Characterization of a high-affinity binding site for a DNA-binding protein from sea urchin embryo mitochondria.

Based on electrophoretic mobility shift assays, DNase I footprinting and modification interference analyses we have identified a sequence-specific DNA-binding protein in blastula stage mitochondria of the sea urchin Strongylocentrotus purpuratus, which interacts with a binding site around the major pause site for DNA replication. This region straddles the boundary of the genes for ATP synthase subunit 6 and cytochrome c oxidase subunit III, and contains also a prominent origin of lagging-strand synthesis. The protein is thermostable, and its natural high-affinity binding site comprises the sequence 5'-AGCCT(N7)AGCAT-3'. Binding studies have demonstrated that two copies of the imperfect repeat, as well as the 7 bp spacing between them, are essential for tight binding. Based on the location of its binding site, we tentatively designate the protein mitochondrial pause-region binding protein (mtPBP) 1.     

Cryopreservation of immature spring wheat zygotic embryos using an abscisic acid pretreatment

Spring wheat (Triticum aestivum L.) zygotic embryos were successfully cryopreserved, without the addition of exogenous cryoprotectants, using only an abscisic acid (ABA) pretreatment. Optimum survival was obtained when embryos were cultured in vitro for 10 days on semisolid Murashige and Skoog (MS) nutrient medium supplemented with 0.5 mg/L (±) ABA prior to cryopreservation. The embryos resumed growth within three days when returned to MS medium devoid of ABA but containing 2mg/L 2,4-dichlorophenoxyacetic acid. The embryogenic calli produced from these embryos exhibited normal plant regeneration on auxin-free media. Changes in dw/fw ratio, as well as the esterified fatty acid and sucrose concentrations correlated positively with the development of tolerance to cryopreservation.NRCC Publication No. 33519     

The rhodopsin system of the squid

Squid rhodopsin (λ_max 493 mµ)—like vertebrate rhodopsins—contains a retinene chromophore linked to a protein, opsin. Light transforms rhodopsin to lumi- and metarhodopsin. However, whereas vertebrate metarhodopsin at physiological temperatures decomposes into retinene and opsin, squid metarhodopsin is stable. Light also converts squid metarhodopsin to rhodopsin. Rhodopsin is therefore regenerated from metarhodopsin in the light. Irradiation of rhodopsin or metarhodopsin produces a steady state by promoting the reactions, See PDF for Equation Squid rhodopsin contains neo-b (11-cis) retinene; metarhodopsin all-trans retinene. The interconversion of rhodopsin and metarhodopsin involves only the stereoisomerization of their chromophores. Squid metarhodopsin is a pH indicator, red (λ_max 500 mµ) near neutrality, yellow (λ_max 380 mµ) in alkaline solution. The two forms—acid and alkaline metarhodopsin—are interconverted according to the equation, Alkaline metarhodopsin + H⁺ acid metarhodopsin, with pK 7.7. In both forms, retinene is attached to opsin at the same site as in rhodopsin. However, metarhodopsin decomposes more readily than rhodopsin into retinene and opsin. The opsins apparently fit the shape of the neo-b chromophore. When light isomerizes the chromophore to the all-trans configuration, squid opsin accepts the all-trans chromophore, while vertebrate opsins do not and hence release all-trans retinene. Light triggers vision by affecting directly the shape of the retinene chromophore. This changes its relationship with opsin, so initiating a train of chemical reactions.     

Metal tolerance in tissue cultures of anthoxanthum odoratum

Tissue cultures were initiated from one non-tolerant (S20) and two zinc and lead tolerant (T92 and T94) clones of Anthoxanthum odoratum. Growth of callus from the non-tolerant clone was reduced by the presence of zinc, lead, copper and nickel, whereas callus from the two tolerant clones showed no reduction of growth in the presence of zinc and lead but growth was reduced by copper and nickel. The specificity of metal tolerance shown by the parental material was maintained in the callus. Tolerant and non-tolerant callus accumulated similar amounts of zinc and lead.     

Incubation in Mice Provides a Signal for the Differentiation of Trypanosoma cruzi Epimastigotes to Trypomastigotes1

The differentiation of Trypanosoma cruzi epimastigotes into trypomastigotes was studied in diffusion chambers sub-cutaneously implanted in mice. Using epimastigotes of the Tulahuén strain, transformation was first evident at 16 h after implantation and reached its maximum (92% trypomastigotes) by 24 h. Shortly before their differentiation into trypomastigotes, epimastigotes were found to develop resistance to lysis by the alternative pathway of complement. Furthermore, implantation of stationary-phase (as opposed to log-phase) parasites resulted in the accumulation of large numbers of complement-resistant epimastigotes in the chambers. These observations suggest that epimastigotes pass through a complement-resistant transitional stage before differentiating into trypomastigotes and that transformation may require cell division. In a further series of experiments, epimastigotes recovered 7 h after implantation in mice were found to differentiate into trypomastigotes when cultured in vitro for an additional 17 h at 37°C. This observation indicates that the events which trigger the morphologic transformation of epimastigotes into trypomastigotes can be dissociated operationally from the differentiation process itself.     

L-histidine induced changes in adenosine 3':5'-monophosphate levels in rat brain and amounts of apo-, holo-a and holo-b fatty acid synthetases in rat liver.

When fasted rats were fed a chow or fat-free diet supplemented 5% with L-histidine for three days, the brain adenosine 3′:5′-monophosphate (cAMP) level increased. A 50% increase occurred in rats fed a chow diet and 20% increase in rats fed a fat-free diet. Purification of liver fatty acid synthetase and the isolation of liver apo-, holo-$\text{a}$ and holo-$\text{b}$ fatty acid synthetases demonstrated that L-histidine feeding caused changes in the relative amounts of these enzymes. Apo- and holo-$\text{b}$ fatty acid synthetases increased while the holo-$\text{a}$ form simultaneously decreased. This effect was observed in rats fed either chow or fat-free diets supplemented with L-histidine.     

L-histidine-induced suppression of lipogenic enzymes.

Diets supplemented 5% with L-histidine produce hypercholesterolemia and increase cholesterol biosynthesis in rat liver. We now report an $\text{inhibitory}$ effect of L-histidine on lipogenic enzymes in the liver. In this study, L-histidine was added to chow and fat-free diets and fed to rats for 18 days. After two days of fasting, the rats were refed the same diet for three days prior to sacrifice. L-histidine decreased fatty acid synthetase activity by 51% when it was added to the chow diet and by 26% when it was added to the fat-free diet. Isocitrate dehydrogenase activity was not altered significantly in rats fed diets supplemented with L-histidine.     

Mutants of Salmonella typhimurium responding to cysteine or methionine: their nature and possible role in the regulation of cysteine biosynthesis.

Nineteen mutants of Salmonella typhimurium responding to either cysteine or methionine (cym) have been identified amongst cysteine (cys) and methionine (met) auxotrophs. Their growth responses to known intermediates in the related pathways of cysteine and methionine biosynthesis and complementation patterns in abortive transduction tests divided the mutants into six groups. Results of conjugation, cotransduction and deletion mapping experiments substantiated these groups, each of which carried a lesion within known cys genes. Enzyme assays on cym mutants from five of the six groups confirmed their cys gene deficiencies. Growth response and enzyme assay data were not consistent with mutants being leaky cys mutants (spared by methionine). None of eight cym mutants tested were able to convert [35S]methionine into [35S]cysteine. Selenate specifically inhibits the early enzymes of cysteine synthesis. In cym mutants this inhibition was relieved by cysteine but not by methionine, indicating that cym mutants require active cys enzymes for growth on methionine. There was evidence that methionine stimulated in vivo activity of cys enzymes in a cym mutant. Resistance to inhibition by 1,2,4-triazole results in reduced levels of the O-acetyl serine sulphydrylase. In cym mutants triazole resistance gave unstable suppression of the cym phenotype. Cym mutants may result from mutation in regulatory regions common to each of the cys genes, with the precise role of methionine as yet unknown.