The influence of 6-hydroxydopamine on the development and inducibility of drug metabolism in rat liver

6-Hydroxydopamine (6-OHDA) pretreatment of rats in the first 4 days of life considerably enhances the low hepatic ethylmorphine N-demethylation rate in 9 and 16 day old rats, whereas the higher rates in adult rats are not influenced. After 6-OHDA treatment the age differences in ethylmorphine N-demethylation rate disappear. The induction of ethylmorphine N-demethylation by phenobarbital is markedly enhanced by neonatal 6-OHDA pretreatment in 16 to 60 day old rats, whereas in 9 day old rats this synergistic action is not detectable. Unlike ethylmorphine N-demethylation, hepatic 7-ethoxy-coumarin 0-deethylation rate is only slightly or not at all influenced by 6-OHDA pretreatment. The basic activities in 9 day and rats are enhanced, the induction by phenobarbital in the same age-group is decreased. In other age-groups neither basic activities nor inducibility are influenced by 6-OHDA pretreatment. Hepatic cytochrome P-450 concentrations in 33- and 60 day old control rats do not differ from those in 6-OHDA-pretreated animals. Phenobarbital treatment increases the P-450 content; this induction effect is enhanced after 6-OHDA pretreatment, but not as markedly as with ethylmorphine N-demethylation.     

Replication of bacteriophage lambda DNA. Examination of variants containing double origins and observation of a bias in directionality

Bacteriophage λ variants have been constructed that possess two λ ori sites. Replicative intermediates resulting from infection with these phages have been investigated. We find that initiation of replication from the ori site on an EcoRI fragment (containing all the DNA sequences from within the red gene to the middle of gene O) cloned in the inverted orientation is predominantly bidirectional but occurs at a decreased frequency. Double initiations were observed at low frequency. However, a second cloned ori fragment (carrying two large deletions and a small insertion) cloned in the normal orientation demonstrated insignificant levels of replication from the cloned site unless the normal ori had already initiated.A bias in directionality of λ replication has been observed. Molecules that replicate unidirectionally propagate to the right more often than to the left. If the cloned ori-containing EcoRI fragment is inserted with reversed polarity, then the bias is towards the left. Bidirectional λ replicative intermediates also appear to show a similar bias but this is superimposed on a large, apparently random, effect that results in asymmetric growing-point propagation.     

Effects of butylated hydroxyanisole on the aryl hydrocarbon hydroxylase of rats and mice

The effects of butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) on the aryl hydrocarbon hydroxylase (AHH) activities in the liver, lung and skin of rats and mice have been studied to examine the possible mechanisms of the anticarcinogenic actions of these compounds. Both compounds inhibit the hydroxylase activities of hepatic microsomes and nuclei, with BHA a more potent inhibitor than BHT. The AHH of lung microsomes is inhibited to a lesser extent by BHA and BHT than that of the liver. The AHH activities of both liver and lung microsomes become less susceptible to the inhibition after pretreatment of the animals with 3-methylcholanthrene (MC) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) but phenobarbital (PB) pretreatment does not produce such an effect. In skin homogenates, however, the AHH activities of control rats and mice are not inhibited by BHA and BHT. The only skin sample which is inhibited by BHA and BHT is that from TCDD-pretreated mice. It has been established that the extent of inhibition with different samples is related to the concentration of BHA in the incubation but not to the amounts or specific activities of microsomes used. Double reciprocal plots suggest that BHA exerts a mixed inhibition on the hydroxylase of liver microsomes with a K_i of 7.7 μM. Analysis of the metabolites of benzo[a]pyrene (BP) shows that BHA inhibits the formation of various metabolites uniformly without changing the regio-selectivity of the enzyme system. The mechanism of inhibition has also been studied with a reconstituted AHH system consisting of cytochrome P-450 (P-450), reductase and phospholipid. The system with P-450 isolated from PB-induced microsomes is inhibited to a much greater extent than that with MC-induced P-450. The results indicate that the inhibitory action of BHA is dependent on the species of the animal, tissue types and treatment with inducers.     

Specificity of Insertion by the Translocatable Tetracycline-Resistance Element Tn10

Genetic analysis of 131 independent transpositions of the tetracycline-resistance element Tn10 from a single site in phage P22 into the histidine operon of Salmonella typhimurium reveals that Tn10 insertions are not randomly distributed along this chromosomal target. The insertions occur in 22 different "clusters"; insertions within each cluster are very tightly linked in recombination tests. Tn10 insertions are not evenly distributed among the identified clusters. The existence of these clusters suggests that this chromosomal target contains particular genetic signals that guide Tn10 to particular preferred positions for insertion. Insertions within each cluster occur in both orientations with roughly equal frequency.--The relationship among different insertions within each cluster has been examined. The resolution of genetic mapping places an upper limit of about 50 basepairs on the distance between different insertions within a cluster. Different insertions within a cluster usually have the same reversion frequency; however, heterogeneity in reversion frequency has been detected in at least two clusters. For most clusters, the available data are consistent with the simple possibility that all insertions within a cluster are at identical positions; however, the data do not exclude other possibilities.     

Transdetermination of Drosophila imaginal discs cultured in vitro

Imaginal discs of Drosophila melanogaster undergo transdetermination when cultured in vivo in the abdominal cavity of adult female hosts. We report here that leg discs cultured in vitro, in a recently developed system, also undergo transdetermination. Whether cultured in vivo or in vitro, leg discs produce a similar range of specific transdetermined structures. Moreover, in comparison to discs cultured in vivo, the discs cultured in vitro exhibit a similar correlation between the amount of growth and the total frequency of transdetermination.     

Protoplast induction inMicrasterias andCosmarium

Summary Protoplasts of the desmidsMicrasterias angulosa, M. denticulata, M. thomasiana andCosmarium turpinii were obtained by incubating cells in Waris' liquid medium + 0.3 M mannitol + 2% Cellulysin for 1–3 hours. One osmotically fragile protoplast was formed at the isthmus from the joint contents of both semicells. The resultant protoplasts were bright green and remained so for more than 5 days in the osmotically protective medium. The protoplast yield was better than 80%. The empty cell walls were not digested by the Cellulysin or by autolytic enzymes.     

The zinc content of yeast alcohol dehydrogenase

Analyses for zinc in high specific activity preparations of yeast alcohol dehydrogenase (YADH) indicate a metal content of 1.8–1.9 moles of zinc per mole of enzyme subunit. This zinc content is observed for YADH prepared from Bakers yeast by recrystallization from Am₂SO₄ containing 1 mM EDTA, followed by chromatography on DE-52 and Sephadex-G-200. YADH obtained from Boehringer-Mannheim is characterized by a variable specific activity: preparations with Sp. Ac. = 380–400 U/mg contain 1.8–1.9 moles of zinc per mole of subunit. Dialysis of YADH against EDTA (pH 8.5, 25°, under N₂) reduces the specific activity and zinc content in an approximately linear fashion down to a Sp. Ac. = 150 U/mg, consistent with the preferential loss of a single, weakly bound zinc per subunit which is essential for catalytic activity. Dialysis of YADH against 1 mM ZnCl₂ (pH 6.5–8.5, 25°, under N₂) does not lead to an increase in the zinc content of the enzyme, indicating that under these conditions zinc does not bind adventitiously to YADH. Dialysis against 50 mM CoSO₄ (pH 5.5, 25°, under N2, 60–90 hr) leads to an exchange of ≈ 40% of the enzyme-bound zinc by cobalt. Our preparations of YADH are consistently characterized by a zinc content of ≈ 2 per subunit and we are unable to reduce the zinc content of YADH by dialysis against EDTA without a concomitant loss in enzyme activity, in contrast to reports of one zinc per subunit [Veillon, C. and Sytkowski, A.J., BBRC $\text{67}$: 1499 (1975); Vallee, B.L. and Hoch, F.L., Proc. Nat. Acad. Sci. USA $\text{41}$: 327 (1955)]. The findings reported here, together with the observed structural similarities between YADH and horse liver alcohol dehydrogenase [Jornvall, H., Woenckhaus, C. and Johnscher, G., Eur. J. Biochem. $\text{53}$: 71 (1975)], suggest a role for zinc at both a structural and catalytic site in YADH.     

Cold-induced [Ca2+]cyt elevations function to support osmoregulation in marine diatoms

Diatoms are a group of microalgae that are important primary producers in a range of open ocean, freshwater, and intertidal environments. The latter can experience substantial long- and short-term variability in temperature, from seasonal variations to rapid temperature shifts caused by tidal immersion and emersion. As temperature is a major determinant in the distribution of diatom species, their temperature sensory and response mechanisms likely have important roles in their ecological success. We examined the mechanisms diatoms use to sense rapid changes in temperature, such as those experienced in the intertidal zone. We found that the diatoms Phaeodactylum tricornutum and Thalassiosira pseudonana exhibit a transient cytosolic Ca²⁺ ([Ca²⁺]_cyt) elevation in response to rapid cooling, similar to those observed in plant and animal cells. However, [Ca²⁺]_cyt elevations were not observed in response to rapid warming. The kinetics and magnitude of cold-induced [Ca²⁺]_cyt elevations corresponded with the rate of temperature decrease. We did not find a role for the [Ca²⁺]_cyt elevations in enhancing cold tolerance but showed that cold shock induces a Ca²⁺-dependent K⁺ efflux and reduces mortality of P. tricornutum during a simultaneous hypo-osmotic shock. As intertidal diatom species may routinely encounter simultaneous cold and hypo-osmotic shocks during tidal cycles, we propose that cold-induced Ca²⁺ signaling interacts with osmotic signaling pathways to aid in the regulation of cell volume. Our findings provide insight into the nature of temperature perception in diatoms and highlight that cross-talk between signaling pathways may play an important role in their cellular responses to multiple simultaneous stressors.

A calcium signaling pathway in marine diatoms is activated by cold temperature and enhances survival during simultaneous hypo-osmotic stress.     

Eudicot primary cell wall glucomannan is related in synthesis,structure, and function to xyloglucan

Hemicellulose polysaccharides influence assembly and properties of the plant primary cell wall (PCW), perhaps by interacting with cellulose to affect the deposition and bundling of cellulose fibrils. However, the functional differences between plant cell wall hemicelluloses such as glucomannan, xylan, and xyloglucan (XyG) remain unclear. As the most abundant hemicellulose, XyG is considered important in eudicot PCWs, but plants devoid of XyG show relatively mild phenotypes. We report here that a patterned β-galactoglucomannan (β-GGM) is widespread in eudicot PCWs and shows remarkable similarities to XyG. The sugar linkages forming the backbone and side chains of β-GGM are analogous to those that make up XyG, and moreover, these linkages are formed by glycosyltransferases from the same CAZy families. Solid-state nuclear magnetic resonance indicated that β-GGM shows low mobility in the cell wall, consistent with interaction with cellulose. Although Arabidopsis β-GGM synthesis mutants show no obvious growth defects, genetic crosses between β-GGM and XyG mutants produce exacerbated phenotypes compared with XyG mutants. These findings demonstrate a related role of these two similar but distinct classes of hemicelluloses in PCWs. This work opens avenues to study the roles of β-GGM and XyG in PCWs.

Patterned β-GGM resembles xyloglucan in structure, biosynthesis, and function.

In a Nutshell Background: Plant primary cell walls (PCWs) need to be rigid enough to define the plant shape and yet allow cell expansion at the same time. Plants achieve this by forming a complex network that is composed of cellulose and various non-cellulosic polysaccharides, such as hemicelluloses. Cell walls differ in the abundance of the various hemicelluloses, and their roles are poorly understood. In contrast to xyloglucan (XyG), which has been the most extensively studied hemicellulose in the PCWs, neither the structure nor functions of glucomannan has been resolved. Question: Are the functions of the glucomannan in PCWs distinct from the roles of the most abundant hemicellulose, XyG? Findings: We discovered a type of glucomannan in eudicot PCWs, which we named β-galactoglucomannan (β-GGM) because of its distinctive structures: disaccharide side chains of β-Gal-α-Gal and alternating repeats of Glc-Man in the backbone. Similarity to XyG in structure and biosynthesis led us to identify a β-galactosyltransferase for the β-GGM biosynthesis. We found that β-GGM contributed to normal cell expansion, in a way that was masked by the presence of XyG. These results suggest related functions of β-GGM to XyG, highlighting the necessity to consider the contribution of multiple hemicelluloses in the functional study of plant cell walls. Next steps: We would like to know how β-GGM binds to cellulose, and how this differs to cellulose binding of XyG. Investigation of the precise arrangements and interactions of cellulose and hemicelluloses including β-GGM and XyG will help further understanding of the enigmatic functions of hemicelluloses.