Genomewide identification of mRNA sites of 2’-O methylation targeted by SNORD116 snoRNAs

Funding Summary

While we know the loss of SNORD116 (a gene that encodes many snoRNA molecules on Chromosome 15) leads to characteristics of PWS, we do not know how this exactly works. We need to understand how SNORD116 functions normally in order to understand why the loss of this region leads to PWS. It is likely that the snoRNAs in the SNORD116 region modify other RNAs, but the exact target genes are not yet known. Dr. Carmichael is an expert in RNA who has developed a new technique to detect and sequence the genes that these snoRNAs are modifying.

This study will help understand what is causing PWS on a molecular level by helping us understand how these critical PWS-region genes normally function, and in the future can help develop therapies that target the effects of the loss of SNORD116.

Dr. Theresa Strong, Director of Research Programs, shares details on this project in this short video clip. 

Watch the full webinar describing the 8 research projects funded in this grant cycle here


Lay Abstract

Prader-Willi syndrome results from deletion of a specific region of the paternal chromosome 15 in humans. This region expresses 29 similar RNA molecules called snoRNAs, which are all relatively short and lack the potential to encode any proteins.  snoRNAs of this type are generally thought to recognize and associate with target RNAs by base complementarity and direct the modification of specific residues by methylating the 2’-hydroxyls of ribose rings. Most cellular snoRNAs target ribosomal RNAs, but those from the Prader-Willi critical region (SNORD116s) do not and their targets are unknown. 

We have recently developed a powerful new method to identify genomewide sites of 2’-O methylation and propose to use this method to identify, for the first time, the actual targets of the SNORD116 snoRNAs.  For this work we have constructed an isogenic pair of stem cells, which differ only in whether they do or do not express RNAs from the PWS critical region. These will be compared by next generation sequencing methods for RNA expression profiles and also for sites of RNA 2’-O methylation.  The cells will also be differentiated into neurons and similar comparisons will be made. Together, these studies should offer us an unprecedented window into the molecular underpinnings of PWS.  

Research Outcomes: Public Summary

Using CRISPR technology, we have generated a PWS model in H9 hESCs and differentiated the line to generate mature neurons. RNA-Seq was then conducted on the H9 and H9-PWS isogenic neuron pair to determine the transcriptomic phenotype. Results were very interesting and provocative. Downregulated genes were mostly involved in neuronal development while upregulated genes were involved in protein synthesis, DNA synthesis, and RNA catabolism. Analysis of alternative splicing revealed that intron retention is the most pronounced event with many H9-PWS transcripts possessing reduced intron retention than H9 transcripts. Dynamic reduction in intron retention has been connected in the literature to neuronal activation, so it is of interest to determine whether this response may be impaired in PWS. Further, a number of genes possibly related to PWS phenotypes showed dramatic differences in RNA expression or processing. The most highly upregulated gene in H9-PWS neurons is TTR (transthyretin), a blood and cerebrospinal fluid transporter of thyroxine and retinol that may play a role in modulating food intake and energy balance. Elevated hypothalamic expression of TTR has been reported to lead to decreased food intake and body weight, possibly related to early failure to thrive in PWS. TRPM3 (transient receptor potential cation channel) is involved in pain sensing and the same gene produces the microRNA miR-204, which is involved in glucose homeostasis and obesity. In H9-PWS neurons, this gene expresses primarily an inactive truncated isoform of TRPM3 but overexpresses miR-204. NTRK2/TrkB is the BDNF (brain-derived neurotrophic factor) receptor, having mutations associated with obesity and mood disorders. In H9-PWS neurons, this gene expresses primarily a truncated isoform lacking its intracellular kinase signaling domain. Finally, lowered expression of GNRH1 (gonadotropin releasing hormone) is associated with hypogonadism. In H9-PWS neurons, this gene expresses an mRNA isoform that is poorly translated.

Research Outcomes: Publications

Carmichael GG. (2020) Unravelling the Biology of snoRNAs Implicated in Prader-Willi syndrome.

Chung MS, Langouët M, Chamberlain SJ, Carmichael GG. (2020) Prader-Willi syndrome: reflections on seminal studies and future therapies. Open Biol. 10: 200195.

Funded Year:


Awarded to:

Gordon Carmichael, Ph.D




University of Connecticut


Gordon Carmichael, Ph.D

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