Breen, P. | bioRxiv Science

bioRxiv2023

Perturbed sulfur homeostasis allows C. elegans to escape growth retardation on Actinobacteria from its natural microbiome

The rate at which organisms grow is influenced by their biotic environment. The nematode Caenorhabditis elegans grows slower in the presence of Actinobacteria, but it is unknown why. Here, we show how perturbed levels of hydrogen sulfide and cysteine modulate the growth rate of C. elegans on Actinobacteria. Using an unbiased forward genetic screen of C. elegans we discovered alleles of the conserved cystathionine gamma-lyase (cth-2/CTH) that improved animal growth rate on Actinobacteria. Conversely, null alleles of cth-2 cause developmental arrest of animals grown on Actinobacteria, which can be rescued by exogenous H2S. We also discovered a leucine rich repeat gene that regulates cysteine and H2S production, lrr-2/LRRC58. A wild isolate of C. elegans that naturally grows well on Actinobacteria has a mutant allele of lrr-2, suggesting this sulfur metabolism pathway is important for the regulation of animal growth rate in its natural ecological context. We propose a model in which wild-type worms use sulfurous compounds to promote growth of their favored bacterial food sources by inhibiting Actinobacteria growth. This strategy becomes a liability when Actinobacteria are the sole food source but can be bypassed by mutations in sulfur metabolism. This study reveals how the homeostasis of sulfurous compounds controls the growth rate of animals in an ecological context.

ecology↗

bioRxiv2020

Induction of RNA interference by C. elegans mitochondrial dysfunction via the DRH-1/RIG-I homologue RNA helicase and the EOL-1/RNA decapping enzyme

RNA interference (RNAi) is an antiviral pathway common to many eukaryotes that detects and cleaves foreign nucleic acids. In mammals, mitochondrially localized proteins such as MAVS, RIG-I, and MDA5 mediate antiviral responses. Here, we report that mitochondrial dysfunction in Caenorhabditis elegans activates RNAi-directed silencing via induction of a pathway homologous to the mammalian RIG-I helicase viral response pathway. The induction of RNAi also requires the conserved RNA decapping enzyme EOL-1/DXO. The transcriptional induction of eol-1 requires DRH-1 as well as the mitochondrial unfolded protein response (UPRmt). Upon mitochondrial dysfunction, EOL-1 is concentrated into foci that depend on the transcription of mitochondrial RNAs that may form dsRNA, as has been observed in mammalian antiviral responses. The enhanced RNAi triggered by mitochondrial dysfunction contributes to the increase in longevity that is induced by mitochondrial dysfunction.

genetics↗

bioRxiv2019

The Klebsiella pneumoniae citrate synthase gene, gltA, influences site specific fitness during infection

Klebsiella pneumoniae (Kp), one of the most common causes of healthcare-associated infections, increases patient morbidity, mortality and hospitalization costs. Kp must acquire nutrients from the host for successful infection. However, the host is able to prevent bacterial nutrient acquisition through multiple systems, including the innate immune protein lipocalin 2 (Lcn2) that prevents Kp iron acquisition by sequestering the siderophore enterobactin. To identify novel Kp factors that mediate evasion of nutritional immunity during lung infection, we undertook an InSeq study using a pool of >20,000 transposon mutants administered to Lcn2+/+ and Lcn2-/- mice. Comparing mutant frequencies between mouse genotypes, this genome-wide screen identified the Kp citrate synthase GltA as potentially interacting with Lcn2, and this novel finding was independently validated. Interestingly, in vitro studies suggest that this interaction is not direct. Given that GltA is involved in oxidative metabolism, we screened the ability of this mutant to use a variety of carbon and nitrogen sources. The results indicated that the gltA mutant has a distinct amino acid auxotrophy and is unable to use a variety of carbon sources. Specifically, we show that gltA is necessary for growth in bronchioloalveolar lavage fluid, which is amino acid-limited, but dispensable in serum, which is amino acid rich. Deletion of Lcn2 from the host leads to increased amino acid levels in bronchioloalveolar lavage fluid, and thus abrogates the loss of gltA during pneumonia in the Lcn2-/- background. GltA was also required for gut colonization, but dispensable in the bloodstream in a bacteremia model, demonstrating that deletion of gltA leads to an organ-specific fitness defect. Together, this study is the first to mechanistically describe a role for gltA in Kp infection and provide unique insight into how metabolic flexibility impacts bacterial fitness during infection.\n\nAuthor SummaryThe bacteria Klebsiella pneumoniae (Kp) is an important cause of infection in healthcare settings. These infections can be difficult to treat, as they frequently occur in chronically ill patients and the bacteria has the ability to acquire multiple antibiotic resistance markers. Kp is a common colonizer of the intestinal tract in hospitalized patients, and can progress to infections of the bloodstream, respiratory and urinary tract. However, the bacterial factors that allow Kp to replicate in these different body sites is unclear. In this study, we found that the Kp citrate synthase, GltA, enables bacterial replication in the lung and intestine by enhancing the ability of Kp to use diverse nutrients, in a mechanism known as metabolic flexibility. Kp lacking GltA require specific amino acids that are abundant in blood, but not other body sites. The work in this study provides novel insight into why Kp is a successful hospital pathogen that can colonize and infect multiple body sites.

microbiology↗