Electronic Thesis and Dissertation Repository

Thesis Format

Monograph

Degree

Master of Science

Program

Microbiology and Immunology

Supervisor

Heit, Bryan

Abstract

MER Tyrosine Kinase (MERTK) is a receptor that mediates efferocytosis - the phagocytosis of apoptotic cells. Efferocytosis is central to homeostasis through removing the ~150 billion apoptotic cells generated each day in the human body. Defects in MERTK function have been shown to contribute to both autoimmune diseases and chronic inflammatory diseases, and MERTK is known to function as the predominant or sole efferocytic receptor in multiple tissues. Despite its clinical importance, very little is understood of how MERTK functions and how it signals. MERTK is known to bind to apoptotic cells via opsonins such as Gas6 and Protein S, with binding activating MERTK’s intrinsic kinase domain. This induces a poorly characterized signaling pathway which is thought to coordinate with integrins to mediate efferocytosis. Using immunoprecipitation and mass spectrometry we have identified a large pre-formed MERTK signalosome on the cell surface containing MERTK, integrins and a number of lipid and protein kinases. Ligation of this complex activates a number of downstream signaling pathways required for efferocytosis, including the canonical Src/Syk/PI3K phagocytic signaling pathway and non-canonical ILK and Erk-dependent signaling pathways. Through ILK and FAK, MERTK signaling activates integrins, and indeed, in macrophages MERTK-mediated efferocytosis requires αxβ2 integrins. Combined, these data demonstrate that MERTK engages in efferocytosis cooperatively with integrins, and mediates this cross-talk through engaging canonical integrin signaling pathways.

Summary for Lay Audience

Everyday our body replaces billions of cells. These cells need to be replaced due to damage or aging. These cells are removed through a special form of cell death called apoptosis, where the cells deliberately disassemble themselves into small “packages” that can be easily removed. These packages are removed by specialized cells, including a type of immune cell called “macrophages”. This removal process is called efferocytosis, and during efferocytosis the macrophage will engulf and break down the dying cell. This process is very important for the maintenance of our bodies, and without it our tissues accumulate dying cells and begin to break down. MERTK is critical to efferocytosis – it is a receptor on macrophages that allows them to detect the presence of dying cells. Defects in MERTK function leads to several inflammatory and immune related diseases, including heart disease. However very little is known about how MERTK functions, and how it allows macrophages to remove dying cells. We have found that MERTK is assembled into large clusters along with multiple other proteins. We have identified several of these MERTK-associated proteins and shown that several of these proteins are required for macrophages to efferocytose dying cells. These critical proteins include ILK and FAK, which allow MERTK to communicate with another receptor known as αxβ2 integrin. MERTK cannot engage in efferocytosis without αxβ2 integrin, and we have shown that MERTK first identifies the dying cell, then activates αxβ2 integrin via ILK and FAK, with αxβ2 integrin then binding to the dying cell strongly enough for the macrophage engulf the dying cell. Together, these experiments demonstrate how MERTK mediates efferocytosis and demonstrates an important role of αxβ2 integrin in this process. Using this information, it may be possible to develop better treatments for diseases driven by defects in efferocytosis, including heart disease – the most common cause of death in the world.

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