A Molecular Mechanism to Regulate Lysosome Motility for Lysosome Positioning and Tubulation.

Abstract

Lysosome is the degradation, recycling, and nutrient sensing center of animal cells. Lysosomal storage diseases are marked with early onset neuron degeneration at organism level and impaired lysosome functions at cellular level. Additionally, insufficient lysosomal clearance of toxic substances is thought to be one major contributing factor for aging-related diseases. Therefore, understanding the pathways regulating lysosomal metabolism is of great importance. Functions of lysosomes rely heavily on the membrane trafficking. Lysosomes travel directionally in cells to their destinations, where they undergo membrane fusion or membrane fission. The major aim of my doctoral work was to investigate regulators of the directional movement and the membrane fission of lysosomes, particularly the role of TRPML1, a lysosomal cation channel, in these processes. I found that TRPML1 is responsible for the perinuclear accumulation of lysosome under acute conditions such as autophagy induction and cytosolic alkalization. In this process, activation of TRPML1 by the lysosomal phosphoinositide PI(3,5)P2 leads to the release of Ca2+ into the cytosol, which in turn recruits a Ca2+ effector protein, ALG-2, to the lysosomal membrane through its Ca2+-dependent interaction with TRPML1. ALG-2 is associated with the dynactin complex, the adaptor for cytoplasmic dynein, and thus promoting microtubule minus-end directed motility of lysosomes. My data also showed that hyper-activity and hypo-activity of TRPML1 both lead to the abolishment of lysosome tubulation, a platform of lysosome membrane fission, and suggested that this is due to the disrupted balance between the plus-end and the minus-end motility of lysosomes. I have also identified that the perinuclear accumulation of lysosomes in lysosome storage disease cells can be caused by the accumulation of lysosomal cholesterol, which constitutively activates minus-end motility of lysosomes. These findings can greatly advance our understanding about the membrane trafficking mechanisms of lysosomes, and may proof valuable towards the treatment of lysosome storage disease patients. Additionally, I also generated and characterized a genetically encode fluorescent probe for PI(3,5)P2, and showed that it could detect the subcellular localization and dynamics of the lipid in living cells. This gives the probe a huge advantage over conventional methods of detecting intracellular PI(3,5)P2 such as HPLC.