Subtle Changes in MicroRNA-218 May Fuel ALS
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Dropping the levels of microRNA-218 (miR-218) — a small molecule that regulates the activity of other genes — below a certain threshold leads to severe motor neuron damage, paralysis, and death in a mouse model of amyotrophic lateral sclerosis (ALS), a study shows.
The findings, combined with the fact that ALS patients usually show lower-than-normal miR-218 levels, highlight how subtle changes in this particular microRNA, instead of an all-or-nothing model, may contribute to ALS.
Gene activity regulation “is increasingly becoming a new and very interesting direction for ALS research,” Samuel Pfaff, PhD, the study’s senior author and a professor at the Salk Institute in California, said in a Salk press release.
“Our study is highly revealing in terms of how gene regulation occurs within neurons,” Pfaff said, adding that “while our experiments were done in mice, we believe these findings will also apply to humans.”
The study, “A hidden threshold in motor neuron gene networks revealed by modulation of miR-218 dose,” was published in the journal Neuron.
MicroRNAs, or miRNAs, are small molecules of RNA that target a particular gene’s messenger RNA (mRNA) — the intermediate molecule derived from DNA and used as template for protein production — to prevent the generation of that protein.
A single miRNA can regulate several mRNAs, and a single mRNA can be regulated by multiple miRNAs. miRNAs are known to play important roles in many important cellular processes, including maintaining nerve cell survival and function.
Notably, many genes known to be associated with ALS, such as TARDBP and FUS, are involved in RNA metabolism, including miRNA processing.
While disruption of miRNAs balance is known to cause neurologic damage, the effects of different miRNA levels, rather than their presence or absolute absence, remain unknown.
In the current study, Pfaff and colleagues first systematically reviewed published studies that measured miRNA levels in people with ALS. They found that a particular miRNA called miR-218 was found at lower-than-normal levels, but was not completely lost, in ALS patients across all studies.
miR-218 is highly present in motor neurons, and was shown previously to regulate these cells’ fate and activity.
The researchers then developed a strategy to finely lower miR-218 levels in a controlled way to evaluate how different levels affected neuromuscular junctions (NMJs) in a mouse model. NMJ is the site of near contact between a motor neuron and a muscle cell and where they release signaling molecules to communicate with each other.
This strategy involved the deletion and/or genetic modification of one or both copies of the miR-218-encoding gene to achieve different levels of the RNA molecule — with the presence of two healthy gene copies representing 100% of levels, and the deletion of both copies representing 0%. Deletion or modification of each gene copy resulted in intermediate levels.
The team found that miR-218 levels above 36% were associated with normal and healthy NMJs, while levels below 7% resulted in lethal neuromuscular deficits, highlighting the presence of a critical threshold somewhere between these levels.
Once miR-218 levels dropped below 36%, the way motor neurons signaled muscles dropped dramatically.
Further analysis using cutting-edge tools showed that miR-218 regulates the activity of about 300 different genes. Many of these genes provided instructions to produce proteins involved in the growth of motor neuron fibers, essential for their communication, and in neuromuscular communication.
Notably, miR-218 was found to regulate these genes in different ways, as modulation of its levels showed distinct dose-response curves of its target mRNAs.
“Instead of acting like a simple switch, the molecule miR-218 is like an orchestra conductor of 300 musicians playing together,” said Neal Amin, MD, PhD, the study’s first author who conducted his doctoral research at Pfaff’s lab and is now a clinical scholar and postdoctoral researcher at Stanford University.
“Instead of gradually telling all of the players to dim the volume of their instruments in unison, it’s telling some musicians to play more quietly and others to stop completely,” Amin said, adding that miR-218 “has a much more dynamic and complex control over gene function than we ever previously appreciated.”
The researchers believe their findings, as well as their method to fine-tune levels of relevant molecules in animal models, may help better understand how genetic mutations that reduce a gene’s activity put patients at risk for developing ALS.
This eventually could lead to new treatments that target the biological mechanisms that drive the disease.
These findings also may have implications in other neurological diseases, such as schizophrenia, which has been associated with changes in miRNA levels.
“We think that these processes may also take place in other diseases related to genes and aging, including cancer,” said Pfaff. “Having a new way to create animal models of how genetic disease begins and how it progresses will allow us to get at the underlying mechanisms and a deeper understanding of these complex activities.”
The study was funded by the National Institutes of Health, TargetALS, and the U.S. Department of Defense.
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