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Behavioral heterogeneity in quorum sensing can stabilize social cooperation in microbial populations
Background: Microbial communities are susceptible to the public goods dilemma, whereby individuals can gain an advantage within a group by utilizing, but not sharing the cost of producing, public goods. In bacteria, the development of quorum sensing (QS) can establish a cooperation system in a population by coordinating the production of costly and sharable extracellular products (public goods). Cooperators with intact QS system and robust ability in producing public goods are vulnerable to being undermined by QS-deficient defectors that escape from QS but benefit from the cooperation of others. Although microorganisms have evolved several mechanisms to resist cheating invasion in the public goods game, it is not clear why cooperators frequently coexist with defectors and how they form a relatively stable equilibrium during evolution.
Results: We show that in Pseudomonas aeruginosa, QS-directed social cooperation can select a conditional defection strategy prior to the emergence of QS-mutant defectors, depending on resource availability. Conditional defectors represent a QS-inactive state of wild type (cooperator) individual and can invade QS-activated cooperators by adopting a cheating strategy, and then revert to cooperating when there are abundant nutrient supplies irrespective of the exploitation of QS-mutant defector. Our mathematical modeling further demonstrates that the incorporation of conditional defection strategy into the framework of iterated public goods game with sound punishment mechanism can lead to the coexistence of cooperator, conditional defector, and defector in a rock-paper-scissors dynamics.
Conclusions: These findings highlight the importance of behavioral heterogeneity in stabilizing the population structure and provide a potential reasonable explanation for the maintenance and evolution of cooperation in microbial communities.
Keywords: Conditional defection Evolution Mathematical modeling Public goods game Quorum sensing Social biology.
In order to measure compositional and functional differences between the gut microbiota of healthy and IBD-affected individuals, 231 fecal and biopsy samples were collected from the Ocean State Crohn's and Colitis Area Registry (OSCCAR) and the Prospective Registry in IBD Study at MGH (PRISM) database. OSCCAR is a state-based, prospective inception IBD cohort, and PRISM is a referral center-based, prospective IBD cohort (see Materials and methods). The samples comprised 136 fecal specimens and 95 colon or small intestinal biopsies, originating from a cross-section of 121 CD patients, 75 UC patients, 27 healthy controls, and 8 indeterminate (Table 1). In addition to general information such as gender and age, data regarding disease characteristics (topography, disease activity as measured by the Harvey-Bradshaw Index (HBI) and the Simple Colitis Activity Index), treatment (antibiotics, corticosteroids, mesalamine, immunosuppressant), and environmental exposure (tobacco use) were collected from each subject and analyzed. DNA was extracted from fecal samples and biopsies, and the 16S rRNA gene was amplified and sequenced using 454 technology. The resulting sequences were then processed using a specific in silico pipeline for sequence cleaning and phylotype assignment (see Materials and methods). At the end of this process, the average sequencing depth was 2,860 reads per sample. These data were first validated by comparison with previous work, recapitulating previously observed changes in microbial community composition during IBD and attributing several to host treatment or environment. They were subsequently associated with reference genomes in order to discover disease-associated modulations of microbial function and metabolism. A subset of 11 samples (7 healthy, 4 CD) were subjected to whole-genome shotgun sequencing using the Illumina MiSeq platform at an average depth of 119 meganucleotides per sample in order to confirm these functional inferences.
Assessing significant covariation of microbiome structure with host IBD status, treatment, and environment
We used a sparse multivariate statistical approach to relate disease phenotype to microbiome structure and function while accounting for potential correlates and confounding factors such as treatment or smoking. Metadata features potentially associated with each clade were first selected using boosting, and the significance of these associations was then assessed using a multivariate linear model with false discovery rate correction (see Materials and methods). We first investigated the resulting association of microbial clades with IBD and with features of our cohorts, testing all available metadata and clades from the genus to phylum levels. Ordination of overall relationships among samples and host status revealed several major combinations of environmental factors that co-varied with the microbiome (Figure 1 Additional file 1). For example, UC covaried in this population with mesalamine treatment, whereas CD patients were more often assessed by biopsy, treated with immunosuppressants, and enriched for Escherichia. Similarity among microbiome compositions in disease subtypes reflects those previously observed [34, 35], with ileal CD (iCD) representing a strong outgroup, UC a generally less-extreme microbial phenotype (less dissimilar from healthy subjects), and non-iCD a broad distribution of microbiome configurations.
Covariation of microbial community structure in IBD with treatment, environment, biometrics, and disease subtype. Fecal and biopsy samples from 228 IBD patients and healthy controls are plotted as squares (ileal CD) or circles (not ileal involved) and colored by disease status. Axes show the first two components of overall variation as determined by multiple factor analysis (see Materials and methods). Covariation in the presence of clinical factors (bold) and in microbial taxa (italic) is shown. Sample origin (biopsy versus stool) is the single most influential factor in determining microbial community structure, accompanied by host age, treatment types, and disease (particularly ileal CD).
An important consideration that informed the remainder of our analysis, and which is often overlooked in studies of the microbiome, was the consistent covariation among disease status, aspects of subject environment, and microbiome structure. For example, the factor most associated with changes in microbiome composition was not disease but whether the sample origin was stool or biopsy. Biopsy location induced minor changes in microbiome composition (Additional files 2, 3 and 4) relative to the extreme differences between stool and biopsy communities, in agreement with previous studies [36, 37]. In this cohort, iCD was always represented by biopsy, whereas 18.4% of non-iCD and 36% of UC samples were biopsies. iCD was also associated with greater likelihood of immunosuppressant treatment: iCD, non-iCD, and UC patients were treated by immunosuppressants in 74.4%, 19.2% and 16% of samples, respectively. In contrast, non-iCD and UC cases were more likely to be treated with mesalamine or antibiotics: mesalamine was used for 30.2% of iCD samples, 69.2% of non-iCD samples, and 77.3% of UC samples, while antibiotics were used in 2.3% of iCD, 17.9% of non-iCD, and 13.3% of UC samples. These associations lead to a range of non-independent covariates. Although disease activity may influence microbiome composition, after adjusting for the other factors, it was not independently associated with a specific shift in the microbiome composition in our analysis, and there were no significant (P < 0.01) associations between microbiome composition and gender (Additional file 5).
The second largely independent factor influencing microbiome composition was age, itself negatively associated with smoking (Figure 1 Additional file 1). Twenty-four (10.4%) of the available subjects were less than 18 years of age and 26 were 60 years or older. Aging is associated with continual changes in the microbiome, primarily a gradual decrease in Bifidobacterium as observed here (Additional file 6) and by others [38, 39]. After observing these overall patterns of covariation among disease, treatment, environment, and gut microbiome composition, we continued our analysis only after assessing the significance of microbiome-disease associations in a multivariate manner to account for host environment and treatment.
Microbial clades differentially abundant specifically in IBD include Roseburia, the Ruminococcaceae, and the Enterobacteriaceae
After adjusting for these covariates, we determined microbial clades differing significantly in abundance between healthy and IBD subjects (Figure 2a Additional file 1). This considered age, smoking, and treatment factors (immunosuppressant, corticosteroids, mesalamine, antibiotics), as well as disease activity at sampling and sample type (stool or biopsy). Two genus-level phylotypes, Roseburia and Phascolarctobacterium, were significantly reduced in both UC and CD, while Clostridium increased, all with false discovery rate q < 0.2. Roseburia is a clade XIVa Clostridia and thus associated with anti-inflammatory regulatory T cell production in the gut . Cultured Roseburia have been described as acetate utilizers and butyrate producers , while cultured Phascolarctobacterium are exclusively succinate consumers, and produce propionate when co-cultured with Paraprevotella . Thus, an IBD-associated decrease in Roseburia and Phascolarctobacterium may reflect a decrease in butyrate and propionate production.
Significant associations of microbial clade abundance and community ecology with IBD and treatment. (a) Taxonomic distribution of clades significant to disease and ileal involvement. Abundant clades not significantly associated with IBD are annotated in gray for context (top 90th percentile of at least 10% of samples and including 5+ genera). Node (non-associated clade) sizes are proportional to the log of the clade's average abundance. (b) Significance of association of sample ecology with disease (CD/UC, ileal/pancolonic), treatment (antibiotics, immunosuppression, mesalamine, steroids), and environment (smoking, stool/biopsy sample origin). Diversity (Simpson's index), evenness (Pielou's index), and richness (Chao1) were calculated for each community (see Materials and methods). False discovery rate q-values are -log10 transformed for visualization, such that values > 0.60 correspond to q < 0.25. Antibiotic treatment is strongly associated with reduced diversity, and stool samples with increased diversity relative to biopsies.
The Ruminococcaceae, which are acetate producers , were decreased in CD, while the Leuconostocaceae, which produce acetate and lactate , were decreased in UC. The only major clade with a significant increase in abundance specific to CD was the Enterobacteriaceae, specifically Escherichia/Shigella. This family has been previously implicated in intestinal inflammation [6, 45–47].
Crohn's disease with ileal involvement presents a distinct microbiome phenotype including reduced Faecalibacterium, and Odoribacter is reduced both in iCD and in pancolonic UC
In CD patients with ileal involvement, sequences of the Ruminococcaceae family and of Faecalibacterium in particular were dramatically reduced compared to other subjects (Figure 2a), confirming previous studies [48, 49]. Faecalibacterium prausnitzii, the only cultured representative of Faecalibacterium, is able to metabolize both diet-derived polysaccharides and host-derived substrates such as N-acetyl glucosamine from intestinal mucus . It is also a major butyrate producer and exhibits anti-inflammatory effects in a colitis setting . The Ruminococcaceae represent the first step of microbiome-linked carbohydrate metabolism, as they degrade several types of polysaccharides in the lower GI tract, including starch, cellulose, and xylan . The Roseburia genus, which is significantly reduced in all IBD patients (including iCD), and the Ruminococcaceae are further functionally connected in that the latter consume hydrogen and produce acetate that can be utilized by Roseburia to produce butyrate [41, 43]. Consistent reductions in all of these clades may thus have functional consequences on the ability of the host to repair the epithelium and to regulate inflammation.
The genera Escherichia/Shigella (indistinguishable as a 16S-based phylotype) were particularly highly enriched in iCD (q < 0.2 Additional file 1) above their general overabundance in CD patients. Lipopolysaccharide produced by Gram-negative bacteria such as Escherichia coli is a canonical microbe-associated molecular pattern, known to activate toll-like receptor 4 (TLR4) signaling  and thus trigger inflammatory cascades. TLR4 expression is highly up-regulated in the intestinal epithelium of IBD patients , and mutations in TLR4 are associated with both CD and UC . Previous culture-based studies have found that E. coli, specifically E. coli exhibiting pathogen-like behaviors such as adhesion and invasiveness , are more frequently cultured from iCD biopsies, and culture-independent studies have found an enrichment in E. coli that contain virulence-associated genes in iCD . This suggests that CD-involved ileum is a favorable milieu for establishment of E. coli with pathobiont features, which may have implications for IBD exacerbations and its chronicity. An inflamed ileum may furnish a specialized niche permissive for microbes with enhanced fitness in inflamed conditions.
The most severe form of UC is pancolitis, in which UC affects the entire colon this condition is associated with greatly increased risk of colon cancer . Patients with pancolitis did not harbor a clear specificity in their dysbiosis. However, both these patients and iCD patients had a reduced abundance of the Odoribacter genus, which belongs to the Porphyromonadaceae family and to the Bacteroidetes phylum. As Odoribacter splanchnus is a known producer of acetate, propionate, and butyrate , decreased Odoribacter may affect host inflammation via reduced SCFA availability.
Microbiome composition is also strongly associated with subject age, treatment, smoking, and sample biogeography
In the process of identifying microbiome perturbations specific to IBD, our multivariate model simultaneously analyzed the surprisingly diverse effects of environmental and treatment factors on GI microbial communities (see selection in Figure 3 complete data in Additional file 1). We observed a significant correlation between increasing age and decreasing Bifidobacterium (Additional file 6). The Firmicutes phylum also significantly decreased while Bacteroides increased with age in this cohort (Additional file 1) this agrees with previous studies [38, 39] and potentially reflects dietary or body mass-related changes with increasing age, which were not directly measured in these subjects, or host metabolism modifications .
Select microbial clades significantly linked to host environment and treatment. Anaerostipes decreased significantly in the gut communities of smokers, and Dorea, Butyricicoccus, and Coriobacteriaceae were among the taxa most reduced in patients receiving antibiotics (Abx). These associations were significant even in a multivariate model accounting for sample biogeography and disease status. Sqrt, square root.
Critical to determining causality in links between IBD and the gut microbiome, IBD treatments were also associated with alterations in microbiome composition. Mesalamine (5-aminosalicylic acid) is a bowel-specific aminosalicylate drug. Although its exact mode of action is unknown, it is thought to act as an antioxidant and to decrease intestinal inflammation, in part by peroxisome proliferator-activated receptor-γ (PPARγ) activation and inhibition of NFκB and pro-inflammatory eicosanoid production. Here, its use was linked to strong reductions in Escherichia/Shigella (> 100% of average abundance, q < 0.04 Additional file 7), in agreement with a recent study . Both 5-aminosalicylic acid and immunosuppressant treatment were associated with modest increases in Enterococcus, the only genus perturbed in immunosuppressant-treated patients with low false discovery rate (also > 100% of average abundance, q < 0.09).
Antibiotics were among the strongest factors associated with a reduction in ecological diversity (Figure 2b). Many individual clades were greatly reduced or nearly absent after administration of antibiotics, including the Collinsella, Dorea, Butyricicoccus, Subdoligranulum, and Acetivibrio (all q < 0.2 Additional file 1). These genera are predominantly from the Clostridiales order, Gram-positive and anaerobic bacteria that are targeted by the antibiotics commonly used in IBD, such as ciprofloxacin and metronidazole.
Smoking is likely the best-known environmental factor that impacts IBD . It is associated with increased risk of CD and is conversely protective towards developing UC . The only common organism to which tobacco usage was linked in these individuals was Anaerostipes (Firmicutes phylum), which decreased (> 60% average abundance, q < 0.15 Figure 3) in current or former tobacco users, beyond any change due solely to smokers' higher average age. The Anaerostipes genera can utilize lactate to produce butyrate , which is beneficial to colonic health.
Finally, as previously mentioned, samples of the stool as opposed to mucosal biopsies differed strongly in microbiome composition (Additional file 2). More than 70 clades were significantly over- or under-enriched in stool samples relative to biopsies at q < 0.2. This effect extended to entire phyla, as the Firmicutes were approximately twofold more abundant in stool (Additional file 1). Microbial habitat dictates the composition of microbial communities  in the GI tract, this has been suggested to occur on biogeographical scales of intestinal regions [37, 63] or even millimeters apart [64, 65], and luminal/mucosal differences may be further perturbed by bowel preparation prior to colonoscopy . The data did not suggest that the luminal and mucosal communities were independent rather, all 14 clades significantly associated with IBD retained the same trend when stratified by sample origin (Additional file 8). The fecal microbiome appeared to convey a consistent but numerically transformed function of mucosal communities, both of which shifted in composition in association with host environment, treatment, and disease.
In a closer analysis of intestinal biogeography as reflected by biopsies drawn from distinct regions, differences in most clades were modest and correlated largely with previously described changes in pH (Additional files 2, 3 and 9) . The clades with the largest regional changes included the Roseburia and Ruminococcaceae, with lower abundance in the low-pH terminal ileum, transverse, and right colon Alistipes, following a similar pattern and the Fusobacteria and Enterobacteriaceae, with an opposite pattern of somewhat increased abundance in the ileum and right colon. Particularly as the former have also been associated with the colorectal cancer microenvironment in previous work [68, 69], it is of note that these variations in the microbiota with respect to biogeography and pH are similar to those we observed with respect to IBD and potentially redox status as detailed below.
The metagenomic abundances of microbial metabolic pathways are more consistently perturbed in IBD than are organismal abundances
We continued our analysis by combining community composition with over 1,200 annotated genomes from the Kyoto Encyclopedia of Genes and Genomes (KEGG) catalog . The genes annotated within each available reference genome were used to provide an approximate gene catalog for each community (see Materials and methods), which we reconstructed into metabolic pathways (Figure 4) and smaller modules and biological processes (Figure 5 Additional file 10) as previously described . Pathway, module, and process abundances were then associated with disease and host environment using the same sparse multivariate model with which microbial abundances were assessed (Additional files 11, 12 and 13).
Microbial metabolic pathways with significantly altered abundances in the gut communities of IBD patients. Abundance of KEGG metabolic pathways in microbiome samples is colored by disease state and, when significant, stratified by ileal involvement. Basic metabolism (for example, most amino acid biosynthesis) and SCFA production were reduced in abundance in disease, while biosynthesis and transport of compounds advantageous for oxidative stress (for example, sulfur, cysteine, riboflavin) and adherence/pathogenesis (for example, secretion) were increased.
Small metabolic modules and biological processes with significantly altered abundances in the IBD microbiome. (a, b) Small (typically 5 to 20 gene) KEGG modules (a) and independently defined biological processes from the Gene Ontology (b) were assessed for significant association with disease and ileal involvement as in Figure 4. Metabolism related to oxidative stress (for example, glutathione and sulfate transport) and for pathobiont-like auxotrophy (for example, N-acetylgalactosamine and amino acid uptake) is increased, while several basic biosynthetic processes are less abundant.
Considering only the contrast between IBD (CD or UC) and healthy subjects, 24 of 200 (12%) total metabolic modules were differentially abundant at q < 0.2. This is in stark contrast to the microbial shifts discussed above, in which only 6 of 263 (2%) genus-level clades reached this significance threshold. Even in the absence of metagenomic or metatranscriptomic data and only leveraging the genes and pathways in reference genomes associated with these communities, changes in microbial function were more consistent than changes in community structure. This has been noted in environmental communities  and suggested with respect to obesity and other biometrics [73, 74], but to date it has not been reported for disease-linked dysbioses or IBD.
We validated these functional shifts by shotgun metagenomic sequencing of the small subset of available samples with appropriate stool DNA, seven healthy controls and four CD patients (Additional file 14). These were sequenced to a shallow depth averaging 119 meganucleotides per sample of 150-nucleotide paired-end Illumina MiSeq reads, reducing our effective limit of detection but otherwise providing close agreement with inferred metabolic shifts in the IBD metagenome. Of the modules highlighted below and in Figure 5, one (cobalamin biosynthesis) fell below the limit of detection, and the remaining six retained the expected trend of over- or under-enrichment in Crohn's disease, as did additional processes detailed below, including glycolysis and bacterial secretion.
Amino acid biosynthesis and carbohydrate metabolism are reduced in the IBD microbiome in favor of nutrient uptake
We observed that even basic GI microbiome metabolism was altered in both UC and CD. Amino acid metabolism showed major perturbation: genes for the metabolism and biosynthesis of nearly all amino acids (particularly histidine and lysine) decreased in abundance (Figure 4), while arginine, histidine, and lysine transport (Figure 5) gene abundance increased. In iCD we also observed a decrease in glutamine-related functional modules, which would lead to a lower amount of glutamate required for gamma-aminobutyric acid, ornithine, and arginine biosynthesis abundance of all three of these modules also decreased. In marked contrast to the other amino acids, genes for metabolism of the sulfur-containing amino acid cysteine significantly increased in abundance, with even greater increase in iCD. This corresponded with an overrepresentation of genes related to sulfate transport in UC and CD (Figure 5), and in increase in sulfur and nitrogen metabolism in CD (Figure 4).
CD was associated with increased abundance of many genes related to carbohydrate transport (Figure 5). There were large increases in pentose phosphate pathway and fructose/mannose metabolism gene abundance in iCD (Figure 4), which were accompanied by increase in carbohydrate metabolism, but they were not significant in UC and CD. In addition, iCD showed increased abundance of transporter genes for glucose, hexoses, maltose, and mono-, di-, and oligosaccharides (Figure 5). We observed a decrease in both butanoate and propanoate metabolism in iCD (Figure 4), suggesting a potential decrease in SCFA production by the microbiome, possibly due to the observed decrease in Roseburia and Faecalibacterium.
We saw an increase in glutathione transport gene abundance in UC and CD (Figure 5) and an increase in glutathione metabolism gene abundance in UC. Glutathione is a tripeptide of cysteine and glutamate, synthesized by Proteobacteria and a few streptococci and enterococci , which allows bacteria to maintain homeostasis during oxidative or acid stress. Inflammatory cascades include production of highly reactive oxygen and nitrogen metabolites, which are greatly increased in active IBD . Lamina propia monocytes also release homocysteine during inflammation, which further contributes to oxidative stress IBD is associated with higher levels of both mucosal and serum homocysteine . Thus, the increases in sulfate transport, cysteine metabolism, and glutathione metabolism may reflect a mechanism by which the gut microbiome addresses the oxidative stress caused by inflammation.
Extreme functional shifts in iCD include changes in redox metabolism, enrichment of signaling/secretion, and suggest a 'pathobiont-like' invasive metagenome
CD with ileal involvement exhibited specific dysfunction at the module level. It was associated with an increase in several modules involved in glycolysis and carbohydrate transport and metabolism (Figure 5). Conversely, iCD exhibited lower abundance of genes involved in lipid metabolism and catabolism, confirming a major imbalance in energy metabolism. We observed a global decrease in nicotinamide, purine, and pyrimidine nucleotide biosynthesis modules in iCD, CD, and UC (Figure 5).
There was a decrease in vitamin biosynthesis associated with iCD, but increases in thiamine and particularly riboflavin metabolism modules (Figure 4). Interestingly, this pathway is fed by the pentose phosphate pathway, which was also overrepresented in iCD. Riboflavin is necessary for regenerating oxidized glutathione back to its reduced form, and is thus essential for pH and oxidative stress homeostasis, as is NADPH, a product of the pentose phosphate pathway. Metabolism of the sulfur-containing amino acids cysteine and methionine was increased in iCD, in marked contrast to the IBD-associated decreases in the non-sulfur-containing amino acids such as lysine and glutamine. As homocysteine is easily convertible to methionine, this may indicate a further mechanism of maintaining redox homeostasis. Alternatively, this may be connected to the iCD-specific increase in carbohydrate metabolism, as cysteine may be metabolized to pyruvate.
Finally, genes involved in pathogenesis processes, such as secretion systems and adherence/invasion, were overrepresented in iCD (Figure 4). For example, genes involved in the shigellosis pathway were more abundant in CD, and type II secretion genes were more abundant in iCD. Type II secretion is involved in the secretion of cell wall-degrading enzymes  and the secretion of toxins such as heat-labile enterotoxin, similar to cholera toxin . These functions are typical of pathobiont adherent-invasive E. coli, which have been observed to increase in iCD in our own study and others [6, 55]. This may be associated with tissue damage, either primarily as a result of toxin secretion, or secondarily as a result of stimulated cytokine production. This tissue destruction is a likely source of metabolites for microbial overgrowth, selecting for auxotrophic specialists able to thrive in this environment and resulting in the microbiome-wide loss of basic biosynthetic processes (Figures 4 and 5). This would in turn lead to further tissue breakdown, bacterial overgrowth, and community structural and functional dysbiosis.
Recent studies using silver- or zinc-containing nanomaterials have recently suggested that the toxicity of these materials partially originates from membrane damage and disruption of ion homeostasis 52,53 . Here, by integrating transcription profiling with genome-scale constraint-based modelling, we found that the impact of radical-mediated TiO2 photocatalysis on genome-wide flux distribution and the metabolism itself is limited and thus does not directly affect PAO1 cell activity in the time frame measured. Rather, gene chip experiments and shotgun proteomics in combination with chemical species identification and quantification support the notion that PAO1 cells respond by attempting to increase their capacity to protect and repair DNA and proteins in addition to promoting the coenzyme-dependent respiratory chain, mobilisation of carbon resources and one-carbon pools, as well as the cycle of fatty acid β-oxidation (the latter being essential for maintenance of membrane integrity). Furthermore, cells react to TiO2 photocatalysis by improving membrane integrity to compensate for the strong alterations caused in the cell wall that covers the cell membrane. Additionally, we observed the activation of metabolic pathways involving production and utilisation of antioxidant coenzymes, such as (ubi)quinol (used by cytochrome oxidases) and 5,10-methenyltetrahydrofolate (produced by GlyA2 and GcvT2 proteins), with the corresponding attenuation of those producing oxidative sub-products, such as H2O2 (produced by Sox enzymes), most likely to compensate for the strong radical stress pressure inside the cell. Supplementary Fig. S3 summarises the major effects in P. aeruginosa cells exposed to TiO2 photocatalysis.
This study provides evidents that cells exposed to TiO2 photocatalysis enxibit i) rapid cell inactivation at the regulatory and signalling levels, ii) a strong decrease of the coenzyme-independent respiratory chains, iii) a lower ability to assimilate and transport iron and phosphorous and iv) a lower capacity for the biosynthesis and degradation of heme (Fe-S cluster) groups. These activities, together with the extensive cell wall modifications, are the main factors that explain the high biocidal performance of TiO2-based nanomaterials. However, it is not known which of these responses precipitate subsequent events. Future dynamic profiling assays are needed to further study these responses. Nonetheless, the fact that titania confers profound biocidal action, even at extremely low doses, under our temporally short assay conditions (2-min intervention), causes dynamic profiling at shorter and longer assay times to be technically challenge due to technical difficulties and extensive cell death and damage, respectively. It should noted that TiO2-based materials possess a high affinity for phospho-proteins and phospho-peptides 54,55 and that phosphorylation of proteins plays a role in the regulation of signalling pathways and the control of enzymatic activities by an “on/off” switch mechanism 56 . Therefore, the fact that after the initiation of TiO2-UV treatment cell lysis is rapidly induced, suggests that this material could be used in situ to bind and enrich phosphoproteins to further explore the level of post-transcriptional protein modification at given times, as a molecular measure of altered protein activities. Whatever the case, the broad-spectrum action at the genetic level illustrated herein allows an investigation of the most significant properties of the photocatalytic disinfection process: universality (i.e., the absence of weakness against any type of microorganisms) and irreversibility (i.e., avoiding cell reparation opportunities and slowing down regulatory/signalling networks). In addition, the high efficiency demonstrated against a clinically relevant pathogen indicates the adequacy of TiO2-based polymer films for a series of technological applications, such as hospital tools and furniture, but also food preservation or wastewater treatment.
Evaluation of mechanisms controlling the priming of soil carbon along a substrate age gradient
Soil organic matter (SOM) decomposition has the potential to radically affect carbon dioxide concentrations in the atmosphere. Priming, the increased decomposition of SOM after the addition of a labile carbon (C) source, may be an important regulator of SOM dynamics, yet little is known about the mechanisms of the priming effect. Two hypotheses generated in the last decade have suggested that priming is caused by either the nutrient conditions in soil or the response of the microbial community to labile C addition. We used a three million year substrate age gradient, with associated changes in nutrient availability and microbial communities, to test these two hypotheses. We added 13 C labeled glucose to soil in quantities similar to increases in root exudation that can be expected in a high carbon dioxide environment, and traced the effect of C addition on soil C, the microbial community, and soil nutrient pools and fluxes. We observed positive priming, negative priming, and no net priming depending on substrate age. Priming was most positive at the youngest sites with the smallest nitrogen (N) pools, and most negative at the site with the most available N. In contrast, we found no significant relationships between priming and phosphorus availability. Though components of the microbial community size and structure (measured by phospholipid fatty acid analysis) changed as a result of C addition, soil N availability was a better explanatory mechanism of priming effects than microbial community dynamics. Our results suggest close linkages between C and N cycles regulate the magnitude and direction of priming. If general, the stability of SOM in temperate ecosystems is likely to be governed by soil N status.
► We tested two existing hypotheses explaining the priming effect. ► We used soil from a naturally occurring nutrient gradient. ► We measured positive, negative, and no net priming effects depending on site. ► The priming effect was correlated with pools and fluxes of nitrogen. ► Priming was not correlated with changes to the available phosphorus pool or the microbial community.
Dr Andrew Edwards, Imperial College London 18/05/21
‘Antibiotics of last resort: from mechanisms to increased efficacy.’
Abstract: Daptomycin and colistin are cyclic-lipopeptide antibiotics of last resort for infections caused by Gram-positive and Gram-negative pathogens, respectively. Unfortunately, despite potent bactericidal activity in vitro, these drugs lack efficacy in vivo, which frequently results in treatment failure. Since there is a lack of new antibiotics in the development pipeline, it is increasingly important to understand the reasons for the poor in vivo efficacy of these drugs and exploit this information to develop new approaches to make these therapeutics work better. This talk will cover new insight into the mode of action of colistin and explain how this information informed the development of a novel combination therapy. This will be followed by a description of work that explains how the host environment can reduce the susceptibility of bacteria to daptomycin and how this may be overcome using a combination therapy approach using a second antibiotic.
Dr Karina Althaus, University of Tuebingen 11/05/21
'Coagulation and COVID-19 – another front in SARS-CoV-2's perfect storm.'
Dr Giulia Zanetti, Birkbeck College, University of London 27/04/21
'Cryo-electron tomography reveals the complex COPII assembly architecture.'
Abstract: COPII mediates Endoplasmic Reticulum to Golgi trafficking of thousands of cargoes. Five essential proteins assemble into a two-layer architecture, with the inner layer thought to regulate coat assembly and cargo recruitment, and the outer coat forming cages assumed to scaffold membrane curvature. Using cryo-electron tomography and subtomogram averaging on in vitro reconstituted budding reactions, we visualise the complete, membrane-assembled COPII coat and revealing the full network of interactions within and between coat layers. We demonstrate the physiological importance of these interactions using genetic and biochemical approaches. Mutagenesis reveals that the inner coat alone can provide membrane remodelling function, with organisational input from the outer coat. These functional roles for the inner and outer coats significantly move away from the current paradigm, which posits membrane curvature derives primarily from the outer coat. We suggest these interactions collectively contribute to coat organisation and membrane curvature, providing a structural framework to understand regulatory mechanisms of COPII trafficking and secretion.
Dr Gabrielle Rudenko, University of Texas 20/04/21
'The competitive world of ‘Synaptic Organizers’ – cell surface molecules implicated in neuropsychiatric disease.'
Abstract: My laboratory focuses on proteins that mediate synapse development, especially the growing class of so-called ‘synaptic organizers’. Many synaptic organizers are implicated in neuropsychiatric disorders such as autism spectrum disorder, schizophrenia, and bipolar disorder. Typically, these proteins form trans-synaptic bridges that span the synaptic cleft, the space between two neurons connected by a synapse. There they mediate adhesion between the presynaptic and postsynaptic membranes, working to facilitate proper neural connections and connect groups of select neurons into discrete neural circuits. Specific synaptic organizers also play a critical role in developing and maintaining excitatory versus inhibitory synapses which are crucial for the excitation/inhibition balance that regulates overall neuronal excitability and communication through neural circuits. While previously, synaptic organizers were thought to simply promote cell adhesion, we now know that they guide the formation of complex protein interaction networks in the synaptic cleft and work as scaffolds to organize macromolecular assemblies that modulate synaptic function. Our laboratory is using a combination of structural biology, biochemical and biophysical methods, and proteomics, to study a portfolio of different synaptic organizers implicated in neuropsychiatric disease. By elucidating structure-function relationships of key molecules that selectively guide synapse development and uniquely impact specific neural circuits, we hope to identify novel therapeutic targets that can be leveraged to design better treatments for brain disorders in future.
Dr Emilia Santos, University of Cambridge 13/04/21
'On the mechanistic basis of morphological change'
Abstract: The evolution of novel morphologies is a key component of organismal diversification, yet the genetic mechanisms and adaptive significance underlying their evolution remain understudied. We address this question in two highly diverse model systems: pigmentation patterns in cichlid fishes and cuticle structures in water striders.
Prof Helen Walden, University of Glasgow 16/03/21
"Regulation of DNA repair by monoubiquitin signals"
Abstract: The Fanconi Anemia DNA repair pathway is needed to fix DNA inter-strand crosslinks. At the heart of the pathway is a single monoubiquitin signal which is attached to two homologous proteins, FANCD2 and FANCI. Both the assembly and the removal of the signal are required for completion of inter-strand crosslink repair. My lab focusses on understanding the mechanisms of assembly, functional consequence, and removal of specific ubiquitin signals. I will present our biochemical and structural data defining each of these steps, from how a single site is targeted for modification, what the addition of ubiquitin does to the ID2 complex, how each ubiquitin signal has a distinct function, and how the signal is removed from a specific site.
Dr Vincent Lynch, University at Buffalo 09/03/21
"On the (im)possibility of elephants"
Abstract: The risk of developing cancer is correlated with body size and lifespan within species. Between species, however, there is no correlation between cancer and either body size or lifespan, indicating that large, long-lived species have evolved enhanced cancer protection mechanisms. Elephants and their relatives (Proboscideans) are a particularly interesting lineage for the exploration of mechanisms underlying the evolution of augmented cancer resistance because they evolved large bodies recently within a clade of smaller bodied species (Afrotherians). Here, we explore the contribution of gene duplication to body size and cancer risk in Afrotherians. Through comparative genomics we identified a duplicate SOD1 gene Proboscideans that we functionally characterize and show may underlie some aspects of their remarkable anti-cancer cell biology. These data suggest that duplication of tumor suppressor genes facilitated the evolution of increased body size by compensating for decreasing intrinsic cancer risk.
Prof Ehab Abouheif, University of McGill 02/03/21
"How Ants and Bacteria Became One"
Abstract: Obligate endosymbiosis, in which distantly related species integrate to form a single replicating individual, represents a major evolutionary transition in individuality. Although such transitions are thought to increase biological complexity, the evolutionary and developmental steps that lead to integration remain poorly understood. Here we show that obligate endosymbiosis between the bacteria Blochmannia and the hyperdiverse ant tribe Camponotini originated and also elaborated through radical alterations in embryonic development, as compared to other insects. The Hox genes Abdominal A (abdA) and Ultrabithorax (Ubx)—which, in arthropods, normally function to differentiate abdominal and thoracic segments after they form—were rewired to also regulate germline genes early in development. Consequently, the mRNAs and proteins of these Hox genes are expressed maternally and colocalize at a subcellular level with those of germline genes in the germplasm and three novel locations in the freshly laid egg. Blochmannia bacteria then selectively regulate these mRNAs and proteins to make each of these four locations functionally distinct, creating a system of coordinates in the embryo in which each location performs a different function to integrate Blochmannia into the Camponotini. Finally, we show that the capacity to localize mRNAs and proteins to new locations in the embryo evolved before obligate endosymbiosis and was subsequently co-opted by Blochmannia and Camponotini. This pre-existing molecular capacity converged with a pre-existing ecological mutualism to facilitate both the horizontal transfer and developmental integration of Blochmannia into Camponotini. Therefore, the convergence of pre-existing molecular capacities and ecological interactions—as well as the rewiring of highly conserved gene networks—may be a general feature that facilitates the origin and elaboration of major transitions in individuality.
Dr Pauline Scanlan, University College Cork, Ireland 23/02/21
"Antagonistic Coevolution between bacteria and phages - from in vitro models to the human gut"
Abstract: Antagonistic coevolution (AC) between bacteria and bacteriophages plays a key role in driving and maintaining microbial diversity. Consequently, AC is predicted to affect all levels of biological organisation, from the individual to ecosystem scales. Nonetheless, we know nothing about bacteria–bacteriophage AC in perhaps the most important and clinically relevant microbial ecosystem known to humankind – the human gut microbiome.
To address this gap in our knowledge I track patterns of resistance and infectivity in naturally occurring populations of bacteria and phages isolated from the human gut, together with modelling their interactions in vitro. In this talk I will focus on emerging data from our in vitro models, which suggest that the dynamics of AC may be constrained by mutation supply and costs of resistance, and that other mechanisms such as phase variation may help explain the coexistence of bacteria and phages in complex environments such as the human gut.
Prof Anna Blom, University of Lund, Sweden. 16/02/21
'Major complement evasion strategy: binding of complement inhibitors to bacterial pathogens'
Abstract: The complement system is a pivotal component of innate immunity. Composed of over 30 plasma proteins and several cellular receptors, complement aims at destruction of invading pathogens. Therefore, the majority of bacterial pathogens developed strategies aiming at inhibition of complement.
We have found that binding of human complement inhibitor C4b-binding protein is a major evasion strategy of Gram-positive Streptococcus pyogenes. Further, this binding is also crucial for development of serum resistance in Gram-negative Neisseria gonorrhoeae. Importantly, N. gonorrhoeae are highly resistant to antibiotics.
Since N. gonorrhoeae evades killing by human serum through binding of C4BP, we previously created a chimeric protein that links the first two domains of C4BP, which bind to gonococci, with Fc domain of IgM (C4BP-IgM) to enhance complement-mediated killing of bacteria. Our findings indicate that C4BP-IgM fusion protein acts as a membrane perturbing agent through complement activation on microbial surface. The rapid and intense complement deposition triggered by C4BP-IgM generates membrane attack complex (MAC) pores, which facilitate the access of antibiotics to their intracellular targets. Consequently, gonococci become more susceptible to antibiotics, which results in an accelerated and enhanced killing. Thus, our data provide insights for the use of complement activators such as C4BP-IgM fusion protein as adjuvant for antibiotic treatment of gonorrhea.
Prof Arwen Pearson, (University of Hamburg) 02/02/21
'Time-resolved structural biology - the next revolution?'
Abstract: With the arrival of ultra-bright free electron X-ray laser sources, diffraction limited synchrotron storage rings, massive automation and the resolution revolution in cryo-electron microscopy structural biology is arguably in a golden age. But even with the large numbers of macromolecules structurally characterised to date, open question remain about how exactly structure is linked to function. Attention is now shifting to adding dynamics to our understanding of structure-function relationships, but time-resolved experiments remain technically challenging and thus are often viewed as a niche technique, where a few specialist groups investigate a subset of well-behaved proteins. In this seminar I will present some of the newest methods and tools for such experiments and show how time-resolved structural studies are becoming within reach of the broader community of structural biologist and molecular cell biologists interested in mechanism.
Dr Mark Webber (Quadram Institute, UK) 12/01/21
“How to understand and predict routes to antimicrobial resistance”
The development of antimicrobial resistance by bacteria remains one of the great challenges to global health. Development of resistance usually involves a change to the DNA content of the strain in question. Development of resistance can however result in a change to the fitness of the organism and the ultimate fate of a resistant organism is dictated by more than just its ability to survive exposure to an antibiotic. Our research is interested in understanding how resistance emerges and evolves under pressure over time. We aim to identify the wider complement of genes involved in defining susceptibility to a stress and studying how development of resistance impacts bacterial fitness. This talk will describe how we have developed a biofilm evolution model to study these issues.
Dr Sarah Coulthurst, (University of Dundee) 08/12/20
"How to kill your rivals: microbial warfare mediated by the Type VI secretion system”
Protein secretion systems are specialised macromolecular machines used to translocate specific proteins out of the bacterial cell, where they can be either released to the external environment or injected into other cells. Secretion systems, and the diverse proteins they secrete, mediate the interaction of bacterial cells with the environment, eukaryotic cells or other bacteria. The Type VI secretion system (T6SS) is a key weapon in the competitiveness and virulence of very many Gram-negative bacteria. Whilst some T6SSs are ‘anti-eukaryotic’, used to target host cells as classical virulence factors, the majority appear to be ‘anti-bacterial’, used to efficiently kill rival bacterial cells and provide a competitive advantage in a variety of polymicrobial niches. T6SSs deliver multiple, diverse toxins (‘effectors’) directly into target cells by a contraction-based firing mechanism. We have used the potent anti-bacterial T6SS of the opportunistic pathogen Serratia marcescens as a model to study the mechanisms and consequences of T6SS effector delivery into competitors. This has revealed a varied portfolio of effector toxins and cognate immunity proteins, used for several forms of inter-microbial competition. In particular, we have characterised several new anti-bacterial effectors and also discovered that the S. marcescens T6SS not only targets bacterial competitors, but can also deploy anti-fungal effector proteins against microbial fungi. Our findings have contributed to the growing appreciation that the role of the T6SS in shaping polymicrobial communities is both important and broad.
Dr Daniel Hebenstreit, (University of Warwick) 01/12/20
"Does RNA polymerase obey lockdown? Spatial and temporal dynamics of Pol2"
Transcription is central to all life, but many things remain unclear. In particular, RNA polymerase II (Pol2) mediated transcription of mRNAs in mammalian systems is characterized by relatively complex, enigmatic phenomena transcripts are made in burst-like fashion rather than at a constant rate, Pol2 appears to cluster at transcriptional hotspots, and the genome is subject to 3D interactions.
Using a broad range of techniques including next gen sequencing, genome-wide analyses, single molecule imaging, and modelling approaches, we investigated how these observations can be integrated into a consistent overall picture of transcription.
Our results reveal mechanistic links between Pol2 localization, transcriptional dynamics and nuclear topology.
Prof Edel O'Toole, (Queen Mary University of London) 24/11/20
"Insights into cell biology from rare skin diseases"
Edel O’Toole is a clinical academic at Barts and the London School of Medicine and Dentistry with an active research group working on rare genetic skin disease biology. She trained in Medicine at University College, Galway, Ireland, followed by general medical and dermatology training in Dublin and London. She was a Howard Hughes Medical Institute Post-Doctoral Fellow with David Woodley at Northwestern University in Chicago from 1994-1998. Her main clinical interests are ichthyosis and palmoplantar keratodermas. She is the current clinical lead for the British Association of Dermatologists Dermatology and Genetic Medicine network. She is also on the steering committee of Pachyonychia Project and is actively involved in 100K Genomes, a gene discovery project within the NHS. In her talk, Professor O'Toole discussed 2 current areas of research interest in her lab: 1) How loss of basement membrane type VII collagen leads to aggressive skin cancer in the rare blistering disorder recessive dystrophic epidermolysis bullosa. 2) Identification of novel therapeutic targets in one of the most severe disorders of the skin barrier, harlequin ichthyosis.
Dr Peter Oliver, (MRC Harwell Institute) 10/03/2020
"Investigating the dual role of TLDc proteins in neurodevelopment, neurodegeneration and neuroprotection"
There is still a great deal to learn regarding the molecular mechanisms that underlie neuronal cell death and dysfunction in disease, in particular the selective vulnerability of cells in the brain - a common theme across many major neurological disorders. Cells have evolved a range of endogenous detection and defence mechanisms against stressful events, and there is hope that these pathways could be exploited in the future for therapeutic benefit.
Oxidation resistance 1 (OXR1) was originally identified from a screen for genes that could prevent oxidative DNA damage, and most recently, disruption of OXR1 has been implicated in human neurodevelopmental disorders characterised by epilepsy and cerebellar ataxia. We have demonstrated that over-expression of Oxr1 can effectively protect against oxidative stress-induced cell death in neuronal cells, while expressing higher levels of Oxr1 in vivo is able to reduce and delay neurodegeneration and neuroinflammation in two independent mouse model of amyotrophic lateral sclerosis (ALS). Conversely, disruption of Oxr1 in the mouse leads to region-specific neurodegeneration and ataxia, as observed in patients. Interestingly, many fundamental properties of this protein appear to be conserved in evolution, from mammals to flies and plants.
Oxr1 contains the TLDc domain, a motif present in a family of proteins including TBC1 domain family member 24 (TBC1D24), a protein mutated in a range of disorders characterised by seizures, hearing loss and neurodegeneration. The TLDc domain itself is highly conserved across species, although the structure-function relationship is unknown. To understand the role of this domain in the stress response and disease, we are carrying out systematic functional analysis of TLDc domain-containing proteins as well as continuing to investigate their neuroprotective properties in mouse models of neurodegeneration. We have also revealed for the first time the essential role of the epilepsy-associated TLDc family member TBC1D24 at the mammalian synapse and are now focussing on a new function for TLDc proteins in vesicular trafficking, combining electrophysiology and three-dimensional structural studies of the synapse with cell biology and biochemical approaches.
Peter graduated from the University of Bath in 1996 (Biochemistry), followed by a PhD at the MRC National Institute for Medical Research and post-doctoral positions in the MRC Functional Genomics Unit, Oxford. In 2013 Peter was awarded a European Research Council Consolidator Grant to establish his own independent group in the Department of Physiology, Anatomy and Genetics at The University of Oxford, and in 2018 moved his group to the MRC Harwell Institute.
Professor Eshwar Mahenthiralingam, (University of Cardiff) 03/03/2020
"Burkholderia bacteria: the ugly, the bad and now the good?"
Burkholderia are a diverse group of antimicrobial resistant Gram-negative bacteria. This talk will cover their ugly and bad traits as transmissible and virulent lung infections in people with cystic fibrosis, plant pathogens, and as problematic contaminants for industrial manufacture. This will be contrasted to the good side of Burkholderia bacteria that within the natural environment can protect plants and insects from attack by fungal pathogens, degrade a range of man-made pollutants, and more recently have been harnessed as producers of novel antibiotics. Further information on the contrasting roles of Burholderia as potential biopesticides versus opportunistic pathogens can be obtained from a recent blog (https://naturemicrobiologycommunity.nature.com/users/207716-eshwar-mahenthiralingam/posts/43397-natural-bacterial-biopesticides-weighing-up-the-risk-of-pathogenic-versus-beneficial-properties)
Professor Ana Caicedo, (University of Massachusetts Amherst) 25/02/2020
"Crops, weeds and wild plants: leveraging the agricultural environment for insight into plant adaptation."
Dr Ellie Harrison, (University of Sheffield)18/02/2020
"Living with bacteriophages - lessons from the lab and from the wild."
Bacteria exist in hugely diverse communities and engage in interactions not just with other species - but also with a menagerie of genetic elements, like plasmids and phages, which infect, kill, benefit and manipulate their bacterial hosts. Interactions between these elements and their bacterial hosts play an important part in shaping the ecology of microbial communities and driving bacterial evolution. A key tool for understanding these interactions has been the use of experimental evolution understanding evolutionary dynamics in simplified, highly controlled communities but limitations on what we can learn from 'abstract' experimental systems is driving a shift to expand this approach into complex, (more) natural microbial communities. I will present my work showing how genetic elements can alter the outcome of bacterial evolution in the lab, as well as introduce new work to understand bacterial - genetic element interactions in the wild.
Professor Sally Ward, (University of Southampton) 11/02/2020
"Targeting subcellular trafficking behaviour for the design of therapeutic antibodies"
The central role of FcRn in regulating IgG persistence and transport provides opportunities for targeting this receptor in multiple different diagnostic and therapeutic situations. The engineering of IgGs with higher affinity for FcRn can be used to produce antibodies with longer in vivo half-lives due to increased recycling within cells, but only if the pH dependence of the IgG-FcRn interaction is retained. Conversely, engineered IgGs with higher affinity for FcRn at both acidic and near neutral pH act as potent inhibitors of FcRn and drive wild type IgG into lysosomes. Consequently, such antibodies (‘Abdegs’, for antibodies that enhance IgG degradation) can lower the levels of endogenous IgG, providing a pathway for the treatment of antibody-mediated autoimmunity. In addition, we have recently generated engineered Fc-antigen fusions that selectively deliver antigen-specific antibodies into lysosomes (called ‘Seldegs’, for selective degradation).
We have also generated engineered, tumour-specific antibodies with altered endosomal trafficking behaviour. Following conjugation to cytotoxic drugs to form antibody-drug conjugates (ADCs), these antibodies are more effective in delivering their toxic payload to target cells, resulting in a potential strategy to circumvent the dose-limiting toxicities that frequently reduce the therapeutic efficacy of current ADCs.
Professor Katie Peichel (University of Bern) 04/02/2020
"Genetics of adaptation: the roles of pleiotropy and linkage"
Despite recent progress, relatively little is known about the specific genetic and molecular changes that underlie adaptation to new environments. Stickleback fish have been at the forefront of research to uncover the genetic and molecular architecture that underlies adaptation and speciation. A wealth of quantitative trait locus (QTL) mapping studies in sticklebacks has provided insight into the distribution of effect sizes during adaptation and has also revealed that several regions of the genome contain more loci than expected for traits involved in adaptation. It is unknown whether these trait clusters result from tight physical linkage of multiple genetic changes responsible for different traits, or from a single genetic change with pleiotropic effects. I will discuss recent research in my group that is focused on disentangling the roles of pleiotropy and linkage in adaptation, using both genome-wide approaches and more focused studies of specific loci with a major effect on adaptation.
Professor Deborah Mackay, (University of Southampton) 10/12/2019
"Genetic Epigenetics: Lessons from Imprinting Disorders"
Prof. Mackay's work has lead to the discovery of novel (epi)genetic disorders that have informed our understanding of the mechanisms underlying imprinted gene expression. Prof. Mackay is Professor of Medical Epigenetics within Medicine at the University of Southampton, and laboratory lead of the Wessex Imprinting Group at Salisbury NHS Trust.
Dr Ronald Jenner, (Natural History Museum) 05/11/2019
"Of complex cocktails and borrowed bullets: the evolution of centipede venom composition"
Venoms have evolved many times in the animal kingdom. Centipedes are one of the oldest groups of venomous land animals. They have a pair of claw-like appendages bearing venom glands, and they use their powerful venoms for predation and defense. Although all centipedes are venomous, our understanding of centipede venoms is based almost exclusively upon a handful of species from 1 of the 5 centipede orders. In this talk I present our research on the evolution of this toxic weapon based on proteotranscriptomic data from all 5 orders. Two of our most surprising findings are that centipedes have evolved complex venoms in parallel multiple times, and that horizontal gene transfer seems to have made important contributions to these toxic arsenals.
Dr Alex Brand (University of Exeter) 29/10/2019
"Understanding the enemy: new perspectives in the pathogenic fungus, Candida albicans"
Candida albicans is a commensal yeast that is carried asymptomatically by most people. However, C. albicans can cause mucosal infections in women of child-bearing age, the elderly, neonates and AIDS patients. Although not life-threatening, billions of people worldwide suffer these irritating infections. In patients undergoing immunosuppressive treatments, C. albicans causes around 200,000 life-threatening bloodstream infections a year, with up to 40 % mortality. In both types of disease, the transformation of yeast to the filamentous hyphal form is a key virulence trait. Blood-born hyphae can invade virtually any human body site to cause inflammation, sepsis and organ failure. My research focuses on the regulation of hyphal growth at the molecular level, and how this translates into defined responses to the physical environment. We combine genetic manipulation and live-cell imaging with a number of applied physical stimuli, such as electric fields and microfabricated topographies, to understand how hyphae detect and respond to their environment. We have identified a key role for a Ras-like GTPase, Rsr1 (human RAP1) in the spatial organisation of apical dominance and cell directionality in hyphae and we have shown that Rsr1 and a Paxillin-like protein are required for contact-dependent behavioural responses. These responses require calcium influx and we have recently developed the first live-cell calcium reporter in C. albicans. This has shown that pH, osmotic and oxidative stress elicit specific signalling and recovery signatures. Together, these approaches are revealing cell processes and behaviours that are important for fungal cell integrity and invasive hyphal growth.
Dr Ian Henderson, (University of Cambridge) 22/10/2019
"Genetic and Epigenetic Control of Recombination in Plant Genomes"
The majority of plants, animals and fungi reproduce via meiosis, which has a profound effect on genetic diversity and adaptation. During meiosis homologous chromosomes pair and recombine, which creates new combinations of alleles. Interestingly, the rate of recombination is highly variable along chromosomes, with hotspots and coldspots. We are interested in defining the mechanisms that cause variation in recombination frequency and the implications this has for genome evolution. I will present our research investigating the roles of genetic and epigenetic factors that shape recombination in plant genomes. This will include discussion of both the role of chromatin in shaping recombination, and mechanisms by which genetic variation can feedback onto the recombination process. Ultimately, I hope that our work will shed light on the enigmatic role of recombination in genome and species evolution.
Dr Roger Williams (University of Cambridge) 08/10/2019
"Structural mechanisms of regulation of enzymes in nutritional sensing"
Eukaryotic cells balance anabolic and catabolic pathways in order to survive in changing environments. The eIF2alpha kinase GCN2 is activated by amino acid starvation, while the protein kinase mTORC1 is activated when amino acids are replete. These nutrition-sensing pathways are linked through their influences on autophagy and endocytic traffic, two broad sorting pathways that are dependent on signalling by lipid second messengers. Our work on GCN2, mTORC1 and the lipid kinase VPS34 is aimed at understanding how these enzymes are regulated in control of growth, pathogenesis and cancer. We have applied a synthesis of hydrogen/deuterium exchange mass spectrometry (HDX-MS), X-ray crystallography, single particle electron cryo-microscopy (Cryo-EM) and electron cryo-tomography (cryo-ET) to understand these mechanisms.
The complex mTORC1 controls cell proliferation by integrating growth factor signals with nutritional availability. Regulation of mTORC1 is controlled by two types of switch-like G-proteins that associate with it, the Rag heterodimers and RHEB. Structures and dynamics of a complex of mTORC1 with active Rags and RHEB explain how oncogenic mutants activate the complex by recruiting it to lysosomes and by allosteric changes in its conformation. An enzyme complex containing phosphatidylinositol 3-kinase (PI3K) VPS34 is also part of the mechanism of mTORC1 activation. Cryo-ET and HDX-MS have helped illuminate the role of another switch-like G-protein, Rab5, in stimulating a VPS34-containing complex involved in endocytic sorting.
While mTORC1 is activated by amino acid abundance and promotes protein translation, nutrient stress activates GCN2, which initiates the Integrated Stress Response (ISR) and inhibits general translation. Human GCN2 is potently stimulated by ribosomes, and using HDX-MS we showed that GCN2 recognises domain II of the uL10 subunit of the ribosomal P-stalk.
Localization and function
AP-2 transcription factors are localized predominantly in the nucleus, where they bind to target sequences and regulate transcription of target genes. AP-2 proteins have also been shown to interfere with other signal transduction pathways for example, it has been proposed that they modulate the pathway downstream of the developmental signaling molecule Wnt by associating with the Adenomatous polyposis coli (APC) tumor suppressor protein in the nucleus .
The activity of AP-2 proteins can be controlled at multiple levels: their transactivation potential, their DNA binding, their subcellular localization [17–19] and their degradation [20, 21] can all be modified. Mechanisms of regulation include post-translational modifications, such as protein kinase A-mediated phosphorylation [22, 23], sumoylation  and redox regulation [25, 26], as well as physical interaction with various proteins (see Table 3 for a comprehensive list). Interacting proteins either modulate the activity of AP-2 proteins or are influenced in their function by binding to AP-2 proteins.
The tissue distribution and developmental functions of AP-2 transcription factors have been studied extensively in several species. Drosophila AP-2 (dAP-2) is expressed in the maxillary segment and neural structures during embryogenesis, and in the central nervous system (CNS) and the leg, antennal and labial imaginal disks during larval development [27, 28]. Mutation of the dAP-2 gene leads to defects in proboscis development and leg-joint formation [29, 30].
The multiple overlapping and diverging expression patterns of AP-2 family proteins suggest that, following the expansion of the family during vertebrate evolution, redundant and non-redundant functions of the individual AP-2 family members evolved. Although the single AP-2 protein in the cephalochordate amphioxus is expressed mainly in non-neuronal ectoderm, in the lamprey, a primitive vertebrate, AP-2 has co-opted a second expression domain, the neural crest . The single AP-2 homolog described so far in Xenopus is expressed in the epidermis and neural crest and has been shown to be critical for the development of these structures [7, 31–33]. In zebrafish, the two AP-2 family members, tfap2a and tfap2b , are coexpressed in the neural tube, the ectoderm and the pronephric ducts of the developing kidney, but only tfap2a is expressed in neural crest cells [35, 36]. Positional cloning revealed that the zebrafish point mutants named mont blanc  and lockjaw  encode tfap2a the mutant animals display impaired development of neural-crest derivatives, such as the facial skeleton, the peripheral nervous system and pigment cells [37, 38]. It is also interesting to note that AP-2 proteins are expressed in the primitive ectoderm of both invertebrates and vertebrates, suggesting an evolutionarily conserved role for the family in the formation of this tissue.
In mice, three of the five AP-2 family members (AP-2α, AP-2β and AP-2γ) are coexpressed in neural-crest cells, the peripheral nervous system, facial and limb mesenchyme, various epithelia of the developing embryo and the extraembryonic trophectoderm [2, 39–41]. AP-2δ expression is restricted mainly to the developing heart, CNS and retina , whereas AP-2ε expression is detected in cells of the olfactory bulb [3, 4]. Despite the overlapping expression patterns of AP-2α, AP-2β and AP-2γ, disruption of these AP-2 genes reveals non-redundant roles during development. Mutation of AP-2α predominantly affects the cranial neural crest and the limb mesenchyme, leading to disturbances of facial and limb development in a manner reminiscent of the defects described in dAP2 mutant flies [42, 43]. AP-2β and AP-2γ, on the other hand, are essential for kidney development [44, 45] or placentation of the embryo [46, 47], respectively. In humans, mutations generating a dominant negative allele of AP-2β have been shown to be the cause of Char syndrome (Online Mendelian Inheritance in Man (OMIM) ID 169100 ) the hallmarks of this syndrome are patent ductus arteriosus (abnormal persistence of a normal fetal heart structure after birth) with facial dysmorphism and abnormal fifth digits [49, 50].
Comparing all mutant phenotypes, it can be seen that loss of AP-2 transcription factor activity generally impairs proliferation and induces premature differentiation and/or apoptosis in various cell types during development. This conclusion is further substantiated by results from a screen for AP-2α target genes  and supported by gain-of-function studies in Xenopus and mice [31, 52, 53]. As uncontrolled proliferation leads to malignancies, AP-2 transcription factors are not only implicated in normal development, but also seem to be involved in cellular neoplasia, and enhanced AP-2 levels have been reported in various types of cancer [19, 54–60]. In a murine breast-cancer model, tumor progression is enhanced after transgenic overexpression of AP-2γ . Thus, AP-2 proteins can be viewed as gatekeepers controlling the balance between proliferation and differentiation during embryogenesis.
P53: the ultimate tumor suppressor gene?
Alterations in the gene encoding the cellular p53 protein are perhaps the most frequent type of genetic lesions in human cancer. At the heart of these alterations is the abrogation of the tumor suppressor activity of the normal p53. In many cases this is achieved through point mutations in p53, which often result in pronounced conformational changes. Such mutant polypeptides, which tend to accumulate to high levels in cancer cells, are believed to exert a dominant negative effect over coexpressed normal p53. Extensive research on p53, especially in the course of the last 3 years, has already provided much insight into the biological and biochemical mechanisms that underlie its capacity to act as a potent tumor suppressor. There are now many indications that p53 may play a central role in the control of cell proliferation, cell survival, and differentiation. Nevertheless, despite the purported importance of p53 for such crucial processes, mice can develop apparently without any defect in the total absence of p53. This raises the possibility that p53 may become critically limiting only when normal growth control is lost.— Oren, M. p53: the ultimate tumor suppressor gene? FASEB J. 6: 3169-3176 1992.
Obg and Membrane Depolarization Are Part of a Microbial Bet-Hedging Strategy that Leads to Antibiotic Tolerance
Within bacterial populations, a small fraction of persister cells is transiently capable of surviving exposure to lethal doses of antibiotics. As a bet-hedging strategy, persistence levels are determined both by stochastic induction and by environmental stimuli called responsive diversification. Little is known about the mechanisms that link the low frequency of persisters to environmental signals. Our results support a central role for the conserved GTPase Obg in determining persistence in Escherichia coli in response to nutrient starvation. Obg-mediated persistence requires the stringent response alarmone (p)ppGpp and proceeds through transcriptional control of the hokB-sokB type I toxin-antitoxin module. In individual cells, increased Obg levels induce HokB expression, which in turn results in a collapse of the membrane potential, leading to dormancy. Obg also controls persistence in Pseudomonas aeruginosa and thus constitutes a conserved regulator of antibiotic tolerance. Combined, our findings signify an important step toward unraveling shared genetic mechanisms underlying persistence.
Rhizobacteria that produce auxins and contain 1-amino-cyclopropane-1-carboxylic acid deaminase decrease amino acid concentrations in the rhizosphere and improve growth and yield of well-watered and water-limited potato (Solanum tuberosum)
A.A. Belimov, Laboratory of Rhizosphere Biology, All-Russia Research Institute for Agricultural Microbiology, Podbelskogo Sh. 3, Pushkin, 196608 St. Petersburg, Russian Federation. Email: [email protected]
Lancaster Environment Centre, Lancaster University, Lancaster, UK
Laboratory of Rhizosphere Biology, All-Russia Research Institute for Agricultural Microbiology, St. Petersburg, Russian Federation
Laboratory of Rhizosphere Biology, All-Russia Research Institute for Agricultural Microbiology, St. Petersburg, Russian Federation
Laboratory of Rhizosphere Biology, All-Russia Research Institute for Agricultural Microbiology, St. Petersburg, Russian Federation
A.A. Belimov, Laboratory of Rhizosphere Biology, All-Russia Research Institute for Agricultural Microbiology, Podbelskogo Sh. 3, Pushkin, 196608 St. Petersburg, Russian Federation. Email: [email protected]bler.ru
Lancaster Environment Centre, Lancaster University, Lancaster, UK
Plant-growth-promoting rhizobacteria (PGPR) utilise amino acids exuded from plant root systems, but hitherto there have been no direct measurements of rhizosphere concentrations of the amino acid 1-amino-cyclopropane-1-carboxylic acid (ACC) following inoculation with PGPR containing the enzyme ACC deaminase. When introduced to the rhizosphere of two potato (Solanum tuberosum) cultivars (cv. Swift and cv. Nevsky), various ACC deaminase containing rhizobacteria (Achromobacter xylosoxidans Cm4, Pseudomonas oryzihabitans Ep4 and Variovorax paradoxus 5C-2) not only decreased rhizosphere ACC concentrations but also decreased concentrations of several proteinogenic amino acids (glutamic acid, histidine, isoleucine, leucine, phenylalanine, serine, threonine, tryptophan, tyrosine, valine). These effects were not always correlated with the ability of the bacteria to metabolise these compounds in vitro, suggesting bacterial mediation of root amino acid exudation. All rhizobacteria showed similar root colonisation following inoculation of sand cultures, thus species differences in amino acid utilisation profiles apparently did not confer any selective advantage in the potato rhizosphere. Rhizobacterial inoculation increased root biomass (by up to 50%) and tuber yield (by up to 40%) in pot trials, and tuber yield (by up to 27%) in field experiments, especially when plants were grown under water-limited conditions. Nevertheless, inoculated and control plants showed similar leaf water relations, indicating that alternative mechanisms (regulation of phytohormone balance) were responsible for growth promotion. Rhizobacteria generally increased tuber number more than individual tuber weight, suggesting that accelerated vegetative development was responsible for increased yield.