Biology subjects

Sridharan, V.

Publications and source records attributed to Sridharan, V..

4 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

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↗

bioRxiv2021

The M2 Gene Is a Determinant of Reovirus-Induced Myocarditis

Although a broad range of viruses cause myocarditis, the mechanisms that underlie viral myocarditis are poorly understood. Here, we report that the M2 gene is a determinant of reovirus myocarditis. The M2 gene encodes outer capsid protein 1, which mediates host membrane penetration during reovirus entry. We infected newborn C57BL/6 mice with reovirus strain type 1 Lang (T1L) or a reassortant reovirus in which the M2 gene from strain type 3 Dearing (T3D) was substituted into the T1L genetic background (T1L/T3DM2). T1L was non-lethal in wild-type mice, whereas greater than 90% of mice succumbed to T1L/T3DM2 infection. T1L/T3DM2 produced higher viral loads than T1L at the site of inoculation. In secondary organs, T1L/T3DM2 was detected with more rapid kinetics and reached higher peak titers than T1L. We found that hearts from T1L/T3DM2-infected mice were grossly abnormal, with large lesions indicative of substantial inflammatory infiltrate. Lesions in T1L/T3DM2-infected mice contained necrotic cardiomyocytes with pyknotic debris, and extensive lymphocyte and histiocyte infiltration. In contrast, T1L induced the formation of small purulent lesions in a small subset of animals, consistent with T1L being mildly myocarditic. Finally, more activated caspase-3-positive cells were observed in hearts from animals infected with T1L/T3DM2 compared to T1L. Together, our findings indicate that substitution of the T3D M2 allele into an otherwise T1L genetic background is sufficient to change a non-lethal infection into a lethal infection. Our results further indicate that T3D M2 enhances T1L replication and dissemination in vivo, which potentiates the capacity of reovirus to cause myocarditis. IMPORTANCEReovirus is a non-enveloped virus with a segmented double-stranded RNA genome that serves as a model for studying viral myocarditis. The mechanisms by which reovirus drives myocarditis development are not fully elucidated. We found that substituting the M2 gene from strain type 3 Dearing (T3D) into an otherwise type 1 Lang (T1L) genetic background (T1L/T3DM2) was sufficient to convert the non-lethal T1L strain into a lethal infection in neonatal C57BL/6 mice. T1L/T3DM2 disseminated more efficiently and reached higher maximum titers than T1L in all organs tested, including the heart. T1L is mildly myocarditic and induced small areas of cardiac inflammation in a subset of mice. In contrast, hearts from mice infected with T1L/T3DM2 contained extensive cardiac inflammatory infiltration and more activated caspase-3-positive cells, which is indicative of apoptosis. Together, our findings identify the reovirus M2 gene as a new determinant of reovirus-induced myocarditis.

microbiology↗

bioRxiv2019

Structure of E3 ligase E6AP with a novel proteasome-binding site provided by substrate receptor hRpn10

Regulated proteolysis by the proteasome involves [~]800 enzymes for substrate modification with ubiquitin, of which [~]600 are E3 ligases. We report here that E6AP/UBE3A is distinguished from other ubiquitin E3 ligases by having a 12 nM binding site at the proteasome contributed by substrate receptor hRpn10/PSMD4/S5a. Intrinsically disordered by itself, and previously uncharacterized, this domain in hRpn10 locks into a novel well-defined helical structure to form an intermolecular 4-helix bundle with the E6AP AZUL domain, which is unique to this E3. We thus name the hRpn10 AZUL-binding domain RAZUL. We further find in human cells that loss of RAZUL by CRISPR-based gene editing leads to loss of E6AP at the proteasome, where associated ubiquitin is correspondingly reduced, suggesting that E6AP ubiquitinates substrates at or for the proteasome. Altogether, our findings indicate E6AP to be a privileged E3 for the proteasome, with a dedicated, high affinity binding site contributed by hRpn10.

molecular biology↗