WHY TURN RED? The autumn leaf is toast, but it still redecorates

BY SUSAN MlLlUS - leaf turning red in the fall makes for a much greater mystery than a leafturning yellow does. The yellowing signals simply a dropping of veils because the yellow pigment has lain hid- den in the leaf during its long, green summer.

When summer ends and the green pigments break down, the yellow shines through. Reds, however, don’t loll around all summer. A leafwith only a few weeks left to hang on its tree summons its faltering resources for a burst of bright- red-pigment making.

What a time to redecorate. Cell physiologists have found a world inside an autumn leafthat resembles the pande- monium on a sinking ship. Metabolic pathways start to fail. Compounds break apart. Doomed cells rush to salvage the valuables, especially nitrogen, by sending them off to safer tis- sues. So in this final crisis, why make a special effort to turn red? Does the red- making machine turn on by accident, or do the red pigments contribute something valuable? Why would passengers fleeing the Titanic stop to repaint their staterooms?

There are plenty of proposed explanations, says David Lee, a trop- ical botanist at Florida International University in Miami. He and other pigment researchers say that modern analytical techniques are enablingthem to test these ideas in new ways-and finally get some answers. The most abun- dant evidence, he says, has revived a 19th- century notion that the red pigments called anthocyanins serve as a protective device for falter- ing photosynthetic chemistry.

RED START Ironically, Lee didn’t get interested in anthocyanins until a job took him to a place without fall. He grew up with the humdrum autumn colors of the relatively dry landscape of east- ern Washington State. “There wasn’t much of an autumn show- a few trees in town turned red,” he says. However, in 1973, he left temperate seasons behind when he joined the faculty at the Uni- versity of Malaya in Kuala Lumpur.

Some of the tropical trees there burst into astonishing reds, though not all at the same time or for the same reason as each other. The Indian almond, for example, blushes brightly just before it sheds its leaves. The leaves of mangos and cacaos do the reverse, turning scarlet when they first sprout.

“A whole tree will quickly flush red,” Lee says. “I saw it and thought, WOW, what’s happening here?”’

Anthocyanins provide the red special effects for much of the plant kingdom. Their fireworks intrigued 19th-century biologists, who discussed the possibility that aleaf might make anthocyanins during a period of vulnerability, to shield the green chlorophyll pigments from sunburn. However, these intensely colorful com- pounds showed up in little walled-off pockets called vacuoles within cells. Since physiologists have o h n considered the vacuole “the cell’s trash bag,” says Lee, the sunscreen proposal faded into disfavor. For much of the past century, he says, physiologists classed antho- cyanins as just some more trash.

Lee suspected the old idea might have something to it, perhaps in the screening of especially vulnerable leaves-the extremely young and the extremely old-from ultraviolet (vv> radiation. Yet anthocyanins have turned out not to absorb UV as well as some

of their own chemical precursors in the leaf do. Making anthocyanins would actually deplete the store of

better UV absorbers. “I became disenchanted with that hypothesis,” Lee r e d s , but he still

wondered whether anthocyanins might shield avulnerable leaffrom some other

In 1992 at a botanists’ meeting in Hawaii, he met plant physiologist Kevin Gould of the University of Auckland in New Zealand. Over a brealrfast in Woolworth’s, they plot- ted a test of the sunscreen hypoth- esis using shade-loving species as examples of light-sensitive plants.

The two researchers focused on certain little plants that dot shaded

forest floors and grow leaves with green tops and red undersides. For example,

the common trout lily of northeastern

Lee had found a Malaysian bego- nia and a Costa Rican melastome thatnatmdyvasyinleafcolor,some individuals sprouting all-green leaves and others putting out leaves with red undersides. Lee and Gould blasted samples of all these leaves with intense light. Physiologists had

already shown that such blasts overload light-gathering chlorophyll and slow it down, a misfortune called photoinhibition.

In Lee and Gould’s experiment, all-green leaves seemed to suffer greater photoinhibition than did two-tone ones of the same species. Reporting their finding in 1995, the two physiol- ogists proposed that random strikes of bright sunlight on the light-dappled forest floor could pose great dangers. A plant with a little protection in the form of anthocyanins could off-load some of that sudden excess energy in the form of its chlorophyll 6 and better withstand a blast.

menace.

forests does this, as do some begonias.

TROUBLED TIMES - What‘s an old leaf to do when its innards just aren’t up to speed any- more and the sun IS still burning down?

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RED SPREAD In the 1990s, other researchers also explored the idea of red pigments as sunscreen. Debate bloomed over how to devise a test that avoids confounding factors, such as different rates of photosynthesis in different-colored leaves.

In 1999, researchers at the University of Queensland in Aus- tralia refined the bright-light tests performed by scientists includ- ing Lee and Gould. In an experiment on the tropical Bauhinia variegata, Robert C. Smillie and Suzan E. Hetherington flashed an assortment of its red or green pods with bursts of white, blue- green, or red light. The red pods tolerated the white and blue- green light flashes better than the green pods did.Yet the red pods didn’t show any superior tolerance to bursts of red light. The researchers contended that in the latter case, anthocyanins, which can’t soak up red wavelengths, weren’t protecting the chlorophyll.

Lee then joined Taylor S. Feild and N. Michele Holbrook of Har- vard University in a similar experiment on autumn leaves. The researchers chose red-osier dogwood shrubs because they end the year in multiple colors. In fall, leaves bathed in brilliant sunlight turn red, but shaded leaves don’t develop anthocyanins and so just turn yellow. The red leaves recovered faster from flashes of intense blue light, the researchers report in the October Plant Physiology. Flashes of red light, the wavelength that anthocyanins can’t absorb, had about the same effect on red leaves as on yellow ones.

The finding dovetails with physiological studies from other labs that suggest that leaves may need special protection during their final weeks. Tests showed that old leaves are more vulnerable to photoinhibition than younger but mature ones are. In a color- changing leaf, the plant’s metabolic pathways for making the ini- tial capture of energy don’t lose their efficiency as fast as the sub- sequent pathways for processing that energy do, a risky imbalance that invites overloads. Seasonal stresses, such as chilling temper- atures, also hobble the leaf metabolism.

Yet during autumn, the aging leaf has to salvage as much nitro- gen as possible and send it to tissues that will survive the winter. So, as decrepit as the photosynthetic mechanism becomes at the end, it has to keep catching and processing sunlight if the leaf is to finish the salvage operation.

That scenario prompted William A. Hoch of the University of Wisconsin-Madison to look at the geographic history of intense red color. He hypothesized that plants would be most likely to manufacture anthocyanins in climes where temperatures often plunge during autumn. So, he ranked the intensity of antho- cyanins in fall-coloring in nine genera of woody plants. Some of these were native to either a cold zone in Canada and the north- ern United States, others to a milder, maritime clime in Europe. Out of 74 species, the 41 that flamed out with reddest leaves all came from the North American chilly zone, he reported in the January 2001 Tree Physiology.

BLUEBERRIES, ETC. The evidence has been building nicely for anthocyanins as safety measures against light overdose, according to Gould. Yet he doesn’t expect that to be their only function. “They’re very talented molecules,” he says.

He got the urge to test for another benefit, he says, while read- ing a newspaper article touting the health benefits of diets that include blueberries. Antioxidant pigments abound in blueberries, and Gould decided to explore whether the antioxidizing powers of the leaf anthocyanins that he was studying benefit their plants.

When purified in the lab, these pigments sop up free radicals, which are alarmingly energized substances that can damage DNA, proteins, and membranes. Anthocyanins in a test tube can corral free radicals four times as well as do the well-known antioxidants vitamin C and E, says Gould. He started planning a test for antho- cyanins’ antioxidant effects inside a living plant.

“It took us along time,”he says, “but I had some very diligent stu- dents.”They borrowed an imaging technique called epifluorescence microscopy from research on animal cells. With it, they could watch bursts of the oxidizing agent hydrogen peroxide as it was released

in a cell. To observe the actions of antioxidants, the researchers had to figure out a way to trigger such oxidizing bursts in plant cells.

Gould remembered that one of their study subjects, a New Zealand piebald shrub called Pseudmintera colorata, developed small red pimples on its leaves where aphids pricked them to suck sap. When the researchers stabbed the leaves with avery fine nee- dle, they triggered bursts of hydrogen peroxide in cells. A steady- handed scientist could induce the bursts and the subsequent red- ness as well as an aphid does. “we could write the word ‘red,’ and it came out red 2 days later,” says Gould.

After patiently perfecting these techniques, Gould’s lab made a movie. The researchers filmed the stabbing ofboth the d-green and the red-splotched leaves of P. colorata. In the October Plant, Cell andEnvironment, Gould and his colleagues report seeing an oxida- tive burst of hydrogen peroxide a minute or less aRer they pushed the needle into the upper layers ofleaftissue. In red tissues, the burst faded quickly In green ones, however, it intensified, and hydrogen peroxide concentrations soared for at least 10 minutes. Gould con-

tends that anthocyanins are the com- Why wo u I d Dassengers

pounds most likelito have quenched the oxidative burst.

Lee welcomes the W D O k enthusiasti- - on the Titanic cally “It’s the first evidince [for antiox- idant behavior] in a living plant,” he says. stop to A suggestion for yet a third function

re pa i nt the i r for anthocyanins in leaves comes from physiologist Linda Chalker-Scott of the

staterooms? University ofWashington in Seattle. She - proposes that the pigments regulate water movement. She’s contributing a

chapter on the idea to the book Anthoqanins in haves (Kevin Gould and David Lee, eds., Academic Press), due out soon.

Anthocyanins dissolve in water, whereas chlorophyll and a lot of other cell pigments don’t, she explains. Water loaded with any dissolved substance has what physiologists call lower osmotic potential, a decreased tendency to flow away. Loading water with dissolved substances also lowers the temperature at which water freezes, potentially an advantage on a frosty fall night.

Chalker-Scott points out that many plants blush red at water- related stresses such as drought, salt buildup, and heat. Her exper- iments testing the idea have been largely on hold since last year, when the building housing her lab was firebombed during aprotest targeting another researcher’s genetic engineering project.

Plenty of other ideas for anthocyanins’ function also remain to be tested. Observers offungus-farming ants, for example, reported in the 1970s that the ants avoid taking red leaves home to feed to their garden. Researchers have speculated that anthocyanins might discourage growth of some fungi.

Another hypothesis states that anthocyanins keep leaves from overheating; an alternative has the pigments protecting leaves from cold. Gould notes that a birch species he encountered in Fin- land holds on to its red leaves year-round, despite temperatures that plunge to -4OOC.

Just last year, a paper by the late theorist W.D. Hamilton and Samuel P. Brown of the University of Montpellier in France mused about whether autumn coloring shares a communication role with the peacock‘s tail. The healthiest birds can grow the most spectacu- lar tails, so a cruising female can get an accurate assessment of a prospective mate’s health by checking out his plumage. In a similar way, Brown and Hamilton speculatedthat the healthiest trees might put on the flashiest f d displays. This leafsignal might give fall-active predatory insects, such as aphids, accurate information about which trees have good defenses and which ones might be easy pickings.

Even if none of these or the abundant other suggestions pans out, researchers already know enough to raise anthocyanins from the category of cellular trash to their deserved status as vital mol- ecules. A big question still remains: If these pigments are so great, why don’t all leaves turn red? rn

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