Repeats in Fragile X Gene Work to Control Protein Production via RNA, Study Finds

Repeats in the FMR1 gene — which cause fragile X syndrome — help to regulate the normal production of the FMRP protein in nerve cells, a new study shows.

This finding has implications for understanding the biology of repeats in the genome, and could pave the way for new therapies for fragile X, the researchers said.

Their study, “A native function for RAN translation and CGG repeats in regulating fragile X protein synthesis,” was published in Nature Neuroscience.

Many areas within the human genome contain repeats of a sequence of nucleotides — the building blocks of DNA. One of the best characterized is in the FMR1 gene, which codes for the protein FMRP.

Typically, this gene has around 30 repeats of CGG (C stands for cytosine and G for guanine). But in people with fragile X, there are generally over 200 repeats, leading to epigenetic silencing of the gene — genetic changes that “turn off” gene expression, resulting in the lack of FMRP.

Although the disease-associated aspects of these repeats are well-characterized, little is known about them in a non-disease setting. Indeed, the biological function of genetic repeats is not well-understood. This poses an obstacle both for overall scientific understanding and for treatment research in certain diseases.

“To develop a new treatment strategy, we really needed to understand the native biology of how these repeats work and why they are there in the first place,” Peter Todd, MD, PhD, the study’s senior author and a professor at the University of Michigan, said in a press release.

His team studied cells to better understand the CGG repeat in FMR1 in a non-disease setting. The researchers first noted that these repeats are generally conserved among humans’ closest biological relatives — primates and other mammals — which supports the idea that they are important biologically.

They then examined the effect of having different numbers of repeats in FMR1. They found that a higher number of repeats correlated with less FMRP being made.

Further experiments revealed that the regulatory effect of repeats was due to their effect on RNA. Specifically, CGG repeats make RNA molecules produced from the FMR1 gene take on a particular 3D structure that is important to regulate protein generation.

In nerve cells, FMRP is produced in response to the activation of receptors of the neurotransmitter glutamate. (Neurotransmitters are key molecules in nerve cell communication.)

The researchers found that, after these receptors activate, less CGG repeats are converted into RNA and more FMRP protein is produced.

“These findings provide initial evidence for physiological roles of repetitive elements in both translational control [protein production] and neurobiology,” the researchers wrote.

This understanding also suggests that blocking the repeats at the RNA level could increase FMRP production, suggesting “a potential target for therapeutic development in fragile X disorders.”

To test this idea, researchers used antisense oligonucleotides (ASOs) — short strands of DNA that can bind to specific sequences of RNA. ASOs have previously been shown to effectively treat other diseases; for example, Spinraza (nusinersen, by Biogen), a treatment for spinal muscular atrophy, contains an ASO that increases the production of the protein SMN through RNA-based mechanisms.

In this case, the researchers created an ASO to block CGG repeats in RNA from FMR1 . This work was done in collaboration with Ionis Pharmaceuticals, a company that specializes in ASOs. Two study researchers are Ionis employees.

Their ASO increased FMRP production in nerve cells. In addition, it also significantly increased cell survival.

Importantly, besides cells from mice, these effects were seen in neurons derived from human stem cells — both cells with a normal numbers of repeats in FRM1 and cells with over 200 repeats as in fragile X.

“Approaches such as the ASO described [in] the present study could thus be useful in combination with [repeat] reactivation strategies to prevent CGG repeat-associated toxicity … and enhance overall FMRP production,” the scientists wrote.

Much more research will be required before these findings might be translated into actual treatments.

“The study was done in dishes, and so there is still a long way before it can be tried in patients, but advancing our understanding of normal nerve cell biology is a crucial step to find cure,” Todd said.