Howard Hughes Medical Institute (HHMI) scientists have discovered how the most common genetic defect in amyotrophic lateral sclerosis kills nerve cells. Their study suggests that the pores that allow molecules into and out of a cell's nucleus get jammed, a finding that could speed the search for other genes that promote this fatal illness.

In people who have amyotrophic lateral sclerosis (ALS), the motor neurons that operate the muscles deteriorate. Over time, the disease deprives patients of the ability to walk, swallow, and breathe, and they usually die within three to five years.

About 10 percent of ALS cases are hereditary, and researchers have pinpointed about 20 defective genes that can cause the disease. The most common gene variant by far is C9ORF72. The faulty version of this gene contains too many copies of a short DNA segment, known as a repeat. Healthy people usually have 23 or fewer copies of this segment, whereas people who develop ALS can carry hundreds or thousands of duplicates. Extra repeats in the gene also cause frontotemporal dementia (FTD), a disease closely related to ALS.

Researchers haven't discovered how the faulty version C9ORF72 induces nerve cell deterioration. "We want to know what is the primary biological defect that is leading to ALS and FTD," says HHMI Investigator J. Paul Taylor of St. Jude Children's Research Hospital, who led the new study with Fen-Biao Gao of the University of Massachusetts Medical School. The research was published August 26, 2015 in the journal Nature.

To gauge the effects of the repeats, Taylor and Gao's teams turned to fruit flies, which researchers often use to study neurodegenerative diseases such as ALS and FTD. The scientists inserted different numbers of repeats into the insects' chromosomes. When the flies received eight copies of the repeat, they appeared normal. That made sense, Taylor says, since people with that number of repeats don't develop ALS or FTD. But when the flies carried 58 copies of the DNA repeat, they showed signs that cells were damaged and dying. For example, in one experiment the researchers inserted the repeats only into the animals' eyes. In these insects, the normally smooth surface of the eye was ragged, a condition known as rough eye that indicates nerve cells have perished.

Those results showed that the extra repeats poisoned cells in the flies, but they didn't explain how. To try to answer that question, Taylor and colleagues performed a genetic screen. They bred flies that carried 58 repeats in their eyes with many different strains of flies that were missing certain genes. The offspring of these crosses lacked the same genes as their parents, allowing the researchers to determine if a gene's absence affected the insects' eyes. "We looked for something that made the original phenotype better or worse," says Taylor. If the flies had rougher eyes when one of the genes was missing, for example, that gene likely helps prevent the death of neurons. In total, the researchers evaluated the roles of 9,000 different fly genes.

Eighteen of the genes that Taylor and colleagues identified through this procedure were involved in the same cellular function: ferrying molecules into and out of the nucleus through nuclear pores. A membrane encloses the cell's nucleus, and nuclear pores are structures in the membrane that allow certain molecules to pass through. Transportation through the pores is crucial for a cell's survival.

One type of molecule that departs the nucleus through nuclear pores is RNA, which carries the instructions for making proteins and performs other functions outside of the nucleus. The researchers asked whether RNA passed through the pores normally in fly cells with eight or 58 repeats. They discovered that RNA failed to exit appropriately and built up in the nuclei of cells with 58 repeats, suggesting that the nuclear pores weren't working properly in these cells.

The team also measured RNA levels in neurons derived from five people with ALS or FTD who have faulty versions of C9ORF72. Compared with cells from four healthy people, neurons from the patients accumulated about 35 percent more RNA inside the nucleus than outside of it. That finding shows that fly and human cells share the same abnormality, Taylor says.

"This study does not reveal a new drug target, but it does reveal the underlying molecular defect" behind many inherited cases of ALS and FTD, he says. A companion paper in the same issue of Nature supports the conclusion that the extra repeats in C9ORF72 disrupt nuclear pores, Taylor says.

Researchers describe about 90 percent of ALS cases as sporadic because they haven't identified which genes are responsible for the illness. "This finding empowers the search for the genetic causes of sporadic ALS," says Taylor. Now, researchers know what to look for: genes that alter transportation through nuclear pores.