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The success of Mycobacterium tuberculosis as a pathogen derives from its facile adaptation to the intracellular milieu of human macrophages. To explore this process, we asked whether adaptation also required interference with the metabolic machinery of the host cell. Temporal profiling of the metabolic flux, in cells infected with differently virulent mycobacterial strains, confirmed that this was indeed the case. Subsequent analysis identified the core subset of host reactions that were targeted. It also elucidated that the goal of regulation was to integrate pathways facilitating macrophage survival, with those promoting mycobacterial sustenance. Intriguingly, this synthesis then provided an axis where both host- and pathogen-derived factors converged to define determinants of pathogenicity. Consequently, whereas the requirement for macrophage survival sensitized TB susceptibility to the glycemic status of the individual, mediation by pathogen ensured that the virulence properties of the infecting strain also contributed towards the resulting pathology.

Survival of Mycobacterium tuberculosis (Mtb) within the host macrophage is mediated through pathogen-dependent inhibition of phagosome-lysosome fusion, which enables bacteria to persist within the immature phagosomal compartment. By employing ultrastructural examination of different field isolates supported by biochemical analysis, we found that some of the Mtb strains were in fact poorly adapted for subsistence within endocytic vesicles of infected macrophages. Instead, through a mechanism involving activation of host cytosolic phospholipase A2, these bacteria rapidly escaped from phagosomes, and established residence in the cytoplasm of the host cell. Interestingly, by facilitating an enhanced suppression of host cellular autophagy, this translocation served as an alternate virulence acquisition mechanism. Thus, our studies reveal plasticity in the adaptation strategies employed by Mtb, for survival in the host macrophage.

Innate immunity is the first line of defense against infections. Pathways regulating innate responses can also modulate other processes, including stress resistance and longevity. Increasing evidence suggests a role for the nucleolus in regulating cellular processes implicated in health and disease. Here we show the highly conserved nucleolar protein, fibrillarin, is a vital factor regulating pathogen resistance. Fibrillarin knockdown enhances resistance in C. elegans against bacterial pathogens, higher levels of fibrillarin induce susceptibility to infection. Pathogenic infection reduces nucleolar size, ribsosomal RNA, and fibrillarin levels. Genetic epistasis reveals fibrillarin functions independently of the major innate immunity mediators, suggesting novel mechanisms of pathogen resistance. Bacterial infection also reduces nucleolar size and fibrillarin levels in mammalian cells. Fibrillarin knockdown prior to infection increases intracellular bacterial clearance, reduces inflammation, and enhances cell survival. Collectively, these findings reveal an evolutionarily conserved role of fibrillarin in infection resistance and suggest the nucleolus as a focal point in innate immune responses.

The daily removal of billions of apoptotic cells in the human body via the process of efferocytosis is essential for homeostasis. To allow for this continuous efferocytosis, rapid phenotypic changes occur in the phagocytes enabling them to engulf and digest the apoptotic cargo. In addition, efferocytosis is actively anti-inflammatory and promotes resolution. Owing to its ubiquitous nature and the sheer volume of cell turnover, efferocytosis is a point of vulnerability. Aberrations in efferocytosis are associated with numerous inflammatory pathologies, including atherosclerosis, cancer and infections. The recent exciting discoveries defining the molecular machinery involved in efferocytosis have opened many avenues for therapeutic intervention, with several agents now in clinical trials.

Chronic non-healing wounds are a major complication of diabetes, which impacts 1 in 10 people worldwide. Dying cells in the wound perpetuate the inflammation and contribute to dysregulated tissue repair 1-3 . Here, we reveal the membrane transporter Slc7a11 as a molecular 'brake' on efferocytosis, the process by which dying cells are removed, and that inhibiting Slc7a11 can accelerate wound healing. First, transcriptomics of efferocytic dendritic cells identified upregulation of several Slc7 gene family members. In further analyses, pharmacological inhibition, siRNA knockdown, or deletion of Slc7a11 enhanced dendritic cell efferocytosis. Interestingly, Slc7a11 was highly expressed in skin dendritic cells, and scRNAseq of inflamed skin showed Slc7a11 upregulation in innate immune cells. In a mouse model of excisional skin wounding, loss of Slc7a11 expression or inhibition accelerated healing dynamics and reduced apoptotic cell load in the wound. Mechanistic studies revealed a link between Slc7a11, glucose homeostasis, and diabetes. Slc7a11-deficient dendritic cells relied on glycogen store-derived aerobic glycolysis for improved efferocytosis, and transcriptomics of efferocytic Slc7a11-deficient dendritic cells identified genes linked to gluconeogenesis and diabetes. Further, Slc7a11 expression was higher in the wounds of diabetic-prone db/db mice, and targeting Slc7a11 accelerated their wound healing. The faster healing was also linked to the release of TGF- family member GDF15 from efferocytic dendritic cells. Collectively, Slc7a11 is a negative regulator of efferocytosis, and removing this brake improves wound healing, with significant implications for diabetic wound management.

sharing the clinical E. coli isolates, A. Wullaert for GasderminD knockout mice, and M. Bertrand for Ripk1 kinase dead mice. We thank the Germ-Free and Gnotobiotic Mouse Facility (UGent/UZ Gent/VIB), VIB Protein Core, VIB Flow Cytometry Core, VIB Bioimaging Core, and VIB Nucleomics Core for their contributions.

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