NPAHs and OPAHs in the atmosphere of two central European cities: Seasonality, urban-to-background gradients, cancer risks and gas-to-particle partitioning.

Derivatives of polycyclic aromatic hydrocarbons (PAHs) such as nitrated- and oxygenated-PAHs (NPAHs and OPAHs) could be even more toxic and harmful for the environment and humans than PAHs. We assessed the spatial and seasonal variations of NPAHs and OPAHs atmospheric levels, their cancer risks and their gas-to-particle partitioning. To this end, about 250 samples of fine particulate matter (PM2.5) and 50 gaseous samples were collected in 2017 in central Europe in the cities of Brno and Ljubljana (two traffic and two urban background sites) as well as one rural site. The average particulate concentrations were ranging from below limit of quantification to 593 pg m-3 for Σ9NPAHs and from 1.64 to 4330 pg m-3 for Σ11OPAHs, with significantly higher concentrations in winter compared to summer. In winter, the particulate levels of NPAHs and OPAHs were higher at the traffic site compared to the urban background site in Brno while the opposite was found in Ljubljana. NPAHs and OPAHs particulate levels were influenced by the meteorological parameters and co-varied with several air pollutants. The significance of secondary formation on the occurrence of some NPAHs and OPAHs is indicated. In winter, 27-47% of samples collected at all sites were above the acceptable lifetime carcinogenic risk. The gas-particle partitioning of NPAHs and OPAHs was influenced by their physico-chemical properties, the season and the site-specific aerosol composition. Three NPAHs and five OPAHs had higher particulate mass fractions at the traffic site, suggesting they could be primarily emitted as particles from vehicle traffic and subsequently partitioning to the gas phase along air transport. This study underlines the importance of inclusion of the gas phase in addition to the particulate phase when assessing the atmospheric fate of polycyclic aromatic compounds and also when assessing the related health risk.

Autophagy in Dictyostelium: Mechanisms, regulation and disease in a simple biomedical model.

Autophagy is a fast-moving field with an enormous impact on human health and disease. Understanding the complexity of the mechanism and regulation of this process often benefits from the use of simple experimental models such as the social amoeba Dictyostelium discoideum. Since the publication of the first review describing the potential of D. discoideum in autophagy, significant advances have been made that demonstrate both the experimental advantages and interest in using this model. Since our previous review, research in D. discoideum has shed light on the mechanisms that regulate autophagosome formation and contributed significantly to the study of autophagy-related pathologies. Here, we review these advances, as well as the current techniques to monitor autophagy in D. discoideum. The comprehensive bioinformatics search of autophagic proteins that was a substantial part of the previous review has not been revisited here except for those aspects that challenged previous predictions such as the composition of the Atg1 complex. In recent years our understanding of, and ability to investigate, autophagy in D. discoideum has evolved significantly and will surely enable and accelerate future research using this model.

ERECTA and BAK1 Receptor Like Kinases Interact to Regulate Immune Responses in Arabidopsis.

ERECTA (ER) receptor-like kinase (RLK) regulates Arabidopsis thaliana organ growth, and inflorescence and stomatal development by interacting with the ERECTA-family genes (ERf) paralogs, ER-like 1 (ERL1) and ERL2, and the receptor-like protein (RLP) TOO MANY MOUTHS (TMM). ER also controls immune responses and resistance to pathogens such as the bacterium Pseudomonas syringae pv. tomato DC3000 (Pto) and the necrotrophic fungus Plectosphaerella cucumerina BMM (PcBMM). We found that er null-mutant plants overexpressing an ER dominant-negative version lacking the cytoplasmic kinase domain (ERΔK) showed an enhanced susceptibility to PcBMM, suggesting that ERΔK associates and forms inactive complexes with additional RLKs/RLPs required for PcBMM resistance. Genetic analyses demonstrated that ER acts in a combinatorial specific manner with ERL1, ERL2, and TMM to control PcBMM resistance. Moreover, BAK1 (BRASSINOSTEROID INSENSITIVE 1-associated kinase 1) RLK, which together with ERf/TMM regulates stomatal patterning and resistance to Pto, was also found to have an unequal contribution with ER in regulating immune responses and resistance to PcBMM. Co-immunoprecipitation experiments in Nicotiana benthamiana further demonstrated BAK1-ER protein interaction. The secreted epidermal pattern factor peptides (EPF1 and EPF2), which are perceived by ERf members to specify stomatal patterning, do not seem to regulate ER-mediated immunity to PcBMM, since their inducible overexpression in A. thaliana did not impact on PcBMM resistance. Our results indicate that the multiproteic receptorsome formed by ERf, TMM and BAK1 modulates A. thaliana resistance to PcBMM, and suggest that the cues underlying ERf/TMM/BAK1-mediated immune responses are distinct from those regulating stomatal pattering.

Transcriptional repression by a bZIP protein regulates Dictyostelium prespore differentiation.

In response to the signaling polyketide DIF-1 DimB directly activates transcription of the ecmB gene in pstB cells; a subset of the prestalk cells that are the precursors of the basal disc. We show that the promoter of pspA, a prespore-specific gene, also contains a DimB binding site. Mutation of this site causes ectopic expression in the prestalk region and ChIP analysis shows that DIF-1 induces binding of DimB to the pspA promoter. DIF-1 represses pspA gene expression in a suspension cell assay but this repression is abrogated in a dimB null strain. These results suggest a coupled control mechanism, whereby the same DIF-DimB signaling pathway that directly activates ecmB gene expression directly represses pspA gene expression.

A SET/MYND chromatin re-modelling protein regulates Dictyostelium prespore patterning.

SmdA is a Dictyostelium orthologue of the SET/MYND chromatin re-modelling proteins. In developing structures derived from a null mutant for smdA (a smdA- strain), prestalk patterning is normal, but using a prespore lacZ reporter fusion, there is ectopic accumulation of beta-galactosidase in the prestalk region. As wild type slugs migrate, there is continual forward movement and re-differentiation of prespore cells into prestalk cells. Thus, a potential explanation for the ectopic reporter localization in smdA null prestalk cells is an increased rate of re-differentiation and anterior movement of prespore cells. In support of this notion, analysis of an unstable lacZ reporter, driven by the prespore promoter, reveals a normal staining pattern in the smdA- strain. We suggest that one or more genes regulated by SmdA acts to repress prespore re-specification.

DIF-1 regulates Dictyostelium basal disc differentiation by inducing the nuclear accumulation of a bZIP transcription factor.

Exposure of monolayer Dictyostelium cells to the signalling polyketide DIF-1 causes DimB, a bZIPtranscription factor, to accumulate in the nucleus where it induces prestalk gene expression. Here we analyse DimB signalling during normal development. In slugs DimB is specifically nuclear enriched in the pstB cells; a cluster of vital dye-staining cells located on the ventral surface of the posterior, prespore region. PstB cells move at culmination, to form the lower cup and the outer basal disc of the fruiting body, and DimB retains a high nuclear concentration in both these tissues. In a dimB null (dimB-) strain there are very few pstB or lower cup cells, as detected by neutral red staining, and it is known that the outer basal disc is absent or much reduced. In the dimB- strain ecmB, a marker of pstB differentiation, is not DIF inducible. Furthermore, ChIP analysis shows that DimB binds to the ecmB promoter in DIF-induced cells. These results suggest that the differentiation of pstB cells is caused by a high perceived level of DIF-1 signalling, leading to nuclear localization of DimB and direct activation of cell type-specific gene expression.

A new protein carrying an NmrA-like domain is required for cell differentiation and development in Dictyostelium discoideum.

We have isolated a Dictyostelium mutant unable to induce expression of the prestalk-specific marker ecmB in monolayer assays. The disrupted gene, padA, leads to a range of phenotypic defects in growth and development. We show that padA is essential for growth, and we have generated a thermosensitive mutant allele, padA(-). At the permissive temperature, mutant cells grow poorly; they remain longer at the slug stage during development and are defective in terminal differentiation. At the restrictive temperature, growth is completely blocked, while development is permanently arrested prior to culmination. padA(-) slugs are deficient in prestalk A cell differentiation and present an abnormal ecmB expression pattern. Sequence comparisons and predicted three-dimensional structure analyses show that PadA carries an NmrA-like domain. NmrA is a negative transcriptional regulator involved in nitrogen metabolite repression in Aspergillus nidulans. PadA predicted structure shows a NAD(P)(+)-binding domain, which we demonstrate that is essential for function. We show that padA(-) development is more sensitive to ammonia than wild-type cells and two ammonium transporters, amtA and amtC, appear derepressed during padA(-) development. Our data suggest that PadA belongs to a new family of NAD(P)(+)-binding proteins that link metabolic changes to gene expression and is required for growth and normal development.