Elucidating the molecular mechanisms underlying defensosome biogenesis and release

About This Grant

ABSTRACT Autophagy, a conserved catabolic process mediated by autophagy-related (ATG) proteins, is integral for maintaining cellular homeostasis. Recent studies suggest that ATG proteins are also necessary for immune responses through processes that are mechanistically distinct from autophagy. The Cadwell lab has identified a novel innate immune mechanism whereby cells produce extracellular vesicles of endosomal origin (exosomes) which can neutralize bacterial toxins and inhibit viral pathogens in the lung and bloodstream. The production of these extracellular vesicles, termed “defensosomes”, is induced by innate immune sensors such as Toll-like receptor 9 (TLR9) and dependent on the ATG protein ATG16L1. However, the molecular pathways governing defensosome biogenesis and release remain elusive. This study aims to elucidate these mechanisms, focusing on the roles of ATG16L1 and its binding partner TMEM59 in defensosome biogenesis, as well as how TLR9 signaling modulates RAB7 activity for defensosome release. We will use a tractable lung epithelial cell culture system and measure defensosomes released in the blood stream of mice as an in vivo readout. Aim 1 investigates how the interaction between ATG16L1 and TMEM59 promotes defensosome biogenesis. We hypothesize that ATG16L1 and TMEM59 facilitate the formation of intraluminal vesicles within multivesicular endosomes. Aim 2 examines how TLR9 signaling influences RAB7 activity to incite defensosome release. We propose that TLR9 activation leads to the upregulation of a GTPase-activating protein, promoting RAB7 dissociation from MVEs, thus diverting defensosome precursors to the plasma membrane for release. This research will provide critical insights into the molecular mechanisms of defensosome biogenesis and release, while also enhancing the understanding of the intersection between autophagy and innate immunity, additionally contributing to the broader field of extracellular vesicle biology.