The clearance of apoptotic cells has been identified as an efficient process under physiological conditions, even in tissues with high rate of cell death such as the thymus. In addition, the removal of cells through apoptosis promotes anti-inflammatory responses and control autoimmune responses, opposite to the clearance of necrotic and pathogen cells. Nevertheless, failure in the clearance of dying cells has been linked with multiple diseases involving inflammation, infection, autoimmunity and even cancer (Poon et al., 2014).
Brain-specific angiogenesis inhibitor 1 (BAI1) is a membrane receptor found to recognize phosphatidylserine (PS) on apoptotic cells. BAI1 was identified to recognize PS via the 5 Thrombospondin type 1 repeat (TSR) domains at the extracellular region of this receptor. Moreover, BAI1 promotes the engulfment of targets upon their recognition by activating intracellular signals that enables the rearrangement of the cytoskeleton membrane of phagocytes (Park et al., 2007). My project aims to characterise how the TSR domains of BAI1 mediate binding to PS on apoptotic cells, as well as identify particular TSRs involved in the recognition of PS.
To date, I have been able to cloned, expressed and purified multiple constructs containing different truncations of the 5 TSRs of BAI1. These purified proteins have been used to perform a competition assay using annexin5 (binds PS). This experiment allowed me to identify that among all the constructs tested, only the construct containing the TSR1-2 of BAI1 was able to significantly inhibit the binding of annexin5 to PS on apoptotic monocytes. Furthermore, mutations at the linker sequence between the TSR1-2 have reduced the ability of the TSR1-2 to inhibit annexin5-PS binding, proving that the linker is particularly involved in the recognition of PS.