Mechanism and in vivo function of LC3-associated phagocytosis: When neighbors help take out the trash
Project management at the University of Würzburg:
LC3-associated phagocytosis (LAP) is a degradative pathway that removes membrane-wrapped cargos within the safety of the phagolysosomal membrane. LAP is involved in removal of intracellular bacteria and clearance of cell corpses and debris to prevent autoimmune reactions. During LAP, the phagosomal membrane is decorated with proteins classically associated with autophagy, including LC3. However, the mechanism of LC3 recruitment to phagosomes and the role of autophagy-related proteins on phagosomes are not clear. Therefore, we propose to determine the mechanisms of LAP-mediated degradation using an in vivo model of cell corpse clearance in C. elegans. Furthermore, based on our expertise with LAP and neurobiology, we will establish C. elegans as a model to define the disease relevant functions of LAP in clearing neuronally released debris.
Our published data shows that Atg8/LC3 family proteins are required for timely breakdown of the corpse membrane inside the phagolysosome. In Aim 1, we will define how the presence of membrane-wrapped cargo is communicated to recruit LC3 to phagosomes and what LC3 does to break down the corpse membrane. This aim depends on our established degron-based reporter system to specifically label the corpse membrane inside the phagosome for time-lapse and correlated light and electron microscopy (CLEM) techniques. Aim 1 will establish exactly how LAP breaks down cargo membranes.
We discovered that phagolysosomes tubulate and vesiculate in an mTOR- and ARL8-dependent manner to accelerate degradation. In Aim 2, we will use RNAi and CLEM to test whether phagolysosome tubulation occurs with the help of motor proteins along microtubules. As mTOR regulates several pathways, we will screen for mTOR effectors that conduct phagolysosome tubulation. This aim will reveal the mechanism of phagolysosome tubulation, providing further insight into phagolysosomal degradation.
Outsourcing degradation may be of pivotal importance for highly differentiated neurons, which need to clear damaged organelles and protein aggregates to stay functional throughout the whole life of the organism. In Aim 3, we will test whether neighboring cells clear unwanted neuronal debris by LAP and determine whether phagolysosomal degradation impacts neurodegenerative motoneuron diseases, such as ALS. This will establish the significance of LAP in the neuronal homeostasis and during disease conditions.
This project will provide mechanistic insight into phagosome maturation in vivo and test the role of LAP in neurodegenerative diseases. LAP-mediated degradation is used by a verity of cells and tissues from worms to mammals. Due to the established links between LAP-deficiency and pathological conditions, our in vivo project may pave the ground for potential clinical studies, and eventually be beneficial for public health.
Projekt period: from 02.2019 to 01.2022