St. Jude Children's Research Hospital investigators have discovered that immune system cells that engulf and destroy germs in the body enlist help for this task from a common housekeeping mechanism that most cells use to keep their interiors healthy, a finding that is likely to help researchers understand how the body defends itself against infections and how cancer cells can resist chemotherapy drugs before they have a chance to work.
The discovery of this link between the two mechanisms--phagocytosis (engulfing germs) and autophagy (housekeeping)--suggests that a common mechanism that triggers both processes individually also links them together using a common set of signals. The processes of phagocytosis and autophagy enclose various cellular structures or germs within a sac that fuses with a bag of digestive enzymes called the lysosome. The lysosome then releases the digestive enzymes into the sac and the enzymes degrade its contents. A report on the discovery appears in the Dec. 20 issue of the journal "Nature."
"Autophagy is a cell-survival jack-of-all-trades, and we're trying to understand the signals that trigger its onset," said Douglas Green, Ph.D., chair of the St. Jude Department of Immunology and the paper's senior author. "This process developed so early in the evolution of life that at least some types of microorganisms must have learned how to avoid this defensive response of the cell to being invaded. We want to know how invading microorganisms avoid being destroyed by autophagy and learn how cancer cells use autophagy to resist chemotherapy drugs before they have a chance to work."
The St. Jude team discovered that TLR, a protein on the surface of cells that phagocytize germs, is the link between this "outside job" of germ engulfment and the housekeeping "inside jobs" done by autophagy. Specifically, TLR recruits special proteins that orchestrate autophagy and uses them to supervise phagocytosis and the formation of the digestive sac called the phagosome.
Autophagy destroys germs that have already forced their way into the cell; isolates and breaks down defective molecules in the cell; destroys structures in the cell that normally carry on important biochemical processes but then begin to produce toxic wastes; and breaks down large molecules to obtain nutrients during times of stress or to obtain building blocks for specific new molecules the cell needs.
Immune system cells called phagocytes use phagocytosis for the "outside job" of environmental cleanup, engulfing germs by trapping them within a depression that forms in the cell's surface. The open end of the depression then closes and the newly formed sac moves to the inside of the cell where it fuses with the lysosome.
The researchers based their study on the findings of previous research that demonstrated specific signals associated with both autophagy and phagocytosis. Scientists already knew that a critical step in autophagy is the binding of many molecules of the protein called LC3 into clumps. Certain phagocytes called macrophages that are on the prowl for germs use the TLR proteins on their surface to sense the presence of their prey. When the macrophage engulfs a germ and forms the depression that traps the microbe, TLR triggers a series of biochemical events that guide development of the phagosome. However, previous research found that TLR also triggered autophagy.
"We suspected that because TLR signaling triggers both phagocytosis and autophagy that this signal might also link these activities together as well as enable each process to occur separately," Green said. "This would make sense, since autophagy kills microorganisms that invade the cell and phagocytosis deliberately brings microorganisms into the cell to destroy them. So having two totally separate and unrelated sets of signals to do a similar job would be like re-inventing the biochemical wheel."
The St. Jude team showed that when macrophages phagocytosed special beads carrying pieces of germs, there was a rapid movement of the autophagy-associated LC3 proteins to the developing phagosome. The scientists then fed particles called zymosan (killed yeast cells) to macrophages that lacked TLR. In those macrophages, the movement of LC3 to phagosomes in response to zymosan ingestion was significantly slowed. This suggested that TLR was required for triggering movement of LC3 to the phagosome, Green said.
The team also showed that two proteins critical for autophagy, ATG5 and ATG7, were key to the ability of macrophages to enable phagosomes to mature. For example, macrophages lacking ATG5 were not able to fuse phagosomes with lysosomes. While macrophages lacking ATG7 were able to phagocytize live yeast cells, the yeast were more likely to survive degradation in macrophages without ATG7 than yeast ingested by normal macrophages. This indicated that ATG7, a protein scientists thought was exclusively associated with autophagy, also participates in the destruction of pathogens internalized by phagocytosis.
"This strongly suggested that TLR recruits parts of the autophagy machinery to enable the phagosome to fully develop and fuse with the lysosome even if autophagy itself doesn't occur simultaneously," Green said.
"We can actually see phagocytosis take place in live cells and we can study it in detail, so we know the steps involved," said Miguel Sanjuan, Ph.D., a staff scientist in Green's laboratory. "But autophagy seems to occur suddenly and we know about it only after it's very obviously underway. Now that we discovered that phagocytosis uses some of the same biochemical signals as autophagy does, we can use that information to solve the mystery of how autophagy works. We hope that we will be able to translate that new knowledge in the future into improved treatments for infections and cancer."
Other authors of this paper include Christopher P. Dillon, Stephen W. G. Tait, Simon Moshiach, Samuel Connell and Sebo Withoff (St. Jude); Masaaki Komatsu and Keiji Tanaka (Tokyo Metropolitan Institute of Medical Science, Japan); Frank Dorsey and John L. Cleveland (The Scripps Research Institute, Jupiter, Florida).
This work was supported by the National Institutes of Health and ALSAC.
St. Jude Children's Research Hospital
St. Jude Children's Research Hospital is internationally recognized for its pioneering work in finding cures and saving children with cancer and other catastrophic diseases. Founded by late entertainer Danny Thomas and based in Memphis, Tenn., St. Jude freely shares its discoveries with scientific and medical communities around the world. No family ever pays for treatments not covered by insurance, and families without insurance are never asked to pay. St. Jude is financially supported by ALSAC, its fundraising organization. For more information, please visit www.stjude.org.