HcRed, a Genetically Encoded Fluorescent Binary Cross-Linking Agent for Cross-Linking of Mitochondrial ATP Synthase in Saccharomyces cerevisiae

Genetically encoded fluorescent cross-linking agents represent powerful tools useful both for visualising and modulating protein interactions in living cells. The far-red fluorescent protein HcRed, which is fluorescent only in a dimer form, can be used to promote the homo-dimerisation of target proteins, and thereby yield useful information about biological processes. We have in yeast cells expressed HcRed fused to a subunit of mitochondrial ATP synthase (mtATPase). This resulted in cross-linking of the large multi-subunit mtATPase complex within the inner-membrane of the mitochondrion. Fluorescence microscopy revealed aberrant mitochondrial morphology, and mtATPase complexes isolated from mitochondria were recovered as fluorescent dimers under conditions where complexes from control mitochondria were recovered as monomers. When viewed by electron microscopy normal cristae were absent from mitochondria in cells in which mATPase complexes were cross-linked. mtATPase dimers are believed to be the building blocks that are assembled into supramolecular mtATPase ribbons that promote the formation of mitochondrial cristae. We propose that HcRed cross-links mATPase complexes in the mitochondrial membrane hindering the normal assembly/disassembly of the supramolecular forms of mtATPase

The HY5-PIF regulatory module coordinates light and temperature control of photosynthetic gene transcription

The ability to interpret daily and seasonal alterations in light and temperature signals is essential for plant survival. This is particularly important during seedling establishment when the phytochrome photoreceptors activate photosynthetic pigment production for photoautotrophic growth. Phytochromes accomplish this partly through the suppression of phytochrome interacting factors (PIFs), negative regulators of chlorophyll and carotenoid biosynthesis. While the bZIP transcription factor long hypocotyl 5 (HY5), a potent PIF antagonist, promotes photosynthetic pigment accumulation in response to light. Here we demonstrate that by directly targeting a common promoter cis-element (G-box), HY5 and PIFs form a dynamic activation-suppression transcriptional module responsive to light and temperature cues. This antagonistic regulatory module provides a simple, direct mechanism through which environmental change can redirect transcriptional control of genes required for photosynthesis and photoprotection. In the regulation of photopigment biosynthesis genes, HY5 and PIFs do not operate alone, but with the circadian clock. However, sudden changes in light or temperature conditions can trigger changes in HY5 and PIFs abundance that adjust the expression of common target genes to optimise photosynthetic performance and growth

The characterisation of microsatellite markers reveals tetraploidy in the Greater Water Parsnip, Sium latifolium (Apiaceae).

BACKGROUND: The Greater Water Parsnip, Sium latifolium (Apiaceae), is a marginal aquatic perennial currently endangered in England and consequently the focus of a number of conservation translocation projects. Microsatellite markers were developed for S. latifolium to facilitate comparison of genetic diversity and composition between natural and introduced populations. RESULTS: We selected 65 S. latifolium microsatellite (MiSeq) sequences and designed primer pairs for these. Primer sets were tested in 32 individuals. We found 15 polymorphic loci that amplified consistently. For the selected 15 loci, the number of alleles per locus ranged from 8 to 17. For all loci, S. latifolium individuals displayed up to four alleles indicating polyploidy in this species. CONCLUSIONS: These are the first microsatellite loci developed for S. latifolium and each individual displayed 1-4 alleles per locus, suggesting polyploidy in this species. These markers provide a valuable resource in evaluating the population genetic composition of this endangered species and thus will be useful for guiding conservation and future translocations of the species

Predictors of well child care adherence over time in a cohort of urban Medicaid-eligible infants

<p>Abstract</p> <p>Background</p> <p>Changes in well child care (WCC) adherence over time have not previously been examined. Our objective is to describe adherence rates to WCC over time in a low-income urban population of infants 0-24 months of age, and to identify predictors of WCC adherence in this population.</p> <p>Methods</p> <p>This is a secondary analysis of a cohort of Medicaid-eligible children followed from birth to 2 years between 2005 and 2008 with structured telephone surveys to assess maternal well-being, social support, and household and demographic information. For the 260 children attending 4 urban pediatric practices, WCC adherence was assessed based on visit data abstracted from electronic medical records. A random-intercept mixed effects logit model clustered on subject was used.</p> <p>Results</p> <p>92% of the mothers were African-American, 27% had not finished high school, 87% were single, and 43% earned < $500/month; mean age was 23. WCC adherence decreased from 88% at 6 months to 47% (12 mo), 44% (18 mo), and 67% (24 mo). The difference across time periods was statistically significant (p < 0.001). Married (OR 1.71, p = 0.02) and primiparous (OR 1.89, p < 0.001) mothers had significantly greater odds of adherence, along with women who reported having been adherent to prenatal care visits (OR 1.49, p = 0.03) and those with the lowest household income (OR 1.40, p = 0.03).</p> <p>Conclusions</p> <p>Maternal education efforts should emphasize the importance of establishing WCC, especially for mothers of more than one child. Further studies using larger, more broadly defined populations are needed to confirm our findings that efforts to increase WCC adherence should be intensified after 6 months of age, particularly for children at higher risk.</p

The Embedding Problem for Markov Models of Nucleotide Substitution

10.1371/journal.pone.0069187PLoS ONE87-POLN

Adaptation and Convergent Evolution within the Jamesonia-Eriosorus Complex in High-Elevation Biodiverse Andean Hotspots

The recent uplift of the tropical Andes (since the late Pliocene or early Pleistocene) provided extensive ecological opportunity for evolutionary radiations. We test for phylogenetic and morphological evidence of adaptive radiation and convergent evolution to novel habitats (exposed, high-altitude páramo habitats) in the Andean fern genera Jamesonia and Eriosorus. We construct time-calibrated phylogenies for the Jamesonia-Eriosorus clade. We then use recent phylogenetic comparative methods to test for evolutionary transitions among habitats, associations between habitat and leaf morphology, and ecologically driven variation in the rate of morphological evolution. Páramo species (Jamesonia) display morphological adaptations consistent with convergent evolution in response to the demands of a highly exposed environment but these adaptations are associated with microhabitat use rather than the páramo per se. Species that are associated with exposed microhabitats (including Jamesonia and Eriorsorus) are characterized by many but short pinnae per frond whereas species occupying sheltered microhabitats (primarily Eriosorus) have few but long pinnae per frond. Pinnae length declines more rapidly with altitude in sheltered species. Rates of speciation are significantly higher among páramo than non-páramo lineages supporting the hypothesis of adaptation and divergence in the unique Páramo biodiversity hotspot

Parasite-Dependent Expansion of TNF Receptor II–Positive Regulatory T Cells with Enhanced Suppressive Activity in Adults with Severe Malaria

Severe Plasmodium falciparum malaria is a major cause of global mortality, yet the immunological factors underlying progression to severe disease remain unclear. CD4+CD25+ regulatory T cells (Treg cells) are associated with impaired T cell control of Plasmodium spp infection. We investigated the relationship between Treg cells, parasite biomass, and P. falciparum malaria disease severity in adults living in a malaria-endemic region of Indonesia. CD4+CD25+Foxp3+CD127lo Treg cells were significantly elevated in patients with uncomplicated (UM; n = 17) and severe malaria (SM; n = 16) relative to exposed asymptomatic controls (AC; n = 10). In patients with SM, Treg cell frequency correlated positively with parasitemia (r = 0.79, p = 0.0003) and total parasite biomass (r = 0.87, p<0.001), both major determinants for the development of severe and fatal malaria, and Treg cells were significantly increased in hyperparasitemia. There was a further significant correlation between Treg cell frequency and plasma concentrations of soluble tumor necrosis factor receptor II (TNFRII) in SM. A subset of TNFRII+ Treg cells with high expression of Foxp3 was increased in severe relative to uncomplicated malaria. In vitro, P. falciparum–infected red blood cells dose dependently induced TNFRII+Foxp3hi Treg cells in PBMC from malaria-unexposed donors which showed greater suppressive activity than TNFRII− Treg cells. The selective enrichment of the Treg cell compartment for a maximally suppressive TNFRII+Foxp3hi Treg subset in severe malaria provides a potential link between immune suppression, increased parasite biomass, and malaria disease severity. The findings caution against the induction of TNFRII+Foxp3hi Treg cells when developing effective malaria vaccines

The Colorectal cancer disease-specific transcriptome may facilitate the discovery of more biologically and clinically relevant information

<p>Abstract</p> <p>Background</p> <p>To date, there are no clinically reliable predictive markers of response to the current treatment regimens for advanced colorectal cancer. The aim of the current study was to compare and assess the power of transcriptional profiling using a generic microarray and a disease-specific transcriptome-based microarray. We also examined the biological and clinical relevance of the disease-specific transcriptome.</p> <p>Methods</p> <p>DNA microarray profiling was carried out on isogenic sensitive and 5-FU-resistant HCT116 colorectal cancer cell lines using the Affymetrix HG-U133 Plus2.0 array and the Almac Diagnostics Colorectal cancer disease specific Research tool. In addition, DNA microarray profiling was also carried out on pre-treatment metastatic colorectal cancer biopsies using the colorectal cancer disease specific Research tool. The two microarray platforms were compared based on detection of probesets and biological information.</p> <p>Results</p> <p>The results demonstrated that the disease-specific transcriptome-based microarray was able to out-perform the generic genomic-based microarray on a number of levels including detection of transcripts and pathway analysis. In addition, the disease-specific microarray contains a high percentage of antisense transcripts and further analysis demonstrated that a number of these exist in sense:antisense pairs. Comparison between cell line models and metastatic CRC patient biopsies further demonstrated that a number of the identified sense:antisense pairs were also detected in CRC patient biopsies, suggesting potential clinical relevance.</p> <p>Conclusions</p> <p>Analysis from our <it>in vitro </it>and clinical experiments has demonstrated that many transcripts exist in sense:antisense pairs including <it>IGF2BP2</it>, which may have a direct regulatory function in the context of colorectal cancer. While the functional relevance of the antisense transcripts has been established by many studies, their functional role is currently unclear; however, the numbers that have been detected by the disease-specific microarray would suggest that they may be important regulatory transcripts. This study has demonstrated the power of a disease-specific transcriptome-based approach and highlighted the potential novel biologically and clinically relevant information that is gained when using such a methodology.</p