Stress resistance in Saccharomyces cerevisiae is strongly correlated with assembly of a novel type of multiubiquitin chain.

The covalent attachment of ubiquitin (Ub) to short-lived or damaged proteins is believed to be the signal that initiates their selective degradation. In several cases, it has been shown that the proteolytic signal takes the form of a multi-Ub chain in which successive Ub molecules are linked tandemly at lysine 48 (K-48). Here we show that Ub molecules can be linked together in vivo at two other lysine positions, lysine 29 (K-29) and lysine 63 (K-63). The formation of these alternative linkages is strongly dependent on the presence of the stress-related Ub conjugating enzymes UBC4 and UBC5. Furthermore, expression of Ub carrying a K-63 to arginine 63 substitution in a strain of Saccharomyces cerevisiae that is missing the poly-Ub gene, UBI4, fails to compensate for the stress defects associated with these cells. Taken together, these results suggest that the formation of multi-Ub chains involving K-63 linkages plays an important role in the yeast stress response. In broader terms, these results also suggest that Ub is a versatile signal in which different Ub chain configurations are used for different functions.     

Regulation of μ-Opioid Receptor mRNA in Rat Globus Pallidus: Effects of Enkephalin Increases Induced by Short- and Long-Term Haloperidol Administration

Abstract: The mRNA encoding μ-opioid receptors is expressed in neurons of the globus pallidus, a region of the basal ganglia that receives a dense enkephalinergic innervation from the striatum. The regulation of the mRNAs encoding the opioid peptide enkephalin in the striatum and the μ-opioid receptor in the globus pallidus was examined with in situ hybridization histochemistry following short- or long-term haloperidol treatments, which alter striatal enkephalin mRNA levels. Animals were administered haloperidol daily for 3 or 7 days (1 mg/kg, s.c.) or continuously for 8 months (1 mg/kg, depot followed by oral). Enkephalin and μ-opioid receptor mRNA levels were unchanged after 3 days of haloperidol treatment. In contrast, the enkephalin mRNA level was increased in the striatum, and μ-opioid receptor mRNA levels were markedly decreased in the globus pallidus after 7 days of haloperidol administration. Similar effects were observed in rats treated with haloperidol for 8 months. The results provide the first evidence of regulation of μ-opioid receptor mRNA in vivo.     

Influences of sea level changes and volcanic eruptions on Holocene vegetation in Tonga

Here, we investigate Mid- to Late-Holocene vegetation changes in low-lying coastal areas in Tonga and how changing sea levels and recurrent volcanic eruptions have influenced vegetation dynamics on four islands of the Tongan archipelago (South Pacific). To investigate past vegetation and environmental change at Ngofe Marsh (‘Uta Vava’u), we examined palynomorphs (pollen and spores), charcoal (fire), and sediment characteristics (volcanic activity) from a 6.7-m-long sediment core. Radiocarbon dating indicated the sediments were deposited over the last 7700 years. We integrated the Ngofe Marsh data with similar previously published data from Avai’o’vuna Swamp on Pangaimotu Island, Lotofoa Swamp on Foa Island, and Finemui Swamp on Ha’afeva Island. Plant taxa were categorized as littoral, mangrove, rainforest, successional/ disturbance, and wetland groups, and linear models were used to examine relationships between vegetation, relative sea level change, and volcanic eruptions (tephra). We found that relative sea level change has impacted vegetation on three of the four islands investigated. Volcanic eruptions were not identified as a driver of vegetation change. Rainforest decline does not appear to be driven by sea level changes or volcanic eruptions. From all sites analyzed, vegetation at Finemui Swamp was most sensitive to changes in relative sea level. While vegetation on low-lying Pacific islands is sensitive to changing sea levels, island characteristics, such as area and elevation, are also likely to be important factors that mediate specific island responses to drivers of change.     

COMPARISON OF PLANT DNA CONTENTS DETERMINED BY FEULGEN MICROSPECTROPHOTOMETRY AND LASER FLOW CYTOMETRY

An improved procedure is reported for determining DNA amounts of plant nuclei. Nuclei stained with propidium iodide, isolated from chopped plant leaves, were passed through an Ortho Cytofluorograph with a Lexel model 95 argon laser (514 nm) and the fluorescence measured, integrated, and recorded using an Ortho 2140 Data Acquisition computer. All nuclear samples were mixed with nuclei of Sultan barley (2C DNA content = 11.12 pg [picogram]) as an internal standard. DNA contents of ten plant species, ranging from 2C = 1.7 pg to 36.1 pg measured by flow cytometry, correlated strongly (r = 0.99, slope = + 1.00) with DNA contents determined from Feulgen-stained nuclei of the same species using microspectrophotometry. The flow cytometric procedures were sufficiently sensitive to detect differences in DNA content between inbred lines of corn and their F1 hybrids. Our results obtained with improved procedures, specifically using propidium iodide as a fluorochrome and plant nuclei instead of chicken erythrocytes as an internal standard, demonstrate that laser flow cytometry can be a precise, rapid, and reliable method for determining nuclear DNA content of plants.     

Limited proteolysis of native proteins: the interaction between avidin and proteinase K.

Avidin is a tetramer of 16-kDa subunits that have a high affinity for biotin. Proteolysis of native apoavidin by proteinase K results in a limited attack at the loop between beta-strands 3 and 4, involving amino acids 38-43. Specifically, sites of proteolysis are at Thr 40-Ser 41 and Asn 42-Glu 43. The limited proteolysis results in an avidin product that remains otherwise intact and which has enhanced binding for 4'-hydroxyazobenzene-2-benzoic acid (HABA), a chromogenic reporter that can occupy the biotin-binding site. Saturation of the biotin-binding site with the natural ligand protects avidin from proteolysis, but saturation with HABA enhances the rate of proteolysis of the same site. Analysis of the three-dimensional structures of apoavidin and holoavidin reveals that the 3-4 loop is accessible to solvent and scores highly in an algorithm developed to identify sites of proteolytic attack. The structure of holoavidin is almost identical to the apoprotein. In particular, the 3-4 loop has the same structure in the apo and holo forms, yet there are marked differences in proteolytic susceptibility of this region. Evidence suggests that the 3-4 loop is rather mobile and flexible in the apoprotein, and that it becomes constrained upon ligand binding. In one crystal structure of the apoprotein, this loop appears constrained by contacts with symmetry-related molecules. Structural analyses suggest that the "lid" to the biotin-binding site, formed by the 3-4 loop, is displaced and made more accessible by HABA binding, thereby enhancing its proteolytic susceptibility.     

The late stage following continuous amphetamine administration to rats is correlated with altered dopamine but not serotonin metabolism

In previous experiments rats pretreated with slow-release d-amphetamine (d-Amp) pellets for $\text{4}\text{1}\text{2}$ days, given a 12-hr drug-free period, and then injected with d-Amp have been found to show a behavioral syndrome which has similarities to that induced by acute injections of the hallucinogens LSD and mescaline. The present results indicate that rats administered this same drug regimen have large decreases in Dopamine (DA), dihydroxyphenyl acetic acid (Dopac), and homovanillic acid (HVA) in caudate nucleus, smaller decreases in DA with no changes in Dopac and HVA levels in nucleus accumbens, but no alterations in 5-hydroxytryptamine (5HT) and 5-hydroxyindole acetic acid (5HIAA) levels in caudate, accumbens, brainstem and hippocampus. Increased 5HIAA levels are found in rats sacrificed with pellets intact following 3 days of continuous d-Amp administration, while sleep deprived and in motor stereotypies. The late and hallucinatory stage following continuous d-amp is correlated more closely with alterations in dopamine than of 5HT.     

Purification and characterization of monkey salivary mucin.

Highly purified mucin was prepared from monkey (Macaca arctoides) extraparotid saliva by sequential chromatography on Sephadex G-200 (followed by reduction and alkylation of void volume materials), Sepharose CL-2B with 6 M urea, and CM52 cellulose with 6 M urea. Purity was critically ascertained by anion exchange chromatography, ultracentrifugal analysis, isoelectric focusing, sodium dodecyl sulfate-polyacrylamide electrophoresis, and crossed immunoelectrophoresis. Use of crossed immunoelectrophoresis to examine mucin preparations has not been previously reported. This technique was useful for assessing purity and displaying charge and size microheterogeneity in the purified S-carboxymethylated mucin. Threonine and serine comprised 37.8% of the total amino acids while the oligosaccharide moiety contained N-acetyl-glucosamine, N-acetylgalactosamine, fucose, galactose, N-acetylneuraminic acid, and sulfate. Following alkaline borohydride treatment, the carbohydrate chains were found to be linked O-glycosidically between N-acetylgalactosamine and threonine (serine).     

Sleeping blood,tremor and paralysis: A trans-cultural approach to an unusual conversion reaction

An unusual conversion reaction and a related folk belief system are described in a Cape Verdean-American woman. A clinical approach is presented which stresses the need to negotiate between professional and popular explanatory models of illness. It is suggested that a significant factor in the successful management of this patient's symptoms was the acceptance of a mutually-understandable idiom for communication and treatment.     

Silver Staining for Nucleolar Organizing Regions of Vertebrate Chromosomes

Refinements to a simple, one-step silver staining technique for nucleolar organizing regions are described. These include fixation of silver stained material with sodium thiosulfate and standardization of silver development conditions for different groups of vertebrates. The central advantages to the method are that it is rapid, reliable, simple, and inexpensive. Additional benefits include (i) consistent and uniform silver staining of nucleolar organizing regions, (ii) few reduced silver deposits elsewhere on the chromosomes or on the slides, (iii) generally unaltered chromosome morphology after silver treatment, and (iv) relative permanence of Permounted preparations. The method works equally well on chromosomes made from cell cultures and from solid tissues of live specimens.     

Normative study of age variation in salivary progesterone profiles.

Daily luteal progesterone levels were measured for 124 regularly menstruating women, aged 18-44, by radioimmunoassay of steroid levels in saliva. A consistent pattern of age variation in luteal function was found, with the lowest levels of progesterone in the 18-19-year-old and 40-44-year-old groups, the highest values in the 25-34 year olds, and intermediate values in the 20-24 and 35-39-year-old groups. The striking similarity of this pattern of age variation with empirically and theoretically generated curves of apparent fecundability, suggests that age-related changes in ovarian function may underlie observed age variation in fecundability.