Dynamic demethylation of the IL2RA promoter during in vitro CD4+ T cell activation in association with IL2RA expression

IL2RA, a subunit of the high affinity receptor for interleukin-2 (IL2), plays a crucial role in immune homeostasis. Notably, IL2RA expression is induced in CD4+ T cells in response to various stimuli and is constitutive in regulatory T cells (Tregs). We selected for our study 18 CpGs located within cognate regulatory regions of the IL2RA locus and characterized their methylation in naive, regulatory, and memory CD4+ T cells. We found that 5/18 CpGs (notably CpG + 3502) show dynamic, active demethylation during the in vitro activation of naive CD4+ T cells. Demethylation of these CpGs correlates with appearance of IL2RA protein at the cell surface. We found no influence of cis located SNP alleles upon CpG methylation. Treg cells show constitutive demethylation at all studied CpGs. Methylation of 9/18 CpGs, including CpG +3502, decreases with age. Our data thus identify CpG +3502 and a few other CpGs at the IL2RA locus as coordinated epigenetic regulators of IL2RA expression in CD4+ T cells. This may contribute to unravel how the IL2RA locus can be involved in immune physiology and pathology.     

Spatial and temporal variations in floral resource availability affect bumblebee communities in heathlands

Modifications of landscape structure and composition can decrease the availability of floral resources, resulting in the decline of many pollinator species, including bumblebees. These declines may have significant ecological consequences, because bumblebees pollinate a large range of plant species. Our study was carried out in heathlands, open semi-natural habitats that have decreased considerably due to human activities. We analysed how floral resources affect bumblebee communities throughout the colony lifetime at three scales: plot scale, heathland patch scale, and landscape scale. Floral density at the plot scale and spruce plantations at the landscape scale influenced bumblebee communities. The abundance of bumblebees on ericaceous species was higher when the landscape included a substantial proportion of unsuitable foraging habitat (i.e., spruce plantations). Both life history traits and colony life cycle stage influenced bumblebee responses to the availability of floral resources. Bumblebees were more affected by floral resources during the colony development phase than during the nest-foundation or mating phases. Moreover, bumblebees of species that form large colonies needed larger quantities of favourable foraging habitat, compared with small-colony bees, and their proportion decreased in habitats dominated by spruce plantations. In conclusion, the conservation of plant–bumblebee interactions will require management at a larger spatial scale than the restricted protected habitats. Moreover, at the landscape scale, both quantity of favourable foraging patches and their ecological continuity are important to conserve both small- and large- colony species.     

Haplotype-specific <Emphasis Type="Italic">MAPT</Emphasis> exon 3 expression regulated by common intronic polymorphisms associated with Parkinsonian disorders

Background

Genome wide association studies have identified microtubule associated protein tau (MAPT) H1 haplotype single nucleotide polymorphisms (SNPs) as leading common risk variants for Parkinson’s disease, progressive supranuclear palsy and corticobasal degeneration. The MAPT risk variants fall within a large 1.8 Mb region of high linkage disequilibrium, making it difficult to discern the functionally important risk variants. Here, we leverage the strong haplotype-specific expression of MAPT exon 3 to investigate the functionality of SNPs that fall within this H1 haplotype region of linkage disequilibrium.

Methods

In this study, we dissect the molecular mechanisms by which haplotype-specific SNPs confer allele-specific effects on the alternative splicing of MAPT exon 3. Firstly, we use haplotype-hybrid whole-locus genomic MAPT vectors studies to identify functional SNPs. Next, we characterise the RNA-protein interactions at two loci by mass spectrometry. Lastly, we knockdown candidate splice factors to determine their effect on MAPT exon 3 using a novel allele-specific qPCR assay.

Results

Using whole-locus genomic DNA expression vectors to express MAPT haplotype variants, we demonstrate that rs17651213 regulates exon 3 inclusion in a haplotype-specific manner. We further investigated the functionality of this region using RNA-electrophoretic mobility shift assays to show differential RNA-protein complex formation at the H1 and H2 sequence variants of SNP rs17651213 and rs1800547 and subsequently identified candidate trans-acting splicing factors interacting with these functional SNPs sequences by RNA-protein pull-down experiment and mass spectrometry. Finally, gene knockdown of candidate splice factors identified by mass spectrometry demonstrate a role for hnRNP F and hnRNP Q in the haplotype-specific regulation of exon 3 inclusion.

Conclusions

We identified common splice factors hnRNP F and hnRNP Q regulating the haplotype-specific splicing of MAPT exon 3 through intronic variants rs1800547 and rs17651213. This work demonstrates an integrated approach to characterise the functionality of risk variants in large regions of linkage disequilibrium.

     

Plant–pollinator interactions: comparison between an invasive and a native congeneric species

Plant–pollinator interactions determine reproductive success for animal-pollinated species and, in the case of invasive plants, they are supposed to play an important role in invasive success. We compared the invasive Senecio inaequidens to its native congener S. jacobaea in terms of interactions with pollinators. Visitor guild, visitation rate, and seed set were compared over 3 years in three sites in Belgium. Floral display (capitula number and arrangement) and phenology were quantified, and visiting insects were individually censused, i.e. number of visited capitula and time per visited capitulum. As expected from capitula resemblance, visitor guilds of both species were very similar (proportional similarity = 0.94). Senecio inaequidens was visited by 33 species, versus 36 for S. jacobaea. For both species, main visitors were Diptera, especially Syrphidae, and Hymenoptera. Visitation rate averaged 0.13 visitor per capitulum per 10 min for S. inaequidens against 0.08 for S. jacobaea. However, insects visited more capitula per plant on S. jacobaea, due to high capitula density (886 m⁻² versus 206 m⁻² for S. inaequidens), which is likely to increase self-pollen deposition considerably. Seed set of S. jacobaea was lower than that of S. inaequidens. We suggest that floral display is the major factor explaining the differences in insect visitation and seed set between the two Senecio species.     

Intracellular delivery of peptides via association with ubiquitin or SUMO-1 coupled to protein transduction domains

Background  

We previously developed small hybrid proteins consisting of SUMO-1 linked to an heptapeptide fused to the Tat protein transduction domain (PTD). The heptapeptide motif was selected from a library of random sequences to specifically bind HIV-1 regulatory proteins Tat or Rev. These constructs, named SHP, are able to enter primary lymphocytes and some of them inhibit HIV-1 replication. Considering these positive results and other data from the literature, we further tested the ability of ubiquitin or SUMO-1 linked to various PTD at their N-terminus to deliver within cells proteins or peptides fused downstream of their diglycine motif. In this system it is expected that the intracellular ubiquitin or SUMO-1 hydrolases cleave the PTD-Ub or PTD-SUMO-1 modules from the cargo polypeptide, thereby allowing its delivery under an unmodified form.     

Reproductive biology of the andromonoecious Cucumis melo subsp. agrestis (Cucurbitaceae)

Background and Aims

Cucumis melo subsp. agrestis (Cucurbitaceae) is cultivated in many African regions for its edible kernels used as a soup thickener. The plant, an annual, andromonoecious, trailing-vine species, is of high social, cultural and economic value for local communities. In order to improve the yield of this crop, the first step and our aim were to elucidate its breeding system.

Methods

Eight experimental pollination treatments were performed during three growing seasons to assess spontaneous selfing, self-compatibility and effects of pollen source (hermaphroditic vs. male flowers). Pollination success was determined by pollen tube growth and reproductive success was assessed by fruit, seed and seedling numbers and characteristics. The pollinator guild was surveyed and the pollination distance determined both by direct observations and by indirect fluorescent dye dispersal.

Key Results

The species is probably pollinated by several Hymenoptera, principally by Hypotrigona para. Pollinator flight distances varied from 25 to 69 cm. No evidence for apomixis or spontaneous self-pollination in the absence of insect visitors was found. The self-fertility index (SFI = 0) indicated a total dependence on pollinators for reproductive success. The effects of hand pollination on fruit set, seed number and seedling fitness differed among years. Pollen tube growth and reproductive success did not differ between self- and cross-pollinations. Accordingly, a high self-compatibility index for the fruit set (SCI = 1·00) and the seed number (SCI = 0·98) and a low inbreeding depression at all developmental stages (cumulative δ = 0·126) suggest a high selfing ability. Finally, pollen origin had no effect on fruit and seed sets.

Conclusions

This andromonoecious species has the potential for a mixed mating system with high dependence on insect-mediated pollination. The selfing rate through geitonogamy should be important.     

Genesis, effects and fates of repeats in prokaryotic genomes

DNA repeats are causes and consequences of genome plasticity. Repeats are created by intrachromosomal recombination or horizontal transfer. They are targeted by recombination processes leading to amplifications, deletions and rearrangements of genetic material. The identification and analysis of repeats in nearly 700 genomes of bacteria and archaea is facilitated by the existence of sequence data and adequate bioinformatic tools. These have revealed the immense diversity of repeats in genomes, from those created by selfish elements to the ones used for protection against selfish elements, from those arising from transient gene amplifications to the ones leading to stable duplications. Experimental works have shown that some repeats do not carry any adaptive value, while others allow functional diversification and increased expression. All repeats carry some potential to disorganize and destabilize genomes. Because recombination and selection for repeats vary between genomes, the number and types of repeats are also quite diverse and in line with ecological variables, such as host-dependent associations or population sizes, and with genetic variables, such as the recombination machinery. From an evolutionary point of view, repeats represent both opportunities and problems. We describe how repeats are created and how they can be found in genomes. We then focus on the functional and genomic consequences of repeats that dictate their fate.