By David N. Leff

When a person afflicted with myotonic dystrophy shakes hands, he or she can't let go. Hand muscles clenched in greeting are unable to relax. This seemingly trivial trait is in fact a diagnostic hallmark of myotonic dystrophy (DM).

"It's not a problem in the nerves," explained molecular biologist Thomas Cooper, at Baylor College of Medicine, in Houston. "It's intrinsic to the skeletal muscle, where it's thought to be a disturbance of electrical conductance. Somehow, the electrical mechanism of contraction gets started and it can't stop.

"Hence the inability of these DM people to release their hand muscles," he continued. "They feel it more as a stiffness than a hard contraction like a muscle spasm. The physiology is unclear."

That's not all that's unclear in the cause and course of DM, which Cooper described as "the most common form of adult-onset muscular dystrophy." A more severe, congenital, form, he added, strikes infants at birth or early in childhood.

Besides torsional contractions of head, neck and arm muscles, DM's symptomatology can cause grave damage to heart, testes, ovaries, eyes and other organ tissues. Rare compared to Duchenne muscular dystrophy, DM numbers one in 8,500 victims in the U.S. — on the order of 32,000 Americans.

Like Huntington's disease (HD), DM is a "triplet-repeat" disease, a genetic defect in which a three-base nucleotide sequence in the patient's genome reiterates itself at great length, until it wreaks havoc in the gene's functions. Also like HD, DM runs in families. Healthy humans walk around with anywhere from five to 35 repeated bases in their skeletal-muscle-cell genes. These consist of cytosine-thymine-guanine (CTG) triplet codons. When these genes start to express their encoded protein program, the unfolding DNA switches hats to its RNA counterpart, wherein uridine replaces thymine as CUG in the triple-base codon.

And that's where myotonic dystrophy's pathological troubles begin: In DM, those proliferating CUG triplets pile up and get stuck in the cells' nuclei.

"Again, like Huntington's," Cooper explained, "the longer the stuttering RNA triplet expands in an individual, the worse the disease gets. And they elongate faster in skeletal muscles than in other tissues."

Cooper is senior author of a paper in the current issue of Science, dated May 1, 1998. Its title: "Disruption of splicing regulated by a CUG-binding protein in myotonic dystrophy."

He and his co-authors studied the muscle cells of two DM patients, a newborn girl and a 30-something man. Both patients presented with classic DM symptoms. The man displayed the myotonia and progressive muscle weakness; the child manifested the disease as a "floppy infant."

A Tale Of Two Patients

"The six-day-old baby," Cooper told BioWorld Today, "who died with congenital DM, "had more than 1,600 triplet repeats in the RNA of one allele [parental variant gene], and only 12 in the other. This is a very long repetitive sequence, which is why she was born with the congenital heterozygous form of the disease.

"The man," Cooper recounted, "was homozygous. That is, he was born unlucky enough to have both of his alleles containing the CTG repeats, with several hundred in each."

Cooper's laboratory at Baylor focuses on the pre-RNA splicing of exons, the segmented coding sequences of genes. (See BioWorld Today, April 29, 1998, p. 1.)

"My co-author, Lubov Timchenko," Cooper recalled, "had purified the protein that's bound to CUG RNA. We found that this CUG-binding protein (CUG-BP) is important for regulating expression of other genes. And the one in particular turned out to be the cardiac treponin gene, which we've been working on for many years in chickens. CUG-BP proved to be involved in regulating splicing of the cardiac treponin gene.

"So, when we looked at the expression of cardiac treponin RNA in these two patients, it was abnormal," Cooper said. "So was the location of their CUG-BP, which accumulated in the nucleus. This enabled us to make the connection between accumulation of this abnormal RNA in the nucleus and abnormal splicing of cardiac treponin."

Treponin, Cooper explained, "is involved in signaling cardiac contraction. It's part of the apparatus that senses changes in the calcium concentration in the cell, which is the sign for cardiac contraction. DM patients do have dilated cardiomyopathy, probably some kind of contraction defect."

The Baylor team then looked at how the RNA from the cardiac treponin gene was spliced in these two patients.

"What we found," Cooper recalled, "was that compared to other individuals who had other cardiac abnormalities, their splicing was quite abnormal, and not only in their heart tissue. We took skeletal muscle cells from the older patient, and put in the gene expressing our version of the cardiac treponin RNA. Sure enough, it was processed abnormally."

DM Gene Defect Suspect In Other Ills, Too

This result, he observed, "is the first example of a human disease caused by an accumulation of RNA. The defective allele expressed an RNA that screwed up the expression of a number of other genes, by altering the function of CUG-BP."

Moreover, the co-authors have found that skeletal muscle isn't the only tissue with genes that contain a binding site for CUG-BP. "So we suspect," Cooper said, "that the reason DM affects so many different organs is that CUG-BP is involved in regulating RNA processing in a variety of bodily tissues."

Looking ahead, he said, researchers want to determine "first in cells, then in transgenic mice, what other such genes are affected by this abnormal CUG-BP.

"This is a new and novel mechanism for human disease," Cooper pointed out. "It's something that people need to keep their minds open to, in terms of other diseases. It could be, as in Huntington's disease, that the triplet expansion in the RNA, not just the mutant protein, actually is contributing to the pathology. So there could be a role for an RNA component in other entities.

"A second thing we are looking into is the importance of RNA processing as a possible way to control genes used for gene therapy. However," he concluded, "I think that that will he the next step, not this step." *