Internalization of a polysialic acid-binding Escherichia coli bacteriophage into eukaryotic neuroblastoma cells

Eukaryotic organisms are continuously exposed to bacteriophages, which are efficient gene transfer agents in bacteria. However, bacteriophages are considered not to pass the eukaryotic cell membrane and enter nonphagocytic cells. Here we report the binding and penetration of Escherichia coli PK1A2 bacteriophage into live eukaryotic neuroblastoma cells in vitro. The phage interacts with cell surface polysialic acid, which shares structural similarity with the bacterial phage receptor. Using fluorescence and electron microscopy, we show that phages are internalized via the endolysosomal route and persist inside the human cells up to one day without affecting cell viability. Phage capsid integrity is lost in lysosomes, and the phage DNA is eventually degraded. We did not detect the entry of phage DNA into the nucleus; however, we speculate that this might occur as a rare event, and propose that this potential mechanism could explain prokaryote-eukaryote gene flow.Peer reviewe

Universal platform for quantitative analysis of DNA transposition

<p>Abstract</p> <p>Background</p> <p>Completed genome projects have revealed an astonishing diversity of transposable genetic elements, implying the existence of novel element families yet to be discovered from diverse life forms. Concurrently, several better understood transposon systems have been exploited as efficient tools in molecular biology and genomics applications. Characterization of new mobile elements and improvement of the existing transposition technology platforms warrant easy-to-use assays for the quantitative analysis of DNA transposition.</p> <p>Results</p> <p>Here we developed a universal <it>in vivo </it>platform for the analysis of transposition frequency with class II mobile elements, i.e., DNA transposons. For each particular transposon system, cloning of the transposon ends and the cognate transposase gene, in three consecutive steps, generates a multifunctional plasmid, which drives inducible expression of the transposase gene and includes a mobilisable <it>lacZ</it>-containing reporter transposon. The assay scores transposition events as blue microcolonies, papillae, growing within otherwise whitish <it>Escherichia coli </it>colonies on indicator plates. We developed the assay using phage Mu transposition as a test model and validated the platform using various MuA transposase mutants. For further validation and to illustrate universality, we introduced IS<it>903 </it>transposition system components into the assay. The developed assay is adjustable to a desired level of initial transposition via the control of a plasmid-borne <it>E. coli </it>arabinose promoter. In practice, the transposition frequency is modulated by varying the concentration of arabinose or glucose in the growth medium. We show that variable levels of transpositional activity can be analysed, thus enabling straightforward screens for hyper- or hypoactive transposase mutants, regardless of the original wild-type activity level.</p> <p>Conclusions</p> <p>The established universal papillation assay platform should be widely applicable to a variety of mobile elements. It can be used for mechanistic studies to dissect transposition and provides a means to screen or scrutinise transposase mutants and genes encoding host factors. In succession, improved versions of transposition systems should yield better tools for molecular biology and offer versatile genome modification vehicles for many types of studies, including gene therapy and stem cell research.</p

Antecedents and consequences of effectuation and causation in the international new venture creation process

The selection of the entry mode in an international market is of key importance for the venture. A process-based perspective on entry mode selection can add to the International Business and International Entrepreneurship literature. Framing the international market entry as an entrepreneurial process, this paper analyzes the antecedents and consequences of causation and effectuation in the entry mode selection. For the analysis, regression-based techniques were used on a sample of 65 gazelles. The results indicate that experienced entrepreneurs tend to apply effectuation rather than causation, while uncertainty does not have a systematic influence. Entrepreneurs using causation-based international new venture creation processes tend to engage in export-type entry modes, while effectuation-based international new venture creation processes do not predetermine the entry mod

Subtropical streams harbour higher genus richness and lower abundance of insects compared to boreal streams, but scale matters

Aim: Biological diversity typically varies between climatically different regions, and regions closer to the equator often support higher numbers of taxa than those closer to the poles. However, these trends have been assessed for a few organism groups, and the existing studies have rarely been based on extensive identical surveys in different climatic regions. Location: We conducted standardized surveys of wadeable streams in a boreal (western Finland) and a subtropical (south-eastern Brazil) region, sampling insects identically from 100 streams in each region and measuring the same environmental variables in both regions. Taxon: Aquatic insects. Methods: Comparisons were made at the scales of local stream sites, drainage basins and entire regions. We standardized the spatial extent of the study areas by resampling regional richness based on subsets of sites with similar extents. We examined differences in genus richness and assemblage abundance patterns between the regions using graphical and statistical modelling approaches. Results: We found that while genus accumulation and rank-abundance curves were relatively similar at the regional scale between Finland and Brazil, regional genus richness was higher in the latter but regional abundance much higher in the former region. These regional patterns for richness and abundance were reproduced by basin and local genus richness that were higher in Brazil than in Finland, and assemblage abundance that was much higher in Finland than in Brazil. The magnitude of the difference in genus richness between Brazil and Finland tended to increase from local through basin to regional scales. Main conclusions: Our findings suggest that factors related to evolutionary diversification might explain differences in genus richness between these two climatically different regions, whereas higher nutrient concentrations of stream waters might explain the higher abundance of insects in Finland than in Brazil.Peer reviewe

Expression of neural cell adhesion molecule and polysialic acid in human bone marrow-derived mesenchymal stromal cells

BACKGROUND: In order to develop novel clinical applications and to gain insights into possible therapeutic mechanisms, detailed molecular characterization of human bone marrow-derived mesenchymal stromal cells (hBM-MSCs) is needed. Neural cell adhesion molecule (NCAM, CD56) is a transmembrane glycoprotein modulating cell–cell and cell–matrix interactions. An additional post-translational modification of NCAM is the α2,8-linked polysialic acid (polySia). Because of its background, NCAM is often considered a marker of neural lineage commitment. Generally, hBM-MSCs are considered to be devoid of NCAM expression, but more rigorous characterization is needed. METHODS: We have studied NCAM and polySia expression in five hBM-MSC lines at mRNA and protein levels. Cell surface localization was confirmed by immunofluorescence staining and expression frequency in the donor-specific lines by flow cytometry. For the detection of poorly immunogenic polySia, a fluorochrome-tagged catalytically defective enzyme was employed. RESULTS: All five known NCAM isoforms are expressed in these cells at mRNA level and the three main isoforms are present at protein level. Both polysialyltransferases, generally responsible for NCAM polysialylation, are expressed at mRNA level, but only very few cells express polySia at the cell surface. CONCLUSIONS: Our results underline the need for a careful control of methods and conditions in the characterization of MSCs. This study shows that, against the generally held view, clinical-grade hBM-MSCs do express NCAM. In contrast, although both polysialyltransferase genes are transcribed in these cells, very few express polySia at the cell surface. NCAM and polySia represent new candidate molecules for influencing MSC interactions.Peer reviewe

Are coronary event rates declining slower in women than in men – evidence from two population-based myocardial infarction registers in Finland?

<p>Abstract</p> <p>Background</p> <p>Studies have suggested that the prevention and treatment of coronary heart disease may not have been as effective in women as in men. Therefore, we aimed to examine whether the incidence, attack rate and mortality of myocardial infarction (MI) events have declined less in women than in men.</p> <p>Methods</p> <p>Two large population-based MI registers, the FINAMI register and the Finnish Cardiovascular Disease Register (CVDR) were used for comparing the event rates among men and women aged ≥35 years in two time periods, 1994–1996 and 2000–2002.</p> <p>Results</p> <p>In the FINAMI register a total of 5,252 events were recorded in men and 4,898 in women. Corresponding numbers in the CVDR were 78,709 and 70,464. Both FINAMI and CVDR data suggested smaller declines in incidence and attack rate of MI events in women than in men. In CVDR data the decline in mortality was also smaller in women than in men, while in FINAMI data this difference did not reach statistical significance. In the large CVDR data set, negative binomial regression models revealed smaller declines in incidence (p = 0.006), attack rate (p = 0.008) and mortality (p = 0.04) in women than in men aged <55 years. In persons ≥55 years no difference was observed between women and men.</p> <p>Conclusion</p> <p>The incidence and attack rate of MI events have declined less in women aged <55 than in men of similar age. In older persons no significant differences were observed. Further studies are warranted to find out the reasons why the development has been less favourable for young women than for men.</p

Stream diatom biodiversity in islands and continents—A global perspective on effects of area, isolation and environment

Aim The species-area relationship (SAR) is one of the most distinctive biogeographic patterns, but global comparisons of the SARs between island and mainland are lacking for microbial taxa. Here, we explore whether the form of the SAR and the drivers of species richness, including area, environmental heterogeneity, climate and physico-chemistry, differ between islands and similarly sized areas on mainland, referred to as continental area equivalents (CAEs). Location Global. Taxon Stream benthic diatoms. Methods We generated CAEs on six continental datasets and examined the SARs of CAEs and islands (ISAR). Then, we compared CAEs and islands in terms of total richness and richness of different ecological guilds. We tested the factors contributing to richness in islands and CAEs with regressions. We used structural equation models to determine the effects of area versus environmental heterogeneity, climate and local conditions on species richness. Results We found a non-significant ISAR, but a significant positive SAR in CAEs. Richness in islands was related to productivity. Richness in CAEs was mainly dependent on area and climate, but not directly on environmental heterogeneity. Species richness within guilds exhibited inconsistent relationships with island isolation and area. Main conclusions Ecological and evolutionary processes shaping diatom island biogeography do not depend on area at the worldwide scale probably due to the presence of distinct species pool across islands. Conversely, area was an important driver of diatom richness in continents, and this effect could be attributed to dispersal. Continents had greater richness than islands, but this was a consequence of differences in environmental conditions such as specific island climatic conditions. We stress the need for more island data on benthic diatoms, particularly from archipelagos, to better understand the biogeography of this most speciose group of algae

Beta diversity of stream insects differs between boreal and subtropical regions, but land use does not generally cause biotic homogenization

Previous studies have found mixed results regarding the relationship between beta diversity and latitude. In addition, by influencing local environmental heterogeneity, land use may modify spatial taxonomic and functional variability among communities causing biotic differentiation or homogenization. We tested 1) whether taxonomic and functional beta diversities among streams within watersheds differ between subtropical and boreal regions and 2) whether land use is related to taxonomic and functional beta diversities in both regions. We sampled aquatic insects in 100 subtropical (Brazil) and 100 boreal (Finland) streams across a wide gradient of land use, including agriculture and exotic planted, secondary, and native forests. We calculated beta diversity at the watershed scale (among 5 streams in each watershed). We found higher taxonomic beta diversity among subtropical than among boreal streams, whereas functional beta diversity was similar between the 2 regions. Total land use was positively correlated with taxonomic and functional beta diversity among subtropical streams, while local environmental heterogeneity was positively correlated with beta diversity among boreal streams. We suggest that different types and intensities of land use may increase among-stream heterogeneity, promoting distinct insect assemblage compositions among streams. Our findings also suggest that beta diversity patterns and their underlying determinants are highly context dependent