A bovine papillomavirus E1-related protein binds specifically to bovine papillomavirus DNA.

The E1 open reading frame of bovine papillomavirus (BPV) was expressed as a RecA-E1 fusion protein in Escherichia coli. The bacterially expressed RecA-E1 protein exhibited sequence-specific DNA binding activity; strong binding to the region from nucleotides 7819 to 93 on the BPV genome (designated region A) and weak binding to the adjacent region from nucleotides 7457 to 7818 (region B) were observed. The interaction between the BPV-derived RecA-E1 protein and region A appeared to be highly specific for BPV DNA, as no comparable binding was detected with heterologous papillomavirus DNAs. Binding to region A was eliminated by digestion of region A at the unique HpaI site, which suggests that the RecA-E1 binding site(s) was at or near the HpaI recognition sequence. Binding to region B but not region A was observed when nuclear extracts from ID13 cells were used as a source of E1 proteins. The absence of region A binding by ID13 extracts may reflect a negative regulation of E1 DNA binding activity.     

Human adenovirus type 9-induced rat mammary tumors.

Following subcutaneous inoculation of newborn Wistar-Furth rats with human adenovirus type 9 (Ad9), 16 of 16 female and 0 of 11 male rats developed mammary tumors. Tumor-positive animals usually developed tumors in multiple glands. Histopathological analyses indicated that three general categories of tumor could be identified. Mammary fibroadenomas were the most common tumor type encountered, but phyllodeslike tumors and solid sarcomas were also frequently found. In situ hybridization and immunohistochemical techniques established that benign fibroadenomas were derived from mammary fibroblasts (collagen type I- and vimentin-positive cells) and that malignant tumors were derived from myoepithelial cells (collagen type IV-, vimentin-, and muscle-specific actin-positive cells). The fact that mammary tumors were limited to female rats suggested that female hormones are essential for tumor growth and development. In this regard, ovariectomy of Ad9-infected female rats prevented tumor development, while subsequent diethylstilbestrol (DES) treatment elicited tumor formation. In addition, Ad9-infected and castrated male rats which received DES also developed mammary tumors. Established male mammary tumors regressed when DES treatment was stopped and reappeared after DES treatment was resumed. Together, these results indicate that estrogen is required for both initiation and maintenance of Ad9-induced mammary tumors. Southern blot analysis of high-molecular-weight tumor DNA showed that mammary tumor cells contained single or multiple integrated copies of the entire Ad9 genome. RNase protection experiments established that estrogen receptor as well as Ad9 E1a and E4 mRNAs were expressed in mammary tumors, but Ad9 E3 and, surprisingly, E1b mRNAs were not expressed at detectable levels.     

Coronary atherosclerosis: current therapeutic approaches and future trends

Invasive cardiovascular procedures, such as percutaneous translumenal coronary angioplasty (PTCA) and aorto-coronary bypass surgery (ACBS), that are currently employed in treating the coronary stenosis or occlusion caused by atherosclerosis represent a major therapeutic advance for managing coronary heart disease (CHD). However, the cellular proliferative response and associated intimal hyperplasia that can follow the damage to blood vessels that occurs with these procedures leads to late complications which cannot be effectively controlled by presently available drugs. Hence, a new approach is required for managing these complications, termed "restenosis" (in the case of PTCA) or "stenosis" (in the case of ACBS). Existing drug therapy is reviewed and some new approaches to this problem are provided herein. Further studies of growth factors and other substances that influence the cellular proliferative response that follows injury to the blood vessel wall could lead to the development of effective therapy. Inhibition of intimal hyperplasia and/or acceleration of endothelial cell re-growth provide a basis for such new approaches. Platelet-derived growth factor (PDGF) and basic fibroblast growth factor (bFGF), as well as endothelium-derived relaxing factor(s) (EDRF) and calcitonin gene-related peptide (CGRP) are among the substances discussed. Modification of certain currently available drugs (e.g. Ca(2+)-antagonists) could also be of value in meeting this therapeutic demand.     

Post-transcriptional regulation in higher eukaryotes: the role of the reporter gene in controlling expression

We have investigated whether reporter genes influence cytoplasmic regulation of gene expression in tobacco and Chinese hamster ovary (CHO) cells. Two genes, uidA encoding beta-glucuronidase (GUS) from Escherichia coli and Luc, encoding firefly luciferase (LUC), were used to analyze the ability of a cap, polyadenylated tail, and the 5'- and 3'-untranslated regions (UTR) from tobacco mosaic virus (TMV) to regulate expression. The regulation associated with the 5' cap structure and the TMV 5'-UTR, both of which enhance translational efficiency, was reporter gene-independent. The poly(A) tail and the TMV 3'-UTR, which is functionally equivalent to a poly(A) tail, increase translational efficiency as well as mRNA stability. The regulation associated with these 3' ends was highly reporter gene-dependent; their effect on GUS expression was almost an order of magnitude greater than that on LUC expression. In tobacco, the tenfold reporter gene effect on poly(A) tail or TMV 3'-UTR function could not be explained by a differential impact on mRNA stability; GUS and LUC mRNA half-life increased only twofold when either the poly(A) tail or TMV 3'-UTR was present. In CHO cells, however, GUS mRNA was stabilized to a greater extent by a poly(A) tail or the TMV 3'-UTR than was LUC mRNA.     

Use of the polymerase chain reaction for typing allelic variants of the human HLA-DQA1 by hybridization with oligonucleotide probes, specific for specific alleles]

Class II HLA molecules are the most useful markers for susceptibility to different autoimmune diseases, including insulin-dependent diabetes mellitus (IDDM) and rheumatoid arthritis (RA). Polymerase chain reaction and hybridization with a set of allele-specific oligonucleotide have been used for analysis of allelic sequence variation. The analysis of frequencies of HLA-DQA1 alleles among 10 patients of the russian population revealed a uneven distribution. We have developed a method for preparing non-radioactive oligonucleotide probes with terminal deoxynucleotidyl transferase and Bio-11-dUTP. Comparison of biotinylated and 32P-labeled hybridization probes gave the same sensitivity for HLA-DQA1 typing of amplified DNA. Amplification of the HLA-DQA1 gene has been successful on 10 pg of total DNA. This amount of DNA is close to the amount of DNA in a single cell. Alternatively, HLA-DQA1 typing could be based on the analysis of buccal cells of saliva that would avoid the problem of individuals who object to giving blood samples.     

Fractionation of eukaryotic DNA in a pulsating electrical field. I. Detection and properties of discrete DNA fragments]

The fractionation of eukaryotic DNA by field inversion gel electrophoresis results in the appearance of discrete DNA-fragments. The set of these fragments is similar to that of different eukaryotic representatives and consists of various chromosomal DNAs, unified by size. The physical properties of DNA-fragments suggest that they can form multimeric structures due to the presence of sticky ends flanking discrete fragments. We suppose that the set of discrete DNA-fragments results in a specific cleavage of intact nuclear DNA and can reflect different levels of chromatin structural organization.     

Size-dependent drift responses of mayflies to experimental hydrologic variation: active predator avoidance or passive hydrodynamic displacement?

Summary Larger nymphs within aquatic insect taxa have been frequently observed to be transported down-stream in the stream drift only at night. Others have hypothesized this pattern results primarily from large nymphs' behavioural avoidance of entering drift during daylight, when size-selective, visually-feeding fish predators are most active. This hypothesis assumes that animals can actively control their entry into the drift, which may not be the case under all flow conditions. We experimentally induced streamflow increases and decreases in adjacent riffles in a hydrologically-stable stream during the daytime to examine whether changes in diel patterns of drift abundance and size-distribution of mayflies were consistent with the hypothesis of active avoidance of diurnal drift. We assessed the likelihood of active vs. passive mechanisms of diurnal drift entry and transport for four taxa that differ with respect to body size, morpho-behavioural attributes, microhabitat use, and general propensity to drift. In each of three seasons, diurnal and nocturnal drift samples were collected in three riffles over two diel cycles. Background drift patterns were established on the first day (no flow manipulation). Six h before sunset on the second day, flow was experimentally increased in one riffle, decreased in the second, and not altered in the third (control). Between-day differences in diurnal and nocturnal drift rate and size composition were then compared among the treatment and reference riffles. Responses of two taxa were consistent with active control over drift entry, transport, or both. For Baetis spp., drift-prone mayflies typically preyed upon by fish, diurnal drift rates immediately increased following both flow reduction and flow elevation in all seasons, but only small individuals comprised the drift. Drift by large individuals was delayed until nighttime. Epeorus longimanus also exhibited significant increases in drift rates following flow reduction and elevation, but responses of this large-bodied species were restricted to nighttime. Drift responses for these two taxa were largely independent of direction of hydrologic change, thus indicating a strong behavioural control over drift. By contrast, numbers and sizes of drifting Paraleptophlebia heteronea and Ephemerella infrequens depended strongly on direction of flow change. Drift rates for both species generally declined after flow reduction and increased after flow elevation. Moreover, after flow elevation, larger individuals often drifted diurnally, a finding consistent with expectations under a passive hydrodynamic model. These experiments indicate that size-dependent mayfly drift reflects not only presumed risk from visual fish predators, but also functional attributes of species such as morphology, behaviour, and microhabitat affiliation, which influence aspects of drift entry and transport under variable hydrologic conditions.     

Trilobite cuticle thickness in relation to palaeoenvironment

100 thickness measurements from thin sections of cephala or pygidia of early Ordovician trilobites occurring across an onshore to offshore environmental gradient show that progressively greater maximum cuticle thickness was characteristic of increasingly inshore sites. There is a 40-fold difference between the thinnest and thickest cuticles, and exclusively thin cuticles are confined to the offshore Olenid Biofacies. Variability in cuticle thickness increases offshore to onshore. Environmental control is shown to be more influential on cuticle thickness than is the overall length of the trilobite: some comparatively large trilobites having thin cuticles and small trilobites thick cuticles. The environmental factors which might be responsible for the pattern are briefly discussed. The thin cuticles dominating the offshore Olenid Biofacies were probably appropriate for dysaerobic conditions. Thick cuticles in the most inshore biofacies may have offered protection against predators and turbulence, but the additional presence there of trilobites with thinner cuticles is considered to reflect the greater heterogeneity of the epeiric habitat.     

Salicyloylaspartate as an endogenous component in the leaves of Phaseolus vulgaris

An amide conjugate of o-methoxybenzoic acid and aspartic acid has been isolated from bean leaves. After extraction and methylation of plant material, this compound was isolated as two isomeric monoethyl monomethyl esters. The ethylation of the aspartyl carboxyl groups was shown to be a likely result of an extraction procedure utilising acidified ethanol, the methylation of the aromatic hydroxy of the methoxy group to be due to the derivatisation procedure. Studies with pentafluorobenzylation confirmed that the endogenous compound is o-hydroxybenzoylaspartate.     

The basic types of scar contractures after burns and methods of eliminating them with trapezeplasty flaps

The study of postburn scar contractures in various locations has revealed four contracture variables: edge, medial, strip, and total. Following the surgical treatment of more than 2000 patients with such contractures, a trapezeplasty flap method has been worked out and applied since 1979. This method allows one to make up for deficient scar tissue of the same shape. Flaps are cut out of sheets of a fold with the maximum use of undamaged skin in adjacent sections. They move toward each other and merge with adjacent sides into a state of tension. This surgery is based on use of the reserve on the width of the surface of a fold and the high tension of skin with pressure on underlying tissue. The extended skin grows quickly, the tension disappears, and the scars dissolve. For each contracture type, there are trapeze-flap variables either in pure form or in combination with the transposition of split-thickness skin with a flap to create a flexible joint zone. The trapezeplasty flap method can be used to treat all edge, medial, strip, and total contractures of joints whose natural position is adduction, the shoulder joint, and contractures between fingers. Skin-fat or skin-fascia trapeze flaps prevent the relapse of contracture and make the weakened scars softer, which, as a rule, ensures a good functional and aesthetic result.