DNA sequence,structure, and phylogenetic relationship of the small subunit rRNA coding region of mitochondrial DNA fromPodospora anserina

Summary DNA sequence analysis and the localization of the 5 and 3 termini by S1 mapping have shown that the mitochondrial (mt) small subunit rRNA coding region fromPodospora anserina is 1980 bp in length. The analogous coding region for mt rRNA is 1962 bp in maize, 1686 bp inSaccharomyces cerevisiae, and 956 bp in mammals, whereas its counterpart inEscherichia coli is 1542 bp. TheP. anserina mt 16S-like rRNA is 400 bases longer than that fromE. coli, but can be folded into a similar secondary structure. The additional bases appear to be clustered at specific locations, including extensions at the 5 and 3 termini. Comparison with secondary structure diagrams of 16S-like RNAs from several organisms allowed us to specify highly conserved and variable regions of this gene. Phylogenetic tree construction indicated that this gene is grouped with other mitochondrial genes, but most closely, as expected, with the fungal mitochondrial genes.     

Lipid-cell interactions: Liposome adsorption and cell-to-liposome lipid transfer are mediated by the same cell-surface sites

The competitive behavior of solid vs. fluid liposomes in liposome-to-cell adsorption and cell-to-liposome lipid transfer processes was investigated with L cells and FBT epithelial sheets. Binding, transfer and ³¹P-NMR experiments have demonstrated that: (i) solid liposomes adhere to the cell surface as integral vesicles retaining the entrapped substances; (ii) fluid liposomes are partly disintegrated at the cell surface with concomitant entry of entrapped substances into the cytoplasm, while their lipids remain on the cell surface; (iii) fluid liposomes that escape lysis dissociate from the cell, taking away cell lipid molecules. The latter process underlies the mechanism of cell-to-fluid liposome lipid transfer. In contrast, no lipid transfer occurs between the plasma membrane and solid liposomes. Cell-bound solid liposomes interfere with the transfer of cell lipids to fluid liposomes, while these in turn inhibit the binding of solid liposomes to the cell surface. Moreover, cell-induced aggregation of both fluid and solid freshly added liposomes is also inhibited by preincubation of the cells with either solid or fluid liposomes. Thus, different types of interaction of both fluid and solid liposomes with the cell are mediated by the same (or closely related) sites on the cell surface.     

A mouse DNA sequence that mediates integration and excision of polyoma virus DNA

In mouse cells transformed by a temperature-sensitive polyoma virus (Py) genome, the integrated viral genome recombines with adjacent chromosomal DNA to yield a small cyclic molecule (RmI) with defined viral and cellular components. We have cloned the cellular component (Ins), determined its sequence, and examined its distribution in normal mouse DNA. The sequence of Ins displays several homologies with that surrounding the replication origin (ori) of Py or SV40 DNA.     

Small scale preparation of skeletal muscle mitochondria, criteria of integrity, and assays with reference to tissue function

Mitochondria prepared in small scale from skeletal muscle were studied with respiration measurements and low temperature spectroscopy. The method of preparation was developed for 25–100 mg tissue with pigeon breast muscle as model organ. The yield was 40%. Data collected during the developmental work were used to evaluate criteria of mitochondrial quality. The cytochrome c conservation, i.e. cytochrome c per mitochondrial quantity in the preparation relative to that in the tissue, is a most useful test parameter. It is bounded between 0–100%. Proportionality between the state 3 rate and the cytochrome c conservation was not rejected by statistical tests. The respiratory control ratio (RCR) was also highly correlated to the cytochrome c conservation. These correlations might be extrapolated to 100% conservation to give hypothetical tissue values. The cause for the correlations is discussed. The P/O ratio showed only weak dependence on the cytochrome c conservation and the state 4 rate s howed no dependence. Other, rather insensitive test parameters are also discussed. The pigeon breast muscle mitochondria isolated by the final method showed cytochrome c conservation of 73 ± 9% (n = 16). They are compared with pig biceps femoris mitochondria prepared by the same method. The two types of mitochondria show many similarities. Some differences may be explained by a different amount of inner mitochondrial membrane relative to mitochondrial protein. The pig tissue contains ten times less mitochondrial protein than the pigeon tissue does. (Mol Cell Biochem 174: 55–60, 1997)     

An improved method for determining enantiomeric excess by 13C‐NMR in chiral liquid crystal media

By using a combination of inverse gated ¹H decoupled ¹³C‐NMR experiments 1 with short acquisition times and NMR Cryo‐probe technology, the sample requirements and experimental times necessary to accurately measure enantiomeric excess of small chiral molecules has been reduced 16‐fold. Quality ¹³C‐NMR spectra can now be obtained from a 1 to 5 mg sample in 12 minutes. The enantiomeric excess determination achieved from the average integration of all the ¹³C‐resonances in the spectrum is comparable to enantiomeric excess measured by chiral SFC. The advantage of the NMR method is that enantiomeric excess can rapidly be measured in situ on practical amounts of enantioselective reaction products without the need for chromatographic separation or chemical modification and with substantially less solvent waste. Chirality, 2010. © 2010 Wiley‐Liss, Inc.     

Shrinkage‐based Diagonal Discriminant Analysis and Its Applications in High‐Dimensional Data

Summary High‐dimensional data such as microarrays have brought us new statistical challenges. For example, using a large number of genes to classify samples based on a small number of microarrays remains a difficult problem. Diagonal discriminant analysis, support vector machines, and k‐nearest neighbor have been suggested as among the best methods for small sample size situations, but none was found to be superior to others. In this article, we propose an improved diagonal discriminant approach through shrinkage and regularization of the variances. The performance of our new approach along with the existing methods is studied through simulations and applications to real data. These studies show that the proposed shrinkage‐based and regularization diagonal discriminant methods have lower misclassification rates than existing methods in many cases.     

Ectoparasites (sucking lice,fleas and ticks) of small mammals in southeastern Kenya

During 1998–2000, at least 14 species (n = 309) of small mammals were live‐trapped and examined for ectoparasites in moist forests of the Taita and Shimba Hills and drier savannah habitats of Nguruman, southeastern Kenya. Ectoparasites were recorded from 11 species of mammals. Five species of sucking lice [Hoplopleura inexpectans Johnson, H. intermedia Kellogg & Ferris, Polyplax reclinata (Nitzsch), P. waterstoni Bedford and Schizophthirus graphiuri Ferris], six species of fleas (Ctenophthalmus leptodactylous Hubbard, Dinopsyllus grypurus Jordan & Rothschild, D. lypusus Jordan & Rothschild, Hypsophthalmus campestris Jordan & Rothschild, Listropsylla basilewskyi Smit and Xiphiopsylla lippa Jordan) and at least six species of ticks (Amblyomma sp., Haemaphysalis sp., Ixodes sp., I. alluaudi Neumann, I. cumulatimpunctatus Schulze, I. muniensis Arthur & Burrow and Rhipicephalus sp.) were recorded from these hosts. Four of the five species of sucking lice were host specific whereas P. reclinata was recorded from two different species of white‐toothed shrews, Crocidura spp. Although fleas and ticks were less host specific, C. leptodactylous, D. grypurus and I. cumulatimpunctatus were only recorded from the murid rodent Praomys delectorum (Thomas), Amblyomma sp. was only recorded from the nesomyid rodent Beamys hindei Thomas, Rhipicephalus sp. was only recorded from the murid Lemniscomys striatus (L.) and I. muniensis was only recorded from the dormouse Graphiurus microtis (Noack). More species of ectoparasites and significantly greater infestation prevalences were recorded from small mammals in moist habitats compared with those from the savannah habitat. At least one of the fleas recorded, D. lypusus, is a known vector of Yersinia pestis Lehmann & Neumann, the causative agent of plague, which is present in the region.     

Simvastatin enhances induction of inducible nitric oxide synthase in 3T3-L1 adipocytes

The present study was designed to determine whether hydroxymethylglutaryl-CoA reductase inhibitors (statins) modulate the NO production via iNOS in adipocytes stimulated by lipopolysaccharide (L) and tumour necrosis factor-α (T). Well-differentiated 3T3-L1 adipocytes significantly produced NO by LT-treatment. Pre-incubation with simvastatin, a lipophilic statin, pravastatin, a hydrophilic one, or Y27632, an inhibitor of Rho kinase, further enhanced the production of NO. The effect of simvastatin was offset by mevalonate and geranylgeranyl pyrophosphate (GGPP) but not by squalene. The mRNA level for iNOS parallelled the NO production. The NF-κB was activated by the LT-treatment and was further enhanced by simvastatin, pravastatin or Y27632 addition. Mevalonate and GGPP completely offset the effect of simvastatin. Statins and Y27632 also further increased the interleukin-6 secretion in the LT-treated 3T3-L1 adipocytes. These results suggest that statins, especially lipophilic type, enhance induction of iNOS by inhibiting the small GTP-binding protein signal in adipocytes.