Physiological Relevance of Novel Lysosomal Ion Channels

Abstract

Impaired lysosomal ion homeostasis can result in a failure of lysosomes to degrade metabolites, export catabolites and traffic correctly within cells. These lysosomal dysfunctions can result in lysosomal storage diseases and neurodegenerative disorders. My thesis is focused on two newly discovered lysosomal ion channels and their contributions to regulating cellular homeostasis. The first data chapter of this thesis reports the discovery and characterization of Lyso-VRACs (lysosomal volume-regulated anion channels), the first ionic-strength-sensitive ion channels of lysosomes. This work provides a molecular foundation upon which to explore the links between lysosome volume regulation, lysosome physiology and human diseases. We determined that these channels are encoded by LRRC8 family genes. Previously, it had been thought that the only site of action of LRRC8 proteins was on the plasma membrane, but these studies revealed an additional essential role played by intracellular LRRC8 proteins in lysosome physiology. We have discovered essential roles of lysosomes in intracellular water homeostasis， and provide compelling evidence that in individual cells, lysosomes sequester, then extrude toxic levels of intracellular water to help alleviate the water crisis in cells under hypo-osmotic stress, comparable with the “charging” and “discharging” phases of the bladder organ in animals. Thus, lysosomes not only function as the cell’s stomach for nutrient processing, but also behave as the cell’s “bladder” for osmoregulation. The second data chapter of this thesis reports the discovery and characterization of the first proton-selective channel of lysosomes. TMEM175, which was previously reported to act as a constitutively-active potassium selective channel, instead acts as a proton selective channel when mammalian lysosomes are studied at their normal pH. By exploring the cellular effects of alterations in TMEM175 activity， I discovered that proton-mediated functions of TMEM175 are essential for lysosomal homeostasis. When TMEM175 is genetically deleted, lysosomes are hyper-acidic, and their protein degradative activity is impaired. When TMEM175 is overexpressed, lysosomes are hypo-acidic. Optimization of lysosomal hyper-acidification pharmacologically or genetically restored degradative activity in lysosomes toward normal. These discoveries about the roles of LRRC8 and TEM175 proteins within lysosomes suggest important roles for these proteins at the level of the physiology and pathophysiology of the whole animal. The role we have demonstrated for Lyso-VRACs in cellular water balance may open a new chapter in understanding the secretion of “watery” body fluids, like saliva, tears, sweat, breast milk, bile acids, and vaginal secretions. Furthermore, the role we have demonstrated for TMEM175 suggests an explanation for why recent Genome Wide Associated Studies have identified TMEM175 as one of the most prominent genetic risk factors for Parkinson’s Disease， and suggest that manipulation of the activity of TMEM175 may have therapeutic applications not only for Parkinson’s Disease but also for many other diseases that have alterations in intracellular pH.