Shedding Light on Luminescence The photoprotein aequorin lights the way for scientists studying calcium in cells

Anyone who's spent time at Stony Beach on a warm August night has seen them: the luminescing ctenophores that twinkle like tiny stars in the moonlit waters. No one knows exactly why these comb jellies flicker and glow like they do, but MBL senior scientist Osamu Shimomura knows a lot about the biochemistry behind it.

In fact, Shimomura's life's work has been devoted to shedding light on luminescence - a complex chemical reaction that occurs within an organism, the end-product of which (energy) is released in the form of light instead of heat.

For 35 years, Shimomura has been studying luminescing jellyfish, ctenophores, krill, shrimps, and, most recently, fungi. Shimomura's passion for understanding how and why organisms emit light began in the early 1960s when, fresh out of graduate school at Japan's Nagoya University, he arrived in the United States to assume a reasearch scholarship at Princeton University.

Almost immediately, Shimomura became fascinated by the luminescent jellyfish Aequorea, which is found in great abundance in Pacific coastal waters. For years his colleagues at Princeton had been attempting to identify the substance that actually causes Aequorea to luminesce. They hypothesized that the biochemical reaction involved the interaction of luciferin and luciterase, an enzyme and a substrate that together cause bioluminescence in a variety of organisms. This popular hypothesis, however, had yet to be confirmed experimentally in Aequorea.

Shimomura decided to join his colleagues in trying to unravel the mystery of Aequorea's luminescent properties. To do so, he needed to collect large numbers of the jellyfish, which meant traveling to the west coast and setting up a laboratory at Friday Harbor. Once in Washington, he spent many long days and nights in the laboratory doing one experiment after the next, testing one chemical and then another, searching exhaustively for the elusive bioluminescent substance. After weeks of work at the lab bench, it soon became apparent that his efforts were getting him nowhere.

Frustrated, Shimomura sought the solitude of his row boat, where he would spend his afternoons adrift in the Harbor and alone with his thoughts. He was perplexed: if not the traditional luciferin/luciferase reaction, what, then, was happening biochemically in this jellyfish to make it luminesce? He spent hours meditating on the problem.

"One afternoon a thought suddenly struck me," he remembers. "It was a thought so simple that I ought to have had it much sooner!" He smiles.

Even though Shimomura had ruled out the luciferin/ luciferase reaction, he suddenly realized that it didn't mean that another protein/enzyme wasn't involved. It was a new hypothesis, and Shimomura knew immediately how to test it. If another protein was responsible for Aequorea's bioluminescence, then Shimomura knew that he could turn the light on or off simply by changing the acidity of the environment surrounding the protein.

Shimomura's hypothesis proved to be correct; another protein was indeed at work in Aequorea. The next step was to identify it. By early the following year, after many more long days in the lab, Shimomura succeeded in purifying the mystery protein. He also discovered that this special protein reacts with calcium to produce light in Aequorea, even in the absence of oxygen. He later named the protein "aequorin" for the jellyfish in which it was found.

"At first biochemistry professors were skeptical about this new protein," says Shimomura. "But now the existence and importance of aequorin has been firmly established."

Just as the Japanese recognized the importance of bioluminescent materials to their war effort (see side bar), cell biologists have now recognized the importance of similar materials to their studies of calcium. Instead of marking soldiers as they make their way through the jungles of the South Pacific, scientists use the photoprotein acquorin to illuminate and then track calcium as it makes its way through the cell.

Scientists now know that calcium and its movement within a cell plays a crucial role in a variety of biological processes, including fertilization, muscle contraction, and the transmission of nerve impulses. Clinicians also recognize that calcium is sign)ficant in the pathology of a number of tragic neurological diseases, including Alzheimer's.

When Osamu Shimomura moved his laboratory from Princeton to the MBL in 1982, he was inspired to focus his research efforts on a very different type of luminescing group of organisms: the fungi. His inspiration came from his own backyard, which abuts Beebe Woods in Sippewissett. As the builders began clearing the lot for his new home, Shimomura found the property littered with the luminescent fungus Panellus. "It was as if I were being ordered by Nature to study that group of organisms next," Shimomura says with a laugh.

Today Shimomura's Lillie laboratory contains samples of jack-o-lantern mushrooms, honey mushrooms, tobacco fungus, and his own local species, Panellus. As members of the fungi they represent the "biggest remaining subject" in bioluminescence research, according to Shimomura. Given his past successes with other difficult questions in bioluminescence, it comes as no surprise that it's a subject that he continues to pursue with a vengeance.