CRISPR Unlocks ‘Sleeping’ Genes—A Potential Cure for Prader-Willi Syndrome?

By Dr. Chinta SidharthanReviewed by Lily Ramsey, LLMFeb 20 2025

A single genetic switch could potentially treat a rare but severe childhood disorder. Prader-Willi Syndrome (PWS) affects approximately 1 in 15,000 newborns, causing insatiable hunger, obesity, and developmental challenges.

This condition stems from the loss of gene activity on a specific section of paternally inherited chromosome 15.

In a recent study published in Cell Genomics, a research team used clustered regularly interspaced short palindromic repeats (CRISPR)-based epigenome editing to reactivate the silenced maternal copy in human stem cells.

This approach offered hope for a lasting therapy to restore essential gene function in individuals with PWS.

​​​​​​​Study: Activation of the imprinted Prader-Willi syndrome locus by CRISPR-based epigenome editing. Image Credit: Prostock-studio/Shutterstock.com

Background

The expression of genes can be switched on or off by chemical modifications to deoxyribonucleic acid (DNA), known as epigenetic changes. This process is crucial for healthy development but can sometimes go wrong.

Imprinting is one such mechanism where genes are expressed from only one parent’s chromosome copy, such as the silencing of maternal chromosomes through imprinting.

In Prader-Willi Syndrome (PWS), genes on a region of chromosome 15 inherited from the father undergo a loss of function, and the mother's version of chromosome 15 remains silenced, leading to problems such as extreme hunger, obesity, and developmental delays.

Scientists have long sought ways to turn on the maternal copy, but traditional methods, including drugs targeting epigenetic enzymes, carry the risk of non-target effects.

However, recent advances in CRISPR-based technologies have allowed researchers to alter gene regulation, offering a more targeted solution precisely.

The Current Study

Here, the researchers used CRISPR-based epigenome editing in human induced pluripotent stem cells (iPSCs) to explore the activation of silenced maternal genes associated with PWS.

They focused on the small nuclear ribonucleoprotein polypeptide N (SNRPN) gene as a marker for the broader PWS region, which is normally active only from the paternal chromosome.

To investigate the regulatory regions controlling gene activation, the team inserted a green fluorescent protein marker into the SNRPN gene, creating cell lines in which the maternal or paternal allele was tagged.

Two complementary screening approaches were employed. First, a CRISPR interference (CRISPRi) system, using a deactivated CRISPR-associated protein 9 (Cas9) protein fused with a repressor, was applied to the paternal allele to map regions needed for gene expression.

Second, a CRISPR activation (CRISPRa) system involving Cas9 fused to an activator was directed at the maternal allele to identify areas capable of turning on the silent gene.

Following these screens, the researchers tested a third tool — a CRISPR-based demethylase (Tet1-dCas9). This enzyme removes methylation, a chemical mark that silences the maternal allele. They targeted the PWS imprinting center (PWS-IC), which is a regulatory DNA region controlling gene activity.

The Tet1 tool was delivered transiently, meaning it was expressed only briefly in the cells. Subsequently, the team monitored gene activity and DNA methylation levels for weeks while the cells matured into neurons.

The team also compared different versions of Tet1 constructs to improve efficiency. Ribonucleic acid (RNA) sequencing, chromatin accessibility assays, and bisulfite sequencing were performed to confirm that the genetic reactivation was specific and stable.

Major Outcomes

The study found that CRISPR-based epigenome editing can activate the normally silent maternal genes on chromosome 15 associated with PWS. The researchers observed that different CRISPR tools achieved this through distinct mechanisms.

The results showed that the maternal SNRPN gene could be turned on using CRISPRa by targeting upstream regulatory regions. This approach boosted gene expression, but only to partial levels compared to the active paternal copy. Furthermore, the activation was not uniform across all cells, and the effects appeared temporary.

In contrast, the most significant breakthrough came from the use of the Tet1 demethylase, which removed DNA methylation marks from the maternal PWS-IC. This resulted in stable and heritable activation of maternal genes, including SNRPN and important downstream noncoding RNAs.

Moreover, the gene expression levels in some cells matched those from the paternal chromosome, and the activation persisted for weeks, even as the cells differentiated into neurons.

Furthermore, maternal activation extended to additional genes in the region, such as melanoma antigen family L2 (MAGEL2) and Necdin or NDN, which are critical for neural function. These results indicated that DNA demethylation can restore long-term gene activity across the entire PWS locus.

However, the researchers noted some limitations. Reactivation efficiency varied among cells, and gene expression levels were not always identical to natural paternal activation.

Additionally, while off-target effects were minimal, they believe that further research is needed to ensure safety in therapeutic applications.

Conclusions

Overall, the findings demonstrated that CRISPR-based epigenome editing, particularly DNA demethylation using Tet1, can reactivate silenced maternal genes in PWS cells.

The activation of the genes in the PWS locus on maternal chromosome 15 was stable and persisted through neuronal development.

While challenges such as efficiency and delivery methods need to be addressed, this approach represents a promising step toward a lasting therapy for PWS. More importantly, targeted epigenetic editing could offer new hope for patients with imprinting disorders.