November 01, 2012
What do Lou Gehrig, Stephen Hawking, David Niven and Mao Zedong have in common? They all suffered (or in Hawking’s case, continue to suffer) terribly from a disease called amyotrophic lateral sclerosis or ALS. And now the humble yeast S. cerevisiae may help scientists find new treatments so that others do not need to suffer similarly.
Patients with ALS gradually lose use of their motor neurons and generally die within 3-5 years of diagnosis. While there are some rare forms that run in families, most are sporadic. There is no history of the disease in the family and then suddenly, it just appears.
The causes of ALS have remained a mystery for many years but recent work has suggested that RNA binding proteins and RNA processing pathways are somehow involved. In particular, an RNA-binding protein called TDP-43 appears to be a key player. Mutations in its gene are associated with ALS, and aggregates of the protein are found in damaged neurons of ALS patients. Unfortunately, since this protein is needed for cell survival it is not an easy target for therapies. This is where yeast can help.
Scientists have managed to mimic the effects of TDP-43 in yeast. When this protein is overexpressed, the yeast cells die just like the motor cell neurons do. In a recent Nature Genetics paper, Armakola and coworkers use this model system for finding better therapeutic targets. And it looks like they may have succeeded.
These authors used two different screens to systematically look for proteins that when deleted or expressed at lower levels rescued yeast overexpressing TDP-43. They found plenty. One screen yielded eight suppressors while the other yielded 2,056 potential suppressors. They decided to focus on one of the stronger suppressors, DBR1.
The first thing they wanted to do was to make sure this wasn’t a yeast specific effect. If lowering the amount of DBR1 has no effect in mammalian models, it is obviously not worth pursuing!
To answer this question, they created a mammalian neuroblastoma cell line with an inducible system for making a mutant version of TDP-43, TDP-43 Gln331Lys, found commonly in ALS patients. As expected, these cells quickly died in the presence of inducer. They could be rescued, though, when DBR1 activity was inhibited with siRNA. The authors confirmed that decreasing the activity of DBR1 in primary neurons decreased TDP-43 toxicity as well.
So decreasing the amount of DBR1 appears to rescue cells that die from the effects of mutant TDP-43. This suggests that targeting DBR1 may be useful as a therapy for ALS. But this study doesn’t stop there. It also tells us a bit about how lowering DBR1 levels might be rescuing the cells.
DBR1 is an RNA processing enzyme involved in cleaning up the mess left behind by splicing. It cleaves the 2’-5’ phosphodiester bond of the spliced-out intron (called a lariat). Previous studies in yeast have shown that when Dbr1p levels are reduced or its catalytic activity is disrupted by a mutation, there is a build up of these lariats. This study showed directly that the accumulated lariats interact with TDP-43 in the cytoplasm to suppress its toxicity. So in ALS, the accumulated lariats may serve as a decoy for the mutant TDP-43 protein, preventing it from binding to and interfering with more essential RNAs.
This last result may also suggest another potential therapy. If scientists can find other ways to increase the amount of decoy RNA, then they may not need to depend on reducing levels of DBR1. There may be many possible approaches to soaking up rogue TDP-43.
by D. Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics