Studying ggdef domain in the act: Minimize conformational frustration to prevent artefacts

GGDEF-containing proteins respond to different environmental cues to finely modulate cyclic diguanylate (c-di-GMP) levels in time and space, making the allosteric control a distinctive trait of the corresponding proteins. The diguanylate cyclase mechanism is emblematic of this control: two GGDEF domains, each binding one GTP molecule, must dimerize to enter catalysis and yield c-di-GMP. The need for dimerization makes the GGDEF domain an ideal conformational switch in multidomain proteins. A re-evaluation of the kinetic profile of previously characterized GGDEF domains indicated that they are also able to convert GTP to GMP: this unexpected reactivity occurs when conformational issues hamper the cyclase activity. These results create new questions regarding the characterization and engineering of these proteins for in solution or structural studies

Molecular insights into RmcA-mediated c-di-GMP consumption: Linking redox potential to biofilm morphogenesis in Pseudomonas aeruginosa

The ability of many bacteria to form biofilms contributes to their resilience and makes infections more difficult to treat. Biofilm growth leads to the formation of internal oxygen gradients, creating hypoxic subzones where cellular reducing power accumulates, and metabolic activities can be limited. The pathogen Pseudomonas aeruginosa counteracts the redox imbalance in the hypoxic biofilm subzones by producing redox-active electron shuttles (phenazines) and by secreting extracellular matrix, leading to an increased surface area-to-volume ratio, which favors gas exchange. Matrix production is regulated by the second messenger bis-(3′,5′)-cyclic-dimeric-guanosine monophosphate (c-di-GMP) in response to different environmental cues. RmcA (Redox modulator of c-di-GMP) from P. aeruginosa is a multidomain phosphodiesterase (PDE) that modulates c-di-GMP levels in response to phenazine availability. RmcA can also sense the fermentable carbon source arginine via a periplasmic domain, which is linked via a transmembrane domain to four cytoplasmic Per-Arnt-Sim (PAS) domains followed by a diguanylate cyclase (DGC) and a PDE domain. The biochemical characterization of the cytoplasmic portion of RmcA reported in this work shows that the PAS domain adjacent to the catalytic domain tunes RmcA PDE activity in a redox-dependent manner, by differentially controlling protein conformation in response to FAD or FADH2. This redox-dependent mechanism likely links the redox state of phenazines (via FAD/FADH2 ratio) to matrix production as indicated by a hyperwrinkling phenotype in a macrocolony biofilm assay. This study provides insights into the role of RmcA in transducing cellular redox information into a structural response of the biofilm at the population level. Conditions of resource (i.e. oxygen and nutrient) limitation arise during chronic infection, affecting the cellular redox state and promoting antibiotic tolerance. An understanding of the molecular linkages between condition sensing and biofilm structure is therefore of crucial importance from both biological and engineering standpoints.The authors would like to acknowledge Sapienza University of Rome [RM120172A7AD98EB to SR, RM1221815D52AB32 to APaiardini and AR12117A63EE6037; AR2221816C44C7B3 to CSR] for financial support. AUC experiments have received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 101004806. We thank Patrick England of the Plateforme de Biophysique Moléculaire of the C2RT (Institut Pasteur) for fruitful discussion

The association between misperceptions around weight status and quality of life in adults in Australia

Objective: Limited evidence supports a possible association between a person’s perception of their weight status and their quality of life (QoL). This study evaluates whether misperception around weight status is associated with QoL and the impact of gender on this association. Methods: A cross-sectional survey of Australian adults (n=1,905 analysed) collected selfreported height and weight (used to estimate BMI), gender and QoL (described using the AQoL-8D). Participants reported whether they perceived their weight status to be ‘underweight’, ‘healthy weight’, ‘overweight’ or ‘obese’. Misperception around weight status was categorised based on perceived weight status and self-reported BMI. Ordinary least squares regression was used to test associations between self-reported overall, physical and psychosocial QoL, misperception of weight status, and gender, across different BMI categories, after controlling for income, education, relationship status and health conditions. Results: Compared to accurate perception, underestimation of weight status was associated with higher overall QoL for obese males and females and for overweight males. Overestimation of weight status was associated with higher overall QoL for underweight females and lower overall QoL for healthy weight males and females. The same pattern was seen for psychosocial QoL. Physical QoL was less sensitive to misperception than psychosocial QoL. Conclusions: Self-reported misperception around weight status is associated with overall, psychosocial and to a lesser extent physical QoL in Australian adults, although its role depends on BMI category and gender. Generally misperception in the direction of “healthy weight” is associated with higher QoL and overestimation of weight status by those who are of healthy weight is associated with lower QoL. Findings should be confirmed in datasets that contain measured as opposed to self-report height and weight

Do the Guidelines for Adolescent Preventive Services (GAPS) Facilitate Mental Health Diagnosis?

Objective: To determine if the Guidelines for Adolescent Preventive Services (GAPS) increases detection or shortens time to diagnosis of mental health (MH) disorders, particularly adolescent depression. Methods: Starting in May 1999, GAPS questionnaires were routinely administered at adolescent annual visits at 1 primary care clinic in a rural health network. Using an administrative database, we enumerated all MH diagnostic codes for outpatient visits of adolescents aged 13 to 15 years. Population based rates were derived using school enrollment data. Using time series, the rates of MH diagnoses were compared pre- and post-GAPS. Using survival analysis, the time to any MH diagnosis subsequent to index annual visits was also compared pre- and post-GAPS. Because the GAPS questionnaire includes questions for depressed mood, anhedonia, and suicidality, ICD-9-CM codes for depression and mood disorder were also analyzed separately. Results: Time series analysis included 8112 adolescents. The rate of MH diagnosis did not change pre- and post-GAPS ( P = .13). Time to any MH diagnosis was similar pre-GAPS (9.0 months) and post-GAPS (7.0 months, log rank P = .30). Time to any first diagnosis of depression or mood disorder was similar post-GAPS (12.2 months) versus pre-GAPS (11.0 months, log rank P = .34). Conclusions: Use of the GAPS was not associated with change in the rate of or time to MH diagnosis. Our results challenge the prevalent expectation that requiring mental health screening will reduce unmet need for MH treatment. Validated MH screening tools, primary care provider training, and access to MH services may also be needed but further study is required

Nutrient sensing and biofilm modulation: the example of L-arginine in Pseudomonas

Bacterial biofilm represents a multicellular community embedded within an extracellular matrix attached to a surface. This lifestyle confers to bacterial cells protection against hostile environ-ments, such as antibiotic treatment and host immune response in case of infections. The Pseudomonas genus is characterised by species producing strong biofilms difficult to be eradicated and by an extraordinary metabolic versatility which may support energy and carbon/nitrogen assimilation under multiple environmental conditions. Nutrient availability can be perceived by a Pseudomonas biofilm which, in turn, readapts its metabolism to finally tune its own formation and dispersion. A growing number of papers is now focusing on the mechanism of nutrient perception as a possible strategy to weaken the biofilm barrier by environmental cues. One of the most important nutrients is amino acid L-arginine, a crucial metabolite sustaining bacterial growth both as a carbon and a nitrogen source. Under low-oxygen conditions, L-arginine may also serve for ATP production, thus allowing bacteria to survive in anaerobic environments. L-arginine has been associated with biofilms, virulence, and antibiotic resistance. L-arginine is also a key precursor of regulatory molecules such as polyamines, whose involvement in biofilm homeostasis is reported. Given the biomedical and biotechnological relevance of biofilm control, the state of the art on the effects mediated by the L-arginine nutrient on biofilm modulation is presented, with a special focus on the Pseudomonas biofilm. Possible biotechnological and biomedical applications are also discussed