to mass spectrometry related experiments and analysis; R.H., Z.Y. and B.R. performed the library construction and next-generation sequencing for ChIP-seq and RNA-seq; M.H. and Y.G.Z. synthesized L-lactyl-CoA. H.H. and D.Z. analyzed ChIP-seq and RNA-seq data. G.Z. provided all primary BMDM cell cultures. D.M.C. carried out the bacterial infection experiments, C.C. carried out TAM experiments. Author Information. Y.Z. is a founder, board member, advisor to, and inventor on patents licensed to PTM Bio Inc. L.B. is co-founder and CSO of rMark Bio Inc., and founder and CEO of Onchilles Pharma Inc. Readers are welcome to comment on the online version of the paper. Data availability. The ChIP-seq and RNA-seq data have been made available at the Gene Expression Omnibus (GEO) repository under the accession number GSE115354. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE 31 partner repository with the dataset identifier PXD014870. All other data are available from the authors upon reasonable request.

The lack of new antibiotics is among the most critical challenges facing medicine. The problem is particularly acute for Gram-negative bacteria. A novel antibiotic strategy is to target bacterial nutrition and metabolism. The metal gallium can disrupt bacterial iron metabolism as gallium can be taken up by bacteria, and replace iron. Here we performed pre-clinical work and a phase 1 human trial to investigate the antibiotic activity of gallium in people with cystic fibrosis (CF) and chronic Pseudomonas aeruginosa airway infections. We found that CF sputum was iron-limited, and that low micromolar concentrations of gallium inhibited P. aeruginosa growth in CF sputum. Ex vivo experiments indicated that gallium inhibited key iron-dependent enzymes, and increased bacterial sensitivity to oxidants. We also found that gallium resistance developed at low rates, its activity was synergistic with some antibiotics, and it did not affect P. aeruginosa killing by human macrophages. Finally, we tested parenteral gallium in murine lung infections, and in CF patients with chronic P. aeruginosa lung infections and found indications of safety and efficacy. These data represent a small step toward targeting iron metabolism, or other nutritional vulnerabilities of bacteria, to treat human infections.

The heat shock response (HSR) is essential to survive acute proteotoxic stress and has been studied extensively in unicellular organisms and tissue culture cells, but to a lesser extent in intact metazoan animals. To identify the regulatory pathways that control the HSR in Caenorhabditis elegans, we performed a genome-wide RNAi screen and identified 59 genes corresponding to 7 positive activators required for the HSR and 52 negative regulators whose knockdown leads to constitutive activation of the HSR. These modifiers function in specific steps of gene expression, protein synthesis, protein folding, trafficking, and protein clearance, and comprise the metazoan heat shock regulatory network (HSN). Whereas the positive regulators function in all tissues of C. elegans, nearly all of the negative regulators exhibited tissue-selective effects. Knockdown of the subunits of the proteasome strongly induces HS reporter expression only in the intestine and spermatheca but not in muscle cells, while knockdown of subunits of the TRiC/CCT chaperonin induces HS reporter expression only in muscle cells. Yet, both the proteasome and TRiC/CCT chaperonin are ubiquitously expressed and are required for clearance and folding in all tissues. We propose that the HSN identifies a key subset of the proteostasis machinery that regulates the HSR according to the unique functional requirements of each tissue.

We sought a new approach to treating infections by intracellular bacteria, namely, by altering host cell functions that support their growth. We screened a library of 640 Food and Drug Administration (FDA)-approved compounds for agents that render THP-1 cells resistant to infection by four intracellular pathogens. We identified numerous drugs that are not antibiotics but were highly effective in inhibiting intracellular bacterial growth with limited toxicity to host cells. These compounds are likely to target three kinds of host functions: (i) G protein-coupled receptors, (ii) intracellular calcium signals, and (iii) membrane cholesterol distribution. The compounds that targeted G protein receptor signaling and calcium fluxes broadly inhibited Coxiella burnetii, Legionella pneumophila, Brucella abortus, and Rickettsia conorii, while those directed against cholesterol traffic strongly attenuated the intracellular growth of C. burnetii and L. pneumophila. These pathways probably support intracellular pathogen growth so that drugs that perturb them may be therapeutic candidates. Combining host- and pathogen-directed treatments is a strategy to decrease the emergence of drug-resistant intracellular bacterial pathogens.