5.3 S RNA is a discrete cleavage product from the 5′-terminus of 18 S RNA of rat liver ribosomes

A 5.3 S RNA species observed in urea-gel electrophoretic analysis of the RNA of the small ribosomal subunit of rat liver has been identified from its sequence as the 5′-terminal 133–134 base fragment of 18 S RNA. Presumably it is cleaved by an endogenous endonuclease when the ribosomal subunits are dissociated, because it usually is not observed in 18 S RNA obtained by direct extraction of cells or tissues.     

Sulfation of mono- and diaryl oximes by aryl sulfotransferase isozymes

Aryl sulfotransferases (3'-phosphoadenylsulfate:phenol sulfotransferase, EC 2.8.2.1) catalyze the sulfonation of a wide variety of hydroxyl-containing substrates, including numerous xenobiotics. The chemical diversity of aryl sulfotransferase substrates is in part attributable to the presence of multiple isozymes, each of which has broad substrate specificity. Of the aryl sulfotransferase isozymes in rat liver cytosol, two (designated isozymes I and II) have previously been shown to sulfonate phenolic compounds exclusively and, moreover, have very similar substrate specificity patterns. The recently reported unusually efficient, rapid isozyme I-catalyzed sulfonation of 9-fluorenone oxime (Mangold, J.B., Mangold, B.L.K. and Spina, A. (1986) Biochim. Biophys. Acta 874, 37-43) was therefore unexpected and suggested that aryl oximes may represent a useful class of model compounds to probe isozymic differences in substrate steric and electronic requirements. In the present study, several mono- and diaryl oximes have been prepared and tested as potential substrates for partially purified aryl sulfotransferases I and II from rat liver cytosol. The results indicate that steric factors, specifically planarity and hydroxyl group position, appear to be important requirements for enzyme-catalyzed sulfonation. In addition, although isozymes I and II had comparable activity with diaryl oximes, some striking differences in the ability of these two isozymes to sulfonate both substituted and unsubstituted monoaryl oximes were observed. This dissimilarity is consistent with distinct differences in the active sites of these isozymes.     

Effects of chronic and acute exercise on cardiovascular beta-adrenergic responses.

beta-Adrenergic receptor density and responsiveness may be increased in experimental animals by physical conditioning, and the opposite effects have been observed after a single bout of exercise. To determine whether the chronic and acute effects of exercise include similar alterations in cardiovascular function in humans, we characterized heart rate, blood pressure, and distal lower extremity blood flow responses to graded-dose isoproterenol infusion in 15 young healthy subjects before and after exercise training and with and without a single preceding bout of prolonged exercise of either low or high intensity (61 +/- 1 or 82 +/- 1% maximal heart rate). VO2max was increased 18% after exercise training (43.2 +/- 2.7 to 51.1 +/- 3.3 ml.kg-1.min-1; P less than 0.001). Despite a concomitant fall in resting heart rate (59 +/- 3 to 50 +/- 2 beats/min; P less than 0.001), chronotropic and lower extremity blood flow responses to isoproterenol remained unchanged. Similarly, 1 h of acute high-intensity treadmill exercise altered baseline heart rate (58 +/- 4 to 74 +/- 5 beats/min; P less than 0.02), but neither low- nor high-intensity acute exercise influenced heart rate or lower extremity blood flow responses to isoproterenol. In contrast, the systolic pressure response to isoproterenol was blunted after high- but not low-intensity prolonged exercise (P less than 0.02). These data indicate that cardiac chronotropic (primarily beta 1) and vascular (beta 2) adrenergic agonist responses are not altered in humans by training or acute exercise. The systolic blood pressure response to beta-adrenergic stimulation is decreased by a single bout of high-intensity prolonged exercise by mechanisms that remain to be defined.     

Attenuation of cardiovascular adaptations to exercise in frail octogenarians.

To determine the mechanisms underlying increased aerobic power in response to exercise training in octogenarians, we studied mildly frail elderly men and women randomly assigned to an exercise group (n = 22) who participated in a training program of 6 mo of physical therapy, strength training, and walking followed by 3 mo of more intense endurance exercise at 78% of peak heart rate or a control sedentary group (n = 24). Peak O2 consumption (V(O2 peak)) increased 14% in the exercise group (P < 0.0001) but decreased slightly in controls. Training induced 14% increase (P = 0.027) in peak exercise cardiac output (Q), determined via acetylene re-breathing, and no change in arteriovenous O2 content difference. The increase in Q was mediated by increases in heart rate (P = 0.009) and probably stroke volume (P = 0.096). Left ventricular stroke work also increased significantly. In the men, the increase in V(O2 peak) was exclusively due to a large increase in peak Q (22%). In the women, the gain in V(O2 peak) was due to small increases in Q and O2 extraction from skeletal muscles. Pulse pressure normalized for stroke volume and arterial elastance during peak effort did not change with training. Controls showed no changes. The results suggest that, although frail octogenarians have a diminished capacity for improvement in aerobic power in response to exercise training, this adaptation is mediated mostly by an increase in Q during peak effort. Furthermore, Q likely plays a greater role in the adaptive increase in V(O2 peak) in old men than old women.     

PCR Detection of Oxytetracycline Resistance Genes otr(A) and otr(B) in Tetracycline-Resistant Streptomycete Isolates from Diverse Habitats

A range of European habitats was screened by PCR for detection of the oxytetracycline resistance genes otr(A) and otr(B), found in the oxytetracycline-producing strain Streptomyces rimosus. Primers were developed to detect these otr genes in tetracycline-resistant (Tc^R) streptomycete isolates from environmental samples. Samples were obtained from bulk and rhizosphere soil, manure, activated sludge and seawater. The majority of Tc^R streptomycetes originated from bulk and rhizosphere soil. Fewer Tc^R streptomycetes were isolated from manure and seawater and none from sewage. By PCR, three out of 217 isolates were shown to contain the otr(A) gene and 13 out of 217 the otr(B) gene. Surprisingly, these genes were detected in taxonomic groups not known as tetracycline-producing strains. The majority of the otr gene–carrying strains was assigned to S. exfoliatus or S. rochei and originated from all habitats from which Tc^R streptomycetes were obtained. Our results indicated that the occurrence of otr(A) and otr(B) genes in natural environments was limited and that otr(B), in comparison to otr(A), seemed to be more common.     

The tumor suppressor CDKN3 controls mitosis

Mitosis is controlled by a network of kinases and phosphatases. We screened a library of small interfering RNAs against a genome-wide set of phosphatases to comprehensively evaluate the role of human phosphatases in mitosis. We found four candidate spindle checkpoint phosphatases, including the tumor suppressor CDKN3. We show that CDKN3 is essential for normal mitosis and G1/S transition. We demonstrate that subcellular localization of CDKN3 changes throughout the cell cycle. We show that CDKN3 dephosphorylates threonine-161 of CDC2 during mitotic exit and we visualize CDC2^pThr-161 at kinetochores and centrosomes in early mitosis. We performed a phosphokinome-wide mass spectrometry screen to find effectors of the CDKN3-CDC2 signaling axis. We found that one of the identified downstream phosphotargets, CKβ phosphorylated at serine 209, localizes to mitotic centrosomes and controls the spindle checkpoint. Finally, we show that CDKN3 protein is down-regulated in brain tumors. Our findings indicate that CDKN3 controls mitosis through the CDC2 signaling axis. These results have implications for targeted anticancer therapeutics.     

A novel improved method for Aspergillus nidulans transformation

We systematically investigated the efficiency of Aspergillus nidulans transformation using protoplasts prepared from different stages of conidiospore germination and young mycelium. Using standard integrative plasmids, increased transformation yields were obtained with protoplasts isolated from a specific stage coincident with germ tube emergence. This increase ranged, on the average, from two- to eightfold depending on different plasmids used. Transformation efficiencies with a replicative plasmid were similar to those obtained using previously described methods. Although this observation suggests that elevated transformation efficiencies might be due to increased efficiency of recombination between plasmid and genomic sequences, we cannot exclude other factors associated with the particular developmental stage used. In the course of this study, we also examined the effect of other parameters that might enhance transformation yields. The method described is also significantly easier and faster than other current methods.     

Fluorescent diethylcarbamazine analogues: sites of accumulation in Brugia malayi

New fluorescein and rhodamine B-labeled antifilarial drug DEC analogues for use in drug localization studies with confocal microscopy have been prepared by a high-yield three-step synthesis. The resulting beta-amine-substituted DEC analogue has a single ethyl substituent which is beta-aminated to accommodate the fluorophore of either fluorescein isothiocyananate or rhodamine B. Confocal microscopy is used to show that the drug accumulates in the adult filarial worms in the pharynx, esophagus, and near the nerve ring of all adults, as well as in the uteri and vulva and the testes of the females and males.