Science Editor

Since it netted its discoverers the Nobel Prize in chemistry this year, ubiquitination probably has become the best-known way in which cells destroy unwanted proteins. But there is a second process for cellular housecleaning: autophagy.

While ubiquitination is similar to an inventory control system for short-living cellular proteins, autophagy deals with the degradation of larger cytoplasmic components and structural features. In fact, Vojo Deretic, professor in the department of molecular genetics at the University of New Mexico at Albuquerque, likens the role of autophagy more to apoptosis than the ubiquitination pathway.

"Apoptosis is like razing a house to build a new one," he told BioWorld Today. "Autophagy is like an internal remodeling job."

Internal remodeling in a home means tossing out faded curtains and dated wallpaper, but two recently published papers suggest that in a cell, autophagy can clean up bacteria.

In a paper published early online in Cell, Deretic and his colleagues at UNM; the Universidad Nacional de Cuyo-CONICET in Mendoza, Argentina; and Duke University in Durham, N.C.; reported that the remodeling job is one defense mechanism cells use against Mycobacterium tuberculosis. The finding comes more or less on the heels of a recent paper in Science showing that autophagy mechanisms operate in the defense against Group A streptococcus.

The Cell paper focuses on autophagy in macrophages, a cell type that routinely uses autophagy-related processes to kill bacteria. Briefly, macrophages engulf invaders in a cellular compartment known as a phagosome, which then fuses with another compartment full of enzymes known as a lysosome, forming what is called the phagolysosome. Bacteria are dismembered in that compartment, and their limbs are presented as antigens to the rest of the immune system.

M. tuberculosis has learned to go straight for the jugular, interfering with the fusion of phagosome and lysosome.

"So now you have these infected long-lived cells. What are you going to do to get rid of the pathogen? Autophagy would be the perfect mechanism," Deretic said.

Using Starvation To Induce Autophagy

Autophagy is induced under conditions of cellular starvation; in fact, in research published in the Nov. 3, 2004, issue of Nature, researchers from the Japan Science and Technology Agency and the Tokyo Metropolitan Institute of Medical Science showed that cells support themselves via autophagy during the period of neonatal starvation that follows the separation of newborn and placenta. In the laboratory, starvation means growing cell cultures under reduced supplies of amino acids.

In the Cell paper, the scientists began by inducing autophagy via starvation in M. tuberculosis-infected cell lines, and checking the effect on phagosomes and bacteria. The take-home message from the research is that autophagy induction killed M. tuberculosis in cell lines, as well as primary cells from mice and humans. The phagosomes became more acidic and matured into phagolysosomes, the process which normally is inhibited in M. tuberculosis infection. Adding autophagy inhibitors partially blocked that maturation. Starvation effects could be mimicked pharmacologically through treatment with rapamycin, an antibiotic and immunosuppressant used in the treatment of kidney transplant patients.

A more thorough characterization of the bacterium-containing compartments showed that autophagy induction through either starvation or rapamycin demonstrated that treated cells had higher levels of a number of markers that are more typical of phagolysosomes than controls. They also had higher levels of a marker localized in autophagic membranes, suggesting that the phagolysosomes were taking on characteristics of autophagic organelles. Control experiments showed that the experimental manipulations stopped short of razing the house; in other words, they did not induce apoptosis.

The findings are extended to other cell types in a paper published in the Nov. 5, 2004, issue of Science by scientists from the Osaka Graduate School of Dentistry, the Japan Science and Technology Agency, the Tokyo Metropolitan Institute of Medical Science, the Nagahama Institute of Bioscience and Technology and the National Institute of Genetics/ SOKENDAI, all located in Japan.

While destroying bacteria is part of macrophages' regular job, the Japanese researchers showed that autophagy is used as a cellular defense mechanism even in cell types - in this case, HeLa cells and mouse embryonic stem cells - that do not regularly suffer invading pathogens. In response to infection with Group A streptococcus (the culprits in strep throat, as well as scarlet fever and other diseases), those cell types also recruited their autophagic machinery to destroy the bacteria. The effect was not seen in knockout cells that lack the ability to form autophagosomes.

Next On The Menu: Viruses

Both papers report on bacterial infections, but theoretically, the method could be effective against viruses as well. In particular, "cytoplasmic viruses would be good targets, one would think. Herpes viruses actually encode a protein that seems to counteract autophagy (albeit somewhat indirectly). We are [also] interested in whether HIV budding into intracellular compartments may be affected by autophagy. If so, one could get 2-for-1 treatment of AIDS and tuberculosis, which often go hand in hand anyway," Deretic said.

But, he added, that remains speculation for now.

In further experiments, Deretic's group linked the mechanisms of autophagy to interferon-gamma (IFN-gamma). The molecule is known to be a key inflammatory cytokine and a "marquee" molecule in the immune response to tuberculosis, but "to this day, people are puzzled by how IFN-gamma actually works," Deretic said. "Well, in our hands, it induces autophagy." Treatment with either IFN-gamma or LRG-47 (a GTPase activated by IFN-gamma) induced autophagy-like processes in macrophage cell lines.

Deretic said he and his colleagues are "very interested in seeing how IFN-gamma applies to other intracellular pathogens," in work that is ongoing. They also are interested in applying their findings clinically, but given that the group also is working on bringing unrelated cystic fibrosis research into the clinic, Deretic thinks it will be a while.

"These things are never simple," he said. "Not even with an already FDA-approved compound like rapamycin."