This article first appeared in the Fall 1992 issue of LabNotes, an MBL publication.

An international team of scientists has discovered a new network that cells use to move molecules about inside their own borders. The result is changing our understanding of how healthy nerve cells work

One afternoon last summer, Drs. George Langford, Dieter Weiss, and Sergei Kuznetsov made an important discovery in very much the manner that scientists usually make important discoveries: they found something they weren't looking for.

Working with material (axoplasm) squeezed out of squid nerve cells, Langford, Weiss, and Kuznetsov-an American, a German, and a Russian-study the puzzle of how proteins are shipped from point to point inside cells. At the time of their discovery last summer, the dogma of intracellular transport dictated that packages of proteins are usually shipped along a network of tracks called microtubules. The three scientists were-and still are-at the forefront of the effort to understand how this transportation system works.

The significance of intracellular transportation is well-known to biologists. Proteins manufactured in the cell's centrally located protein factories must somehow arrive at the cell's distant outposts, where highly specialized molecules are needed as building blocks for cell growth. Other specialized protein molecules are needed at the tips of nerve cells to carry out the signaling process by which the cells send chemical messengers across intercellular spaces to communicate with neighboring nerve cells.

The cell's internal transport system must either be a two-way system or must include a separate return network, so that some of the spent proteins can be sent back to the cell's manufacturing centers for recycling. The result is a microscopic transportation system that is constantly in the process of moving uncountable thousands of protein packages about inside a single cell.

Anyone who has seen gridlock in Manhattan, or ferry traffic in Woods Hole, will not be surprised to learn that the very much busier intracellular transport system occasionally breaks down. In nerve cells, a breakdown in the intracellular transport system is thought to be involved in various degenerative diseases, most notably Lou Gehrig's disease (amyotrophic lateral sclerosis). How, or why, does the transport system break down?

Unfortunately, the answer to that important question is not yet known. But the first step in answering that question is to ask the questions Langford and his colleagues are asking: how does the normal, healthy process work?

Understanding the normal intracellular transportation system may also help physicians treat people whose muscles are wasting away as a result of athletic or automobile injuries to peripheral nerves. Peripheral nerves, unlike nerves of the central nervous system, grow back after injury, although the process is naturally slow and muscle cells may waste away before they are innervated by new nerve cells. The ability to speed the process of nerve growth, perhaps by speeding the transportation of proteins to the growth sites, might thus protect against muscle atrophy following injury.

The July afternoon Langford and his colleagues made their discovery, they were planning to look for clues about what regulates the movement of proteins along microtubules. But before they started their afternoon's experiments, the three scientists noticed something odd on the television monitor biologists now use to view the microscopic world.

At the edge of the axoplasm, in an area devoid of microtubules, little packages of proteins were zipping happily along, moving over what the fascinated biologists could only assume was an alternate system of tracks small enough to be invisible to their high-powered light microscope.

"We stayed up until the wee hours of the morning," Langford says, very much in the tradition of the MBL's summer scientists who must scramble to unravel the implications of their discoveries before the summer ends and they have to pack up their labs and return to their home institutions where, in many cases, the marine organisms they have been working on are no longer available.

Langford, Weiss, and Kuznetsov had a string of questions they wanted to answer before they had to return to their home bases of Dartmouth, Munich, and Moscow. What are the invisible tracks along which the cell's transport vehicles move? What sort of vehicles, or organelles, move on this alternate system? At what speed and in what direction? What motor powers the movement, and what chemicals regulate the operation of that motor?

Part of the explanation, Langford says, has to do with the preparation his team uses to study microtubules. Langford, Weiss, and Kuznetsov study "native microtubules," which means simply that they study microtubules in axoplasm. Other very good labs remove their microtubules from the axoplasm, and in the process discard the second, invisibly small network.

Then, too, a quirk of timing was involved in the movement Langford and colleagues noticed that July afternoon. When the scientists squeeze axoplasm out of the squid axon onto a cover slip, the second transportation network is not immediately in place. The afternoon the scientists discovered the new system, they had fortuitously let their preparation sit longer than normal on the slide before setting to work, a delay that gave the axoplasmic construction crews time to assemble the network.

Finally, there was the matter of location. "We had to look in just the right place, where there are no microtubules, to notice it," Langford explains.

Once they noticed the new type of movement, the scientists quickly identified the invisible network as a fine mesh of actin filaments. It was already known that the interaction of actin filaments and myosin motors causes movement inside plant cells, amoebas, and muscle cells. And although actin filaments are too small to see with even the most powerful light microscopes, they can be tagged with a fluorescent marker molecule that reveals the location of the filaments (see above). While these fluorescing actin networks were known to exist in various animal cells, they had been-like a surprisingly large number of other cellular discoveries-structures in search of a function.

Of course the earlier observations about intracellular transport are also correct. Packages of protein do move along microtubules, powered by the molecular motors (kinesin and dynein) identified in the 1980s by scientists working in part at the MBL. In recent video tapes made by Langford and colleagues with high-powered light microscopes, packages could be seen traveling along microtubules, then hopping off to travel short distances in areas where no tracks were visible. Why has nature evolved two intracellular transportation systems capable of carrying the very same packages of material?

Possibly the cell has duplicate systems as a back up-if one system breaks down the other takes over. Or perhaps the packages move on one system during certain physiological states and along the other tracks at other times. Langford, Weiss, and Kuznetsov have not ruled out these explanations, but they currently lean more toward a complementary explanation.

"We think the two systems may work together, much like express trains and local trains," Langford explains. The microtubules, which are longer and stiffer, and arranged parallel to the long axis of the axon, form the express network. The shorter, more flexible actin filaments are a cross-hatched meshwork that reaches into areas not served by microtubules. Presumably some packages of protein move long distances on the microtubule tracks and then switch to the actin filaments as they near their destinations.

With the identity of the second network revealed, Langford, Weiss, and Kuznetsov set out to identify the molecular motor or motors that power traffic along the tracks. The molecular motor would be a protein itself-one capable of turning stores of chemical energy into the mechanical energy needed to push or pull protein packages along the tracks. Locating such a protein, which may be present in relatively small amounts, among the thousands of still unknown proteins in the axoplasm, is a daunting task. By way of illustration, the search for the motors associated with microtubules was one of the most hotly competitive areas of science at the MBL in the early and mid- 1980s, requiring several summers of full-tilt effort by a number of large and small labs.

In addition to isolating the motor, Langford and colleagues would like to identify the regulatory proteins that control the working of the motor. Regulatory proteins are only somewhat less tricky to identify than motor proteins; some of the proteins that regulate known motors are still elusive-these, in fact, were what Langford and his colleagues were looking for last summer when they inadvertently discovered the second transportation network.

Down the road, Langford foresees the possibility of a fascinating non-biomedical use for the protein motors that power intracellular transport. "These motor proteins are highly specialized molecules that can convert chemical energy into mechanical work," he points out. "We don't understand yet how they work, but when we understand them, we may be able to use these proteins to store information in a molecular computer," Langford says.

At the close of the 1992 summer research season, Langford, Weiss, and Kuznetsov report that the actin-associated motors are still elusive. The international team is closing in on at least one of the important regulatory proteins, but the rest of that story will have to await the summer of 1993.