat but the severity of take-all is increased in host plants whose tissues are deficient for manganese (Mn). Take-all of wheat will be decreased by all techniques which lift Mn concentrations in shoots and roots of Mn-deficient hosts to adequate levels. Wheat seedlings were grown in a Mn-deficient calcareous sand in small pots and inoculated with four field isolates of Ggt. Infection by three virulent isolates was increased under conditions which were Mn deficient for the wheat host but infection by a weakly virulent isolate, already low, was further decreased. Only the three virulent isolates caused visible oxidation of Mn in vitro. The sensitivity of Ggt isolates to manganous ions in vitro did not explain the extent of infection they caused on wheat hosts. In a similar experiment four Australian wheat genotypes were grown in the same Mn-deficient calcareous sand and inoculated with one virulent isolate of Ggt. Two genotypes were inefficient at taking up manganese and were very susceptible to take-all, one was very efficient at taking up manganese and was resistant to take-all, and the fourth genotype was intermediate for both characters. All genotypes were equally resistant under Mn-adequate conditions.