Biology subjects

Wodzanowski, K. A.

Publications and source records attributed to Wodzanowski, K. A..

5 recordsLinked to original sources

bioRxiv2026

Human caspase-4 and caspase-5 assemble into non-canonical inflammasomes in response to overlapping and distinct LPS structures

Innate immune sensing of bacterial lipopolysaccharide (LPS) is critical to host defense, but a dysregulated response can lead to sepsis, a life-threatening condition associated with multiple organ failure and shock. LPS is sensed at the cell surface by Toll-Like Receptor 4 (TLR4) and in the cytosol by mouse caspase-11 (Casp11) or its human orthologs, caspase-4 and 5 (CASP4/5). In response to cytosolic LPS, Casp11 oligomerizes in a catalytically dependent manner into supramolecular organizing centers (SMOCs). We demonstrate that CASP4 and CASP5 also form catalytically dependent SMOCs in response to bacterial LPS in macrophages. Furthermore, we find that CASP4 and CASP5 differentially respond to distinct LPS structural variants that fail to activate CASP11 and that only activating LPS structures induce SMOC formation by the corresponding caspase. Notably, transgenic mice expressing CASP4 in lieu of Casp11 exert more effective control of Francisella novicida, a rodent pathogen whose LPS evades TLR4 and Casp11 detection. Conversely, CASP4-expressing mice are more susceptible to sepsis when exposed to E. coli expressing a lipid A variant that fails to activate TLR4 and Casp11. Together, these findings reveal human CASP4 and CASP5 have evolved to assemble into SMOCs in response to a broader range of LPS structures than mouse Casp11 and highlight the potential tradeoff between systemic inflammation and control of bacterial infection that may underline these differences. Our findings may inform the development of targeted therapeutic strategies for gram-negative bacterial infection and sepsis that are effective in humans.

immunology↗

bioRxiv2025

Dendritic cells activate pyroptosis and effector-triggered apoptosis to restrict Legionella infection

The innate immune system relies on pattern recognition receptors (PRRs) to detect pathogen-associated molecular patterns (PAMPs) and guard proteins to monitor pathogen disruption of host cell processes. How different immune cell types engage PRR- and guard protein-dependent defenses in response to infection is poorly understood. Here, we show that macrophages and dendritic cells (DCs) respond in distinct ways to bacterial virulence activities. In macrophages, the bacterial pathogen Legionella pneumophila deploys its Dot/Icm type IV secretion system (T4SS) to deliver effector proteins that facilitate its robust intracellular replication. In contrast, T4SS activity triggers rapid DC death that potently restricts Legionella replication within this cell type. Intriguingly, we found that infected DCs exhibit considerable heterogeneity at the single cell level. Initially, a subset of DCs activate caspase-11 and NLRP3 inflammasome-dependent pyroptosis and release IL-1{beta} early during infection. At later timepoints, a separate DC population undergoes apoptosis driven by T4SS effectors that block host protein synthesis, thereby depleting the levels of the pro-survival proteins Mcl-1 and cFLIP. Together, pyroptosis and effector-triggered apoptosis robustly restrict Legionella replication in DCs. Collectively, our work suggests a model where Mcl-1 and cFLIP guard host translation in DCs, and that macrophages and DCs distinctly employ innate immune sensors and guard proteins to mount divergent responses to Legionella infection.

immunology↗

bioRxiv2023

Synthesis and validation of click-modified of NOD1/2 agonists

NOD1 and NOD2 sense small bacterial peptidoglycan fragments often called muropeptides. These muropeptides include iE-DAP and MDP, the minimal agonists for NOD1 and NOD2, respectively. Here, we synthesized and validated alkyne-modified muropeptides, iE-DAP-Alk and MDP-Alk, for use in click-chemistry reactions. While it has long been known that many cell types respond to extracellular exposure to muropeptides, it is unclear how these innate immune activators access their cytosolic innate immune receptors, NOD1 and NOD2. The subcellular trafficking and transport mechanisms by which muropeptides access these cytosolic innate immune receptors are a major gap in our understanding of these critical host responses. The clickchemistry-enabled agonists developed here will be particularly powerful to decipher the underlying cell biology and biochemistry of NOD1 and NOD2 innate immune sensing.

immunology↗

bioRxiv2020

Multiscale Invasion Assay for Probing Macrophage Response to Bacteria

The immune system is a complex network of various cellular components that must differentiate between pathogenic bacteria and the commensal bacteria of the human microbiome, where misrecognition is linked to inflammatory disorders. Fragments of bacterial cell wall peptidoglycan bind to pattern recognition receptors within macrophages, leading to immune activation. To study this complex process, an approach for three-dimensional (3D) culture of human macrophages and their invasion with relevant bacteria in a well-defined hydrogel-based synthetic matrix inspired by the gut was established. Workflows were developed for monocyte encapsulation and differentiation into macrophages in 3D culture with high viability. Bacteria invaded into macrophages permitted in situ peptidoglycan labeling. Macrophages exhibited biologically-relevant cytokine release in response to bacteria. This multi-dimensional bacteria-macrophage co-culture system will prove useful in future studies to observe bacterial fragment production and localization in the cell at the carbohydrate level for insights into how our immune system properly senses bacteria. TOC Figure O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=125 SRC="FIGDIR/small/385617v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@45cdf1org.highwire.dtl.DTLVardef@d3c3d3org.highwire.dtl.DTLVardef@dd74f2org.highwire.dtl.DTLVardef@128d1da_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2020

Bacterial Peptidoglycan Fragments Deferentially Regulate Innate Immune Signaling

The human innate immune system responds to both pathogen and commensal bacteria at the molecular level using bacterial peptidoglycan (PG) recognition elements. Traditionally, synthetic and commercially accessible PG monosaccharide units known as muramyl dipeptide (MDP) and N-glycolyl MDP (ng-MDP) have been used to probe the mechanism of innate immune activation of pattern recognition receptors (PRRs) such as NOD-like receptors (NLRs). However, bacterial PG is a dynamic and complex structure, with various chemical modifications and trimming mechanisms that result in the production of disaccharide containing elements. These molecules pose as attractive targets for immunostimulatory screening; however, studies are limited due to their synthetic accessibility. Inspired by disaccharide containing compounds produced from the gut microbe, Lactobacillus acidophilus, a robust and scalable chemical synthesis of PG-based disaccharide ligands was implemented. Together with a monosaccharide PG library, compounds were screened for their ability to stimulate proinflammatory genes in bone marrow derived macrophages (BMDMs). The data reveal a diverse gene induction pattern between monosaccharide and disaccharide PG units, suggesting that PG innate immune signaling is more complex than a one-activator-one pathway program, as biologically relevant fragments induce distinct transcriptional programs. These disaccharide molecules will serve as critical immunostimulatory tools to more precisely define specialized innate immune regulatory mechanisms that distinguish between commensal and pathogenic bacteria residing in the microbiome.

immunology↗