Biology subjects

Zhong, B. L.

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

4 recordsLinked to original sources

bioRxiv2026

Cytosolic MagLOV Magnetofluorescence in Mammalian Cell Lines and Primary Neurons

Magnetic fields can influence the outcome of photochemical reactions through the radical pair mechanism, but whether this sensitivity extends to standard experimental conditions in mammalian cells has remained unclear. Here, we quantify the cytosolic magnetofluorescence of MagLOV, a genetically encoded, flavin-binding fluorescent protein, in mammalian cells, validated against extensive artifact controls. Excitation intensity and magnetic field strength differentially tune response kinetics and amplitude, respectively, with amplitude saturating above approximately 8 mT. Magnetic field-effect amplitude is further modulated by cellular culture state. This response generalizes across HEK293T, HeLa, U2OS, and A549 cells and primary mouse cortical neurons, with plateau amplitudes ranging from 1.4% to 2.7%. Together, these results establish that genetically encoded spin-dependent photochemistry can be quantitatively interrogated under standard mammalian live-cell imaging conditions, and generalizes across mammalian cell types.

biophysics↗

bioRxiv2023

Piconewton forces mediate GAIN domain dissociation of the latrophilin-3 adhesion GPCR

Latrophilins are adhesion G-protein coupled receptors (aGPCRs) that control excitatory synapse formation. aGPCRs, including latrophilins, are autoproteolytically cleaved at their GPCR-Autoproteolysis Inducing (GAIN) domain, but the two resulting fragments remain associated on the cell surface. It is thought that force-mediated dissociation of the fragments exposes a peptide that activates G-protein signaling of aGPCRs, but whether GAIN domain dissociation can occur on biologically relevant timescales and at physiological forces is unknown. Here, we show using magnetic tweezers that physiological forces dramatically accelerate the dissociation of the latrophilin-3 GAIN domain. Forces in the 1-10 pN range were sufficient to dissociate the GAIN domain on a seconds-to-minutes timescale, and the GAIN domain fragments reversibly reassociated after dissociation. Thus, mechanical force may be a key driver of latrophilin signaling during synapse formation, suggesting a physiological mechanism by which aGPCRs may mediate mechanically-induced signal transduction.

biophysics↗

bioRxiv2022

STReTCh: a strategy for facile detection of mechanical forces across proteins in cells

Numerous proteins experience and respond to mechanical forces as an integral part of their cellular functions, but measuring these forces remains a practical challenge. Here, we present a compact, 11 kDa molecular tension sensor termed STReTCh (Sensing Tension by Reactive Tag Characterization). Unlike existing genetically encoded tension sensors, STReTCh does not rely on experimentally demanding Forster resonance energy transfer (FRET)-based measurements and is compatible with typical fix-and-stain protocols. Using a magnetic tweezers assay, we calibrate the STReTCh module and show that it responds to physiologically relevant, piconewton forces. As proof-of-concept, we use an extracellular STReTCh-based sensor to visualize cell-generated forces at integrin-based adhesion complexes. In addition, we incorporate STReTCh into vinculin, a cytoskeletal adaptor protein, and show that STReTCh reports on forces transmitted between the cytoskeleton and cellular adhesion complexes. These data illustrate the utility of STReTCh as a broadly applicable tool for the measurement molecular-scale forces in biological systems.

bioengineering↗

bioRxiv2020

Complete Reconstitution and Deorphanization of the 3 MDa NOCAP (NOCardiosis-Associated Polyketide) Synthase

Several Nocardia strains associated with nocardiosis, a potentially life-threatening disease, house a nonamodular assembly-line polyketide synthase (PKS) that presumably synthesizes an unknown natural product. Here, we report the discovery and structure elucidation of the NOCAP (NOCardiosis-Associated Polyketide) aglycone by first fully reconstituting the NOCAP synthase in vitro from purified protein components followed by heterologous expression in E. coli and spectroscopic analysis of the purified products. The NOCAP aglycone has an unprecedented structure comprised of a substituted resorcylaldehyde headgroup linked to a 15-carbon tail that harbors two conjugated all-trans trienes separated by a stereogenic hydroxyl group. This report is the first example of reconstituting a trans-acyltransferase assembly-line PKS either in vitro or in E. coli, and of using these approaches to "deorphanize" a complete assembly-line PKS identified via genomic sequencing. With the NOCAP aglycone in hand, the stage is set for understanding how this PKS and associated tailoring enzymes confer an advantage to their native hosts during human Nocardia infections.

biochemistry↗