Gene expression in autumn leaves

Two cDNA libraries were prepared, one from leaves of a field-grown aspen (Populus tremula) tree, harvested just before any visible sign of leaf senescence in the autumn, and one from young but fully expanded leaves of greenhouse-grown aspen (Populus tremula x tremuloides). Expressed sequence tags (ESTs; 5,128 and 4,841, respectively) were obtained from the two libraries. A semiautomatic method of annotation and functional classification of the ESTs, according to a modified Munich Institute of Protein Sequences classification scheme, was developed, utilizing information from three different databases. The patterns of gene expression in the two libraries were strikingly different. In the autumn leaf library, ESTs encoding metallothionein, early light-inducible proteins, and cysteine proteases were most abundant. Clones encoding other proteases and proteins involved in respiration and breakdown of lipids and pigments, as well as stress-related genes, were also well represented. We identified homologs to many known senescence-associated genes, as well as seven different genes encoding cysteine proteases, two encoding aspartic proteases, five encoding metallothioneins, and 35 additional genes that were up-regulated in autumn leaves. We also indirectly estimated the rate of plastid protein synthesis in the autumn leaves to be less that 10% of that in young leaves.     

Dispersal biology of Liatris scariosa var. novae-angliae (Asteraceae), a rare New England grassland perennial

Propagule dispersal biology is a crucial avenue of research for rare plant species, especially those adapted to disturbance, such as northern blazing star (Liatris scariosa var. novae-angliae), a rare, early-successional New England grassland perennial. We examined the dispersal ability of northern blazing star propagules collected from 14 populations covering the entire latitudinal range of the taxon. Multiple regression demonstrated that dispersal ability, as measured by drop time in still air and flight distance in a low-speed wind tunnel, decreased significantly with propagule size and achene length, and increased with achene width and (for flight distance) pappus length. We used this multiple regression model to test for differences in predicted dispersal capability among maternal families, populations, and inland, coastal, and island habitats. Dispersal capability differed significantly among families and populations but not regions, and allometric relationships between morphological measurements were consistent across populations. Overall, dispersal capability was negatively correlated with germination success in a common greenhouse environment. However, germination success for a given dispersal ability, as well as achene shape, differed among populations. These results suggest specific populations to be targeted for management efforts promoting dispersal and establishment.     

Influence of nutrient status and grazing pressure on the fate of Francisella tularensis in lake water

The natural reservoir of Francisella tularensis , the causative agent of tularaemia, is yet to be identified. We investigated the possibility that Francisella persists in natural aquatic ecosystems between outbreaks. It was hypothesized that nutrient-rich environments, with strong protozoan predation, favour the occurrence of the tularaemia bacterium. To investigate the differences in adaptation to aquatic environments of the species and subspecies of Francisella , we screened 23 strains for their ability to survive grazing by the ciliate Tetrahymena pyriformis . All the Francisella strains tested were consumed at a low rate, although significant differences between subspecies were found. The survival and virulence of gfp -labelled F. tularensis ssp. holarctica were then studied in a microcosm experiment using natural lake water, with varying food web complexities and nutrient availabilities. High nutrient conditions in combination with high abundances of nanoflagellates were found to favour F. tularensis ssp. holarctica . The bacterium was observed both free-living and within the cells of a nanoflagellate. Francisella tularensis entered a viable but nonculturable state during the microcosm experiment. When studied over a longer period of time, F. tularensis ssp. holarctica survived in the lake water, but loss of virulence was not prevented by either high nutrient availability or the presence of predators.     

Variation in predator species abundance can cause variable selection pressure on warning signaling prey

Predation pressure is expected to drive visual warning signals to evolve toward conspicuousness. However, coloration of defended species varies tremendously and can at certain instances be considered as more camouflaged rather than conspicuous. Recent theoretical studies suggest that the variation in signal conspicuousness can be caused by variation (within or between species) in predators' willingness to attack defended prey or by the broadness of the predators' signal generalization. If some of the predator species are capable of coping with the secondary defenses of their prey, selection can favor reduced prey signal conspicuousness via reduced detectability or recognition. In this study, we combine data collected during three large-scale field experiments to assess whether variation in avian predator species (red kite, black kite, common buzzard, short-toed eagle, and booted eagle) affects the predation pressure on warningly and non-warningly colored artificial snakes. Predation pressure varied among locations and interestingly, if common buzzards were abundant, there were disadvantages to snakes possessing warning signaling. Our results indicate that predator community can have important consequences on the evolution of warning signals. Predators that ignore the warning signal and defense can be the key for the maintenance of variation in warning signal architecture and maintenance of inconspicuous signaling.     

Nucleosome Spacing Generated by ISWI and CHD1 Remodelers Is Constant Regardless of Nucleosome Density

Arrays of regularly spaced nucleosomes are a hallmark of chromatin, but it remains unclear how they are generated. Recent genome-wide studies, in vitro and in vivo, showed constant nucleosome spacing even if the histone concentration was experimentally reduced. This counters the long-held assumption that nucleosome density determines spacing and calls for factors keeping spacing constant regardless of nucleosome density. We call this a clamping activity. Here, we show in a purified system that ISWI- and CHD1-type nucleosome remodelers have a clamping activity such that they not only generate regularly spaced nucleosome arrays but also generate constant spacing regardless of nucleosome density. This points to a functionally attractive nucleosome interaction that could be mediated either directly by nucleosome-nucleosome contacts or indirectly through the remodelers. Mutant Drosophila melanogaster ISWI without the HAND-SANT-SLIDE (HSS) domain had no detectable spacing activity even though it is known to remodel and slide nucleosomes. This suggests that the role of ISWI remodelers in generating constant spacing is not just to mediate nucleosome sliding; they actively contribute to the attractive interaction. Additional factors are necessary to set physiological spacing in absolute terms.     

Functional and molecular analysis of the double-positive stage-specific CD8 enhancer E8III during thymocyte development

Several developmental stage-, subset-, and lineage-specific Cd8 cis-regulatory regions have been identified. These include the E8(III) enhancer, which directs expression in double-positive (DP) thymocytes, and E8(II), which is active in DP cells and CD8(+) T cells. Using a transgenic reporter expression assay, we identified a 285-bp core fragment of the E8(III) enhancer that retains activity in DP thymocytes. In vitro characterization of the core enhancer revealed five regulatory elements that are required for full enhancer activity, suggesting that multiple factors contribute to the developmental stage-specific activity. Furthermore, deletion of E8(III) in the mouse germline showed that this enhancer is required for nonvariegated expression of CD8 in DP thymocytes when E8(II) is also deleted. These results indicate that E8(III) is one of the cis-elements that contribute to the activation of the Cd8a and Cd8b gene complex during T cell development.     

In vivo analysis of proteomes and interactomes using Parallel Affinity Capture (iPAC) coupled to mass spectrometry

Affinity purification coupled to mass spectrometry provides a reliable method for identifying proteins and their binding partners. In this study we have used Drosophila melanogaster proteins triple tagged with Flag, Strep II, and Yellow fluorescent protein in vivo within affinity pull-down experiments and isolated these proteins in their native complexes from embryos. We describe a pipeline for determining interactomes by Parallel Affinity Capture (iPAC) and show its use by identifying partners of several protein baits with a range of sizes and subcellular locations. This purification protocol employs the different tags in parallel and involves detailed comparison of resulting mass spectrometry data sets, ensuring the interaction lists achieved are of high confidence. We show that this approach identifies known interactors of bait proteins as well as novel interaction partners by comparing data achieved with published interaction data sets. The high confidence in vivo protein data sets presented here add new data to the currently incomplete D. melanogaster interactome. Additionally we report contaminant proteins that are persistent with affinity purifications irrespective of the tagged bait.     

Genetic Loci Associated with Alzheimer’s Disease and Cerebrospinal Fluid Biomarkers in a Finnish Case-Control Cohort

Objectives

To understand the relation between risk genes for Alzheimer’s disease (AD) and their influence on biomarkers for AD, we examined the association of AD in the Finnish cohort with single nucleotide polymorphisms (SNPs) from top AlzGene loci, genome-wide association studies (GWAS), and candidate gene studies; and tested the correlation between these SNPs and AD markers Aβ_1–42, total tau (t-tau), and phosphorylated tau (p-tau) in cerebrospinal fluid (CSF).

Methods

We tested 25 SNPs for genetic association with clinical AD in our cohort comprised of 890 AD patients and 701-age matched healthy controls using logistic regression. For the correlational study with biomarkers, we tested 36 SNPs in a subset of 222 AD patients with available CSF using mixed models. Statistical analyses were adjusted for age, gender and APOE status. False discovery rate for multiple testing was applied. All participants were from academic hospital and research institutions in Finland.

Results

APOE-ε4, CLU rs11136000, and MS4A4A rs2304933 correlated with significantly decreased Aβ_1–42 (corrected p<0.05). At an uncorrected p<0.05, PPP3R1 rs1868402 and MAPT rs2435211 were related with increased t-tau; while SORL1 rs73595277 and MAPT rs16940758, with increased p-tau. Only TOMM40 rs2075650 showed association with clinical AD after adjusting for APOE-ε4 (p = 0.007), but not after multiple test correction (p>0.05).

Conclusions

We provide evidence that APOE-ε4, CLU and MS4A4A, which have been identified in GWAS to be associated with AD, also significantly reduced CSF Aβ_1–42 in AD. None of the other AlzGene and GWAS loci showed significant effects on CSF tau. The effects of other SNPs on CSF biomarkers and clinical AD diagnosis did not reach statistical significance. Our findings suggest that APOE-ε4, CLU and MS4A4A influence both AD risk and CSF Aβ_1–42.