Biology subjects

Chapman, M. R.

Publications and source records attributed to Chapman, M. R..

4 recordsLinked to original sources

bioRxiv2026

The Microglial Protein sTREM2 Inhibits the Bacterial Functional Amyloid CsgA and Suppresses Amyloid-Dependent Biofilm Formation

Protein misfolding and aggregation, including amyloid fibril formation, underlie a large class of human diseases including prominent neurological disorders such as Alzheimers and Parkinsons disease. A small number of human proteins have been identified that inhibit amyloidogenesis. One such protein is sTREM2, a soluble receptor liberated from microglia, the resident macrophages of the central nervous system. The extracellular domain of TREM2 is shed upon proteolytic cleavage to create sTREM2, which has previously been shown to inhibit amyloid-{beta} aggregation in vitro. TREM2 is also expressed by intestinal macrophages, which are known to directly bind the bacterial amyloid curli and mount cytokine responses upon exposure. Here we show that sTREM2 is a sub-stoichiometric inhibitor of CsgA amyloidogenesis, CsgA being the major protein component of curli that drives biofilm formation in uropathogenic Escherichia coli and many other proteobacteria. In vitro, sTREM2 potently and sub-stoichiometrically inhibited CsgA amyloidogenesis in a dose-dependent manner. Kinetic modeling indicated that sTREM2 slowed primary and secondary nucleation, rather than altering fiber elongation. When added exogenously to bacterial growth medium, sTREM2 significantly suppressed curli-dependent pellicle biofilm formation without affecting bacterial growth. These findings establish sTREM2 as a member of the small group of human proteins capable of inhibiting bacterial functional amyloidogenesis, suggesting that gut-resident TREM2-expressing macrophages, which are already known to interact with curli, may employ sTREM2 as a physiologically relevant defense against bacterial amyloid formation.

biochemistry↗

bioRxiv2025

Transient binding facilitates super-resolution imaging of functional amyloid fibrils on living bacteria

Curli, which are the major proteinaceous component of the Escherichia coli biofilm extracellular matrix, help protect cells against environmental stressors including dehydration and antibiotics. Composed of the amyloid proteins CsgA and CsgB, curli self-assemble as these protomers are secreted into the extracellular space. The mechanisms of curli assembly and their functional roles within the extracellular matrix are incompletely understood. High-resolution imaging tools compatible with live-cell conditions provide a critical means to investigate the assembly and function of curli in their native context. In this study, we use super-resolution imaging to visualize curli fibrils on living bacterial cells. Transient amyloid binding of the fluorogenic dye Nile blue facilitates two complementary super-resolution fluorescence microscopy approaches: single-molecule localization microscopy and super-resolution optical fluctuation imaging. Additionally, imaging fluorescence correlation spectroscopy was used to measure the characteristic relaxation times associated with Nile blue binding to CsgA fibrils. Together, these approaches offer a framework for imaging-based biophysical characterization of curli structures on living cells. Importance/impact statementEscherichia coli and other enteric bacteria secrete amyloid proteins that self-assemble into fibrillar structures called curli, forming a key component of biofilm extracellular matrices. Bacterial biofilms confer resilience to harsh environments with broad implications for human health. In this study, we extend transient amyloid binding to the novel application of super-resolution fluorescence microscopy of curli on living cells, offering promising approaches to gain structural and mechanistic insights.

biophysics↗

bioRxiv2025

The bacterial chaperone CsgC inhibits functional amyloid CsgA formation by promoting the intrinsically disordered pre-nuclear state

E. coli assembles a functional amyloid called curli during biofilm formation. The major curlin subunit is the CsgA protein, which adopts a beta-sheet rich fold upon fibrillization. The chaperone-like protein CsgC inhibits CsgA amyloid formation. CsgA undergoes a 3-stage aggregation process: an initial lag phase where beta-rich nuclei form, an exponential elongation phase, and a plateau phase. It is currently not known if CsgC inhibits amyloid formation by inhibiting formation of a pre-fibril nucleus, or if CsgC inhibits a later stage of amyloid formation by blocking monomer addition. Here, CsgC homologs from C. youngae, C. davisae, and H. alvei were purified and characterized for their ability to interrogate CsgA amyloid formation. Each of the CsgC homologs prolonged the lag phase of E. coli CsgA amyloid formation similar to E. coli CsgC. Additionally, we found E. coli CsgC interacted transiently and weakly with a monomeric, pre-nucleus species of CsgA which delayed amyloid formation. A transient CsgC-CsgA heterodimer was observed using ion mobility-mass spectrometry. When CsgC was added to actively polymerizing CsgA, exponential growth commonly associated with nucleation-dependent amyloid formation was lost. Adding preformed CsgA seeds did not rescue exponential growth, indicating that CsgC also has inhibitory activity during fibril elongation. Indeed, CsgC interacted strongly with CsgA fibers, suggesting the interaction between CsgC and CsgA fibers can slow new fiber growth. CsgC displays unique inhibitory activity at multiple stages of amyloid formation. CsgC acts as an energy-independent chaperone that transiently interacts with prefibrillar CsgA and an amyloid fiber.

biochemistry↗

bioRxiv2022

Cell Surface-localized CsgF Condensate is a Gatekeeper in Bacterial Curli Subunit Secretion

Curli are functional amyloids present on the outer membrane of E. coli. Cell-surface aggregation of CsgA, the major subunit of curli, is a well-orchestrated process. CsgB, the minor subunit of curli, nucleates the aggregation of CsgA while CsgF, a curli accessory protein, ensures proper anchoring of CsgB to the cell surface. The molecular basis of the interactions between CsgF and curli subunits is unclear. Here, we show that CsgF undergoes phase separation in vitro and that the ability of CsgF variants to phase separate tightly correlated with CsgF function in cells during curli biogenesis. Substitution of phenylalanine residues in the CsgF N-terminus both reduced the propensity of CsgF to phase-separate and impaired curli biogenesis. Exogenous addition of purified CsgF complemented csgF - cells. This exogenous addition assay was used to assess the ability of CsgF variants to complement csgF - cells. The presence of CsgF on the cell surface modulated the secretion of CsgA to the cell surface. We also found that the CsgB nucleator protein is a CsgF client. CsgB can form SDS-insoluble aggregates within the dynamic CsgF condensate, and we propose that these multi-component CsgF-B condensates form a nucleation-competent complex that templates CsgA amyloid formation on the cell surface. Together, our study provides insight into the ability of CsgF to phase separate, regulate CsgA secretion, and promote CsgB aggregation in curli assembly.

biochemistry↗