Tseng, P. L. | bioRxiv Science

bioRxiv2025

An unbiased screening approach for identifying proteins with a mechanosensitive nuclear localization

Mechanical forces profoundly influence a wide range of cellular behaviors. However, the mechanisms by which these forces are sensed and transduced into specific biological signals remain incompletely understood. One established principle of mechanotransduction is the mechanical regulation of subcellular protein localization, which enables control over the activity of key regulators such as transcription and splicing factors. In this protocol, we present a screening approach to identify proteins that exhibit mechanosensitive nuclear localization. We describe the engineering of cell lines with tuneable actomyosin contractility and the application of a spatially restricted in situ biotinylation strategy. This method, combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS), enables the identification of proteins whose nuclear localization changes in response to mechanical cues. For a detailed application of this protocol, please refer to Tseng et al. (2025) [1]

cell biology↗

bioRxiv2024

Mechanical control of the splicing factor PTBP1 regulates extracellular matrix stiffness-induced cell proliferation and mechanomemory

Cells sense and respond to mechanical cues from their environment. Mechanical cues are important for many biological processes, including embryonic development, ageing, cellular homeostasis, and diseases. Cells translate mechanical cues into cellular biochemical signals that govern cellular behaviour, like cell proliferation or migration, via a process called mechanotransduction. However, this process and the proteins involved remain incompletely understood. Here, we present an unbiased and large-scale approach to identify proteins involved in mechanotransduction. The screen revealed that the splicing factor PTBP1 is a novel mechanotransducer. We show that the nuclear localisation of PTBP1 depends on extracellular matrix stiffness, cell density, and the actomyosin-based contractility of the cell. Furthermore, we demonstrate that PTBP1 promotes the mechanosensitive splicing of the adapter protein Numb and that alternative splicing of Numb is crucial for matrix stiffness-induced cell proliferation and mechanomemory. Our results support the idea that changes in alternative splicing are an integral part of mechanotransduction and provide a mechanism by which matrix stiffness regulates cell proliferation and the formation of a mechanomemory in cells.

cell biology↗