Characterization of the microtubule movement produced by sea urchin egg kinesin

We have used an in vitro assay to characterize some of the motile properties of sea urchin egg kinesin. Egg kinesin is purified via 5'-adenylyl imidodiphosphate-induced binding to taxol-assembled microtubules, extraction from the microtubules in ATP, and gel filtration chromatography (Scholey, J. M., Porter, M. E., Grissom, P. M., and McIntosh, J. R. (1985) Nature 318, 483-486). This partially purified kinesin is then adsorbed to a glass coverslip, mixed with microtubules and ATP, and viewed by video-enhanced differential interference contrast microscopy. The microtubule translocating activity of the purified egg kinesin is qualitatively similar to the analogous activity observed in crude extracts of sea urchin eggs and resembles the activity of neuronal kinesin with respect to both the maximal rate (greater than 0.5 micron/s) and the direction of movement. Axonemes glide on a kinesin-coated coverslip toward their minus ends, and kinesin-coated beads translocate toward the plus ends of centrosome microtubules. Sea urchin egg kinesin is inhibited by high concentrations of SH reagents ([N-ethylmaleimide] greater than 3-5 mM), vanadate greater than 50 microM, and [nonhydrolyzable nucleotides] greater than or equal to [MgATP]. The nucleotide requirement of sea urchin egg kinesin is fairly broad (ATP greater than GTP greater than ITP), and the rate of microtubule movement increases in a saturable fashion with the [ATP]. We conclude that the motile activity of egg kinesin is indistinguishable from that of neuronal kinesin. We propose that egg kinesin may be associated with microtubule-based motility in vivo.     

The cytosol origin of macromolecules extruded by cultured chick embryo fibroblast cells

The DNA and protein extruded by chick embryo fibroblast cells has been analysed by chromatography. A high proportion of the DNA is in the form of a protein-complex of size around 5 X 10(5) dalton. The patterns of the DNA and protein extruded into the supernatant are closely similar in many respects to those found in the cell cytosol. It is concluded that the macromolecular material extruded by cells in culture is of cytosol origin: a possible function in terms of "information" carriage is proposed.     

Ultrastructural detection of the onset of pituitary thyrotroph sensitivity to lowered thyroid hormone concentrations in the fetal sheep

The goitrogen methylthiouracil was administered orally to pregnant ewes of known gestational ages to induce hypothyroidism in both mother and fetus. Developing pituitary thyrotrophic cells were studied using electron microscopy to detect the earliest gestational age at which morphological changes occurred in response to lowered plasma thyroid hormone concentrations. At 50 days of gestation, the pituitaries of fetuses exposed to the goitrogen were indistinguishable from untreated control glands. However, at 58 days and subsequent ages, "thyroidectomy' cells were observed in pituitaries of all hypothyroid fetuses. These findings indicate that fetal sheep pituitary thyrotrophs are sensitive to lowered thyroid hormone concentrations by 58 days of gestation, suggesting that thyroid-thyrotroph interaction exists at this early stage of development.     

Interdependence of inoculum size and trace element supply in growth of Aspergillus oryzae

Summary Further to earlier reports indicating that the effects of inoculum size on growth of Aspergillus oryzae in synthetic media are abolished by certain trace element compositions, it has now been shown that those effects can be made more pronounced, but can also be reversed, by varying the balance of available trace elements.Addition of Ca to substrate A ₁ increases the usual effects of inoculum size whereas Cu abolishes them with a tendency for reversal.Supplementation of substrate A ₁ with suitable amounts of ethylenediaminetetraacetic acid results in a stronger inhibition of small-inoculum cultures than large ones, thus increasing the usual effects of inoculum size. These effects are reversed if the sugar of substrate A ₁ is purified by Al(OH)₃ co-precipitation method, but only if the treatment is carried out at neutral or alkaline reaction. This reversal of the phenomena is due to an almost complete absence of Zn after purification and presence of Fe in substrate A ₁.The Al(OH)₃ treatment of the sugar results in substrates so poor in Zn that addition of as little as 5 g ZnSO₄ · 7 H₂O/l has an effect. This influence, however, results in an inhibition of the growth rate, and higher concentrations of Zn are required in order to obtain stimulation with both large and small-inoculum cultures.The above-mentioned reversal of the phenomena was abolished in a substrate with slightly caramelized glucose.

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Zusammenfassung Diese Arbeit ist eine Erweiterung vorhergehender Berichte über den Einfluß der Größe der Impfmenge auf das Wachstum von Aspergillus oryzae, wo u. a. festgestellt wurde, daß die beobachteten Effekte durch bestimmte Spurenelement-Zusammensetzungen aufgehoben werden.Es wurde nun festgestellt, daß Zugabe von Ca zu Substrat A ₁ die gewohnten Effekte erhöht, während Cu diese verringert mit einer Tendenz zur Umkehrung.Werden geeignete Mengen Äthylendiamintetraessigsäure zu Substrat A ₁ zugegeben, so werden Kulturen, die aus kleiner Impfmenge erwachsen sind, stärker in ihrer Wachstumsgeschwindigkeit und maximaler Ausbeute an Mycel gehemmt als Kulturen von großer Impfdichte, was einer Verstärkung der gewohnten Effekte gleich kommt. Diese Effekte werden umgekehrt, wenn die Glucose des Substrates A ₁ durch Al(OH)₃-Fällung gereinigt wird, jedoch nur wenn diese Behandlung bei neutraler oder alkalischer Reaktion erfolgt. Diese Umkehrung ist auf nahezu vollständige Entfernung von Zn und gleichzeitige Anwesenheit von Fe in Substrat A ₁ zurückzuführen.Schon Zugabe sehr kleiner Mengen Zn (5 g ZnSO₄ · 7 H₂O/l) zum gereinigten Substrat übt eine Wirkung aus, welche aber in einer Hemmung der Wachstumsgeschwindigkeit besteht. Höhere Zn-Konzentrationen sind nötig, um das Wachstum der Kulturen von großen und kleinen Impfmengen zu fördern.Die vorerwähnte Umkehrung der Phänomene wurde aufgehoben, wenn ein Substrat mit leichter Caramelisierung der Glucose angewendet wurde.