Investigating the effects of the membrane lipid composition on ATG9A structure and dynamics.

Autophagy is a cellular self-eating process critical for maintaining cellular homeostasis. During this process organelles and proteins are sequestered by the formation of a double-membrane vesicle, the autophagosome, and delivered to the lysosome for degradation and recycling. Modulation of autophagy has been highlighted as an attractive therapeutic avenue towards diseases associated with cellular dyshomeostasis, such as cancers. The membrane embedded protein, ATG9A, plays a pivotal role in autophagosome formation. It is shown to continuously traffic across different organelles, such as the plasma membrane, trans-Golgi network, endosomes, and autophagic membranes. Moreover, ATG9A trafficking is reported to be sensitive to the lipid composition of the membrane. Sphingolipids act as regulators of autophagy, and are altered in cancers. It has been demonstrated that sphingolipids can regulate ATG9A trafficking and autophagosome maturation. Recently, the structure of the membrane embedded portion of ATG9A was solved by cryo-EM, and it was revealed to exist in two distinct conformations. In addition, ATG9A was described as a phospholipid scramblase. Despite these recent advances, many aspects of the ATG9A function and dynamics as well as its exact role in autophagosome maturation, remain elusive. We are working with a cross-disciplinary approach, combining structural, cellular, and computational modeling methods to address open questions on how the lipid environment affects ATG9A structure, function, and dynamics. We investigate the effect of the membrane lipid composition on the structure, as well as the interconversion between ATG9A states. In addition, we identify putative binding motifs for certain classes of sphingolipids to elucidate the molecular details of their interaction with ATG9A. This will provide novel insights into the mechanisms underlying autophagosome formation by strengthening our understanding of the function and regulation of ATG9A in lipid environments.