The traveler who commutes between the high-tech molecular biology labs at the Massachusetts General Hospital and the steamy wooden porches of Laguneta crosses distances not easily measured in miles. Yet, by the spring of 1983, powerful bonds of common interest linked these two sites, each marked in different ways by Huntington's disease. For certain scientists at Massachusetts General, the people of Laguneta, Barranquitas, and San Luis were becoming a focus of intense interest. Many difficult technical obstacles to mapping the Huntington's disease gene had been solved, problems relating to cell lines and storage. Researchers were now convinced that the Venezuelan communities offered the kind of families needed for such a study. In addition, the number of RFLP markers was rapidly growing, confirming the predictions of Botstein, White, Housman, and others that RFLPs would be numerous and distributed evenly enough throughout the genome to serve as useful markers for mapping genes. Back in 1980, only two such markers had been known. By now there were nearly two hundred RFLP markers, and RFLP mapping was well under way. Some geneticists estimated that a linkage map of the entire human genome would be completed within three or four years. [1]

Still, one of the major challenges for Gusella lay in generating more RFLP markers. Even more daunting was the prospect of testing each newly developed marker for a possible correlation with Huntington's. Gusella expected that they might have to test 150 to 200 markers - an immense amount of work - before they found one close to the Huntington's gene. [2] As one of Gusella's technicians, Rudolph Tanzi, put it, "We were really in the random boat."

For several months, Gusella's group had been testing each newly created marker against two different Huntington's disease pedigrees, hoping to find a consistent difference between people with Huntington's and those without it. They were using DNA from the Iowa family, which was theoretically large enough to yield statistically significant results. [3] They were also using DNA from the rapidly expanding Venezuela pedigree, which now numbered some 3,000 people, of whom about 1,200 were living. [4] By the spring, they had constructed thirteen new genetic markers and had tested eleven of them for linkage, without coming up with significant results. [5] Gusella wasn't discouraged. He knew he was still in the initial phase of the work.

Sometime in April, they decided to test the twelfth probe, called G-8 after the technician named Ginger Weeks, who had designed this probe from DNA plucked arbitrarily out of a human DNA library - a collection of short segments of DNA. G-8 was a little swatch of DNA about 17,600 base pairs in length. It was an anonymous sequence, which meant that no one had yet identified the particular chromosome from which it came. And it was ideal as a probe since it was one of a kind - a sequence of nucleotides that occurred only once on one chromosome. (In fact, G-8 was only the third such "single copy" probe of the twelve they tested.) As they did with each probe, they first tested G-8 to determine if it picked up any variable regions in the DNA. When they mixed the G-8 probe together with DNA fragments from the Iowa family-fragments that had been chopped up by the restriction enzyme HindIII, they saw that G-8 revealed fragments of several different sizes. That meant that G-8 was indeed picking up two different RFLPs-that is, two places in the DNA where individuals varied. These variations, which were extremely close together, occurred at spots where the HindIII enzyme happened to cut. Hence they gave rise to four different fragment lengths.

Some individuals produced fragments that were 15,000 nucleotides in length; these people possessed the first sequence recognized and cut by the HindIII enzyme. Others produced longer fragments, of 17,500 nucleotides; these people lacked the first HindIII cutting site. There was a second variable spot as well. At this spot, those individuals whose DNA was cut by HindIII produced two different fragments, of 3,700 and 1,200 nucleotides. Other people, who lacked the second cutting site, produced one longer fragment, of 4,900 nucleotides. Altogether, G-8 picked up two sites of variation, with four possible combinations determined by the presence or absence at each site of a sequence cut by HindIII.

It sounds complicated, but it was actually straightforward: four possible patterns of fragments, technically called haplotypes, that Gusella's group called A, B, C, and D. Each individual in a family inherits two haplotypes, one from the father, one from the mother. This is true in families without Huntington's as well as those with the disease. The critical question for Gusella was whether any of these haplotypes might be correlated with the illness infamilies affected by the disease. (In unaffected families, the haplotype would be insignificant.) In other words, do individuals who have the disease consistently have a particular haplotype? Does a particular pattern of fragments always travel together with the disease?

When Gusella looked at the distribution of patterns with G-8 in the Iowa family it did look as if there might be a relationship. Individuals who were known to have the disease seemed consistently to have the A haplotype, though there were many individuals without the disease who also had the A haplotype. (Since these four haplotypes occur in people with and without Huntington's in the family, and since everyone has two haplotypes, it is quite possible for healthy people in a family with Huntington's to have an A pattern as well. It is also possible for someone with Huntington's to have two A haplotypes, one from the parent with the illness and one from the other parent.)

The LOD score for the Iowa family came out of the computer at 1.8. A LOD score is a statistical calculation geneticists use to determine the likelihood that two genes - or any two segments of DNA - are actually linked, that is, physically located on the same chromosome, and not merely inherited together by chance; they consider a LOD score of 3 as establishing genuine linkage. (Technically, a LOD score is the logarithm of the ratio of the odds for or against linkage.) The higher the LOD score, the closer together on the chromosome the two points are likely to be. A score of 3 means the odds are a thousand to one that linkage is not a chance result.

So a score of 1.8 was good, but not startling: it meant that the odds were 65 to 1 against chance that G-8 was linked with Huntington's in this family Gusella was not impressed. "There's not a geneticist in the world, especially in a case like Huntington's disease," Jim told me later, "who wouldn't say, well, that score will go away with more data. So we weren't particularly excited about it." He wanted a LOD score of at least 3. Still, after Mike Conneally in Indianapolis got the data from Jim and did a statistical analysis, he called Boston and said, "Jim, it looks promising. Why don't you stick with that one, G-8, and do the Venezuelan family." At that point, though, Gusella did not have enough DNA extracted from any of the Venezuelan families, and isolating the DNA from the nuclei of white blood cells takes time. He and Rudi Tanzi decided to prepare a large subset of the Venezuelans that they knew would be informative because of all the people with Huntington's in it. That process continued until the beginning of July.

Meanwhile, they already had some DNA ready from a much smaller subset of the Venezuelan family that they had been using for something else. In early June they decided to run gels on this group with G-8. "It was a very simple little pedigree," Jim recalled. "All it involved was a mother and father and four children. And it was the only case, the only set of people where we had a grandparent available and already made into DNA. By itself it wouldn't give you very much of a LOD score. But it's one that's very easy to look at and see what's going on." In this little pedigree there were two children with the disease and two without it. Both children with Huntington's had the pattern or haplotype called C, while the unaffected children had a different haplotype. (That the individuals with Huntington's in the Iowa family had an A haplotype while the Venezuelans with Huntington's had a C haplotype was insignificant: the critical question was whether, in each family only one haplotype traveled consistently with the disease.)

"At this point," recalled Mike Conneally "the LOD score went up to 2.2. Now, that's odds of about a hundred twenty to one against chance." It still wasn't highly significant; but at least the scores were going up. "I had hardly heard about it," said Mike, "hardly gotten the run out of the computer, when Nancy called. And I said to Nancy, things are looking good, they're looking promising, I think we're getting somewhere. I think we've got a marker. And Nancy said, oh terrific! And when I hung up I thought, Mike, you damn fool, you should never have said that, you really put your foot in it, you'll have to eat crow later. I thought, why didn't I have the fortitude to hold back and not tell her that? because I'd been so frustrated in the past, I really didn't believe it myself."

But Jim too was impressed by the results. "Of course it was not definitive," he admitted later; the pedigree they had used was too small. And one of the unaffected children was quite young and could still develop the disease in the future. "But I looked at that," said Gusella later, "and my mind went from saying that the 1.8 was merely being tested to thinking, maybe the 1.8 is really right. And we better get this tested fast."

Feeling a new sense of urgency, Jim and Rudi rushed to prepare the DNA from the other Venezuelan subset, which was quite large and therefore more informative. Gusella felt increasingly pressured, since he was scheduled to attend a week-long meeting in Aspen, Colorado, at the end of July and really wanted to get the data out to Mike before he departed. "On the Friday afternoon before I had to leave we got the autoradiograms, got them all dried, and Rudi and I sat down and read them all," Jim recalled later. "And then we took the data and put it on the pedigree to send to Mike. And it was just obvious, from the pedigree, that this was going to make the LOD score go up rather than down, because it matched perfectly Everybody who was affected had a C [haplotype], and virtually nobody else did." Looking at the pattern on the pedigree spread out in front of him, Gusella knew he had found the linkage even before the data went through the computer. He could see from the pedigree that G-8 picked up two RFLPs that were right next door to the Huntington's gene.

As soon as he realized the linkage result, Jim tried to call Nancy in Washington, but he was unable to reach her. He decided to wait until he got the LOD score back from Mike Conneally before trying her again. Meanwhile, in Indianapolis, Mike Conneally took a fast look at the data Jim had sent him and saw that, superficially, it looked good. He too was about to leave for the West, for a vacation in the Grand Canyon, so he instructed his graduate students to feed the new data into the computer. "Then I'm in Grand Canyon National Park on Tuesday night," Mike recalled, "sitting by my campfire having a beer with my wife and daughter. And I see this cop coming back and shining a light on our car. At least I thought he was a cop. He was really a park ranger. When he saw we had Indiana plates he came over and said, are you Dr. Connally? They always call me Connally! I said, yes I am. And he said, I have an emergency, an important message for you to call Peggy [Wallace] or Beth [Jones] - these were my graduate students. It was now eleven o'clock at night. At that time, it would have been one in the morning in Indiana. I thought, well, of course I can't call them tonight. Little did I know they were waiting up for me to call. So I didn't call until the next morning. But I knew. I was convinced. I said to my wife, we've got it! At last it has happened! And the next morning when I called, of course, there it was. The LOD score was six point something, and there was absolutely now no doubt, the odds were a million to one, and you know, that was it."

"How'd you feel when you heard the news?" I asked Mike Conneally later, though the answer was pretty obvious. "Oh, elated!" said Mike. "Absolutely elated. I had a few more beers that night, I can tell you. I was almost crying, because I thought, at long, long last, the search is over. I felt so good. And you know who I felt good for? Nancy I thought, that's great for Nancy because she worked so hard on it. Then I talked to her, two nights later. It was the greatest feeling I've ever had, I know, in my life."

This issue of and our previous issue of HMS Beagle feature a number of articles on neurodegenerative diseases and neuroscience, with numerous additional endlinks to relevant online resources. For more on neurodegenerative disease, see the Cutting Edge debate, "Neurodegenerative Diseases" and In Situ's "Adult-Onset Neurodegenerative Diseases". For more on neuroscience, see our Site Review of Neurosciences on the Internet and the Meeting Brief "Brains and Hurricanes: Highlights of the 1997 Society for Neuroscience Meeting."

Mapping Fate - a brief description of Wexler's book, and a review.

The Hereditary Disease Foundation - a "nonprofit, basic-science organization dedicated to the cure of genetic disease," the foundation focuses on Huntington's disease as a model. The foundation provides grants and fellowships, and sponsors interdisciplinary workshops. The site covers testing protocol and ethical, legal, and social issues; recommended reading; news; research funding; and links to related sites.

Huntington's Disease Society of America - a national voluntary health agency promoting HD research and supporting HD sufferers and their families. Site sections include: Publications and Videotapes; Genetic Testing for HD; HDSA's Research Grant and Fellowship Program; HD on the Internet; Where to Find Help, which provides information on HDSA chapters; Residential Care Facilities; HD/Movement Disorder Clinics; HD Research Roster and DNA Bank; Brain Tissue Banks; and the International Huntington Association. Most information is organized by state.

Huntington's Disease Sites - links to lists of links. Sections are Caring for Huntington's Disease; Facing Huntington's Disease; and Research Highlights on Huntington's Disease. The last section links to the NIH's National Institute of Neurological Disorders and Stroke.

"Huntington's Disease Genetics" - a brief article on the genetics of the disease, with a graphic of affected DNA and the resultant protein huntingtin.

Online Mendelian Inheritance in Man (OMIN) Database - a catalog of human genes and genetic disorders, including text, images, and references. In addition to OMIN resources, links are also provided to "allied resources" including Entrez MEDLINE, the Davis Human/Mouse Homology Map; Online Mendelian Inheritance in Animals, and the Alliance of Genetic Support Groups.

The Genetics of Neurodegenerative Diseases - links to recent key citations and to a short review of the field. Maintained by Cambridge Scientific Abstracts.