Expression of normal and mutant GFP-tagged y(+)L amino acid transporter-1 in mammalian cells

Lysinuric protein intolerance (LPI; MIM 222700) is an autosomal recessive disorder characterized by defective transport of cationic amino acids lysine, arginine and ornithine. The defect is localized in the basolateral membrane of polar epithelial cells of the renal tubules and intestine. The SLC7A7 (solute carrier family 7, member 7) gene that encodes y(+)LAT-1 (y(+)L amino acid transporter-1) is mutated in LPI, and leads to the malfunction of the heterodimer composed of y(+)LAT-1 and 4F2hc (4F2 heavy chain) responsible for the system y(+)L amino acid transport activity at the membrane. In this study, the intracellular trafficking and membrane expression of wild type and four mutant y(+)LAT-1 proteins (LPI(Fin), G54V, 1548delC, W242X) was studied in two human cell lines by expressing green fluorescent protein (GFP) tagged proteins. Different SLC7A7 mutations influenced the trafficking of y(+)LAT-1 in the cells differently, as the wild type and missense mutant fusion proteins localized to the plasma membrane, while the frameshift and nonsense mutants sequestered to the cytoplasmic membranes, never reaching the target areas of the cell.     

The KlPGS1 gene encoding phosphatidylglycerolphosphate synthase in Kluyveromyces lactis is essential and assigned to chromosome I

The phosphatidylglycerolphosphate synthase (CDP-diacylglycerol:sn-glycerol-3-phosphate 3-phosphatidyltransferase, EC 2.7.8.5) is an essential enzyme in biosynthesis of cardiolipin. In this work we report the isolation, heterological cloning, molecular characterization and physical mapping of the Saccharomyces cerevisiae PEL1/PGS1 homologue from Kluyveromyces lactis. The pel1 mutant strain of S. cerevisiae was used to isolate this homologue by screening a K. lactis genomic library. The novel cloned gene was named KlPGS1. Its coding region was found to consist of 1623 bp. The corresponding protein exhibits 55% amino acid identity to its S. cerevisiae counterpart. The presence of the mitochondrial presequence indicates its mitochondrial localization. Sporulation and ascus dissection of diploids heterozygous for single-copy disruption of KlPGS1 revealed that the KlPGS1 gene, is essential in K. lactis. Using a DIG-dUTP-labeled DNA probe-originated from the KlPGS1 gene and Southern hybridization of contour-clamped homogeneous electric field (CHEF)-separated K. lactis chromosomal DNA, the KlPGS1 gene was assigned to chromosome I. The nucleotide sequence data reported in this paper were submitted to GenBank and assigned the Accession No. AY176328.     

Intracardiac injection of a capsid-modified Ad5/35 results in decreased heart toxicity when compared to standard Ad5

Background

Human immunodeficiency virus type 1 (HIV-1) Nef-encoded protein plays key functions at almost all stages of the viral life cycle, but its role in translation is largely unknown.

Methods

To determine the effect of Nef on translation we used an in vitro translation assay. The detection of Nef/RPS10 complexes and the presence of 18S rRNA and tRNAs in the complexes were performed by coimmunoprecipitation and RT-PCR assay.

Results

We observed that the HIV-1 Nef protein specifically impaired translation in vitro. We observed the interaction of Nef with RPS10 by coimmunoprecipitation assay. In addition 18S rRNA and tRNAs were present in the Nef/RPS10 complexes.

Conclusions

Our results are consistent with a model in which the Nef protein by binding to two components of the 40S small ribosomal subunit, RPS10 and 18S rRNA, and to a lesser extent to tRNAs, could lead to decreased protein synthesis.     

Differential costs of reproduction in females and hermaphrodites in a gynodioecious plant

Background and Aims

Plants exhibit a variety of reproductive systems where unisexual (females or males) morphs coexist with hermaphrodites. The maintenance of dimorphic and polymorphic reproductive systems may be problematic. For example, to coexist with hermaphrodites the females of gynodioecious species have to compensate for the lack of male function. In our study species, Geranium sylvaticum, a perennial gynodioecious herb, the relative seed fitness advantage of females varies significantly between years within populations as well as among populations. Differences in reproductive investment between females and hermaphrodites may lead to differences in future survival, growth and reproductive success, i.e. to differential costs of reproduction. Since females of this species produce more seeds, higher costs of reproduction in females than in hermaphrodites were expected. Due to the higher costs of reproduction, the yearly variation in reproductive output of females might be more pronounced than that of hermaphrodites.

Methods

Using supplemental hand-pollination of females and hermaphrodites of G. sylvaticum we examined if increased reproductive output leads to differential costs of reproduction in terms of survival, probability of flowering, and seed production in the following year.

Key Results

Experimentally increased reproductive output had differential effects on the reproduction of females and hermaphrodites. In hermaphrodites, the probability of flowering decreased significantly in the following year, whereas in females the costs were expressed in terms of decreased future seed production.

Conclusions

When combining the probability of flowering and seed production per plant to estimate the multiplicative change in fitness, female plants showed a 56 % and hermaphrodites showed a 39 % decrease in fitness due to experimentally increased reproduction. Therefore, in total, female plants seem to be more sensitive to the cost of reproduction in terms of seed fitness than hermaphrodites.     

Educational Inequalities in Acute Myocardial Infarction Incidence in Norway: A Nationwide Cohort Study

Background

Increasing differences in cardiovascular disease (CVD) mortality across levels of education have been reported in Norway. The aim of the study was to investigate educational inequalities in acute myocardial infarction (AMI) incidence and whether such inequalities have changed during the past decade using a nationwide longitudinal study design.

Methods

Data on 141 332 incident (first) AMIs in Norway during 2001–2009 were obtained through the Cardiovascular Disease in Norway (CVDNOR) project. Educational inequalities in AMI incidence were assessed in terms of age-standardised incidence rates stratified on educational level, incidence rate ratios (IRR), relative index of inequality (RII) and slope index of inequality (SII). All calculations were conducted in four gender and age strata: Men and women aged 35–69 and 70–94 years.

Results

AMI Incidence rates decreased during 2001–2009 for all educational levels except in women aged 35–69 among whom only those with basic education had a significant decrease. In all gender and age groups; those with the highest educational level had the lowest rates. The strongest relative difference was found among women aged 35–69, with IRR (95% CI) for basic versus tertiary education 3.04 (2.85–3.24)) and RII (95% CI) equal to 4.36 (4.03–4.71). The relative differences did not change during 2001–2009 in any of the four gender and age groups, but absolute inequalities measured as SII decreased among the oldest men and women.

Conclusions

There are substantial educational inequalities in AMI incidence in Norway, especially for women aged 35–69. Relative inequalities did not change from 2001 to 2009.     

Community composition of butterflies and bumblebees in fallows: niche breadth and dispersal capacity modify responses to fallow type and landscape

Fallows (i.e. fields temporarily taken out of production) provide important habitat for flower-visiting insects in intensively cultivated agricultural landscapes. Cost-efficiency of fallowing schemes could be enhanced through improved understanding of key characteristics of fallows and surrounding landscape that determine community composition and provide support for species of conservation concern. Impacts of fallow characteristics and landscape structure on the species composition of butterflies and bumblebees were studied in two types of perennial fallows in boreal farmland. To understand species’ responses to environmental factors from a conservation perspective, community composition was examined with respect to two species traits—niche breadth and dispersal capacity. Whereas overall species composition of butterflies and bumblebees was strongly affected by forest cover in the surrounding landscape, the studied species traits were most related to fallow type and the cover of perennial grasslands. Habitat breadth of butterflies was narrowest in long-term grassland fallows in landscapes with high grassland cover. Dispersal capacity of butterflies was also lowest in grassland-rich landscapes. Diet breadth of bumblebees was narrower in long-term grassland fallows than in short-term fallows. The results confirm that the diversity of butterflies and bumblebees can be enhanced by establishing and managing fallows both in open and forested landscapes. For conservation of habitat specialists and less mobile species, retention of long-term fallows in grassland-rich landscapes is apparently the best option. The results provide no justification for exempting forested regions or farms with high grassland cover from the ecological focus area requirement under the European Union’s current agricultural policy.     

HaploRec: efficient and accurate large-scale reconstruction of haplotypes

Background  

Haplotypes extracted from human DNA can be used for gene mapping and other analysis of genetic patterns within and across populations. A fundamental problem is, however, that current practical laboratory methods do not give haplotype information. Estimation of phased haplotypes of unrelated individuals given their unphased genotypes is known as the haplotype reconstruction or phasing problem.     

Specific absorption rate and electric field measurements in the near field of six mobile phone base station antennas

In this article, the exposure to radio frequency electromagnetic fields was studied in close proximity (distances of 10, 100, 300, and 600 mm) to six base station antennas. The specific absorption rate (SAR) in 800 mm × 500 mm × 200 mm box phantom as well as unperturbed electric field (E) in air was measured. The results were used to determine whether the measurement of local maximum of unperturbed electric field can be used as a compliance check for local exposure. Also, the conservativeness of this assessment method compared to the ICNIRP basic restriction was studied. Moreover, the assessment of whole‐body exposure was discussed and the distance ranges presented in which the ICNIRP limit for local exposure could be exceeded before the limit for whole‐body SAR. These results show that the electric field measurement alone can be used for easy compliance check for the local exposure at all distances and for all antenna types studied. However, in some cases when the local peak value of E was compared directly to the ICNIRP reference level for unperturbed E, the exposure was overestimated only very slightly (by factor 1.1) compared to the basic restriction for localized SAR in a human, and hence these results can not be generalized to all antenna types. Moreover, it was shown that the limit for localized exposure could be exceeded before the limit for the whole‐body average SAR, if the distance to the antenna was less than 240 mm. Bioelectromagnetics 30:307–312, 2009. © 2009 Wiley‐Liss, Inc.     

Large-volume cytocentrifuge for processing alcohol-fixed cytologic specimens. Application in urinary cytology

The use of a large-volume cytocentrifuge for the routine processing of urine specimens was investigated. Most specimens arrived in the laboratory mixed with 50% ethanol. Polyethylene glycol was added to the fixative supplied to the clinicians or to the cell suspension received in the laboratory. The slides used in the cytocentrifuge were coated with gelatin or poly-L-lysine to minimize cell loss. The resulting monolayers of cells, with occasional true tissue fragments, were stained by the Papanicolaou method. When indicated, immunocytochemical and histochemical stains were used. The technique gave good morphologic details, good cell yield and consistent staining quality. This method has also been applied to other cytologic specimens, such as serous fluids and fine needle aspirates, with good results. The method saves the time of cytotechnologists, and slides prepared by this technique are suitable for automatic staining.