Methyltransferase Contingencies in the Pathway of Everninomicin D Antibiotics and Analogues

Everninomicins are orthoester oligosaccharide antibiotics with potent activity against multidrug-resistant bacterial pathogens. Everninomicins act by disrupting ribosomal assembly in a distinct region in comparison to clinically prescribed drugs. We employed microporous intergeneric conjugation with Escherichia coli to manipulate Micromonospora for targeted gene-replacement studies of multiple putative methyltransferases across the octasaccharide scaffold of everninomicin effecting the A₁, C, F, and H rings. Analyses of gene-replacement and genetic complementation mutants established the mutability of the everninomicin scaffold through the generation of 12 previously unreported analogues and, together with previous results, permitted assignment of the ten methyltransferases required for everninomicin biosynthesis. The in vitro activity of A₁- and H-ring-modifying methyltransferases demonstrated the ability to catalyze late-stage modification of the scaffold on an A₁-ring phenol and H-ring C-4’ hydroxy moiety. Together these results establish the potential of the everninomicin scaffold for modification through mutagenesis and in vitro modification of advanced biosynthetic intermediates.     

A simple handling technique for mammalian oocytes and embryos during preparation for transmission electron microscopy

Due to their small size, mammalian oocytes and embryos pose unique problems during preparation for transmission electron microscopy. This paper outlines a method which combines protein embedding with centrifugation to locate the specimens on the face of a Beem capsule mould. This method facilitates both the processing of oocytes with minimal loss and rapid location of the specimens within the block for simultaneous sectioning, staining and examination.     

Instructional texts rewritten by five expert teams: Revisions and retention improvements.

700 undergraduates were tested in 3 experiments on original or rewritten versions of 52 instructional texts about Army job tasks, general science, philosophy, and history. Five experts had rewritten various sets of the texts and stated hypotheses about the efficacious features of their revisions. We tested their hypotheses and several others. Recall and recognition tests were given immediately and after a 24-hr delay. Results showed that revisions made by 3 of the 5 experts improved retention of text information. The kind and number of revisions and improvements varied across the text sets. Most expert hypotheses were not supported, and they made many revisions they were declaratively unaware of. Some of our hypotheses about the revision features were supported, but different features were effective for different sets of texts. It was concluded that some experts have effective knowledge about improving instructional text, but it exists primarily in procedural form. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Relationships between urbanization and the oak resource of the Minneapolis/St. Paul Metropolitan area from 1991 to 1998

Urbanization was associated with loss and transformation of the oak forest in the Twin Cities (Minneapolis and St. Paul) metropolitan area (TCMA) over a recent 7-year interval. Between 1991 and 1998, urbanization increased based on several indicators: population density, area of developed land, and area of impervious surface—total impervious area and area within three classes of increasing degree of imperviousness (protected, affected, and degraded). We quantified relationships between changes in urbanization and changes in several parameters describing the oak forest at the scale of ecological subsection. Increased total and affected impervious area were strongly correlated with decreased area of oak forest when changes of the urbanization indicators and oak were expressed as percentages of the subsection area. Relationships were reversed when changes were expressed as percentages of the 1991 values. Increased population density was strongly correlated with increased loss in numbers and increased isolation of oak patches, but weakly correlated with loss of oak forest area. This is the first study to quantify relationships between changes in urbanization and changes in a specific forest cover type. Our results demonstrate complexities of urbanization impacts on a metropolitan forest resource, and highlight the importance of selected variables, spatial and temporal scales, and expressions of change when quantifying these relationships.     

The ethics of asking about abuse and the harm of "don't ask, don't tell."

Replies to comments by J. Read (see record 2007-07130-012), D. Gleaves et al (see record 2007-07130-013), V. Edwards et al (see record 2007-07130-014), M. Black and R. Black (see record 2007-07130-015), and S. Ullman (see record 2007-07130-016), which raised important points about the authors' original article (see record 2006-03947-003). Those comments extend our thinking about how to ask participants about abuse in an ethical way. Together, the comments point to the importance of researchers examining our own reasons for asking--or not asking--about abuse and of paying attention to how we respond when we ask. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Metastatic parathyroid carcinoma in the MEN2A syndrome

We report a patient with a metastatic parathyroid carcinoma and medullary carcinoma of the thyroid. This patient represents a variation of the multiple endocrine neoplasia syndrome (MEN) type 2A. There was no evidence of a phaeochromocytoma. The case illustrates the difficulties that may be encountered in localising the source of PTH secretion; the patient underwent four unsuccessful exploratory operations of the neck and mediastinum before further investigations revealed a single metastatic deposit of parathyroid carcinoma involving the first thoracic vertebra. PCR amplification and sequencing of the RET oncogene from the metastatic parathyroid carcinoma and genomic DNA revealed a heterozygous mutation (Cys634Tyr) in exon 11, as has previously been described to occur in MEN 2A. In addition, loss of tumour heterozygosity was demonstrated at loci from chromosomes 1, 2, 3p, 13q and 16p. This represents the first report of a parathyroid carcinoma in a MEN2A patient, in which the multiple allelic deletions are consistent with the generalised losses observed in aggressive tumours.     

Effects of anxiety arousal and mental stress on the vestibulo-ocular reflex

Although the subjective reports of patients suggest that anxiety may aggravate vertigo and imbalance, there has been little research into how anxiety might directly affect balance system functioning. We conducted two studies to examine the effect of anxiety and arousal on the vestibulo-ocular reflex (VOR). In the first study, pre-lest fear ratings were obtained from 20 normal subjects and 36 anxious subjects immediately prior to rotation and caloric testing. Fear ratings were significantly correlated with the maximum slow-phase velocity (SPV) of nystagmus induced by caloric testing. In the second study, we assessed the VOR response to rotation of 36 normal subjects under 3 task conditions: a) minimal alerting (counting backwards during rotation), b) physical arousal (induced by exertion prior to rotation); c) mental arousal (induced by performance of stressful mental tasks during rotation). Both the physical and mental tasks induced a significant increase in heart rate compared with the alerting condition. The maximum SPV of the nystagmus induced by rotation was significantly greater during performance of the mental task than in the other two conditions. These combined results indicate that anxiety may influence the gain of the VOR.     

Chlorisondamine inhibits the nicotine-induced stimulation of c-fos in the pigeon brain for up to 2 weeks.

Chlorisondamine is a charged molecule that acts as long-acting nicotinic antagonist in many species, including pigeon. Evidence indicates that, despite the charged nature of chlorisondamine, it blocks some central effects of nicotine. The present study examined the time course of chlorisondamine's blockade of nicotine-induced c-fos expression in the pigeon brain. Chlorisondamine's central blockade was examined from 1 hr to 28 days prior to nicotine administration. Nicotine stimulated increases in c-fos mRNA in the hippocampus, hyperstriatum accessorium, hyperstriatum ventrale, nucleus accumbens, bulbus olfactorius, paleostriatum augmentatum, and stratum griseum et fibrosum superficiale. Nicotinic receptors labeled by [(125)I]-epibatidine were not always found in the same regions as nicotine-induced increases in c-fos expression. Acute chlorisondamine increased the level of c-fos mRNA in the cerebellum, hippocampus, hyperstriatum accessorium, locus parolfactorius, nucleus accumbens, tectum opticum, paleostriatum augmentatum, and stratum griseum et fibrosum superficiale but had no effect on its own 24 hr after administration. Chlorisondamine blocked nicotine-induced increases in c-fos RNA for 4 days in the nucleus accumbens, a week in the bulbus olfactorius, and 2 weeks in the stratum griseum et fibrosum superficiale. The time course of chlorisondamine's blockade of nicotine-induced c-fos expression is consistent with the time course of the ability of chlorisondamine to block behavioral and physiological responses to nicotine.