The COMPASS family of H3K4 methylases in Drosophila

Methylation of histone H3 lysine 4 (H3K4) in Saccharomyces cerevisiae is implemented by Set1/COMPASS, which was originally purified based on the similarity of yeast Set1 to human MLL1 and Drosophila melanogaster Trithorax (Trx). While humans have six COMPASS family members, Drosophila possesses a representative of the three subclasses within COMPASS-like complexes: dSet1 (human SET1A/SET1B), Trx (human MLL1/2), and Trr (human MLL3/4). Here, we report the biochemical purification and molecular characterization of the Drosophila COMPASS family. We observed a one-to-one similarity in subunit composition with their mammalian counterparts, with the exception of LPT (lost plant homeodomains [PHDs] of Trr), which copurifies with the Trr complex. LPT is a previously uncharacterized protein that is homologous to the multiple PHD fingers found in the N-terminal regions of mammalian MLL3/4 but not Drosophila Trr, indicating that Trr and LPT constitute a split gene of an MLL3/4 ancestor. Our study demonstrates that all three complexes in Drosophila are H3K4 methyltransferases; however, dSet1/COMPASS is the major monoubiquitination-dependent H3K4 di- and trimethylase in Drosophila. Taken together, this study provides a springboard for the functional dissection of the COMPASS family members and their role in the regulation of histone H3K4 methylation throughout development in Drosophila.     

River ecosystem resilience to extreme flood events

Floods have a major influence in structuring river ecosystems. Considering projected increases in high‐magnitude rainfall events with climate change, major flooding events are expected to increase in many regions of the world. However, there is uncertainty about the effect of different flooding regimes and the importance of flood timing in structuring riverine habitats and their associated biotic communities. In addition, our understanding of community response is hindered by a lack of long‐term datasets to evaluate river ecosystem resilience to flooding. Here we show that in a river ecosystem studied for 30 years, a major winter flood reset the invertebrate community to a community similar to one that existed 15 years earlier. The community had not recovered to the preflood state when recurrent summer flooding 9 years later reset the ecosystem back to an even earlier community. Total macroinvertebrate density was reduced in the winter flood by an order of magnitude more than the summer flood. Meiofaunal invertebrates were more resilient to the flooding than macroinvertebrates, possibly due to their smaller body size facilitating greater access to in‐stream refugia. Pacific pink salmon escapement was markedly affected by the winter flood when eggs were developing in redds, compared to summer flooding, which occurred before the majority of eggs were laid. Our findings inform a proposed conceptual model of three possible responses to flooding by the invertebrate community in terms of switching to different states and effects on resilience to future flooding events. In a changing climate, understanding these responses is important for river managers to mitigate the biological impacts of extreme flooding effects.     

piggyBacis an effective tool for functional analysis of thePlasmodium falciparumgenome

Background  

Much of thePlasmodium falciparumgenome encodes hypothetical proteins with limited homology to other organisms. A lack of robust tools for genetic manipulation of the parasite limits functional analysis of these hypothetical proteins and other aspects of thePlasmodiumgenome. Transposon mutagenesis has been used widely to identify gene functions in many organisms and would be extremely valuable for functional analysis of thePlasmodiumgenome.     

The periplasmic chaperone SurA exploits two features characteristic of integral outer membrane proteins for selective substrate recognition

The Escherichia coli periplasmic chaperone and peptidyl-prolyl isomerase (PPIase) SurA facilitates the maturation of outer membrane porins. Although the PPIase activity exhibited by one of its two parvulin-like domains is dispensable for this function, the chaperone activity residing in the non-PPIase regions of SurA, a sizable N-terminal domain and a short C-terminal tail, is essential. Unlike most cytoplasmic chaperones SurA is selective for particular substrates and recognizes outer membrane porins synthesized in vitro much more efficiently than other proteins. Thus, SurA may be specialized for the maturation of outer membrane proteins. We have characterized the substrate specificity of SurA based on its natural, biologically relevant substrates by screening cellulose-bound peptide libraries representing outer membrane proteins. We show that two features are critical for peptide binding by SurA: specific patterns of aromatic residues and the orientation of their side chains, which are found more frequently in integral outer membrane proteins than in other proteins. For the first time this sufficiently explains the capability of SurA to discriminate between outer membrane protein and non-outer membrane protein folding intermediates. Furthermore, peptide binding by SurA requires neither an active PPIase domain nor the presence of proline, indicating that the observed substrate specificity relates to the chaperone function of SurA. Finally, we show that SurA is capable of associating with the outer membrane. Together, our data support a model in which SurA is specialized to interact with non-native periplasmic outer membrane protein folding intermediates and to assist in their maturation from early to late outer membrane-associated steps.     

Pseudomonas aeruginosa polysaccharide Psl supports airway microbial community development

Pseudomonas aeruginosa dominates the complex polymicrobial cystic fibrosis (CF) airway and is a leading cause of death in persons with CF. Oral streptococcal colonization has been associated with stable CF lung function. However, no studies have demonstrated how Streptococcus salivarius, the most abundant streptococcal species found in individuals with stable CF lung disease, potentially improves lung function or becomes incorporated into the CF airway biofilm. By utilizing a two-species biofilm model to probe interactions between S. salivarius and P. aeruginosa, we discovered that the P. aeruginosa exopolysaccharide Psl promoted S. salivarius biofilm formation. Further, we identified a S. salivarius maltose-binding protein (MalE) that is required for promotion of biofilm formation both in vitro and in a Drosophila melanogaster co-infection model. Finally, we demonstrate that promotion of dual biofilm formation with S. salivarius is common among environmental and clinical P. aeruginosa isolates. Overall, our data supports a model in which S. salivarius uses a sugar-binding protein to interact with P. aeruginosa exopolysaccharide, which may be a strategy by which S. salivarius establishes itself within the CF airway microbial community.Subject terms: Bacteriology, Biofilms, Microbiome, Clinical microbiology     

Bronchiectasis-Associated Hospitalizations in Germany, 2005–2011: A Population-Based Study of Disease Burden and Trends

Background

Representative population-based data on the epidemiology of bronchiectasis in Europe are limited. The aim of the present study was to investigate the current burden and the trends of bronchiectasis-associated hospitalizations and associated conditions in Germany in order to inform focused patient care and to facilitate the allocation of healthcare resources.

Methods

The nationwide diagnosis-related groups hospital statistics for the years 2005–2011 were used in order to identify hospitalizations with bronchiectasis as any hospital discharge diagnosis according to the International Classification of Diseases, 10th revision, code J47, (acquired) bronchiectasis. Poisson log-linear regression analysis was used to assess the significance of trends. In addition, the overall length of hospital stay (LOS) and the in-hospital mortality in comparison to the nationwide overall mortality due to bronchiectasis as the primary diagnosis was assessed.

Results

Overall, 61,838 records with bronchiectasis were extracted from more than 125 million hospitalizations. The average annual age-adjusted rate for bronchiectasis as any diagnosis was 9.4 hospitalizations per 100,000 population. Hospitalization rates increased significantly during the study period, with the highest rate of 39.4 hospitalizations per 100,000 population among men aged 75–84 years and the most pronounced average annual increases among females. Besides numerous bronchiectasis-associated conditions, chronic obstructive pulmonary disease (COPD) was most frequently found in up to 39.2% of hospitalizations with bronchiectasis as the primary diagnosis. The mean LOS was comparable to that for COPD. Overall, only 40% of bronchiectasis-associated deaths occurred inside the hospital.

Conclusions

The present study provides evidence of a changing epidemiology and a steadily increasing prevalence of bronchiectasis-associated hospitalizations. Moreover, it confirms the diversity of bronchiectasis-associated conditions and the possible association between bronchiectasis and COPD. As the major burden of disease may be managed out-of-hospital, prospective patient registries are needed to establish the exact prevalence of bronchiectasis according to the specific underlying condition.     

β-Glucosidase 2 (GBA2) Activity and Imino Sugar Pharmacology

β-Glucosidase 2 (GBA2) is an enzyme that cleaves the membrane lipid glucosylceramide into glucose and ceramide. The GBA2 gene is mutated in genetic neurological diseases (hereditary spastic paraplegia and cerebellar ataxia). Pharmacologically, GBA2 is reversibly inhibited by alkylated imino sugars that are in clinical use or are being developed for this purpose. We have addressed the ambiguity surrounding one of the defining characteristics of GBA2, which is its sensitivity to inhibition by conduritol B epoxide (CBE). We found that CBE inhibited GBA2, in vitro and in live cells, in a time-dependent fashion, which is typical for mechanism-based enzyme inactivators. Compared with the well characterized impact of CBE on the lysosomal glucosylceramide-degrading enzyme (glucocerebrosidase, GBA), CBE inactivated GBA2 less efficiently, due to a lower affinity for this enzyme (higher K_I) and a lower rate of enzyme inactivation (k_inact). In contrast to CBE, N-butyldeoxygalactonojirimycin exclusively inhibited GBA2. Accordingly, we propose to redefine GBA2 activity as the β-glucosidase that is sensitive to inhibition by N-butyldeoxygalactonojirimycin. Revised as such, GBA2 activity 1) was optimal at pH 5.5–6.0; 2) accounted for a much higher proportion of detergent-independent membrane-associated β-glucosidase activity; 3) was more variable among mouse tissues and neuroblastoma and monocyte cell lines; and 4) was more sensitive to inhibition by N-butyldeoxynojirimycin (miglustat, Zavesca®), in comparison with earlier studies. Our evaluation of GBA2 makes it possible to assess its activity more accurately, which will be helpful in analyzing its physiological roles and involvement in disease and in the pharmacological profiling of monosaccharide mimetics.     

Protein microarray signature of autoantibody biomarkers for the early detection of breast cancer

Cancer patients spontaneously generate autoantibodies (AAb) to tumor-derived proteins. To detect AAb, we have probed novel high-density custom protein microarrays (NAPPA) expressing 4988 candidate tumor antigens with sera from patients with early stage breast cancer (IBC), and bound IgG was measured. We used a three-phase serial screening approach. First, a prescreen was performed to eliminate uninformative antigens. Sera from stage I-III IBC (n = 53) and healthy women (n = 53) were screened for AAb to all 4988 protein antigens. Antigens were selected if the 95th percentile of signal of cases and controls were significantly different (p < 0.05) and if the number of cases with signals above the 95th percentile of controls was significant (p < 0.05). These 761 antigens were screened using an independent set of IBC sera (n = 51) and sera from women with benign breast disease (BBD) (n = 39). From these, 119 antigens had a partial area under the ROC curve (p < 0.05), with sensitivities ranging from 9-40% at >91% specificity. Twenty-eight of these antigens were confirmed using an independent serum cohort (n = 51 cases/38 controls, p < 0.05). Using all 28 AAb, a classifier was identified with a sensitivity of 80.8% and a specificity of 61.6% (AUC = 0.756). These are potential biomarkers for the early detection of breast cancer.