Biology subjects

Ross, B. L.

Publications and source records attributed to Ross, B. L..

7 recordsLinked to original sources

bioRxiv2026

Molecular crowding differentially regulates magnetic field-dependent fluorescence, recovery kinetics, and photostability of the magnetosensitive protein MagLOV2

Macromolecular crowding and solvent composition can alter protein conformation, diffusion, hydration, and cofactor photophysics, yet their influence on magnetically sensitive flavoproteins is poorly understood. We tested seven molecular environments in MagLOV2-overexpressing Escherichia coli lysate using a custom-built, high-intensity magnetofluorescence platform. Five-point concentration series of Dextran 70, Ficoll PM70, PEG6000, PEG3350, glycerol, ethylene glycol, and bovine serum albumin (BSA) were measured at 22 mT under continuous 6.12 W/cm2 blue-light excitation. Dextran 70, Ficoll PM70, and PEG6000 progressively attenuated the negative magnetic field effect (MFE), whereas PEG3350 shifted toward mild enhancement at higher concentrations. Glycerol and ethylene glycol enhanced the negative MFE despite producing the strongest slowing of recovery upon the field being switched off (OFF-recovery), while BSA modestly enhanced MFE and accelerated OFF-recovery. Across 105 treatment cuvettes, the change in MFE was not globally correlated with changes in the OFF-recovery time constant (r = -0.07, p = 0.449). The initial pre-field illumination period revealed a second phenotype: Dextran and Ficoll increased late fluorescence loss, whereas glycerol and BSA were photoprotective. Pre-field fluorescence loss correlated moderately with MFE change across the full dataset (r = 0.474, p = 3.28 x 10-7) but not consistently within crowder classes. These findings show that MagLOV2 spin-dependent magnetofluorescence is shaped by crowder chemistry, polymer size, solvent environment, recovery kinetics, and photostability rather than by a single generic crowding or viscosity mechanism.

biochemistry↗

bioRxiv2026

Spatial profiling of Kawasaki disease cardiac inflammation identifies STING as a therapeutic target

The cellular subsets and in situ mechanisms underlying the development of coronary arteritis in Kawasaki disease (KD), the leading cause of acquired heart disease in children, remain poorly defined. We characterized the cellular composition and spatial organization of cardiac inflammation in autopsy coronary artery (CA) specimens from KD patients using 64-plex immunofluorescence. We identified spatially organized immune infiltrates of neutrophils, macrophages, and T cells associated with arterial wall destruction and remodeling, as well as disrupted stromal/muscle cell colocalization and diffuse myocardial inflammation. Spatial analysis also revealed expression of the innate immune sensor STING in myeloid cells in KD CA tissues. Pharmacological and genetic inhibition of STING reduced vasculitis development in a mouse model of KD. These findings define the spatial immune architecture of KD cardiac inflammation, reveal inflammatory injury beyond the coronary arteries, and identify STING as a potential therapeutic target for treatment-resistant or severe KD.

immunology↗

bioRxiv2026

Naturally cysteine-less LOV domains from halophilic archaea exhibit magnetic field effects on their fluorescence

Magnetic fields can modulate the function of certain flavoproteins through the radical pair mechanism, in which they influence the spin evolution of a coherent pair of radicals. This process can result in magneto-fluorescence, in which magnetic fields modulate the intensity of fluorescence emitted from the flavin cofactor. The prevalence of this property across protein families and domains of life, however, remains poorly characterized. In canonical Light-Oxygen-Voltage (LOV) domains, a conserved cysteine forms an adduct with the flavin that leads to downstream signaling. Mutating this cysteine instead yields signaling through the neutral semiquinone radical form, and the same mutation was crucial for enhancing magneto-fluorescence in the engineered protein MagLOV2. We therefore hypothesized that natural LOV domains that lack this conserved cysteine may exhibit magneto-fluorescence. Using a custom magneto-fluorescence imaging platform, we measured the fluorescence of E. coli colonies expressing three such domains, as well as a single mutant of one of them, under a switched external field. HsuLOV, from a halophilic archaeon, exhibited magneto-fluorescence, as did a single mutant of BAT-LOV, from a second halophilic archaeon. The domain amb2291 from magnetotactic bacteria, on the other hand, showed no detectable response under our illumination conditions. Magneto-fluorescence is therefore not exclusive to engineered proteins but is present in natural cysteine-less LOV domain sequences. The proteins reported here are, to our knowledge, the first magnetosensitive proteins reported from archaea. This suggests that radical-pair magnetosensitivity may be more widespread across LOV domains than previously appreciated.

biophysics↗

bioRxiv2026

An Open-Source Magnetofluorescence Imaging Platform forPlate-Scale Screening of Magnetic Field Effects in LiveBacteria

Magnetic field effects (MFEs) in biological systems are typically small and experimentally challenging to measure reproducibly across large sample populations. Existing approaches to measure such effects often rely on low-throughput microscopy or custom-built magnetic stimulation systems that provide limited control over magnetic field geometry, synchronization, or experimental automation. Here, we present an open-source magnetofluorescence imaging platform designed for bacterial plate-scale screening of MFEs in live colonies. The instrument integrates a programmable three-axis vector electromagnet, synchronized fluorescence excitation and imaging, and integrated acquisition software with per-frame metadata logging on a hardware-synchronized data acquisition card. An extensive calibration procedure enables accurate generation of arbitrary magnetic field vectors, while synchronized triggering ensures deterministic alignment between field application, illumination, and image acquisition. The system images an entire 100 mm Petri dish in a single acquisition. Typical experiments monitor hundreds of bacterial colonies simultaneously over multi-hour acquisition sequences. Control software, calibration routines, mechanical design files, and acquisition workflows are provided openly to facilitate replication. Instrument performance is demonstrated through detection of magnetic field-dependent fluorescence changes in E. coli expressing the engineered magnetosensitive fluorescent protein MagLOV2. This instrument provides a flexible and scalable platform for high-throughput magnetobiology, synthetic biology, and quantum biology experiments.

bioengineering↗

bioRxiv2026

Measuring magnetic field effects in fluorescent flavoproteins via spin-dependent fluorescence intensity requires photoexcitation to be faster than spin-independent ground state recovery

Weak magnetic fields affect many biological processes across the tree of life, though the precise molecular sensors and pathways involved in such magnetoresponses remain mostly uncharacterized. Fluorescence is a useful tool for investigating magnetic field effects in flavoproteins, as their chromophores fluorescence intensity can be shown to depend on the spin states of electronic radical pairs. Here, we describe a four-state ordinary differential equation model to understand what parameter sets result in fluorescence contrast between spin states in photocycles with singlet and triplet radical pairs. We conclude that only certain sets of parameters result in the fluorescence intensity being a good proxy measurement for singlet yield. In particular, we observe that the illumination intensity required to obtain fluorescence contrast depends on the rate of the slow spin-independent radical termination reactions that recover ground-state oxidized fluorophores. Moreover, to observe a magnetic field effect in fluorescence intensity when an external magnetic field modulates the singlet yield, the illumination intensity must be strong enough such that photoexcitation is not the rate-limiting step. This understanding suggests that flavoproteins that do not exhibit magnetic field effects in their fluorescence emission under certain experimental setups may still be sensitive to weak magnetic fields in terms of function, as magnetosensitivity in fluorescence depends strongly on illumination conditions.

biophysics↗

bioRxiv2026

The magnetic field-dependent fluorescence of MagLOV2 in live bacterial cells is consistent with the radical pair mechanism

MagLOV2 is an engineered flavoprotein designed to have large changes in fluorescence intensity in response to weak magnetic fields. Here, we characterize the magnitude of these fluorescence changes, known as the "magnetic field effect," as a function of the strength of an externally applied magnetic field in E. coli colonies expressing MagLOV2. We observe that the magnetic field effect is positive at low magnetic fields, reaches a maximum positive value near 1 mT, and then decreases, reversing sign at approximately 2 mT. Furthermore, the effect starts to plateau above approximately 70 mT, with a decreased sensitivity of fluorescence changes to magnetic fields above this range. The non-monotonic behavior, as well as the diminished responsiveness to higher magnetic fields, are consistent with the changes in fluorescence being driven by electron spin-dependent chemical processes governed by the radical pair mechanism.

biophysics↗

bioRxiv2025

Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

BACKGROUNDSirtuin 1 (SIRT1), a NAD+-dependent protein deacetylase, regulates cardiovascular inflammation by modulating cellular stress, inhibiting NLRP3 activation, and promoting the clearance of damaged mitochondria. However, its precise role in the pathogenesis of Kawasaki disease (KD), a pediatric systemic vasculitis and the leading cause of acquired heart disease in children, remains unclear. METHODSUsing the Lactobacillus casei cell wall extract (LCWE) murine model of KD, we evaluated the severity of vasculitis in mice supplemented with NAD+ precursors, as well as transgenic mice overexpressing SIRT1, and mice with specific deletion of Sirt1 in vascular smooth muscle cells (VSMCs) and myeloid cells. Proteomics analysis was performed on the abdominal aortas of WT and SIRT1-overexpressing mice. We performed immunofluorescent staining of cardiovascular tissues to assess the expression of proteins related to the autophagy/mitophagy pathway and the pathogenic switch of VSMCs. Western blot analysis was performed on primary VSMCs and cardiovascular tissues to determine the impact of SIRT1 on autophagic flux. The production of pro-inflammatory cytokines was measured in bone marrow-derived macrophages and peritoneal lavage of transgenic mice using ELISAs. RESULTSSIRT1 expression was downregulated in cardiovascular lesions of LCWE-injected mice, which was associated with a significant reduction of circulating levels of nicotinamide. Supplementation of mice with NAD+ precursors or genetic overexpression of SIRT1 significantly reduced the development of LCWE-induced KD, while the specific deletion of Sirt1 in VSMCs or myeloid cells exacerbated vasculitis. Proteomics analysis indicated impaired mitophagy/autophagy and the pathogenic synthetic switch of VSMCs in LCWE-injected mice, which was rescued with SIRT1 overexpression and associated with reduced production of proinflammatory cytokines. CONCLUSIONSThis study reveals the presence of an impaired NAD+-SIRT1 axis in the pathogenesis of LCWE-induced KD vasculitis and the therapeutic potential of targeting this axis to reduce cardiovascular lesions and inflammation.

immunology↗