Conservation of heterochromatin protein 1 function

Heterochromatin represents a cytologically visible state of heritable gene repression. In the yeast, Schizosaccharomyces pombe, the swi6 gene encodes a heterochromatin protein 1 (HP1)-like chromodomain protein that localizes to heterochromatin domains, including the centromeres, telomeres, and the donor mating-type loci, and is involved in silencing at these loci. We identify here the functional domains of swi6p and demonstrate that the chromodomain from a mammalian HP1-like protein, M31, can functionally replace that of swi6p, showing that chromodomain function is conserved from yeasts to humans. Site-directed mutagenesis, based on a modeled three-dimensional structure of the swi6p chromodomain, shows that the hydrophobic amino acids which lie in the core of the structure are critical for biological function. Gel filtration, gel overlay experiments, and mass spectroscopy show that HP1 proteins can self-associate, and we suggest that it is as oligomers that HP1 proteins are incorporated into heterochromatin complexes that silence gene activity.     

Testing Commercial Sex Workers for Sexually Transmitted Infections in Victoria,Australia: An Evaluation of the Impact of Reducing the Frequency of Testing

Background

The frequency of testing sex workers for sexually transmitted infections (STIs) in Victoria, Australia, was changed from monthly to quarterly on 6 October 2012. Our aim was to determine the impact of this change to the clients seen at the Melbourne Sexual Health Centre (MHSC).

Methods

Computerised medical records of all clients attending at MHSC from 7 October 2011 to 7 October 2013 were analysed.

Results

Comparing between the monthly and quarterly testing periods, the number of consultations at MSHC with female sex workers (FSW) halved from 6146 to 3453 (p<0.001) and the consultation time spent on FSW reduced by 40.6% (1942 h to 1153 h). More heterosexual men (p<0.001), and women (p<0.001) were seen in the quarterly testing period. The number of STIs diagnosed in the clinic increased from 2243 to 2589 from the monthly to quarterly period, respectively [15.4% increase (p<0.001)]. Up to AU$247,000 was saved on FSW testing after the shift to quarterly testing.

Conclusions

The change to STIs screening frequency for sex workers from monthly to quarterly resulted in a 15% increase in STI diagnoses in the clinic and approximate a quarter of a million dollars was diverted from FSW testing to other clients. Overall the change in frequency is likely to have had a beneficial effect on STI control in Victoria.     

Modulation of ion transporter expression in gill mitochondrion-rich cells of eels acclimated to low-Na(+) or-Cl(-) freshwater

In this study, we aimed to establish an experimental model to study the role of the gill mitochondrion-rich cells (MRCs) of freshwater fish in Na(+) uptake and to examine the effect of adjusting external Na(+) and Cl(-) ions on selected ion transporters in gill MRCs. Japanese eels (Anguilla japonica) acclimated to deionized (DI) water for 2 weeks were transferred directly to (a) ion-supplemented artificial freshwater (AF), (b) Na(+) -deficient AF, or (c) Cl(-) -deficient AF for 2 days. The effects of the transfer on the expression levels of ion transporters in isolated gill cells were investigated. Our data demonstrated that the 2-day acclimation in ion-supplemented AF, Na(+) -deficient AF, or Cl(-) -deficient AF led to a significant increase in serum osmolarity attributed mainly to an increase in serum Na(+) and/or Cl(-) levels when compared with DI-acclimated eel. Significant inductions of V-type H(+) -ATPase (V-H(+) -ATPase) and cotransporter (NBC1) mRNA expression in gill MRCs were detected in AF-acclimated fish. In fish acclimated to Na(+) -deficient AF, mRNA expression levels of V-H(+) -ATPase, NBC1, and Na(+) /H(+) -exchanger-3 (NHE3) were significantly increased in MRCs. Fish acclimated to Cl(-) -deficient AF showed no observable change in expression levels of ion transporters in gill MRCs. In addition, expression levels of ion transporters in pavement cells were stable throughout the 2-day experiments. These data indicate that the level of Na(+) in freshwater is important for altering the mRNA expression of ion transporters in gill MRCs, which supports the notion that gill MRCs play important roles in freshwater Na(+) uptake.     

Dendritic cells and macrophages are productively infected by poliovirus

Expression of the poliovirus receptor (PVR) on cells is a major host determinant of infection by poliovirus. Previously, the only immune cell type known to express PVR was the blood-derived monocyte, which is susceptible to infection at very low frequency. We demonstrate that professional antigen-presenting cells-macrophages and dendritic cells, generated upon differentiation of monocytes-retain expression of PVR and are highly susceptible to infection by type 1 Mahoney strain of poliovirus. Maximal cell-associated titers of virus are obtained within 6 to 8 h postinfection, and cell death and lysis occurs within 24 h postinfection. Similar kinetics are observed in cells infected with the Sabin 1 vaccine strain. Although protein synthesis and receptor-mediated endocytosis are inhibited upon poliovirus infection of these critical antigen-presenting cells, we demonstrate for the first time that functional presentation of antigen occurs in these infected cells via the HLA class II pathway.     

Age-related decrease of protein kinase G activation in vascular smooth muscle cells

The p21 (cip1/waf1) protein induces cell cycle arrest through inhibition of the activity of cdk (cyclin dependent kinase)/cyclin complexes. Expression of p21 is induced in a p53-dependent manner by DNA damage. p21 can also be induced independently of p53 by phorbol ester or okadaic acid. In this study, we have addressed the role of the PKC (protein kinase C) signaling pathway in the induction of p21 in response to PMA (phorbol myristate acetate) and okadaic acid. Levels of p21 (protein and mRNA) rapidly increased (within approximately 4 h) in U937 cells treated with PMA. The PKC-specific inhibitors RO 31-8220 and GF109203X down-regulated PMA or okadaic acid-induced p21 expression. Following persistent PKC activation, p21 mRNA levels remained elevated, indicating an enhanced stability of the mRNA. Using actinomycin D to measure mRNA stability and p21 promoter luciferase assays to measure activity, we provide evidence to support a role for the PKC signaling pathway in p21 mRNA stability. Thus, PKC regulates the amount of p21 in U937 cells at the level of mRNA accumulation and translation.     

Environmental Profile of a Community's Health (EPOCH): An Ecometric Assessment of Measures of the Community Environment Based on Individual Perception

BACKGROUND: Public health research has turned towards examining upstream, community-level determinants of cardiovascular disease risk factors. Objective measures of the environment, such as those derived from direct observation, and perception-based measures by residents have both been associated with health behaviours. However, current methods are generally limited to objective measures, often derived from administrative data, and few instruments have been evaluated for use in rural areas or in low-income countries. We evaluate the reliability of a quantitative tool designed to capture perceptions of community tobacco, nutrition, and social environments obtained from interviews with residents in communities in 5 countries. METHODOLOGY/ PRINCIPAL FINDINGS: Thirteen measures of the community environment were developed from responses to questionnaire items from 2,360 individuals residing in 84 urban and rural communities in 5 countries (China, India, Brazil, Colombia, and Canada) in the Environmental Profile of a Community's Health (EPOCH) study. Reliability and other properties of the community-level measures were assessed using multilevel models. High reliability (>0.80) was demonstrated for all community-level measures at the mean number of survey respondents per community (n?=?28 respondents). Questionnaire items included in each scale were found to represent a common latent factor at the community level in multilevel factor analysis models. CONCLUSIONS/ SIGNIFICANCE: Reliable measures which represent aspects of communities potentially related to cardiovascular disease (CVD)/risk factors can be obtained using feasible sample sizes. The EPOCH instrument is suitable for use in different settings to explore upstream determinants of CVD/risk factors.     

Developmental regulation of the gene for formate dehydrogenase in Neurospora crassa.

We have isolated and characterized a gene, fdh, from Neurospora crassa which is developmentally regulated and which produces formate dehydrogenase activity when expressed in Escherichia coli. The gene is closely linked (less than 0.6 kb apart) to the leu-5 gene encoding mitochondrial leucyl-tRNA synthetase; the two genes are transcribed convergently from opposite strands. The expression patterns of these genes differ: fdh mRNA is found only during conidiation and early germination and is not detectable during mycelial growth, while leu-5 mRNA appears during germination and mycelial growth. The structure of the fdh gene was determined from the sequence of cDNA and genomic DNA clones and from mRNA mapping studies. The gene encodes a 375-amino-acid-long protein with sequence similarity to NAD-dependent dehydrogenases of the E. coli 3-phosphoglycerate dehydrogenase (serA gene product) subfamily. In particular, there is striking sequence similarity (52% identity) to formate dehydrogenase from Pseudomonas sp. strain 101. All of the residues thought to interact with NAD in the crystal structure of the Pseudomonas enzyme are conserved in the N. crassa enzyme. We have further shown that expression of the N. crassa gene in E. coli leads to the production of formate dehydrogenase activity, indicating that the N. crassa gene specifies a functional polypeptide.     

Experimental and Computational Investigation of Biofilm Formation by Rhodopseudomonas palustris Growth under Two Metabolic Modes

We examined biofilms formed by the metabolically versatile bacterium Rhodopseudomonas palustris grown via different metabolic modes. R. palustris was grown in flow cell chambers with identical medium conditions either in the presence or absence of light and oxygen. In the absence of oxygen and the presence of light, R. palustris grew and formed biofilms photoheterotrophically, and in the presence of oxygen and the absence of light, R. palustris grew and formed biofilms heterotrophically. We used confocal laser scanning microscopy and image analysis software to quantitatively analyze and compare R. palustris biofilm formation over time in these two metabolic modes. We describe quantifiable differences in structure between the biofilms formed by the bacterium grown heterotrophically and those grown photoheterotrophically. We developed a computational model to explore ways in which biotic and abiotic parameters could drive the observed biofilm architectures, as well as a random-forest machine-learning algorithm based on structural differences that was able to identify growth conditions from the confocal imaging of the biofilms with 87% accuracy. Insight into the structure of phototrophic biofilms and conditions that influence biofilm formation is relevant for understanding the generation of biofilm structures with different properties, and for optimizing applications with phototrophic bacteria growing in the biofilm state.     

The effect of different spectral light quality on the photoinhibition of Photosystem I in intact leaves

Photosynthesis Research - Light energy causes damage to Photosystem I (PSI) and Photosystem II (PSII). The majority of the previous photoinhibition studies have been conducted with PSII, which...     

Influence of Formulation and Processing Variables on Properties of Itraconazole Nanoparticles Made by Advanced Evaporative Precipitation into Aqueous Solution

Nanoparticles, of the poorly water-soluble drug, itraconazole (ITZ), were produced by the Advanced Evaporative Precipitation into Aqueous Solution process (Advanced EPAS). This process combines emulsion templating and EPAS processing to provide improved control over the size distribution of precipitated particles. Specifically, oil-in-water emulsions containing the drug and suitable stabilizers are sprayed into a heated aqueous solution to induce precipitation of the drug in form of nanoparticles. The influence of processing parameters (temperature and volume of the heated aqueous solution; type of nozzle) and formulation aspects (stabilizer concentrations; total solid concentrations) on the size of suspended ITZ particles, as determined by laser diffraction, was investigated. Furthermore, freeze-dried ITZ nanoparticles were evaluated regarding their morphology, crystallinity, redispersibility, and dissolution behavior. Results indicate that a robust precipitation process was developed such that size distribution of dispersed nanoparticles was shown to be largely independent across the different processing and formulation parameters. Freeze-drying of colloidal dispersions resulted in micron-sized agglomerates composed of spherical, sub-300-nm particles characterized by reduced crystallinity and high ITZ potencies of up to 94% (w/w). The use of sucrose prevented particle agglomeration and resulted in powders that were readily reconstituted and reached high and sustained supersaturation levels upon dissolution in aqueous media.