Myddosome Signaling: Defining the spatiotemporal rules of innate immune activation.

Mammalian immune systems are comprised of interconnected layers: Barriers, the innate and the adaptive immune system. We study the cell biology of innate immune cells and how these special cells contribute to the detection of microbes and defense against pathogens. For this, innate immune cells employ a range of pattern-recognition receptors, that upon their activation “ring the alarm” and initiate inflammation and an immune response. One class of pattern-recognition receptors, the Toll-like receptors (TLRs) are central to this innate immune activation. Canonical models of TLR signaling describe ligand-induced assembly of MyD88-dependent signaling complexes (“myddosomes”) at the plasma membrane or endosomal compartments. This receptor-proximal clustering of MyD88 nucleates a highly complex signaling assembly and activates diverse signaling pathways (e.g. NF-κB and MAPKs). Emerging evidence suggests that myddosomes are not limited to the receptor-proximal compartment where they were seeded (proto-myddosomes). Instead, the updated model of TLR signaling supports that myddosomes can assemble and persist in the cytosol. The mechanistic basis, molecular composition, and functional consequences of these cytosolic assemblies remain incompletely understood.

This project seeks to further characterize cytosolic myddosomes and define the rules and factors that govern their assembly. The central hypothesis is that cytosolic myddosomes are structurally and compositionally distinct from receptor-bound proto-myddosomes and that their spatial reorganization modulates signaling amplitude, duration, and transcriptional output; altogether shaping the subsequent immune response. To test this, the project will combine quantitative (live-cell) imaging, genetic engineering to externally control MyD88 and its complexes, and transcriptional profiling:

1. We will investigate the assembly dynamics of cytosolic myddosomes using CRISPR-engineered knock-in innate immune cell lines (e.g. macrophages) and high-resolution live-cell imaging. By comparing ligand-induced receptor-proximal complexes with cytosolic puncta that emerge after receptor disengagement, we will determine how the cytosolic assemblies arise from membrane-proximal complexes. Quantitative and biochemical analysis will further characterize their properties.

2. We will determine and manipulate the molecular composition of cytosolic myddosomes using inducible clustering of MyD88 combined with biochemical and imaging-based quantifications. We will explore biosynthetic approaches to control MyD88 independently of the receptors and to target MyD88-assemblies to different cellular compartments. By spatially restricting MyD88, we will identify proteins selectively enriched in each context and define whether canonical components such as IRAK kinases are differentially recruited.

3. The functional consequences of cytosolic MyD88 signaling will be assessed through time-resolved transcriptomic analysis and quantitative measurements of signaling dynamics and cytokine production. Comparing acute receptor-driven activation with persistent cytosolic clustering will determine the impact of cytosolic myddosomes on innate immune cell activation.

By integrating mechanistic cell biology with systems-level analysis, we will learn how TLRs and their associated signaling pathways govern the inflammatory output of innate immune cells. Given the central role of TLRs and MyD88 in infection, autoimmunity, and inflammatory diseases, uncovering these principles may reveal new strategies for modulating pathological inflammation.

Relevant reading:

“Molecular definition of the endogenous Toll-like receptor signalling pathways”

Fisch et al , Nature 2024; PMID: 38961291; DOI: 10.1038/s41586-024-07614-7

Daniel Fisch is joining GIMM as a Group Leader in 2026
- currently at the Boston Children’s Hospital & Harvard Medical School, Boston MA, US

ABOUT THE LAB

Innate Immune Cell Biology Laboratory

Our group investigates how the innate immune system detects danger, processes information, and executes protective responses. We are interested in the molecular “machines” that allow cells to sense infection and translate these signals into inflammation, antimicrobial defense, and communication with the adaptive immune system. By uncovering how these systems work at the molecular level, we aim to better understand immunity in health and disease, with implications for infection, autoimmunity, and cancer.

Toll-like Receptor Signaling – Detectors and Computers of Immune Responses

Our research focuses on how innate immune cells (e.g. macrophages) process danger signals through Toll-like receptors (TLRs). These receptors act as “detectors” for microbial and damage-associated molecules and initiate inflammatory signaling programs that instruct the broader immune response. Rather than signaling through simple linear pathways, TLRs assemble large, dynamic protein platforms (e.g. Myddosomes) that function as molecular “computers,” integrating inputs and generating specific outputs.

We investigate how these signaling complexes assemble at endogenous levels inside cells, how their composition and structure change over time, and how these properties determine the quality and magnitude of inflammation. We are also developing synthetic tools that allow us to build and control these signaling machines on demand. By dissecting how TLR signaling is organized at the molecular level, we aim to uncover principles that explain how distinct immune responses are computed and how they might be selectively tuned.

Guanylate-Binding Proteins (GBPs) – Guarding the Cell from Within

Another focus of our research is understanding how cells detect and eliminate intracellular pathogens. We study a family of interferon-inducible proteins known as Guanylate-binding proteins (GBPs), which function at the frontline of cell-intrinsic immunity. GBPs can directly sense microbial invasion, target pathogens that enter the cytosol, and trigger inflammatory defense programs, including forms of programmed cell death to eliminate infected cells. In this way, they act as both detectors and executors of immune defense.

Beyond classical pattern-recognition systems, GBPs participate in “guard” mechanisms that monitor the integrity of key cellular processes and respond when pathogens disrupt them. This strategy provides flexibility: rather than recognizing only conserved microbial molecules, guard pathways detect the consequences of infection itself. Our work seeks to define how GBPs are activated, how their activity is carefully restrained to prevent damage to healthy tissue, and how these regulatory circuits shape inflammation. We are particularly interested in whether manipulating GBP pathways could enhance antimicrobial immunity or cancer immunotherapy.

Our lab uses genetics, cell biology, biochemistry and quantitative imaging to uncover the principles that govern innate immune cell biology. By studying the detectors, computers, and executors of immunity as integrated molecular machines, we seek to build a mechanistic framework for how immune responses are initiated, computed, and controlled and how they can be manipulated for therapeutic benefit.

ABOUT THE GROUP LEADER

Education & Training

2022-2026: Postdoc, Boston Children’s Hospital & Harvard Medical School, Boston MA, US

2020-2021: Postdoc, The Francis Crick Institute & University of Birmingham, UK

2016-2020: PhD; The Francis Crick Institute & Imperial College London, UK

2011-2016: BSc Molecular Medicine & MSc Biochemistry and Biophysics; Albert-Ludwigs University Freiburg, Germany

Publications

Jastrab JB, Tseng A, Fisch D, Ragland SA, Kagan JC (2025) “Wall teichoic acid is required for DNA-triggered innate immune receptor activation by Staphylococcus aureus.” Biorxiv preprint at https://doi.org/10.1101/2025.09.25.678139.

Holley CL, Monteleone M, Fisch D, Libert AES, Ju RJ., Choi JH, Condon ND, Emming S, Crawford J, Lawrence GMEP, et al. (2024) “Pyroptotic cell corpses are crowned with F-actin-rich filopodia that engage CLEC9A signaling in incoming dendritic cells” Nat. Immunol.: 1–11.

Fisch D, Zhang T, Sun H, Ma W, Tan Y, Gygi SP, Higgins DE, Kagan JC (2024) “Molecular definition of the endogenous Toll-like Receptor signaling pathway” Nature: 1-10

Fisch D, Pfleiderer MM, Anastasakou E, Mackie GM, Wendt F, Liu X, Clough B, Lara-Reyna S, Encheva V, Snijders AP, Bando H, Yamamoto M, Beggs AD, Mercer J, Shenoy AR, Wollscheid B, Maslowski KM, Galej WP, Frickel E-M (2023) “PIM1 controls GBP1 activity to limit self-damage and to guard against pathogen infection” Science: 382, eadg2253

Clough B, Fisch D, Mize TH, Encheva V, Snijders A, Frickel EM (2023) “p97/VCP targets Toxoplasma gondii vacuoles for parasite restriction in interferon-stimulated human cells.” mSphere, e00511-23

Fisch D, Clough B, Khan R, Healy L & Frickel E-M (2021) “Toxoplasma-proximal and distal control by GBPs in human macrophages” Pathogens & Disease: ftab058

Brandel A, Aigal S, Lagies S, Schlimpert M, Meléndez AV, Xu M, Lehmann A, Hummel D, Fisch D, Madl J, Eierhoff T, Kammerer B & Römer W (2021) “The Gb3-enriched CD59/flotillin plasma membrane domain regulates host cell invasion by Pseudomonas aeruginosa.” Cellular and Molecular Life Sciences 78: 3637–3656

Fisch D, Evans R*, Clough B, Byrne S.K, Channell W.M, Dockterman J & Frickel EM (2021) “HRMAn 2.0: Next generation artificial intelligence‐driven analysis for broad host‐pathogen interactions.” Cellular Microbiology.: e13349.

Fisch D, Clough B, Domart M-C, Encheva V, Bando H, Snijders AP, Collinson LM, Yamamoto M, Shenoy AR & Frickel E (2020) “Human GBP1 Differentially Targets Salmonella and Toxoplasma to License Recognition of Microbial Ligands and Caspase-Mediated Death.” Cell Reports 32: 1–14

Fisch D, Yakimovich A, Clough B, Mercer J & Frickel EM (2020) “Image-Based Quantitation of Host Cell–Toxoplasma gondii Interplay Using HRMAn: A Host Response to Microbe Analysis Pipeline.” In Toxoplasma gondii: Methods and Protocols pp 411–433.

Fisch D, Clough B & Frickel EM (2019) Human immunity to Toxoplasma gondii. PLOS Pathogens 15: e1008097

Clough B, Finethy R, Khan RT, Fisch D, Jordan S, Patel H, et al. (2019). “C57BL/6 and 129 inbred mouse strains differ in Gbp2 and Gbp2b expression in response to inflammatory stimuli in vivo.” Wellcome Open Res. 2019 Aug 20;4:124.

Fisch D, Bando H, Clough B, Hornung V, Yamamoto M, Shenoy AR & Frickel EM (2019). “Human GBP1 is a microbe‐specific gatekeeper of macrophage apoptosis and pyroptosis.” EMBO J.: e100926

Fisch D, Yakimovich A, Clough B, Wright J, Bunyan M, Howell M, Mercer J & Frickel EM (2019). “Defining host–pathogen interactions employing an artificial intelligence workflow.” Elife 8

Awards

2025: HRIA Charles A. King Trust Postdoctoral Research Fellowship

2022: Human Frontiers Science Program (HFSP) Postdoctoral Long-term Fellowship

2022: EMBO Postdoctoral Fellowship

2017: Boehringer Ingelheim Fonds (BIF) PhD fellowship