Unraveling the Dual-Stretch-Mode Impact on Tension Gauge Tethers' Mechanical Stability.

Tension gauge tethers (TGTs), short DNA segments serving as extracellular tension sensors, are instrumental in assessing the tension dynamics in mechanotransduction. These TGTs feature an initial shear-stretch region and an unzip-stretch region. Despite their utility, no theoretical model has been available to estimate their tension-dependent lifetimes [τ(f)], restricting insights from cellular mechanotransduction experiments. We have now formulated a concise expression for τ(f) of TGTs, accommodating contributions from both stretch regions. Our model uncovers a tension-dependent energy barrier shift occurring when tension surpasses a switching force of approximately 13 pN for the recently developed TGTs, greatly influencing τ(f) profiles. Experimental data from several TGTs validated our model. The calibrated expression can predict τ(f) of TGTs at 37 °C based on their sequences with minor fold changes, supporting future applications of TGTs.

In situ single-molecule investigations of the impacts of biochemical perturbations on conformational intermediates of monomeric α-synuclein.

α-Synuclein aggregation is a common trait in synucleinopathies, including Parkinson's disease. Being an unstructured protein, α-synuclein exists in several distinct conformational intermediates, contributing to both its function and pathogenesis. However, the regulation of these monomer conformations by biochemical factors and potential drugs has remained elusive. In this study, we devised an in situ single-molecule manipulation approach to pinpoint kinetically stable conformational intermediates of monomeric α-synuclein and explore the effects of various biochemical factors and drugs. We uncovered a partially folded conformation located in the non-amyloid-ß component (NAC) region of monomeric α-synuclein, which is regulated by a preNAC region. This conformational intermediate is sensitive to biochemical perturbations and small-molecule drugs that influencing α-synuclein's aggregation tendency. Our findings reveal that this partially folded intermediate may play a role in α-synuclein aggregation, offering fresh perspectives for potential treatments aimed at the initial stage of higher-order α-synuclein aggregation. The single-molecule approach developed here can be broadly applied to the study of disease-related intrinsically disordered proteins.

Alternative molecular mechanisms for force transmission at adherens junctions via ß-catenin-vinculin interaction.

Force transmission through adherens junctions (AJs) is crucial for multicellular organization, wound healing and tissue regeneration. Recent studies shed light on the molecular mechanisms of mechanotransduction at the AJs. However, the canonical model fails to explain force transmission when essential proteins of the mechanotransduction module are mutated or missing. Here, we demonstrate that, in absence of α-catenin, ß-catenin can directly and functionally interact with vinculin in its open conformation, bearing physiological forces. Furthermore, we found that ß-catenin can prevent vinculin autoinhibition in the presence of α-catenin by occupying vinculin´s head-tail interaction site, thus preserving force transmission capability. Taken together, our findings suggest a multi-step force transmission process at AJs, where α-catenin and ß-catenin can alternatively and cooperatively interact with vinculin. This can explain the graded responses needed to maintain tissue mechanical homeostasis and, importantly, unveils a force-bearing mechanism involving ß-catenin and extended vinculin that can potentially explain the underlying process enabling collective invasion of metastatic cells lacking α-catenin.

Tension Gauge Tethers as Tension Threshold and Duration Sensors.

Mechanotransduction, the process by which cells respond to tension transmitted through various supramolecular linkages, is important for understanding cellular behavior. Tension gauge tethers (TGTs), short fragments of double-stranded DNA that irreversibly break under shear-stretch conditions, have been used in live cell experiments to study mechanotransduction. However, our current understanding of TGTs' mechanical responses is limited, which limits the information that can be gleaned from experimental observations. In this study, we quantified the tension-dependent lifetime of TGTs to better understand their mechanical stability under various physiologically relevant stretching conditions. This work has broad applications for using TGTs as tension threshold and duration sensors and also suggests the need to revisit previous interpretations of experimental observations.