Exploring the Miller Worm Farm

By Leigh MacMillan
March 11, 2002

It seems like any other laboratory door sign, until the words on it make you look twice. “Miller Worm Farm,” announces the small blue placard next to David Miller’s laboratory. Worm Farm?, you wonder, expecting to glimpse dirt-filled aquariums boasting intricate tunnels of wriggling worms. Instead, a standard lab scene greets your inquiring eyes — microscopes and computers are as exotic as it gets.

Miller's laboratory is indeed a place dedicated to cultivating worms, but not the juicy kind that nourish gardens. These worms are tiny, about the thickness of an eyelash and only a millimeter long. And Miller doesn't raise them just for kicks — he's in the business of scientific discovery, tackling in particular the tough questions of how nerve cells choose partners.

“The worm,” as it is fondly known, is an increasingly popular organism for biomedical research. Many of the fundamental cellular processes found in the worm are shared across millions of years of evolution with human beings. By studying a simpler “model” organism like the worm, investigators hope to learn about the processes at work in human cells.

The making of a good model

Scientifically, the worm is called Caenorhabditis elegans, C. elegans for short. It is a nematode, a member of a large phylum of worms that live in soil or water, or as parasites in plants or animals.

C. elegans got its start as a model system for biological research in 1965. Sydney Brenner, an investigator at the Medical Research Council in Cambridge, England, selected the worm as a promising animal for investigating the development and function of a simple nervous system.

Brenner had already participated, with Francis Crick, in the identification of messenger RNA and the elucidation of the triplet genetic code. In the early 1960’s, he and Crick felt that the future of molecular biology “lay in tackling more complex biological problems,” especially in the fields of development and the nervous system, Brenner wrote in a Foreword to the 1988 book The Nematode Caenorhabditis elegans.

Brenner originally selected a C. elegans cousin, C. briggsiae, as a promising candidate for rapid genetic and biochemical analysis of the mechanisms of cellular development. He was looking for a multicellular organism that had a short life cycle, could be easily cultivated, was small enough to be handled in large numbers, had relatively few cells so that cell lineage and patterns could be studied, and was amenable to genetic analysis. Both nematodes fit the bill, and C. elegans was eventually selected in preference.

In the nearly four decades since Brenner first proposed studying a nematode, hundreds of investigators have joined the cause. The development of every cell, starting with the single egg and progressing to the 959 body cells of the adult worm, has been mapped. “We have in essence a very large family tree diagram that maps out the parents of every single cell,” Miller explains.

The worm field also benefits from a complete “wiring diagram” of the animal’s 302-celled nervous system. Investigators spent more than 10 years “slicing up the worm-like a salami-into thousands of thin sections, looking at each one with an electron microscope, tracing out all of the neurons, and reconstructing the whole circuitry,” Miller says. He shows off the tattered volume of the journal that published the work. “In here are catalogued the morphology and connections of every single worm neuron,” he says. “It’s the only organism for which this kind of information exists.”

The fully charted cell lineage and nervous system wiring maps are invaluable tools for investigators, who can easily compare cell patterns in genetically manipulated and normal worms.

Most recently, the worm became one of the first multicellular organisms to have its entire genome sequenced. Since that development in 1998, even more investigators have joined the ranks of those interested in studying fundamental biological processes in a simple animal.

“The completion of the genome really jumpstarted worm research,” Miller says. “It has made the worm accessible to focused molecular approaches that weren’t necessarily available before then.”

Uncoordinated worms shed light on nerve cell connections

Miller was a graduate student at Rice University in Houston when he heard that Sydney Brenner was giving a presentation at nearby Baylor College of Medicine. It was 1978, about five years after Brenner’s first paper using the worm. The seminar charted Miller’s future course, prompting him to pursue postdoctoral research in the worm field — eventually with Brenner himself.

“He presented the idea of using this organism to understand biology, of using genetic approaches to figure out the key molecules in biochemical pathways,” Miller recalls. “I was really swept away by the notion that you could use genetics to dissect a complex process without making any assumptions about what the players are.”

Using chemicals to induce genetic mutations, Brenner had isolated worms with unusual or uncoordinated movements. His collection of “unc” mutants is still being studied today. Miller’s group, in fact, studies the unc-4 mutant, a worm unable to crawl backward. In unc-4 mutant worms, the pattern of connections between nerve cells is changed — specific motor neurons are incorrectly “wired” with the wrong synaptic inputs.

Miller and colleagues cloned and characterized the unc-4 gene and the protein it encodes (UNC-4). UNC-4 is a protein called a transcription factor; it works in the cell nucleus to turn other genes “off.” “UNC-4 turns off the ‘wrong’ genes so that neurons make the ‘right’ connections,” Miller says. In fact, this negative mechanism of gene regulation to specify motor neuron connections appears to be evolutionarily conserved in the vertebrate spinal cord.

In unc-4 mutant worms, Miller says, the “wrong” genes that would normally be turned off by UNC-4 are being left “on.” And somehow this is changing nerve cell connections. Miller hopes that figuring out which genes UNC-4 regulates will answer “fundamental questions about how neurons choose synaptic partners.”

To find the genes UNC-4 controls, Miller and colleagues will take advantage of an advanced technology called DNA microarrays, which are tiny glass “chips” with thousands of DNA spots dotting their surfaces. Scientists can use them to probe which genes are on or off in a certain kind of cell, for example the cells of an unc-4 mutant worm.

Miller and collaborators including Kevin Strange recently developed a method for culturing worm cells in a dish and separating the unc-4-expressing neurons from other cells on the basis of a fluorescent protein (GFP). Comparing the pattern of gene expression in unc-4 mutant neurons and normal control neurons should reveal the genes regulated by UNC-4, Miller says.

Worms provide key insights to cell death, axon guidance

Other investigators have pursued studies of uncoordinated worms or have carried out genetic screens for other kinds of defects. For at least two areas, cell death and axon guidance, “genetic approaches in the worm have yielded key insights into biological pathways that are highly conserved between worms and human beings,” Miller says.

During the course of worm development, approximately 300 extra cells are generated and then self-destruct. Because the worm is transparent, investigators can actually watch worm cells as they divide and as they die. Programs of cell suicide are normal and happen on a grand scale in the developing human body. The classic example, Miller says, is the programmed death of cells that, in the fetus, web together the fingers and toes.

Robert Horvitz at the Massachusetts Institute of Technology pioneered the search for genes controlling cell death. Like Brenner, he used chemicals to cause genetic mutations in the worm. Instead of looking for uncoordinated movements though, Horvitz picked out mutant worms with abnormal patterns of cell death. The cell death — “ced” — genes he and colleagues identified have defined a core cell death pathway and accelerated understanding of cell death in mammalian systems.

These insights have direct bearing on the molecular understanding of cancer, which can result from abnormal control of cell death pathways. Similarly, the worm has accelerated studies of axon guidance-how the communicating processes of nerve cells find their way to the right target cells during development.

In just one example, a series of the “unc” mutants led to the identification of a biochemical signaling pathway required for certain neurons to send their axons in the correct direction. The conserved molecules were cloned more quickly from vertebrates by using the known DNA sequences of the worm molecules. “This is one of many cases where the genetics of C. elegans predicted a paired signal-receptor set important for steering axons,” Miller says.

Understanding the molecular signaling pathways that guide nerve cell growth and connections could lead to strategies for repairing nervous system damage.

Studies using the worm as a model system will undoubtedly continue to illuminate the molecular participants in biological pathways that are simply too cumbersome to tackle in a mammalian system. Miller and other Vanderbilt investigators are along for the exciting ride, eager to probe fundamental questions of cell biology that will inform understanding of human health and disease.

The Miller lab is supported by grants from the National Institutes of Health.

The Miller Lab