CRISPR-Assisted Study Works Out Mechanistic Details of ALS
Our Genetic Superweapon has now helped glean insights into the genetic underpinnings of amyotrophic lateral sclerosis, a neurodegenerative disease that’s notoriously tricky to parse.
Like many neurodegenerative diseases, the inner workings of amyotrophic lateral sclerosis (ALS, also known as Lou Gehrig’s disease) are largely cryptic. ALS degrades neurons that operate voluntary muscle movements — patients who suffer from the disease become immobile and unable to talk, or even breathe, on their own. Scientists know that in the brains of these patients large clumps of anomalous proteins build up around healthy neurons, and as a result, the cells die off. What’s less clear, however, is how or why this happens.
The new Stanford study is a step towards demystifying this disease and could even help lay the groundwork for new therapeutic targets.
Accounting for nearly 40 percent of inherited cases of ALS and 25 percent of inherited FTD cases, disease-causing mutations in C9orf72 insert extra sequences of DNA, called hexanucleotide repeats, into the gene. These repeats produce potentially toxic RNA and protein molecules that kill neurons resulting in problems with movement and eventually paralysis for ALS patients and language and decision-making problems for FTD patients.
The
team used a genomewide screening approach that uses CRISPR-Cas9 to alter the function of every single human gene simultaneously. They used the technology to produce ‘gene knockouts’ that target genes with a kind of molecular scissors that makes accurate cuts, rendering the genes unable to function normally.These gene knockouts were found to be helpful in identifying genes that either increase or prevent toxicity, meaning they can be used in the detection of specific genes involved in protecting neurons from degeneration.
The results confirmed a set of genes as strong protecters against the toxic proteins, but there was a particular one – Tmx2 – that caught the researchers’ attention: They saw that after knocking-out Tmx2 of the mouse neurons cells in vitro, the neurons survived almost 100 percent of the time, while normally only 10 percent survive.
“We could imagine that Tmx2 might make [a] good drug target candidate,” said co-author Michael Haney in a press release. “If you have a small molecule that could somehow impede the function of Tmx2, there might be a therapeutic window there.”
The Tmx2 protein resides in an intracellular organelle called the endoplasmic reticulum, but its function is still a mystery. However, some studies suggest it may work with other genes to trigger cells’ death.
“We’re still in early phases, but I think figuring out exactly what Tmx2 normally does in a cell is a good place to start — that would hint at what functions are disturbed when these toxic species kill the cell, and it could point to what pathways we should look into,” said Nicholas Kramer, another first author of the study.
Previously such studies needed a few months to find candidate genes and could only be performed on yeast, worm, and fly genomes. With CRISPR, the researchers in this study needed just about two weeks to conduct a complete search of the human genome. The results suggest that this faster and more comprehensive approach may be used to rapidly identify genes that may be involved in other neurological disorders.
