When CAG spells trouble: DNA repeats may turn good proteins into bad - repeated nucleotide triplet underlying many genetic diseases

Science News, June 10, 1995 by John Travis

When Huda Y. Zoghbi, a pediatric neurologist at Baylor College of Medicine in Houston, spoke to a local family marked for seven generations by the inherited disease spinocerebellar ataxia type 1 (SCA1), no one recalled having heard anything dramatic about the long-dead man to whom she had traced the first obvious sign of the illness. He had lived well past his 80th birthday and "just tumbled around a bit late in life. Nobody thought much of it,'' current family members told Zoghbi.

By the beginning of this decade, geneticists thought they had a firm handle on most of the ways DNA could get nasty. Then in 1991 came an unexpected report about the genetic flaw underlying fragile X syndrome. The most common inherited form of mental retardation, fragile X afflicts 1 in every 2,500 people, most of them men.

On the X chromosome of affected individuals, a small stretch of DNA had seemingly taken on a life of its own and copied itself over and over. As researchers traced this unstable DNA through family trees, they found that the repeating portion was often larger in each new generation.

Suddenly, the puzzling but well-documented trait of "anticipation," in which a disease strikes earlier and more severely in successive generations, had a possible explanation. Researchers quickly established that other conditions exhibiting anticipation, such as myotonic dystrophy and Huntington's disease, also owed their unusual inheritance pattern to expanding DNA.

More than a half dozen genetic illnesses have now been pinned on this phenomenon, which still goes by various names: unstable DNA, dynamic mutations, triplet repeats. "This is a whole new area of medical genetics. We didn't even know about [these repeats] a few years ago,'' says Marian DiFiglia of Boston's Massachusetts General Hospital (MGH).

Once past the initial shock of discovering a new mechanism behind inherited diseases, researchers began delving into how repeating DNA wreaks its havoc. Recent work provides some insight into three deadly inherited neurodegenerative illnesses--Huntington's disease, SCA1, and dentatorubral and pallidoluysian atrophy (DRPLA)--caused by these genetic stutters.

Each of the diseases initially destroys only a specific group of brain cells. SCA1 targets cerebellar neurons called Purkinje cells, while DRPLA devastates different neurons in the cerebellum and other regions of the brain. Huntington's disease ravages so-called spiny neurons in the striatum.

All three are also CAG-repeat diseases, a description that refers to the strip of DNA that proliferates. The building blocks of DNA are four versions of a complex organic molecule called a nucleotide. Designating these nucleotides as C, A, G, and T, geneticists spell out the sequence of any DNA strand using just these letters.

When researchers identified the genes responsible for the three diseases, they found that in unaffected individuals, each cell's two copies of the relevant gene had stretches of DNA where CAG repeated anywhere from half a dozen to nearly 40 times. But in affected people, the CAG triplet occurred from 40 to more than 100 times on one or both copies of the gene.

How does this genetic profusion lead to a disease? In fragile X syndrome (where the repeated nucleotide triplet is CCG rather than CAG), the position of the extra DNA within a gene appears to work by squelching the manufacture of a protein at the transcriptional level. Transcription is the process in which the DNA sequence of a gene is converted into messenger RNA, a molecule that the cell then uses to construct the gene's protein.

If the number of CCG repeats exceeds a still poorly understood threshold, cells produce less and less messenger RNA from the fragile X gene and correspondingly less protein. Geneticists therefore attribute the syndrome to a loss of function, since it appears to result from the absence of a vital protein.

Three generations after that first family member was affected, some members were bedridden by their sixties and dead by their seventies. SCA1 first destroys neurons in the cerebellum, an area of the brain that helps control balance and coordination. "It eventually kills patients because they can't swallow or breathe,'' says Zoghbi.

Not all DNA-repeat diseases work this way. Researchers have shown that in a fourth CAG-repeat illness, Kennedy's disease, the expanded gene pumps out flawed versions of its normal protein, a receptor for the hormone androgen. Since each nucleotide triplet determines a specific amino acid for a protein, the extra CAGs appear to add a long stretch of one amino acid, glutamine, to the receptor's normal sequence.

Groups from France, Japan, and the United States report in the May Nature Genetics that Huntington's disease, DRPLA, and SCA1 follow in the footsteps of Kennedy's disease, not fragile X. The CAG repeats on the disease genes apparently translate into extra glutamines, creating larger than normal proteins.

Research teams led by Ichiro Kanazawa of the University of Tokyo and Masao Yamada of the National Children's Medical Research Center in Tokyo collaborated on the DRPLA research. From the sequence of the normal DRPLA gene, they predicted the form of small bits, or peptides, of the resulting protein and synthesized them. They then inoculated rabbits with the peptides, producing antibodies that bind to those fragments.