Molecular mechanisms of cell death and cell clearance

Abstract: Programmed cell death (PCD) is a naturally occurring event in multicellular organisms. It is part of the development as well as ensuring tissue homeostasis and cellular turnover. The cell death process and the removal of the dying cells has to be tightly regulated since dysregulated cell death and cell clearance pathways are correlated with various pathological conditions. Notwithstanding the critical importance of individual cell death modes and of cell clearance for the well-being of the organism, our knowledge regarding the underlying mechanisms as well as its consideration in various toxicological or medical settings is still rather limited. The overall aim of this thesis is to shed light on different aspects regarding the mechanisms of programmed cell death and cell clearance. Emphasis was put on the exposure of phosphatidylserine (PS) – a well studied ‘eat-me’ signal that is known to facilitate recognition and engulfment of dying cells by phagocytes and is known to be an evolutionarily conserved signal. Moreover, we combined a nanotoxicological study with the elucidation of the underlying cell death pathways. Paper I addresses the effect of different single point mutations on the function of the Caenorhabditis elegans aminophospholipid translocase TAT-1 – a protein that was shown to prevent the externalization of PS in the membranes and that is important for endocytic transport. This in vivo study shows for the first time that two conserved motifs of TAT-1 – located in the transmembrane domain four and in the following intracellular domain, respectively – are critical for proper protein function. In paper II, apoptosis, necroptosis or ferroptosis was induced in Jurkat cells and cell death was further characterized regarding morphological and biochemical properties. Cell clearance by primary human macrophages was investigated. Interestingly, all three forms of PCD express PS – even though this was previously thought to be a signal specific for apoptotic cells. Apoptotic cells were phagocytosed more efficiently compared to cells undergoing other cell death modes. In paper III, we studied the effect of size and surface functionalization on the toxic properties of gold nanoparticles using multi-omics studies in combination with validation experiments. We found that only the cationic gold nanoparticles caused toxicity and mitochondrial dysfunction. These particles triggered apoptosis, but at high doses cells died by necrosis. The cationic particles also caused in vivo lethality in C. elegans while carboxylated particles were non-toxic. Taken together, the ‘eat-me’ signal PS and the mechanisms leading to its exposure are of central importance for clearance of dying cells. With this work we elucidate the relevance of conserved regions in P4-type ATPases for its PS transport function. Moreover, we highlight that PS externalization is not unique for apoptotic cells and that macrophages differ in the recognition and uptake of different forms of PCD. Finally, we point out the importance to study the mechanisms of cell death in a toxicological setting using multi-omics approaches combined with validation experiments both in vitro and in vivo.