Jumbo phages protect their genomes from DNA-sensing bacterial defense systems by enclosing them within vesicles and nucleus-like compartments. Very little is known about defense systems specialized to counter these phages. Here, we show that AVAST type 5 (Avs5) systems, part of the signal transduction ATPases of numerous domains (STAND) superfamily, confer conserved immunity against jumbo phages. Using fluorescence microscopy and biotin proximity labeling, we demonstrate that Avs5 localizes to early infection vesicles, where it senses an essential, early-expressed phage protein named JADA (Jumbo phage Avs5 Defense Activator). Recognition of phage infection triggers the Sir2-like effector domain of Avs5 across three Avs5 clades, resulting in rapid NAD + hydrolysis, disruption of phage nucleus formation, and arrest of infection. These findings reveal a spatially coordinated bacterial immune strategy that targets an early vulnerability in jumbo phage infection. ... Read more

Exogenous fatty acids are directly incorporated into bacterial membranes, heavily influencing cell envelope properties, antibiotic susceptibility, and bacterial ecology. Here, we quantify fatty acid biosynthesis metabolites and enzymes of the fatty acid synthesis pathway to determine how exogenous fatty acids inhibit fatty acid synthesis in Escherichia coli. We find that acyl-CoA synthesized from exogenous fatty acids rapidly increases concentrations of long-chain acyl-acyl carrier protein (acyl-ACP), which inhibits fatty acid synthesis initiation. Accumulation of long-chain acyl-ACP is caused by competition with acyl-CoA for phospholipid synthesis enzymes. Furthermore, we find that transcriptional regulation rebalances saturated and unsaturated acyl-ACP while maintaining overall expression levels of fatty acid synthesis enzymes. Rapid feedback inhibition of fatty acid synthesis by exogenous fatty acids thus allows E. coli to benefit from exogenous fatty acids while maintaining fatty acid synthesis capacity. We hypothesize that this indirect feedback mechanism is ubiquitous across bacterial species. ... Read more

All free-living microorganisms homeostatically maintain the fluidity of their membranes by adapting lipid composition to environmental temperatures. Here, we quantify enzymes and metabolic intermediates of the Escherichia coli fatty acid and phospholipid synthesis pathways, to describe how this organism measures temperature and restores optimal membrane fluidity within a single generation after a temperature shock. A first element of this regulatory system is a temperature-sensitive metabolic valve that allocates flux between the saturated and unsaturated fatty acid synthesis pathways via the branchpoint enzymes FabI and FabB. A second element is a transcription-based negative feedback loop that counteracts the temperature-sensitive valve. The combination of these elements accelerates membrane adaptation by causing a transient overshoot in the synthesis of saturated or unsaturated fatty acids following temperature shocks. This strategy is comparable to increasing the temperature of a water bath by adding water that is excessively hot rather than adding water at the desired temperature. These properties are captured in a mathematical model, which we use to show how hard-wired parameters calibrate the system to generate membrane compositions that maintain constant fluidity across temperatures. We hypothesize that core features of the E. coli system will prove to be ubiquitous features of homeoviscous adaptation systems. ... Read more

Prokaryotes have evolved a multitude of defense systems to protect against phage predation. Some of these resemble eukaryotic genes involved in antiviral responses. Here, we set out to systematically project the current knowledge of eukaryotic-like antiviral defense systems onto prokaryotic genomes, using Pseudomonas aeruginosa as a model organism. Searching for phage defense systems related to innate antiviral genes from vertebrates and plants, we uncovered over 450 candidates. We validated six of these phage defense systems, including factors preventing viral attachment, R-loop-acting enzymes, the inflammasome, ubiquitin pathway, and pathogen recognition signaling. Collectively, these defense systems support the concept of deep evolutionary links and shared antiviral mechanisms between prokaryotes and eukaryotes. ... Read more