Structural alterations in synaptosomal membrane-associated proteins and lipids by transient middle cerebral artery occlusion in the cat

We have previously reported that ischemia reperfusion injury results from free radical generation following transient global ischemia, and that this radical induced damage is evident in the synaptosomal membrane of the gerbil. [Hall et al, (1995) Neuroscience 64: 81–89] In the present study we have extended these observations to transient focal ischemia in the cat. We prepared synaptosomal membranes from frontal, parietal-temporal, and occipital regions of the cat cerebral cortex with reperfusion times of 1 and 3 hours following 1 hour right middle cerebral artery occlusion. The membranes were selectively labeled with protein and lipid specific paramagnetic spin labels and analyzed using electron paramagnetic resonance spectrometry. There were significant motional changes of both the protein and lipid specific spin labels in the parietal-temporal and occipital regions with 1 hour reperfusion; but, both parameters returned to control values by 3 hours reperfusion. No significant changes were observed in the normally perfused frontal pole at either reperfusion time. These results support the argument that free radicals play a critical role in cell damage at early reperfusion times following ischemia.     

Characterization of heat-shock response of the marine bacterium Vibrio harveyi

We have investigated heat-shock response in a marine bacterium Vibrio harveyi. We have found that 39 C was the highest tempature at which V. harveyi was able to grow steadily. A shift from 30° C to 39° C caused increased synthesis of at least 10 proteins, as judged by SDS-PAGE, with molecular masses of 90, 70, 58, 41, 31, 27, 22, 15, 14.5 and 14kDa. The 70, 58, 41 and 14.5 kDa proteins were immunologically homologous to DnaK, GroEL, DnaJ and GroES heat-shock proteins of Escherichia coli, respectively. V. harveyi GroES protein had a lower molecular mass (14.5 kDa) than E. coli GroES, migrating in SDS-PAGE as 15 kDa protein. We showed that a protein of ～43 kDa, immunologically reactive with antiserum against E. coli sigma 32 subunit (σ³²) of RNA polymerase, was induced by heat-shock and co-purified with V. harveyi RNA polymerase. These results suggest that the 43 kDa protein is a heat-shock sigma protein of V. harveyi. Preparation containing the V. harveyi sigma 32 homologue, supplemented with core RNA polymerase of E. coli, was able to transcribe heat-shock promoters of E. coli in vitro.     

Correlation of tumor metastasis with sterol carrier protein and plasma membrane sterol levels

Squalene and sterol carrier protein (SCP) levels and sterol/phospholipid molar ratios of whole cells and plasma membranes were measured in cultured primary tumor and metastatic cell lines. SCP is abundant in all cell lines. However, metastatic lines have significantly lower SCP levels and plasma membrane sterol/phospholipid ratios than do primary lines. The results indicate that extremely malignant, metastatic cells are unable to produce or maintain adequate levels of both SCP and plasma membrane sterols when grown in lipoprotein deficient media. This defect, in vivo, probably causes excess uptake of SCP and lipid.     

The effects of recurrent inhibition on the cross-correlated firing patterns of motoneurones (and their relation to signal transmission in the spinal cord-muscle channel)

Cross-correlation histograms (CCH) were computed for discharge sequences of pairs of motoneurones which were excited by sinusoidal muscle stretches. These CCH's were compared before and after opening of the recurrent inhibitory loop by Renshaw cell blocking agents. Periodic patterns in the CCH's indicative of specifically timed phase relations between discharges of different motoneurones were enhanced after Renshaw cell blockage. This was confirmed by power spectra computed for the CCH's. They contained power peaks about 50Hz which tended to increase after depression of recurrent inhibition. The correlation was thus due predominantly to line current interference which seemed to act as a common entrainment input at the spinal level. It is concluded that Renshaw cells de-correlate discharge patterns of different motoneurones of the same pool by injecting uncorrelated signals into them. This de-correlation is an important prerequisite for distortion suppression of signal transmission in a multi-channel system, like that of stretch reflex, and for its linearization.     

Biological activity of gibberellin analogues

In order to determine the significance of the C-6 carboxyl group for the biological activity gibberellin A₃, 6-epigibberellin A₃, 7-norgibberellin A₃, 6-methyl-7-norgibberellin A₃, and 7-homogibberellin A₃ were studied using dwarf pea, dwarf maize, dwarf rice, dwarf barley and -amylase bioassays. All gibberellin A₃(GA₃)derivatives tested were considerably less active than GA₃. In all biossays, 6-epi-GA₃ showed a low activity of the same order, whereas 6-methyl-7-nor-GA₃ was inactive. Surprisingly, 7-nor-GA₃ had some activity in the dwarf rice (root application), dwarf barley, and -amylase bioassay, in contrary to its low potency in the dwarf pea, dwarf maize, and dwarf rice (micro drop) bioassay. 7-Homo-GA₃ was primarily active in the dwarf maize, dwarf barley and dwarf rice bioassay. It also caused antigibberellin effects in dwarf rice. The results demonstrate that the C-6 carboxyl group is not absolutely essential for biological activity of gibberellins. The different activities of 7-nor-GA₃ observed in the various test systems may indicate that the C-6 carboxyl group is a structural requirement more for uptake and/or transport processes than for receptor affinity.Abbreviation GA₃ gibberellic acid     

Two major regulatory steps in cholesterol synthesis by human renal cancer cells.

Two major mechanisms regulating cholesterol biosynthesis exist in a human renal cancer cell line, Caki-1. Caki-1 is a newly established cell line whose characteristics of rapid growth and active cholesterol synthesis qualify it as a potentially valuable tool for elucidation of regulatory mechanism of cholesterol synthesis and transport. In the absence of exogenous cholesterol, cholesterol is the dominant sterol arising from labeled acetate and mevalonate. As expected, in the presence of exogenous cholesterol, the conversion of acetate to cholesterol and the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase (EC 1.1.1.34) is markedly reduced and this inhibition is released when cholesterol is removed from the medium. An unexpected and possibly unique finding is the inhibition of the conversion of mevalonate to cholesterol in the presence of exogenous cholesterol. This second major control process results in the accumulation of squalene and may involve additional late steps in cholesterol biosynthesis or metabolism. The occurrence of two major mechanisms regulating cholesterol synthesis may be a unique property of renal cancer cells or a previously unrecognized characteristic of a variety of cultured cells.     

Lambda transducing phages derived from a FinO- R100::lambda cointegrate plasmid: proteins encoded by the R100 replication/incompatibility region and the antibiotic resistance determinant.

Summary Three transducing phages have been isolated from pEDR20, an R100:: cointegrate plasmid in which the insertion inactivated the R100 finO gene. Physical analysis of the three phages showed that the is inserted at kilobase coordinate 81.3 of R100. All three phages carry different amounts of R100 DNA in the left arm of . Each phage contains IS1b, the mer genes and the region between coordinate 81.3 and 88.6; thus, all contain the genes necessary for R100 replication. One phage, VA73, contains the entire r-determinant of R100 in addition to the above DNA. Five proteins coded by the region between 81.3 and 88.6 were detected. These had subunit molecular weights of 10,400; 12,200; 16,200; 19,600; and 38,300. The first was made constitutively and the other four only from a promoter. Other constitutive proteins were one from the cml fus region with a molecular weight of 22,400 (cml) and two from the str sul region with molecular weights of 31,500 (str?) and 30,100 (sul?). Mercuric ion induced synthesis of at least 10 proteins. Six of these were known from earlier work. The total size of the proteins which appear to derive from the mer genes exceeds by a factor of 1.5, the coding capacity of this region without overlapping genes. Some, or all of these extra proteins may be chromosomal in origin, possibly derepressed in response to mercury gene products.