Interaction of the release factors with the Escherichia coli ribosome: structurally and functionally-important domains.

There are two major domains of interaction between the Escherichia coli release factors (RF-1 and RF-2) and each subunit of the ribosome. RF-2 has a binding domain on the shoulder and lower head region of the small subunit at the small lobe distant from the decoding site. This is in close proximity to one of the domains on the large subunit which includes the body dimer of L7/L12 and L11. The other domains of interaction, at the decoding site on the small subunit, and at the peptidyltransferase centre of the large subunit of the ribosome, are some distance from the first two, although the evidence for direct contact with the ribosome is less comprehensive. The release factors may therefore have two distinct structural domains, and in support of this concept RF-1 and RF-2 can both be cleaved into two fragments by papain. Region-specific antibodies, and antibodies against defined peptide within the RF sequences have given an indication that a significant part of an interacting RF molecule is in close proximity to the ribosome surface, confirming an observation by immunoelectron microscopy which suggested that the RF penetrates deeply into the cleft between the two subunits. A region of highly conserved primary sequence between the two release factors from E coli is also conserved in those from B subtilis suggesting it forms an important structural or functional domain. Antibodies against peptides from the N-terminal end of this region strongly inhibit binding of the RF to the ribosome.     

Effect of nickel on oxygen free radical metabolism

The effect of nickel on superoxide dismutase activity (SOD), as well as on rate of hydroxydopamine oxidation, was studied in vitro since lipid peroxidation has been implicated in cell damage by nickel, whose toxicity and carcinogenicity are well established. Nickel strongly inhibits SOD activity. The degree of inhibition is directly proportion to the nickel concentration (tested range 0.066 to 0.33 microgram/mL in the reaction mixture); to the substrate concentration (tested range 0.4 x 10 4M to 1.1 x 10 4M 6-hydroxydopamine); and to reaction mixture. Autoxidation of 6-hydroxydopamine was increased by nickel concentrations higher than 15 micrograms/mL. The combination of excessive oxygen free radical production and inhibition of their elimination by inhibition of SOD activity may contribute to the nickel toxicity that has been reported in industrial accidents, as well as to the high incidence of cancer occurring in nickel workers. It may also contribute to many complications in uremic patients, in whom increased serum nickel levels were reported to be in a similar range to those inhibiting SOD.     

Syntheses, structures and anorectic effects of human and rat amylin.

Amylin, a 37-residue polypeptide with a single disulfide bond originally isolated from the pancreas of type-II diabetic patients, has been shown to cause peripheral insulin resistance and to attenuate the inhibition of hepatic glucose output by insulin. We have also shown that amylin is present in the rat hypothalamus and that it inhibits food intake by rats. In order to further investigate the anorectic properties we synthesized both human and rat amylin by the solid phase method and purified to homogeneity in an overall yield of 10-20%. Structural analyses indicated that human amylin exhibited predominantly a beta-sheet structure at both acidic and alkaline pH, whereas no ordered structure was evident in the case of rat amylin. Intrahypothalamic injection of rat amylin resulted in a potent dose-dependent inhibitory effect on the food intake by rats adapted to eat their daily ration of food in an eight-hour period. Human amylin was less effective as an anorectic agent. Furthermore, rat amylin completely blocked the potent orexigenic effect of neuropeptide Y (NPY). These investigations show that there is a fundamental difference in the secondary structures of human and rat amylin and that rat amylin is a potent inhibitor of both basal and NPY-induced feeding by rats.     

Interaction of soil moisture and elevated CO2 on the above-ground growth rate, root length density and gas exchange of turves from temperate pasture

Interactions between water availability and elevated atmosphericCO₂ concentrations have the potential to be important factorsin determining future forage supply from temperate pastures.Using large turves from an established pasture, the responseof these communities at 350 or 700 l l^–1 CO₂ to a soilmoisture deficit and to recovery from the deficit in comparisonto turves that were well-watered throughout was measured. Priorto this experiment the turves had been exposed to the CO₂ treatmentsfor 324 d. Net CO₂ exchange continued at elevated CO₂ even when the volumetricsoil moisture content was less than 0.10 m³ m^–3 soil;at the same moisture deficit gas exchange at ambient CO₂ waszero. The additional carbon fixed by the elevated CO₂ turveswas primarily allocated below-ground as shown by the maintenanceof root length density at the same level as in well-wateredturves. When the dry turves were rewatered there was compensatorygrowth at ambient CO₂ so that the above-ground growth rate exceededthat of turves that had not experienced a moisture deficit.At the start of this experiment, the turves that were growingat 700 l I^–1 CO₂ had a greater proportion of legume (principallywhite clover, Trifolium repens L.) in the harvested herbage.There was a trend for the legume content at elevated CO₂ tobe reduced under a soil moisture deficit. The results indicate different strategies in response to soilmoisture deficits depending on the CO₂ concentration. At ambientCO₂, growth stopped, but plants were able to respond stronglyon rewatering; while at elevated CO₂ growth continued (particularlybelow-ground), but no additional growth was evident on rewatering.Ecosystem gas exchange measurements taken at the end of theexperiment (after 429 d of exposure to CO₂) showed 33% moreCO₂ was fixed at elevated CO₂ with only a small (12%) and nonsignificantdownward regulation. Key words: Carbon dioxide, climate change, grassland, gas exchange, soil moisture deficit     

Alterations of auxin perception in rolB-transformed tobacco protoplasts. Time course of rolB mRNA expression and increase in auxin sensitivity reveal multiple control by auxin.

Expression and physiological effects of the root-inducing rolB gene of Agrobacterium rhizogenes T-DNA were studied simultaneously in tobacco (Nicotiana tabacum) mesophyll protoplasts. The kinetic study of the expression of rolB mRNA following exogenous auxin application showed that auxin transiently stimulated rolB expression, with mRNA levels starting to accumulate 6 to 9 h after auxin was supplied and increasing 300-fold after 12 to 18 h. The parallel study of the auxin sensitivity of rolB-transformed protoplasts, as assayed by their electrical response to the hormone, showed that the auxin treatment generated an increase in sensitivity by a factor of up to 100,000, whereas in untransformed protoplasts the same auxin treatment induced an increase in auxin sensitivity that never exceeded 30- to 50-fold. This reflects a strong cooperative effect of auxin and rolB in transformed protoplasts. Surprisingly, the maximal increase in sensitivity was observed several hours before the maximal accumulation of rolB mRNA, suggesting that the dramatic control of auxin sensitivity by auxin in rolB-transformed protoplasts requires only low levels of rolB expression. Antibodies directed against ZmER-abp1, the major auxin-binding protein from maize, differentially altered the auxin sensitivity of the electrical response of rolB-transformed and normal protoplasts. This suggests that alterations of the auxin reception-transduction pathway at the plasma membrane of rolB-transformed protoplasts may account for their increased auxin sensitivity.     

Linkage of bovine erythrocyte antigen loci B, C, L, S, Z, R' and T' and the serum protein loci post-transferrin 2 (PTF 2), vitamin D binding protein (GC) and albumin (ALB) to DNA microsatellite markers

Seven bovine erythrocyte antigen loci and three serum protein loci were tentatively assigned to chromosomes or synteny groups by linkage analysis to previously assigned microsatellite DNA markers. The erythrocyte antigen locus EAB was mapped to synteny group U27; EAC to chromosome 18, synteny group U9; EAL to chromosome 3, synteny group U6; EAS to chromosome 21, synteny group U4; EAZ to chromosome 10, synteny group U5; EAR' to chromosome 16, synteny group U1; and EAT' to chromosome 19, synteny group U21. The vitamin D binding protein (GC) and albumin (ALB) loci were assigned to chromosome 6, synteny group U15 and post-transferrin 2 (PTF 2) to chromosome 19, synteny group U21.     

Integrating the porcine physical and linkage map using cosmid-derived markers

An essential part in the development of informative linkage maps is to include genetic markers that have been anchored by physical mapping. Here a set of 18 porcine cosmid-derived genetic markers are reported that have been mapped by linkge analysis, and that also have been physically localized by fluorescence in situ hybridization (FISH). Three different strategies were used to establish polymorphic markers from the cosmid clones. Firstly, dinucleotide microsatellite loci were derived by sequencing cosmid subclones containing (CA), repeats. Secondly, variable SINE 3′ poly(A) tracts (SINEVA) were identified by direct SINE-PCR amplification of cosmid clones. Thirdly, the cosmids were used in Southern blot hybridization to detect restriction fragment length polymorphisms (RFLPs). Compared with the most recent consensus compilation of the porcine gene map, the present assignment of markers to chromosomes Zp, 3, 4, 10, 12q, and 16 represents the first loci mapped to these chromosomes, for which linkage as well as in situ data are now available.     

Regulated food recruitment through individual behavior of scouts in the ant,Myrmica sabuleti (Hymenoptera: Formicidae)

By presenting different kinds of food sources to colonies ofM. sabuleti, we have demonstrated that this species regulates its foraging activity by altering the proportion of scouts that return to the nest to recruit nestmates after discovering a food source and by varying the number of nestmates recruited by a scout. These two parameters are related to the kind of food discovered. Our behavioral experiments showed that the probability that a scout returned to the nest decreased with a decreasing quantity of sucrose solution. In contrast, the number of returned scouts that elicited recruitment from the nest and the mean number of nestmates recruited by one of these scouts were independent of the quantity of the sucrose solution. Recruitment even occurred toward a 1- or 0.25-µl droplet of sucrose solution. When a scout discovered a large dead prey, a large drop of prey juice, a cluster of 30 dead fruit flies, or 1 isolated fruit fly, it always went back to the nest, but it elicited recruitment only when the food source was a large dead prey or a large drop of prey juice. No recruitment occurred when the food source was a single fruit fly and recruitment occurred only once in 30 trials when a cluster of 30 fruit flies was discovered.