Human Gasdermin D and MLKL disrupt mitochondria, endocytic traffic and TORC1 signaling in budding yeast

Abstract

Gasdermin D (GSDMD) and mixed lineage kinase domain-like protein (MLKL) are the pore-forming effectors of pyroptosis and necroptosis, respectively, with the capacity to disturb plasma membrane selective permeability and induce programmed cell death. The budding yeast Saccharomyces cerevisiae has long been used as a simple eukaryotic model for the study of proteins associated with human diseases by heterologous expression. In this work, we expressed in yeast both GSDMD and its N-terminal domain [GSDMD(NT)] to characterize their cellular effects, and compare them to those of MLKL. GSDMD(NT) and MLKL inhibited yeast growth, formed cytoplasmic aggregates, and fragmented mitochondria. Loss-of-function point mutants of GSDMD(NT) showed affinity for this organelle. Besides, GSDMD(NT) and MLKL caused an irreversible cell cycle arrest through TORC1 inhibition, and disrupted endosomal and autophagic vesicular traffic. Our results provide a basis for a humanized yeast platform to study GSDMD and MLKL, a useful tool for structure-function assays and drug discovery.