91av

A FEW weeks from now, a tremor of anticipation will be running through North America’s leaf peepers. More than a million people will set off on the annual pilgrimage to one of nature’s finest spectacles: the transformation of the northern greenwoods into a mosaic of yellows, golds and reds. It’s the reds that are the main attraction. From bright signal-red to deep burgundy, they blaze in apparent defiance of the coming cold and dark of winter.

The reddening of the cool northern forests is all too brief. But you can see crimson-crowned trees all year round. In tropical forests the new leaves of many species unfurl to reveal spectacular scarlets. Near the forest floor, there are more subtle displays: turn over any green leaf and the chances are the underside will be as red as a maple in Maine. Elsewhere, red leaves might be less eye-catching, but they are surprisingly common once you start to look. Red pigments colour both young leaves and old, shade-loving plants and those that prefer bright sunshine. Some plants even have leaves of both colours and some are patterned with bright red spots or stripes.

There must be a reason plants produce these biologically expensive pigments. People have puzzled over what that might be since the time of Aristotle. But only now has anyone come up with a convincing explanation for such widespread redness: red pigments are nature’s stress-busters.

They protect the delicate structures of a cell from the molecular mayhem that threatens stressed-out plants. Water shortages, intense sunlight, lack of nutrients, attacks by herbivores and pathogens – all of these generate dangerous free radicals that can attack membranes and disrupt DNA. The pigments are there to clean up the mess. “Protection from stress is not a luxury,” says Kevin Gould, a plant physiologist at the University of Auckland. “It is critical for survival.”

There’s been no shortage of suggestions for why some plants have red leaves, each advanced by a respected scientist with data to back them up. Red pigments keep leaves warm in cold climates, they help protect plants from drought and they warn off insects – or attract them. One of the most persistent ideas is that red pigments block out hazardous ultraviolet radiation. But anthocyanins are generally confined to the photosynthetic tissues of the mesophyll (see Graphic). This would leave the leaf’s uppermost cells exposed to danger. What’s more, other pigments are easier to produce and make better sunscreens against UVB. Each suggestion seems to fit a few cases of red leaves, but none can explain them all. “What holds up for some species doesn’t work for others,” says Gould.

Almost all the red colour in leaves is the result of a small group of flavonoid molecules called anthocyanins, which are manufactured in the cytoplasm and then transported into the cell vacuole. From colourless raw materials, the addition of sugar molecules and a few chemical tweaks produces every shade of red right through to maroons, purples and blues. The manufacturing process is complicated and energy-consuming, so it’s unlikely a plant would invest so much in these pigments unless they served some good purpose. Bright petals attract passing pollinators. Luscious-coloured fruits advertise their ripeness to animals that will disperse their seeds. But plants were making red pigments long before flowers and fruits had evolved.

Plants such as grasses make anthocyanins, even though they have inconspicuous flowers. So do conifers, which don’t have flowers at all, and even some of the more ancient ferns and mosses. “They must have an important biological or physiological function,” says David Lee, a botanist at Florida International University in Miami who has been investigating red leaves since the 1970s.

Gould had also pondered the age-old question for several years. As he sipped a glass of cabernet sauvignon one evening, he thought about the claims of some researchers that a glass or two of red wine a day is good for you. The most obviously healthy components of red wine are the flavonoids – precisely the group of pigments that includes the anthocyanins.

Flavonoids are powerful antioxidants. They mop up free radicals and reactive forms of oxygen which, if left unchecked, can destroy membranes and DNA. Health shops sell extracts of plant pigments to people hoping to stave off wrinkles and rheumatism or avoid cancer – all of which are blamed in part on free radicals. Doctors and public health experts exhort us to eat more fruit and vegetables, especially red ones. Not only are they full of vitamins, which are themselves good antioxidants, they are also packed with anthocyanins. These red pigments are four times as effective at mopping up free radicals as vitamins C or E.

Because free radicals are as bad for plant cells as human ones, Gould wondered if red pigments might offer the same protection to plants as they do to us. Plants are constantly exposed to free radicals and reactive forms of oxygen, some of which are lethal to cells. Plant cells are equipped with a suite of enzymes and phenolic pigments to deal with these, but there are times when they can’t cope with the onslaught. Gould speculated that anthocyanins might provide an extra level of protection when the plant needs it most.

Indeed, the same stresses that generate free radicals also trigger the manufacture of anthocyanins. “When an insect bites into the leaf of New Zealand’s horopito tree, a red blotch spreads around the wound,” says Gould. “And during drought, green leaves of native parataniwha shrubs turn red.” What’s more, anthocyanins are often particularly prominent during the most stressful periods of a leaf’s life, including its first flush and its final, dying days.

Although there’s plenty of evidence that extracts containing anthocyanins are potent antioxidants, no one had checked whether the same was true in a plant. Gould was ideally placed to investigate, because New Zealand has an extraordinary range of red-leafed flora. “The occurrence of red leaves is very variable,” says Gould. “Red-leaved specimens grow next to green ones. Some plants have both red and green leaves, sometimes on the same branch. And some species have leaves that are mottled green and red.” These plants are perfect for investigating the role of red pigments because their red and green tissues are identical in all respects bar one: the levels of anthocyanins.

Sam Neill, a PhD student in Gould’s lab, took extracts of both red and green leaves from parataniwha (Elatostema rugosum) and compared how potent they were as antioxidants. On average, extracts from red leaves were 14 times as powerful as extracts from green leaves (Plant, Cell and Environment, vol 25, p 539). “It was kind of neat to see the results so different when these plants grow right next to each other in the bush,” says Neill. So why aren’t all parataniwha leaves red? The answer seems to be that the plant produces red pigment only in leaves that are exposed to more intense light, while leaves in slightly cooler, shadier spots stay green.

It worked in the test tube, but Gould wanted to see anthocyanins in action in living cells. This time, he chose red-and-green blotched leaves from the horopito (Pseudowintera colorata). With the help of undergraduate James McKelvie, he simulated the piercing bites of an insect and watched the plant’s response. First he had to make sure an insect bite would trigger the formation of free radicals. McKelvie removed thin sections of green and red tissue from the leaves and impregnated them with a dye that glows under UV light when free radicals are present. Gould and McKelvie then faked an insect attack by stabbing the leaf tissue with a needle. The effect was instant. Under the microscope they could see bright haloes forming around the wounds – whatever colour the tissue. The difference was that in the red areas, the bright glow faded in less than five minutes, when all the free radicals had gone. “You can see these things being sponged up by red cells,” says Gould. In the green cells, the attack lasted twice as long (Plant, Cell & Environment, vol 25, p1261).

But there was still the question of how anthocyanins could be effective as a defence against free radicals when they are tucked away in cell vacuoles. Neill, however, now has good evidence that anthocyanins fresh off the production line in the cytoplasm are just as good at scavenging radicals as those in the vacuoles. “This suggests that they could provide widespread protection to cellular membranes, organelles and DNA,” he says.

Anthocyanins do more than scavenge free radicals. They limit their production too. One of the biggest hazards plants face is, ironically, sunlight. When plants are exposed to too much light, the chlorophyll molecules in the photosynthetic tissues can transfer the excess energy to molecular oxygen, breaking it up to form singlet oxygen, hydrogen peroxide and the extremely toxic hydroxyl radical. All of these can damage vital parts of a cell. The light doesn’t always have to be strong to be harmful: when plants are under stress, quite moderate intensities can be enough.

Red pigments protect photosynthetic tissue from this kind of damage by blocking out green light. Although chlorophyll reflects most of the green light that reaches it, these wavelengths are highly energetic and more likely to trigger production of free radicals than others. Green light also penetrates deeper into the leaf than other wavelengths, reaching down into the spongy tissue (see Graphic).

Neill found that in lollo rosso lettuces – the ones that are reddish purple around the crinkly outer edges and green towards the centre – the red tissue coped better with intense light than the green. Strong light generated fewer free radicals in the red tissues. “When things are going badly wrong, red pigments act directly by scavenging radicals and indirectly by acting as a shield,” says Neill.

Gould, with the help of Tom Vogelmann at the University of Wyoming, was able to see for himself how effective that shield is. They studied leaves from the tawheowheo (Quintinia serrata), a tree that has both red and green leaves and some that are blotched with both colours. If chlorophyll is exposed to green light it fluoresces. When Gould and Vogelmann shone green light on green leaves, the glow extended from top to bottom. When they did the same to red leaves, none of the chloroplasts in cells below the red pigmented tissue glowed (Physiologia Plantarum, vol 116, p 127). “Simply having red pigments reduces the amount of light falling on the chloroplasts,” says Gould. “With anthocyanins placed in the mesophyll, they can both screen light and are also optimally placed to scavenge free radicals – twice the protection for your money,” says Gould.

The pigment’s dual role as light-shield and free radical scavenger would explain why the young leaves of so many tropical species are brilliant red. New shoots high in the canopy are exposed to high temperatures and intense light. “The red layer shields the developing chloroplasts, which are very susceptible to damage,” says Neill. By the time the leaves have unfolded fully they can produce more antioxidant enzymes and other flavonoids. They no longer need high-level protection and the red disappears.

The shield-and-scavenger theory probably also applies to those plants on the rainforest floor that have red undersides. Only a tiny fraction of the light hitting the treetops reaches ground level, and plants that grow there are adapted to deep shade. But just occasionally a shaft of sunlight penetrates the gloom and hits plants with light a thousand times as intense as they’re used to. These shade-adapted leaves lack the waxy coatings, hairs and abundance of antioxidant enzymes that protect leaves used to life in the sunny canopy. Instead they have red pigments. But why do they have them only in the underside of the leaf?

Bottom dealing

There is a good reason. If the upper tissues were red the plant would find it difficult to capture enough light to photosynthesise at all. Putting the anthocyanins in the lower layers of the leaf seems to be a compromise, says Lee: it creates a gradient of light through the tissue, protecting the most light-sensitive chloroplasts in the lower layers while allowing photosynthesis to take place in the less sensitive cells nearer the surface.

Could this shield-and-scavenger idea also explain the dazzling reds of a New England autumn? For many years the accepted view among plant physiologists was that autumn colours are simply what’s left over after the plant breaks down its chloroplasts and salvages what it can before it throws off its leaves. That’s true of the yellow and orange colours, says Lee, but not the red ones.

With Michele Holbrook and Taylor Feild from Harvard University, Lee examined 89 species with red autumn leaves from the Harvard Forest in Massachusetts. “In 70 per cent of the species we looked at, anthocyanins were produced during senescence,” says Lee. That is, the leaves actively pump out more red pigments even as they begin to die. What’s more, when these trees are exposed to extra stress, such as a sudden frost, their leaves turn an even more intense red.

Anthocyanins do a good job of protecting autumn leaves. Lee and his Harvard colleagues looked at how effective the red shield is in the red-osier dogwood (Cornus stolonifera). The more exposed leaves of this shrubby dogwood turn reddish-purple in autumn, while the more shaded leaves stay greenish. Given a 30-minute blast of high intensity light, green leaves suffered more damage than red ones, and recovered more slowly, the researchers found (Plant Physiology, vol 127, p 566).

But what’s the point of protecting dying leaves? Lee believes the red pigment shields chloroplasts from excess sunlight long enough for the tree to retrieve what nutrients it can from the leaves. These extra nutrients help the tree produce new growth the following spring. In autumn as the leaves begin to dismantle their chloroplasts, they become more susceptible to light damage. The chlorophyll molecules liberated in this breakdown are highly dangerous: if light hits them they can generate a profusion of potentially lethal oxygen radicals. “If the leaf cells were damaged during senescence, that would reduce the success of the salvage operation,” says Lee.

So have Gould and Lee finally solved the puzzle of why some leaves are red? Their evidence is compelling, even if they haven’t won over all their colleagues yet. But the final answer might come from a more lowly plant, a liverwort. Liverworts were the first plants to grow on land, taking up terrestrial life some 450 million years ago. They colonised a hostile world where they were exposed to high light levels and intense ultraviolet radiation. Survival depended on finding a means of dealing with free radicals. Did they cope by making red pigments?

Today, most liverworts grow in damp, shady places and they’re green. Jamesoniella colorata is different. It grows on Rangitoto Island, a volcano in the middle of Auckland Harbour. It’s a nasty place for plants, with intense light, few nutrients and lots of UV – probably not so different from the world the first liverworts inhabited. There’s something else that’s different about this particular liverwort. It’s red.