At six foot three, Philip Benfey towers incongruously over the bedraggled-looking collection of shriveled plants that he displays with considerable pride. The plants languish in plastic pots on a shelf in his laboratory's closet-like plant-growth room. They remain stubbornly, even ungratefully stunted, although they bask in brilliant artificial sunlight, ensconce their roots in the best soil, and imbibe the scientifically determined optimal measure of water and fertilizer.
However, to Benfey, who is chair and professor of biology at Duke, the puny plants constitute the intellectual equivalent of giant redwoods. These particular mustard plants, scientific name Arabidopsis thaliana, harbor a fascinating gene mutation that eliminates a key growth-regulating gene. The mutation interferes with subtle biochemical signals between cells in their growing roots--stunting them, and, thus, the entire plant. In contrast, the normal plants nearby reach for the sun--or rather the brilliant artificial light in the growth room. They stretch their gangly stems upward about a foot, supported by clear plastic cylinders. Benfey studies what happens when arabidopsis genes known as "Short Root" and "Scarecrow" are mutated, in effect, broken so that they don't work properly. His work has yielded extraordinary insights into how these growing roots develop.
While Arabidopsis might seem an obscure bit of foliage, the little plant is celebrated among geneticists as the laboratory mouse of the plant kingdom. A relative of cabbage and radishes, Arabidopsis is small and prolific and grows easily and quickly.
"The root has a fairly complex structure, with lots of different cell types. And it all begins from a single cell," says Benfey. But unlike the impossibly intricate convolutions and migrations of developing animal bodies, each new Arabidopsis root cell arises conveniently from its neighbor. "When you look at the anatomy of the root, the origins of the entire structure are right there in front of you," he says. "You can see all the stages of development. For genomics, this is an enormously simplifying feature."Benfey's studies of the plant's tiny tangled roots might be considered just a minor botanical curiosity if they applied to only one species. But his research is helping science get to the ... well ... root of one of the central questions in all of biology: the immensely complex puzzle of how entire tissues, whether plant roots or human brains, blossom from a single cell. The solution would advance a vast range of disciplines from agriculture to medicine. And the Arabidopsis root has afforded Benfey and his colleagues a ringside seat at the biological spectacle of the development of living tissue.
Thus, says Benfey, exploring the consequences of mutations in just a single gene such as Short Root or Scarecrow can yield a world of insight into tissue development. Biologists, including Benfey and his cohorts, are gleeful scientific saboteurs, mutating genes to make them malfunction and keenly observing the resulting biological havoc. (The scientists, perhaps perversely, often name genes according to the ill effects of breaking them. The origin of the name Short Root is rather obvious; the mutation of the Scarecrow gene produces roots missing a critical layer of root cells--like the missing brain in the Wizard of Oz character.)
Sabotaging genes is especially informative because they are the blueprints for the multitude of proteins that make up the machinery that keeps cells--from plant roots to hair roots--functioning. A sabotaged blueprint produces a nonfunctional protein, disrupting that machinery in interesting and instructive ways. Benfey's research has revealed that Scarecrow and Short Root are blueprints for proteins that help form the same growth machinery pathway in the plant root.
One of the holy grails in modern biology is understanding these pathways in order to learn to control them. The machinery of every living cell consists of a host of such molecular signaling pathways, like the systems that make up a car's machinery--the fuel system, cooling system, electrical system, drive train, and all-important entertainment system that keeps the kids quiet in the back seat. By assiduously breaking one component or another--like, say, taking a hammer to a carburetor--researchers can deduce which pathway each component belongs to.
The proteins made by the Short Root and Scarecrow genes are more than modest cogs in the cell's machinery--they are themselves master controllers of other genes. Such controllers, called transcription factors, orchestrate the activation of a multitude of other genes that themselves build the cell's machinery. The two transcription factors are the molecular equivalents of the Wizard of Oz--not the "great and powerful" public Oz, but the little man hidden behind the curtain, yanking genetic levers and poking genetic buttons that help orchestrate the amazing phenomenon of development. It's a phenomenon every bit as spectacular as the flame and folderol created by the little wizard--with myriad genes creating the miraculous, elegant symphony of growth that induces a single embryonic cell to burgeon into an intricate living organism.
In the quest that led to Short Root and Scarecrow, Benfey began, as have so many other scientists, with fingers tightly crossed. "When we did the first genetic screen of mutant plants, we didn't set out to answer a specific question and design a fancy, elegant screen that would get to an answer. Rather, I said, 'I'll take anything I can get,' and then go after it." But they were seeking mutations that affected plant growth, and they found many stunted plants among the mutants they created using gene-damaging chemicals. When the scientists analyzed the stunted plants, they pinpointed two particularly intriguing genes--Short Root and Scarecrow--with mutations that appeared to cause a very surprising pathology in their roots.
"Something I had no way of predicting was that the Short Root mutants were missing entire cell layers," says Benfey. The missing layers hinted tantalizingly at the failure of a critically important step in tissue development--the differentiation of a whole class of cells from general purpose into those with particular specialties and functions. In animals, a parallel might be the disappearance of a whole layer of the brain. The differentiation of cells from the general to the specialized is the hallmark of development from embryo to adult, and understanding differentiation is a central aim of biologists.
Benfey knew he had stumbled onto a key developmental gene. Further study of Short Root mutants yielded yet another scientific surprise: The gene makes a protein transcription factor that appears to activate genes in an entirely different set of cells from the ones in which it is made. "And then the big surprise was that the protein transcription factor, in fact, moves from cell to cell," says Benfey. That migration ran counter to all understanding of how transcription factors work--as if the Wizard of Oz had jumped books and somehow contrived to take the controls of the Little Engine that Could. "If you asked me ten years ago which of those I would have predicted," Benfey adds, "I would say neither of them."
Benfey's research has revealed that the Short Root gene produces a biologically bossy protein that instructs neighboring root cells in three ways. It tells a neighboring cell's genes what kind of root cell to become, how to divide to make new cells, and even where to position itself in the burgeoning root. The researchers have also discovered that Short Root and Scarecrow are developmental partners, with Short Root's protein apparently activating Scarecrow's to govern cell division. Now the researchers are searching for the "downstream" genes that Short Root and Scarecrow control, and the "upstream" genes that control them. It's a process not unlike opening a mystery novel at the middle and trying to deduce how the mystery began and how it will end.
If Short Root and Scarecrow exemplify the traditional "retail" gene-by-gene study of root development, Benfey and his colleagues have now gone into the "wholesale" research business as well. Last year, they announced that they had created the first detailed "gene expression map." (Gene expression is the process by which a gene activates itself to make protein.) The map reveals when and where some 22,000 genes are activated in each of the intricate mosaic of cells of the growing Arabidopsis root. "This is the first time anybody has achieved this level of resolution of gene expression on a global basis for any organism," says Benfey. Other researchers had ground up whole plant tissues and analyzed their gene expression, he says. However, in such studies "critical information on the mechanisms of development was lost. Development occurs at the single-cell level, and there's a dramatic difference from one cell to the next, in terms of its gene expression."
The results of the painstaking analyses, published in the journal Science, in essence told of a scientific race against time in the Benfey laboratory. Once the scientists had labeled an individual root-cell type with a fluorescent marker gene, they had only about ninety minutes to isolate the particular type of cell from the rapidly growing root for genetic analysis. "The cells are intimately connected to one another and constantly signaling to one another," says Benfey. "And, if you wait much longer, they begin to change their gene expression." Once the cells were isolated, and their genes extracted, the scientists used so-called "gene chips"--massive arrays of genes on fingernail-sized glass chips--to measure the activity of each of about 22,000 genes in each cell type.
The Arabidopsis gene profile exemplifies a new era of "systems biology," says Benfey, in which scientists graduate from studying individual genes to mapping interacting networks of genes and the proteins they produce. "A key to understanding development of plant tissues and organs is determining how whole networks of genes are regulated during development," he says. Such knowledge will bring profoundly important practical benefits. "In applying systems biology to agriculture, we can progress from altering one or two genes at a time. We can graduate from just putting a foreign gene from a bacterium into a plant to change one trait such as herbicide resistance. Rather, we can change broad traits that are already there, because we will know how regulatory networks function. We'll be able to modify plants to resist drought, grow in salty soil, resist higher temperatures due to global warming, or resist pathogens."
Benfey says he also believes that lessons learned from the Arabidopsis plant will likely apply to higher organisms, even humans. "It would surprise me if the particular processes that we are finding with Short Root and Scarecrow exist only in plants," he says. "The history of biology as I read it is that just about everything that seemed to be peculiar to one particular system or stage of an organism has turned out to be much more general."
His career will always reflect the search for scientific truths in straightforward structures such as roots, says Benfey. "In retrospect, my quest for simple systems is a theme throughout my scientific career. In the Arabidopsis root, I saw a tissue in which I could immediately identify the component parts, where its growth properties offered a simple model of development and differentiation."
And with the scientific truths from the plant root have come an appreciation of its unconventional beauty. "Most people think of the root as this very unaesthetic part of the plant. They focus on distinctive flowers or beautiful leaves," he says. "But if you think of aesthetics in the sense of simplicity, when you slice through a root and you show people that they can really see its various parts, they do find an immediate attraction there."
October 1, 2004