Species‐level, metagenomic and proteomic analysis of microbe‐immune interactions in severe asthma

Background: The airway microbiome in severe asthma has not been characterised at species‐level by metagenomic sequencing, nor have the relationships between specific species and mucosal immune responses in ‘type‐2 low’, neutrophilic asthma been defined. We performed an integrated species‐level metagenomic data with inflammatory mediators to characterise prevalence of dominant potentially pathogenic organisms and host immune responses. Methods: Sputum and nasal lavage samples were analysed using long‐read metagenomic sequencing with Nanopore and qPCR in two cross‐sectional adult severe asthma cohorts, Wessex (n = 66) and Oxford (n = 30). We integrated species‐level data with clinical parameters and 39 selected airway proteins measured by immunoassay and O‐link. Results: The sputum microbiome in health and mild asthma displayed comparable microbial diversity. By contrast, 23% (19/81) of severe asthma microbiomes were dominated by a single respiratory pathogen, namely H. influenzae (n = 10), M. catarrhalis (n = 4), S. pneumoniae (n = 4) and P. aeruginosa (n = 1). Neutrophilic asthma was associated with H. influenzae, M. catarrhalis, S. pneumoniae and T. whipplei with elevated type‐1 cytokines and proteases; eosinophilic asthma with higher M. catarrhalis, but lower H. influenzae, and S. pneumoniae abundance. H. influenzae load correlated with Eosinophil Cationic Protein, elastase and IL‐10. R. mucilaginosa associated positively with IL‐6 and negatively with FGF. Bayesian network analysis also revealed close and distinct relationships of H. influenzae and M. catarrhalis with type‐1 airway inflammation. The microbiomes and cytokine milieu were distinct between upper and lower airways. Conclusions: This species‐level integrated analysis reveals central, but distinct associations between potentially pathogenic bacteria and airways inflammation in severe asthma

Suppression of protein aggregation by chaperone modification of high molecular weight complexes

Protein misfolding and aggregation are associated with many neurodegenerative diseases, including Huntington's disease. The cellular machinery for maintaining proteostasis includes molecular chaperones that facilitate protein folding and reduce proteotoxicity. Increasing the protein folding capacity of cells through manipulation of DNAJ chaperones has been shown to suppress aggregation and ameliorate polyglutamine toxicity in cells and flies. However, to date these promising findings have not been translated to mammalian models of disease. To address this issue, we developed transgenic mice that over-express the neuronal chaperone HSJ1a (DNAJB2a) and crossed them with the R6/2 mouse model of Huntington's disease. Over-expression of HSJ1a significantly reduced mutant huntingtin aggregation and enhanced solubility. Surprisingly, this was mediated through specific association with K63 ubiquitylated, detergent insoluble, higher order mutant huntingtin assemblies that decreased their ability to nucleate further aggregation. This was dependent on HSJ1a client binding ability, ubiquitin interaction and functional co-operation with HSP70. Importantly, these changes in mutant huntingtin solubility and aggregation led to improved neurological performance in R6/2 mice. These data reveal that prevention of further aggregation of detergent insoluble mutant huntingtin is an additional level of quality control for late stage chaperone-mediated neuroprotection. Furthermore, our findings represent an important proof of principle that DNAJ manipulation is a valid therapeutic approach for intervention in Huntington's disease