Biology subjects

Rapp, A.

Publications and source records attributed to Rapp, A..

3 recordsLinked to original sources

bioRxiv2026

A bistable type III secretion system is primed and loaded in Pseudomonas aeruginosa hypervirulent isolates with positively selected codon changes in exsA or exsD

Pseudomonas aeruginosa has multiple virulence factors that promote airway infections in people with immune deficiencies. Most clinical isolates from human infections have functional type III secretion systems (T3SSs) that are subject to bistable regulation and secrete the cytotoxic effector ExoS into neutrophils. During airway infection in people with cystic fibrosis (CF), P. aeruginosa maintains bistable regulation or genetically adapts by inactivating or hyperactivating T3SS expression. CF clinical isolates with hyperactive T3SSs have codon changes in the transcriptional regulators, exsA or exsD. It is not known how these mutations: 1) arise, 2) promote hypervirulence, or 3) impact bistable T3SS regulation. To understand how these mutants arise, ~24,000 genomes in the NCBI Pathogens database were searched for positively selected mutations in exs genes based on nonsynonomous to synonymous codon change ratios. Missense and nonsense mutations were detected in exsA (47) or exsD (45) the majority of which are predicted to be positively selected. To determine how one of these codon changes promotes hypervirulence, an exsAT48I isolate was used to establish lung infections in mice engineered with a CF genotype. The exsAT48I mutant and an exsA+ control had similar lung burdens, but the former caused exacerbated weight loss and lung inflammation, suggesting immunopathology from phagocyte death as the cause of hypervirulence. Two additional hyperactive mutants were identified by screening a small collection of isolates from children with CF. Immunoblotting and fluorescence microscopy showed that hyperactive exsA or exsD mutants increased production of T3SS injectisomes and effectors, indicating they have locked on (primed and loaded) phenotypes. ExsA dimer modeling revealed that codon changes like T48I in the {beta}-barrel domain cause a conformational change that hyperactivates the transcription factor. These results define that hyperactive mutants are positively selected, hypervirulent due to lung inflammation, and have primed and loaded T3SSs.

microbiology↗

bioRxiv2026

Inflammasome Activation and IL-1β Release in Alveolar Macrophages Infected with Pseudomonas aeruginosa is Reduced in Hypoxia

Pseudomonas aeruginosa is an important agent of acute or chronic airway infections. A complex set of genetic and environmental factors determine the outcome of a P. aeruginosa airway infection. P. aeruginosa can undergo genetic adaptation to fine tune the expression of a type III secretion system (T3SS) and other virulence factors in response to selective pressures encountered during infection. Genetic and environmental factors predispose many patient groups for P. aeruginosa infections, including people with cystic fibrosis (pwCF). CF is a genetic disorder resulting in areas of hypoxia and thick mucus that fosters P. aeruginosa airway infection. Resident alveolar macrophages (AMs) help coordinate immune responses to P. aeruginosa in airways by producing proinflammatory cytokines such as IL-1{beta}, which can be important for infection resistance. AMs infected with laboratory strains of P. aeruginosa detect the T3SS and activate the NLRC4 inflammasome, resulting in IL-1{beta} release. Studies of inflammasome responses in AMs to a clinical CF isolate of P. aeruginosa have not been reported. Here, we characterized the CF clinical isolate DH1137 and found that it has a downregulated but functional T3SS, is adapted to grow in hypoxia, and induces significant production of the inflammasome cytokines IL-1{beta} and IL-1 during lung infection in a CF mouse model. Inflammasome gene expression was primed in AMs by LPS stimulation, and upon DH1137 infection these cells released IL-1[beta]. Intriguingly, we found that infection of AMs with DH1137 in hypoxia resulted in significantly dampened inflammasome activation compared to normoxia. These data describe a new mechanism that P. aeruginosa may exploit to evade immune detection by tissue-resident lung macrophages in the context of CF.

immunology↗

bioRxiv2022

SAMHD1 controls innate immunity by regulating condensation of immunogenic self RNA

Recognition of pathogen-derived foreign nucleic acids is central to innate immune defense. This requires discrimination between structurally highly similar self and nonself nucleic acids to avoid aberrant inflammatory responses as in the autoinflammatory disorder Aicardi-Goutieres syndrome (AGS). How vast amounts of self RNA are shielded from immune recognition to prevent autoinflammation is not fully understood. Here we show that SAM domain and HD domain-containing protein 1 (SAMHD1), one of the AGS-causing genes, functions as a single-stranded RNA (ssRNA) 3exonuclease, the lack of which causes cellular RNA accumulation. Increased ssRNA in cells leads to dissolution of RNA-protein condensates, which sequester immunogenic double-stranded RNA (dsRNA). Release of sequestered dsRNA from condensates triggers activation of antiviral type I interferon via retinoic acid-inducible gene I-like receptors. Our results establish SAMHD1 as a key regulator of cellular RNA homeostasis and demonstrate that buffering of immunogenic self RNA by condensates regulates innate immune responses.

immunology↗