By David N. Leff

In movies, TV, fiction and real life, the double agent - who plays both ends against the middle - makes for a thriller. As usual, Nature got there first.

It was back in 1955 that a Japanese electron microscopist named Yamada described flask-shaped indentations in the plasma membranes of cells. He named these hyper-minute invaginations "caveolae" - little caves.

Time went by, and cell biologists puzzled over what role, if any, these tiny lacunae could be playing. They found caveolae paving the outer surfaces of cells all over the human body. On epithelia - skin cells - as many as 10,000 of them crowded into 1 cubic millimeter. They measured from 50 to 200 nanometers in diameter, but could expand and elongate upon demand.

That demand came from infectious microorganisms. Microbial pathogenesist and immunologist Soman Abraham, at Duke University Medical Center in Durham, N.C., tells how he and his colleagues fingered caveolae as cellular safe houses that offered refuge to invading bacteria, even while setting them up to be killed by their victim's immune defenses.

"We have discovered that certain potentially pathogenic bacteria can bind to this distinct caveolae structure," he said, "and then trigger entry into its host cell. So basically, the bacteria seem to be co-opting these little caves for their own benefit - to get into the immune cells that are normally present in our bodies to kill them. So these guys trigger entry into these phagocytic cells, but the key fact is they are not killed.

"From the caveolae," Abraham continued, "they get into these immune system cells and remain inside them. In that environment they're protected from antibiotics. They would be ineffective. Also, the various other immune cells in the body, besides phagocytes, are unable to get to these bacteria. So it's of a very nice niche or refuge site for those bugs to hide in. They can remain in there for days on end, and some time later, when we stop giving the antibiotic, they come back out and reinfect the body.

"The other aspect," he went on, "is that nobody really knows how diverse the functions of these caveolae really are. So our discovery shows that these structures can take very large particles into your cells. Relative to an immune cell, a bacterium is a very large particle. So caveolae are able to encase, to capture, huge particles and take them into a cell."

How Cave Dwellers Escape The Chop

Abraham is senior author of a paper in today's issue of Science, dated Aug. 4, 2000, titled: "Involvement of cellular caveolae in bacterial entry into mast cells."

"Our findings," he told BioWorld Today, "are relevant to a family of bacteria that are found in the human gut, and also some pathogens of the urinary tract. So I could name a whole bunch of enteric [intestinal] pathogens that have utilized this caveolae system - the common E. coli, Klebsiella pneumoniae, and some species of Salmonella. But it's conceivable that other bacteria that we have not investigated also use this hideaway system. So I don't know how broad the scope is."

Abraham and his co-authors have tested for caveolae in the two main types of antibacterial immune system cells. "Both macrophages and mast cells," he explained, "can recognize bacteria coated with antibody, or not coated. The antibody-coated bacteria are taken up through the classical phagocytic pathway, where they are engulfed and then killed. The uncoated bacteria interact with macrophages and mast cells via uptake in which caveolae are involved. In this case the bacteria remain viable."

Mast cells, which have a lot to do with allergic reactions and asthma, recruit other immune cells to the sites of bacterial infection. There they tuck the invading germs into caveolae, where they snuggle down in latency, safe from their victim's other defense mechanisms.

As recent research has shown, bacteria aren't the only molecules playing this double-agent game. It's becoming increasingly clear that viruses use caveolae. "For example," Abraham recounted, "there is now some documentation that HIV, the AIDS virus, also seems to bind to caveolae. And the protein coat on that viral surface is also composed of caveolae material. So this is just the tip of the iceberg; more and more things are about to come.

Abraham recounted his experiment: "What we did was expose mast cells to our model bacterium, E. coli. A key receptor molecule, CD48, sits inside that little cave, and is able to signal the mast cell to take up the bacteria. They bound to the surface of the mast cells, then triggered entry. We followed what happened to the bacteria that were inside the mast cells, in terms of their viability. So when we cut sections through the mast cells to see what had happened to these bacteria, we found them encased in very tight-fitting vesicles, and looking perfectly viable. They didn't seem degraded; they were not killed. This suggested to us that the bacteria were entering the cell via a route - the caveolae - that is distinct from the classical phagocytic pathway.

"Once we found that they had not been killed, we thought about caveolae, based on the CD48 bacterial receptor being present in them. So that's how we identified caveolae's involvement in the entry."

Abraham made the point, "Caveolae are present in our cells to undertake very natural physiological functions, not just shelter invading pathogens. They respond to hormones - because hormones bind to their receptor, and induce physiological changes in that cell. For example," he pointed out, "when insulin binds to its receptor - which is within the caveolae - you get sugar-control function.

Spelunking Approach To Drug Discovery

"One thing that we are looking at right now," Abraham said, "is the possibility that since we have demonstrated this nice phagocytic pathway, we can think of actually delivering therapeutic drugs into infected cells, using this pathway. So one can link a potential target to these caveolae, and then get it into the cell. That's what we are trying to move into," he concluded, "but it's very preliminary work."

Accompanying the Duke article in today's Science is an editorial titled, "Bacterial spelunkers." It concludes: " . . . the mysterious recesses of caveolae will keep researchers spelunking further into the depths of these fascinating cellular domains for years to come."

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