Lysosomal membrane permeabilization (LMP) is observed under many pathological conditions, leading to cellular dysfunction and death. However, the mechanisms by which lysosomal membranes become leak...

Perivascular stromal cells (PSCs) are a recently identified cell type that comprises a small percentage of the platelet derived growth factor receptor-β+ cells within the CNS perivascular space. PSCs are activated following injury to the brain or spinal cord, expand in number and contribute to fibrotic scar formation within the injury site. Beyond fibrosis, their high density in the lesion core makes them a potential significant source of signals that act on neural cells adjacent to the lesion site. Our developmental analysis of PSCs, defined by expression of Collagen1a1 in the maturing brain, revealed that PSCs first appear postnatally and may originate from the meninges. PSCs express many of the same markers as meningeal fibroblasts, including expression of the retinoic acid (RA) synthesis proteins Raldh1 and Raldh2. Using a focal brain ischemia injury model to induce PSC activation and expansion, we show a substantial increase in Raldh1+/Raldh2+ PSCs and Raldh1+ activated macrophages in the lesion core. We find that RA levels are significantly elevated in the ischemic hemisphere and induce signaling in astrocytes and neurons in the peri-infarct region. This study highlights a dual role for activated, non-neural cells where PSCs deposit fibrotic ECM proteins and, along with macrophages, act as a potentially important source of RA, a potent signaling molecule that could influence recovery events in a neuroprotective fashion following brain injury.

Iron is a critical nutrient for most microbial pathogens, and the immune system exploits this requirement by sequestering iron. The opportunistic pathogen Pseudomonas aeruginosa exhibits a high requirement for iron yet an exquisite ability to overcome iron deprivation during infection. Upon iron starvation, P. aeruginosa induces the expression of several high-affinity iron acquisition systems, as well as the PrrF small regulatory RNAs (sRNAs) that mediate an iron-sparing response. Here, we used liquid chromatography-tandem mass spectrometry to conduct proteomics of the iron starvation response of P. aeruginosa. Iron starvation increased levels of multiple proteins involved in amino acid catabolism, providing the capacity for iron-independent entry of carbons into the tricarboxylic acid (TCA) cycle. Proteins involved in sulfur assimilation and cysteine biosynthesis were reduced upon iron starvation, while proteins involved in iron-sulfur cluster biogenesis were increased, highlighting the central role of iron in P. aeruginosa metabolism. Iron starvation also resulted in changes in the expression of several zinc-responsive proteins and increased levels of twitching motility proteins. Subsequent analyses provided evidence for the regulation of many of these proteins via posttranscriptional regulatory events, some of which are dependent upon the PrrF sRNAs. Moreover, we showed that iron-regulated twitching motility is partially dependent upon the prrF locus, highlighting a novel link between the PrrF sRNAs and motility. These findings add to the known impacts of iron starvation in P. aeruginosa and outline potentially novel roles for the PrrF sRNAs in iron homeostasis and pathogenesis. IMPORTANCE Iron is central for growth and metabolism of almost all microbial pathogens, and as such, this element is sequestered by the host innate immune system to restrict microbial growth. Here, we used label-free proteomics to investigate the Pseudomonas aeruginosa iron starvation response, revealing a broad landscape of metabolic and metal homeostasis changes that have not previously been described. We further provide evidence that many of these processes, including twitching motility, are regulated through the iron-responsive PrrF small regulatory RNAs. As such, this study demonstrates the power of proteomics for defining stress responses of microbial pathogens.

Tissue injury induces metabolic changes in stem cells, which likely modulate regeneration. Using a model of organ regeneration called wound-induced hair follicle neogenesis (WIHN), we identified skin-resident bacteria as key modulators of keratinocyte metabolism, demonstrating a positive correlation between bacterial load, glutamine metabolism, and regeneration. Specifically, through comprehensive multiomic analysis and single-cell RNA sequencing in murine skin, we show that bacterially induced hypoxia drives increased glutamine metabolism in keratinocytes with attendant enhancement of skin and hair follicle regeneration. In human skin wounds, topical broad-spectrum antibiotics inhibit glutamine production and are partially responsible for reduced healing. These findings reveal a conserved and coherent physiologic context in which bacterially induced metabolic changes improve the tolerance of stem cells to damage and enhance regenerative capacity. This unexpected proregenerative modulation of metabolism by the skin microbiome in both mice and humans suggests important methods for enhancing regeneration after injury.