Biology subjects

Kota, D.

Publications and source records attributed to Kota, D..

5 recordsLinked to original sources

bioRxiv2026

Counteraction of HMGB1 at ss-dsDNA junctions maintains liquidity of protamine-DNA co-condensates

In the sperm nucleus, protamine replaces histones to mediate extreme DNA compaction. The histone-to-protamine transition involves the occurrence of double-strand breaks, and is facilitated by transition proteins including those containing high-mobility-group (HMG) boxes. Here we used optical tweezers and microscopy to study the actions of HMGB1 and protamine on DNA. Confocal scans of GFP-HMGB1 on overstretched {lambda}-DNA show 2-3 foci that spread on the DNA upon retraction. Spreading of foci coincides with reannealing of ssDNA tracks, confirming their localization at ss-dsDNA junctions. Whereas the force-extension curves of protamine-bound {lambda}-DNA show tangles that withstand forces > 60 pN, premixing protamine with HMGB1 produces only bends and bridges ([~] 20 pN). The counteraction of HMGB1 involves its acidic C-terminal tail, as HMGB1-{Delta}C fails to prevent tangle formation. In line with these single-molecule results, brightfield and confocal imaging shows that HMGB1 converts protamine-dsDNA aggregates into liquid droplets whereas HMGB1-{Delta}C fails to do so. Together, these observations support our hypothesis that chromatin-associated proteins like HMGB1 help maintain early protamine-mediated DNA condensates in a liquid state, enabling the recruitment of the repair machinery to restore the duplex structure.

biophysics↗

bioRxiv2023

ATP Mediates Phase Separation of Disordered Basic Proteins by Bridging Intermolecular Interaction Networks

ATP is an abundant molecule with crucial cellular roles as the energy currency and a building block of nucleic acids and for protein phosphorylation. Here we show that ATP mediates the phase separation of basic intrinsically disordered proteins (bIDPs). In the resulting condensates, ATP is highly concentrated (apparent partition coefficients at 200-5000) and serves as bridges between bIDP chains. These liquid-like droplets have some of the lowest interfacial tension ([~]25 pN/m) but high zero-shear viscosities (1-15 Pa s) due to the bridged protein networks, and yet their fusion has some of the highest speeds ([~]1 m/ms). The rapid fusion manifests extreme shear thinning, where the apparent viscosity is lower than zero-shear viscosity by over 100-fold, made possible by fast reformation of the ATP bridges. At still higher concentrations, ATP does not dissolve bIDP droplets but results in aggregates and fibrils.

biophysics↗

bioRxiv2021

Shear Relaxation Governs Dynamic Processes of Biomolecular Condensates

Phase-separated biomolecular condensates must respond agilely to biochemical and environmental cues in performing their wide-ranging cellular functions, but our understanding of condensate dynamics is lagging. Ample evidence now indicates biomolecular condensates as viscoelastic fluids, where shear stress relaxes at a finite rate, not instantaneously as in viscous liquids. Yet the fusion dynamics of condensate droplets has only been modeled based on viscous liquids, with fusion time given by the viscocapillary ratio (viscosity over interfacial tension). Here we used optically trapped polystyrene beads to measure the viscous and elastic moduli and the interfacial tensions of four types of droplets. Our results challenge the viscocapillary model, and reveal that the relaxation of shear stress governs fusion and other dynamic processes of condensates.

biophysics↗

bioRxiv2019

Myosin II Tension Sensors Visualize Force Generation within the Actin Cytoskeleton in Living Cells

Type II myosin motors generate cytoskeletal forces that are central to cell division, embryogenesis, muscle contraction, and many other cellular functions. However, at present there is no method that can directly measure the forces generated by myosins in living cells. Here we describe a Forster resonance energy transfer (FRET)-based tension sensor that can measure forces generated by Nonmuscle Myosin IIB (NMIIB) in living cells with piconewton (pN) sensitivity. Fluorescence lifetime imaging microscopy (FLIM)-FRET measurements indicate that the forces generated by NMIIB exhibit significant spatial and temporal heterogeneity, with inferred tensions that vary widely in different regions of the cell. This initial report highlights the potential utility of myosin-based tension sensors in elucidating the roles of cytoskeletal contractility in a wide variety of contexts.

cell biology↗

bioRxiv2018

Transparent Titanium Dioxide Nanotubes: Processing, Characterization, and Application in Establishing Cellular Response Mechanisms

The therapeutic applications of titanium dioxide nanotubes (TiO2 NTs) as osteogenic surface treatments for titanium (Ti)-based implants are largely due to the finely tunable physical characteristics of these nanostructures. As these characteristics change, so does the cellular response, yet the exact mechanisms for this relationship remains largely undefined. We present a novel TiO2 NT imaging platform that is suitable for use with live-cell imaging techniques, thereby enabling, for the first time, dynamic investigation of those mechanisms. In this work, fabrication methods for producing transparent TiO2 NTs with diameters of 56 {+/-} 6 nm, 75 {+/-} 7 nm, 92 {+/-} 9 nm, and 116 {+/-} 10 nm are described. To demonstrate the diagnostic potential of these TiO2 NT imaging platforms, the focal adhesion protein vinculin and actin cytoskeletal filaments were fluorescently tagged in osteoblasts and real-time, high-resolution fluorescent microscopy of live-cell interactions with TiO2 NT substrates were observed. The scope of such a platform is expected to extend far beyond the current proof-of-concept, with great potential for addressing the dynamic response of cells interacting with nanostructured substrates.

bioengineering↗