Elucidating the Biomechanics of MERTK-Mediated Efferocytosis

Brandon Hayato Dickson, Western University

Abstract

Macrophages are key mediators of efferocytosis – the phagocytic engulfment and removal of apoptotic cells. During engulfment, the coordinated activity of efferocytic receptors induces the remodeling of the actin cytoskeleton, which facilitates the envelopment of the cell by the plasma membrane. Mer receptor tyrosine kinase (MERTK) is a crucial efferocytic receptor, but its role during actin remodeling is not well understood. Previously, our lab showed that MERTK is an activator of β₂ integrins – which are comprised of receptors known to induce the actin polymerization that is required for engulfment. We hypothesized that MERTK is an indirect stimulator of actin polymerization via the stimulation of the actin-remodeling β₂ integrins. Herein, the biomechanics of efferocytosis were investigated by fluorescence imaging of the efferocytic synapse. In a frustrated efferocytosis model – where macrophages attempted to engulf a planar substrate that mimicked apoptotic cells, MERTK formed discrete clusters within the centre of the efferocytic synapse, where the periphery of the synapse was delineated by an expanding ring of actin-associated β₂ integrins. This suggests that integrins, not MERTK, are local directors of actin polymerization. Single particle tracking revealed that MERTK was less restricted in its free diffusion compared to the highly confined β₂ integrins, indicating these receptors play unique biomechanical roles during engulfment. Finally, we began the development of a traction force microscopy assay to characterize the biophysics of MERTK-dependent engulfment. Preliminary data revealed that frustrated efferocytosis on opsonized hydrogels required the simultaneous activity of MERTK and efferocytic integrin receptors. We expect that integrins transduce the forces required for engulfment via the actin cytoskeleton, whereas MERTK stimulates the initial activation of the integrins. Further elucidation of the mechanism of MERTK-mediated engulfment will be critical in our understanding of pathologies where efferocytosis is defective, such as systemic lupus erythematosus and atherosclerosis.