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Packard Center Research Expands Search for ALS Genes

RNA-binding proteins show strong links to ALS

A new study, led by Packard Center scientist Aaron Gitler, has identified a strategy that uses yeast to help identify new genes that may be responsible for amyotrophic lateral sclerosis (ALS). Gitler, a cell biologist at the University of Pennsylvania, and his colleagues show that a class of genes that bind to RNA may be responsible for more cases of ALS than previously thought.

In recent years, scientists have begun to identify some of the genes associated with ALS, including SOD1, C9ORF72, TDP-43, FUS, VCP, and ATXN2. Even with this recent flurry of genetic discovery, the genetic cause of many ALS cases remains unknown. Researchers and patients need better, more efficient ways to find the genes that cause ALS. The similar functions of the proteins coded by TDP-43 and FUS gave Gitler and colleagues a hint. Both of these proteins bind to RNA, and numerous studies have shown that problems with RNA metabolism are common in ALS patients.

“The evidence supporting defects in RNA metabolism in ALS is really compelling,” Gitler said.

Gitler and his colleagues have been using the simple yeast model system to study basic properties of the TDP-43 and FUS proteins. Like in human ALS patient neurons, they found that TDP-43 and FUS also formed clumps in the cytoplasm of yeast cells. “We reasoned that if TDP-43 and FUS, both with proven links to ALS, form clumps in yeast cells, we might be able to find other human RNA-binding proteins that do the same thing, and then these might also be associated with ALS,” explained Gitler. To test this idea, Gitler’s group used yeast to probe 133 human proteins that can bind to RNA. Thirty eight of these proteins (including TDP-43 and FUS) formed toxic clumps in the cytoplasm of the yeast. The researchers selected one protein that formed clumps most easily and showed other similarities to TDP-43 and FUS to test further. The researchers sequenced this protein, known as TAF15, and looked for TAF15 mutations in ALS patient blood samples from the US, Australia, and Sweden. Overall, the researchers identified six patients with sporadic ALS who had mutations in TAF15.

The lab of James Shorter, PhD, assistant professor of Biochemistry and Biophysics at the University of Pennsylvania, and Packard Center investigator, provided more clues about TAF15. In an aggregation assay using purified TAF15 protein, they found that mutant versions of TAF15 clumped faster. The purified TAF15 protein also harbored a prion-like portion in its amino acid sequence, much like TDP-43 and FUS, which might be a future predictor of neurodegenerative qualities in mutant proteins.

To see how these TAF15 genetic variants behaved in other organisms, Gitler and colleagues expressed the variants in a cell culture of rat embryo spinal cord neurons. The ALS-associated variants clumped together in the cytoplasm of the neurons in a manner similar to FUS and TDP-43. When the scientists inserted the ALS-linked TAF15 gene into the fruit fly Drosophila melanogaster, the TAF15 proteins caused damage to the eyes and motor neurons and shortened the fly’s lifespan, with the ALS-linked mutant forms of TAF15 having a more severe effect.

 “When it comes to RNA-binding proteins and ALS, TDP-43 and FUS might just be the tip of the iceberg. Many new additional RNA-binding proteins could also contribute to the disease, with some having stronger and some having weaker effects. Hopefully, the list of genes we’ve generated from the simple yeast screen could be a kind of guide or a resource for the ALS community to focus on,” Gitler said.

These findings open up a class of proteins that may lead to neurodegeneration and ALS. A better understanding of the genetic causes of ALS may one day lead to better detection and therapeutics, researchers say.

Read Aaron Gitler's article in PNAS

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