ApoL6 Induces Dichotomous Cell Death Phenotype Involving Both Apoptosis and Necroptosis in Cancer Cells

Abstract

Apolipoprotein L6 (ApoL6) is a pro-death, BH3-only and phospholipid-binding member of the Bcl-2 family. Previously, we showed that ectopic expression of ApoL6 induces mitochondria-mediated apoptosis. In this study, we hypothesized that ApoL6 plays a dichotomous role in regulated cell death (RCD) pathways that involves both apoptosis and necroptosis. Necroptosis, or programmed necrosis, can occur simultaneously or alternatively to apoptosis as a caspase-independent pathway, and is characterized by the formation of intracellular necrosomes and plasma membrane pores. The pseudokinase mixed lineage kinase domain-like protein (MLKL) interacts with the necrosome to become phosphorylated and activated. Phosphorylated (p-) MLKL molecules migrate to the plasma membrane to form pores, disrupt membrane integrity, and selectively release intracellular components. The proinflammatory cytokine IL-1β is amongst a number of molecules released in this process. To investigate the role of ApoL6 in RCD, ApoL6 was overexpressed via a Tet-Off gene inducible system in a p53 null colorectal cancer cell line DLD-1. Necroptotic cell death was marked by an increased level of intracellular and plasma membrane p-MLKL and extracellular release of IL-1β along with the disruption of the plasma membrane. We found that in the presence of apoptotic inhibitors, levels of necroptotic cell death increased, indicating an enhanced push away from apoptosis. We also showed that ApoL6 preferentially binds with phosphatidylinositol phosphate species (PIPs), which include PI (4,5) P2, PI4P and PI5P. Thus, we expand on the role of ApoL6 in RCD to induce a mixed cell-death phenotype that includes both apoptosis and necroptosis.