Mutant ATP-binding RNA aptamers reveal the structural basis for ligand binding

A nuclear mutant of maize, tha1, which exhibited defects in the translocation of proteins across the thylakoid membrane, was described previously. A transposon insertion at the tha1 locus facilitated the cloning of portions of the tha1 gene. Strong sequence similarity with secA genes from bacteria, pea and spinach indicates that tha1 encodes a SecA homologue (cp-SecA). The tha1-ref allele is either null or nearly so, in that tha1 mRNA is undetectable in mutant leaves and cp-SecA accumulation is reduced > or = 40-fold. These results, in conjunction with the mutant phenotype described previously, demonstrate that cp-SecA functions in vivo to facilitate the translocation of OEC33, PSI-F and plastocyanin but does not function in the translocation of OEC23 and OEC16. Our results confirm predictions for cp-SecA function made from the results of in vitro experiments and establish several new functions for cp-SecA, including roles in the targeting of a chloroplast-encoded protein, cytochrome f, and in protein targeting in the etioplast, a nonphotosynthetic plastid type. Our finding that the accumulation of properly targeted plastocyanin and cytochrome f in tha1-ref thylakoid membranes is reduced only a few-fold despite the near or complete absence of cp-SecA suggests that cp-SecA facilitates but is not essential in vivo for their translocation across the membrane.     

Changes in renal phosphate reabsorption in the aged rat

PURPOSE: We tested whether exudative ARM was associated with low whole blood levels of selenium (Se). METHODS: Blood samples, drawn from 10 exudative ARM patients (61.2-76.1 yr) and 9 healthy-eyed (66.9-75.1 yr) subjects, were analyzed by atomic absorption spectroscopy. RESULTS: Selenium concentration was significantly lower in the ARM group (186.6 microg/l) than in controls (207.0 microg/l). Because many ARM patients took Se supplements, we tested the effect on blood Se of 80 microg per day of sodium selenate. We found no enduring effects of supplementation for healthy-eyed, younger adults. CONCLUSIONS: Significant group differences in this preliminary study indicate a larger-scale study of blood Se concentration in exudative ARM patients is warranted. If the effect of Se supplementation on the progression of exudative ARM is tested in future trials, it will be important to use organic Se, to identify the components of blood affected, and to observe protocol for at least six months.     

Cine film replacement: digital archival requirements and remaining obstacles

The acceptance of the Digital Imaging and Communication in Medicine (DICOM) standard and the Compact Disk-Recordable (CD-R) as the interchange medium have been critical developments for laboratories that need to move forward on the cine replacement front, while at the same time retain a means to communicate with other centers. One remaining essential component which has not been satisfactorily addressed is the issue of how digital image data should be archived within an institution. Every laboratory must consider the diverse issues which affect the choice of a digital archiving system. These factors include technical and economic issues, along with the clinical routines prevailing in their laboratory. A complete understanding of the issues will lead to the formulation of multiple options which may prove acceptable and will help to overcome the last obstacle which remains for the complete replacement of cine film in the cardiac catheterization laboratory.     

Clavaric acid and steroidal analogues as Ras- and FPP-directed inhibitors of human farnesyl-protein transferase

We have identified a novel fungal metabolite that is an inhibitor of human farnesyl-protein transferase (FPTase) by randomly screening natural product extracts using a high-throughput biochemical assay. Clavaric acid [24, 25-dihydroxy-2-(3-hydroxy-3-methylglutaryl)lanostan-3-one] was isolated from Clavariadelphus truncatus; it specifically inhibits human FPTase (IC50 = 1.3 microM) and does not inhibit geranylgeranyl-protein transferase-I (GGPTase-I) or squalene synthase activity. It is competitive with respect to Ras and is a reversible inhibitor of FPTase. An alkaline hydrolysis product of clavaric acid, clavarinone [2,24,25-trihydroxylanostan-3-one], lacking the 3-hydroxy-3-methylglutaric acid side chain is less active as a FPTase inhibitor. Similarly, a methyl ester derivative of clavaric acid is also inactive. In Rat1 ras-transformed cells clavaric acid and lovastatin inhibited Ras processing without being overtly cytotoxic. Excess mevalonate reversed the effects of lovastatin but not of clavaric acid suggesting that the block on Ras processing by clavaric acid was due to inhibition of FPTase and not due to inhibition of HMG-CoA reductase. Despite these results, the possibility existed that clavaric acid inhibited Ras processing by directly inhibiting HMG-CoA reductase. To directly examine the effects of clavaric acid and clavarinone on HMG-CoA reductase, cholesterol synthesis was measured in HepG2 cells. No inhibition of HMG-CoA reductase was observed indicating that the inhibition of Ras processing by this class of compounds is due to inhibition of FPTase. To date, clavaric acid is the second reported nitrogen-free compound that competes with Ras to inhibit FPTase activity. A series of related compounds derived from computer-based similarity searches and subsequent rational chemical synthetic design provided compounds that exhibited a range of activity (0.04 --> 100 microM) against FPTase. Modest changes in the structures of these inhibitors dramatically change the inhibitory activity of these inhibitors.     

Ultraviolet B irradiation modulates the immune system of fish (Rutilus rutilus, Cyprinidae). I. Phagocytes

When western blotting analysis of Japanese eel neutrophil lysate was performed using antibody against the synthetic peptide corresponding to the carboxyl terminal region of human cytochrome b558 large subunit, a broad band was specifically detected at approximately 90 kDa. The antibody recognized the epitope present in eel neutrophils only after the cells were permeabilized with detergent. These results indicate that the large subunit of cytochrome b exists in fish neutrophils and the epitope is exposed to the cytoplasmic side of membrane, and suggest that cytochrome b is conserved across species.     

Diagnosing coronary arterial stent thrombosis and arterial closure

This single-center review of a consecutive series of patients requiring reexamination by angiography within 1 week of a coronary stent placement due to chest pain reveals that patients treated with a poststent anticoagulation regimen of warfarin and aspirin, and those with lower poststent deployment dilation pressures, have an increased risk of subacute stent thrombosis. Repeat cardiac catheterization within the first week after coronary artery stent implantation should be reserved for patients with significant electrocardiographic changes.     

A complex program of striatal gene expression induced by dopaminergic stimulation

Dopamine acting in the striatum is necessary for normal movement and motivation. Drugs that change striatal dopamine neurotransmission can have long-term effects on striatal physiology and behavior; these effects are thought to involve alterations in gene expression. Using the 6-hydroxydopamine lesion model of Parkinson's disease and differential display PCR, we have identified a set of more than 30 genes whose expression rapidly increases in response to stimulation of striatal dopamine D1 receptors. The induced mRNAs include both novel and previously described genes, with diverse time courses of expression. Some genes are expressed at near-maximal levels within 30 min, whereas others show no substantial induction until 2 hr or more after stimulation. Some of the induced genes, such as CREM, CHOP, and MAP kinase phosphatase-1, may be components of a homeostatic response to excessive stimulation. Others may be part of a genetic program involved in cellular and synaptic plasticity. A very similar set of genes is induced in unlesioned animals by administration of the psychostimulant cocaine or the antipsychotic eticlopride, although in distinct striatal cell populations. In contrast to some previously described early genes, most of the novel genes are not induced in cortex by apomorphine, indicating specificity of induction. Thus we have identified novel components of a complex, coordinated genetic program that is induced in striatal cells in response to various dopaminergic manipulations.     

Myxobolus cerebralis in Tubifex tubifex from a whirling disease epizootic in Montana

Prolonged suppressive antibiotic therapy may be an alternative to removal of infected orthopedic prostheses in some patients. However, the efficacy of prolonged suppressive antibiotics is not well established. We retrospectively reviewed 18 patients with infected orthopedic prostheses who had been treated with prolonged antimicrobial suppression during the last 10 years. Eighteen episodes of infection were identified in these 18 patients. There were nine men and nine women, and the mean age was 66 years (range, 31-83 years). All patients had a functional prosthesis and were treated with surgical debridement, retention of the prosthesis, and administration of intravenous antibiotics for 6-8 weeks, followed by prolonged oral antibiotic suppression. Fifteen of the 18 patients appear to have had a good response and have been able to retain a functional prosthesis. Complications related to antibiotic suppression occurred in 22% but did not necessitate discontinuation of the antibiotic therapy. Prolonged antibiotic suppression is a reasonable alternative to surgery in selected patients with infected orthopedic prostheses.