LYsosomes are tiny sacs of digestive enzymes that declutter cells by breaking down waste. But they can also be troublesome: when their outer surface is damaged, their destructive proteins begin to spill into the cytoplasm and damage the cell. Indeed, the frequency of these leaks increases as a person ages and likely plays a role in diseases associated with aging such as neurodegenerative conditions. However, a study published on September 7 in Nature discovers a previously unknown pathway that cells use to repair leaky lysosomes, which may have implications for the treatment of these diseases.
It’s a “very comprehensive and well-designed” study, and the first to link lipid transport to a non-metabolic biological process, says Marja Jäättelaprofessor of cell death and metabolism at the Danish Cancer Society Research Center, who was not involved in the work.
Research had already established a way for cells to repair leaky lysosomes. Previously, a collection of proteins known as the ESCRT machinery was shown to be able to plug holes in the membranes of organelles. However, “something was missing”, according to the co-author Jay Tan, a cell biologist at the University of Pittsburgh. Up to 90% of lysosomal damage is repaired even when the ESCRT complex is inhibited, suggesting that a different pathway carries out most of the repair, says Tan The scientist.
To identify the individual components of the missing pathway, Tan used a lentivirus to express an enzyme called Turbo-ID in human cells, which sticks a biotin tag on any protein within ten nanometers. Using a version of the enzyme that localizes to lysosomes and a chemical known to perforate the lysosomal membrane, he was able to biotinylate all of the proteins surrounding the damaged organelle.
Tan isolated the biotin-tagged proteins and identified them by mass spectrometry. Among the purified molecules were the components of the ESCRT complex, but also proteins known to interact with phosphoinositides, a family of lipids known to regulate key cellular processes, including proliferation and migration.
Further experiments revealed that when the lysosome membrane is compromised, an enzyme called phosphatidylinositol-4-kinase type 2a, or PI4K2A, is recruited to the surface of the organelle, possibly in response to calcium ions that s escape from the lysosome, says Toren Finkel, study co-author and professor of medicine at the University of Pittsburgh. PI4K2A generates a lipid called phosphatidylinositol-4-phosphate (PI4P), which acts as a danger signal and recruits several proteins called ORPs (for oxysterol-binding protein-related protein), which bind the endoplasmic reticulum to the lysosome.
These ORPs then exchange PI4P with endoplasmic reticulum lipids, including phosphatidylserine, which recruits the lipid transporter ATG2, the final component of the pathway. “ATG2 is like a fire hose” for lipids, Finkel says, pumping molecules into the membrane to plug the hole.
Lipids are transported from the endoplasmic reticulum (green) to seal holes in damaged lysosomes (red). Cell nuclei are shown in blue.
JAY XIAOJUN TAN
The researchers then mutated the different subunits of ATG2 to confirm that the lipid transporter is an essential part of the pathway. “Instead of a smooth tunnel, we put in place a bunch of molecular toll booths by changing amino acids, which makes it harder for lipids to move,” says Finkel. These changes in effect blocked lipid transport in the cell culture and blocked lysosomal repair.
As an ode to the University of Pittsburgh, the two scientists dubbed the phosphoinositide-initiated membrane tethering and lipid transport pathway, or PITT for short.
Tan believes the two mechanisms have evolved to repair different types of damage, with the ESCRT complex repairing small pores while the PITT pathway repairs larger holes.
The new track can do most of the manual work of the cell. The researchers found that cells typically take about an hour to repair damaged lysosomes, but this healing takes up to 11 hours in cells that lack PI42KA. “It looks like a pretty big lane, maybe more than the ESCRT lane,” says Antoine Galionea pharmacologist from the University of Oxford in the UK who was not involved in the study.
Tan says the results could point to drug targets for neurodegenerative diseases, such as Alzheimer’s disease, because aggregates of proteins like tau escape into the lysosome membrane, preventing their destruction and facilitating their spread between neurons, says Tan. Indeed, the researchers found that depletion of PI4K2A, the initiator of the PITT pathway, increased the spread of tau in cell culture.
It’s “definitely an important process to study further in disease models of neurodegeneration,” says molecular biologist Caroline Mauvezin at the University of Barcelona, Spain, who did not participate in the study. But the pathway might work differently in other cells, including neurons, so more work is needed, she adds.
The researchers plan to screen drugs for their ability to activate the pathway, starting with currently available drugs that could be repurposed. One intriguing lead is ginseng, an herb used in traditional Chinese medicine, whose components appear to activate PI42KA, Tan says.