The reported point centromeres of Scheffersomyces stipitis are retrotransposon long terminal repeats

Point centromeres, found in some ascomycete yeasts such Saccharomyces cerevisiae, are very different in structure from the centromeres of other eukaryotes. They are tiny and nonrepetitive and contain only two short conserved sequence motifs. Until recently, point centromeres were thought to have a single evolutionary origin, in the budding yeast family Saccharomycetaceae. Most yeasts outside this family have centromeres that are many kilobases in size. Some have centromeres consisting of a large inverted repeat sequence, others have centromeric clusters of retrotransposons, and a third group including Candida albicans has centromeres with no conserved sequence features. It was recently reported that Scheffersomyces stipitis has point centromeres with a strongly conserved 125-bp core sequence, which is unexpected because S. stipitis is only distantly related to the known point-centromere species. We show here that the 125-bp core sequence is actually part of the long terminal repeat (LTR) of the Ty5-like retrotransposon Tps5, which forms a cluster in the centromeric region of each S. stipitis chromosome. Thus, the LTR of a centromere-associated retrotransposon confers centromere-like mitotic stability when cloned into a plasmid. The centromeric regions of S. stipitis contain three types of Tps5 element (Tps5a, Tps5b, and Tps5c) and a noncoding nonautonomous large retrotransposon derivative.     

Purification of the cardiac sarcoplasmic reticulum membrane protein phospholamban from recombinant Escherichia coli

Phospholamban (PLN) was expressed in Escherichia coli as a protein fusion with glutathione S-transferase (GST). GST-PLN was mostly present in the insoluble protein fraction and accounted for approximately 50% of total insoluble protein. Attempts to suppress inclusion body formation or to use GST as an affinity-purification tag failed. A successful purification method is based on preparative SDS/PAGE and electrodialysis. From 1 g cells we typically purified 13.5 mg fusion protein with a PLN content of 2.8 mg. We genetically inserted an enterokinase (EK) protease site just in front of the PLN sequence and demonstrated the proteolytical liberation of PLN from the carrier protein. The approach described represents a substantial advancement in PLN expression and purification.     

Human stem cell assays in immune-deficient mice

Much of our understanding of the organization of the cells that comprise the hematopoietic system and the cellular and molecular mechanisms that regulate their development is derived from mouse models. However, knowledge of the human hematopoietic system and identification of human stem cells have, until recently, been hampered by the absence of in vivo assays that measure their repopulation capacity. The development of methods to transplant normal and leukemic human hematopoietic cells into immune-deficient SCID mice provides the foundation for human stem cell assays. This review will focus on recent evidence that normal and leukemic human stem cells can be assayed in these systems.     

Epilepsy, retarded growth and onchocerciasis, in two areas of different endemicity of onchocerciasis in Burundi

A high prevalence of epilepsy (up to 1.3%) was observed in an area hyperendemic for onchocerciasis in the province of Bururi, Burundi. Following this observation, epileptics and controls were examined in 2 communes (administrative units) with different endemicity for onchocerciasis. Altogether, onchocerciasis was more frequent in epileptics (81.8%) than in controls from the same households (68.3%; P < 0.05). This difference was more marked in the hyperendemic area. Other possible causes of epilepsy, including cysticercosis, were infrequent (18 possible cases among 110 epileptics). During this survey, 9 epileptics with growth retardation were seen. They showed most characteristics of Nakalanga syndrome, which was described from Uganda. All 9 such cases were suffering from onchocerciasis. These findings give more evidence of a possible association between onchocerciasis and epilepsy, and between onchocerciasis and Nakalanga syndrome.     

A method for functional magnetic resonance imaging of olfaction

A method for generating olfactory stimuli for humans within a functional magnetic resonance imaging (fMRI) experimental design is described. The system incorporates a nasal-mask in which the change from odorant to no-odorant conditions occurs in less than 500 ms and is not accompanied by visual, auditory, tactile, or thermal cues. The mask provides an ordorant-free environment following prolonged ordorant presence. Specific imaging parameters that are conducive to the study of the human olfactory system are described. In a pilot study performed using these methods, the specific patterns of activation observed converged with published experimental and clinical findings.     

Differential divalent cation requirements uncouple the assembly and catalytic reactions of human immunodeficiency virus type 1 integrase

Previous in vitro analyses have shown that the human immunodeficiency virus type 1 (HIV-1) integrase uses either manganese or magnesium to assemble as a stable complex on the donor substrate and to catalyze strand transfer. We now demonstrate that subsequent to assembly, catalysis of both 3' end processing and strand transfer requires a divalent cation cofactor and that the divalent cation requirements for assembly and catalysis can be functionally distinguished based on the ability to utilize calcium and cobalt, respectively. The different divalent cation requirements manifest by these processes are exploited to uncouple assembly and catalysis, thus staging the reaction. Staged 3' end processing and strand transfer assays are then used in conjunction with exonuclease III protection analysis to investigate the effects of integrase inhibitors on each step in the reaction. Analysis of a series of related inhibitors demonstrates that these types of compounds affect assembly and not either catalytic process, therefore reconciling the apparent disparate results obtained for such inhibitors in assays using isolated preintegration complexes. These studies provide evidence for a distinct role of the divalent cation cofactor in assembly and catalysis and have implications for both the identification and characterization of integrase inhibitors.     

Predictors of underlying coronary artery disease in cocaine associated myocardial infarction: a meta-analysis of case reports

A preliminary report presenting the results of fibular strut grafting in the severely resorbed mandibular and maxillary region is presented. Thirteen patients were treated due to severe resorption of alveolar and basilar bone of 49 segments of the mandible and the maxilla. Two patients additionally had pathological fractures of the mandible. In 10 cases the strut graft was harvested by means of a new minimally invasive technique. After modelling the fibular bone it was fixed to the recipient site by miniscrews or implants. After a mean follow-up period of 20 months (max. 31, min. 11 months) a retrospective analysis of clinical and radiological findings was carried out. It showed that a mean augmentation of 16 mm was achieved. Compared to other studies the fibular strut graft was resorbed less, and due to the primary stability it could be used for the treatment of fractures of the mandible. No more than natural resorption was observed when the patients received their prostheses fixed to dental implants.