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The Effect of Thymosin Beta 4 on Cell Mechanics and Motility
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Dissertation
The Effect of Thymosin Beta 4 on Cell Mechanics and Motility
2014
Overview
Thymosin Beta 4 (TB4), a multi-functional 5 kDa peptide found at up to 500 µM concentration in virtually every mammalian cell, is the main G-actin sequestering protein. Extracellular concentrations are generally in the low nM range. Although no receptor for TB4 is known, higher extracellular levels of TB4 have been linked to enhanced cell migration and epithelial-mesenchymal transition. As these cellular effects involve substantial changes in the cytoskeleton, we measured the effect of TB4 on cytoskeletal mechanics, contractility, and motility. SW480 cells showed a distinct biphasic dose response with increasing concentrations of extracellular TB4. Cells stimulated with 0.2 µM TB4 showed the highest level of stiffness and contractility, displayed a pronounced elongated, mesenchymal morphology, migrated with higher directional persistence on flat 2D surfaces, and became highly invasive in a 3D collagen invasion assay. All parameters return to baseline levels at concentrations larger than 1 µM. Differential scanning calorimetry (DSC) and fluorescence correlation spectroscopy (FCS) measurements using fluorescently labelled TB4 and synthetic lipid membranes demonstrated that TB4 binds to the cell membrane (Kd=72 µM for a 1:5 DMPS/DMPC lipid ratio). The binding of TB4 to the cell membrane presents a novel mechanism linking small extracellular TB4 concentrations to pronounced cell mechanical responses.
Publisher
ProQuest Dissertations & Theses
Subject
ISBN
9798597013190
