The SNORD116-NHLH2 pathway: insights into the molecular genetic basis of Prader-Willi Syndrome

Prader-Willi Syndrome (PWS) is a genetic condition resulting from paternal inheritance of a deletion within an imprinted region of chromosome 15q. The smallest known deleted region encompasses a small nucleolar non-coding RNA locus called SNORD116 (SNORD116), but very little is known about how deletion of SNORD116 leads to PWS. As shown using pluripotent stem cells from PWS patients and control individuals, expression of the NHLH2 and PCSK1 genes are significantly reduced in PWS patients. NHLH2 is a transcription factor that transcriptionally-regulates the PCSK1 gene, while PSCK1 acts as a protease that cleaves and activates neuropeptides. In mouse models, deletion of NHLH2 and PSCK1 leads to obesity and other conditions that overlap with phenotypes of PWS. Human PSCK1 deletion or variants that inactivate PSCK1 protein lead to in human obesity. The identification of NHLH2 and PSCK1 as potential SNORD116@ regulatory targets allows us to begin to address a major unanswered question for PWS patients and their families: How does deletion of this region of chromosome 15q lead to PWS? What is known is that SNORD116 is a C/D box small RNA that potentially acts as an RNA guide for proteins that methylate RNA. Methylation difference could change the stability of the RNA, or could lead to differential RNA splicing of target mRNAs. We proposed that if SNORD116 RNAs serve as guides for methyltransferases, loss of SNORD116 from the deleted region in patients could explain differences in NHLH2 and PSCK1 expression, and possibly the phenotype of PWS patients. Dr. Good’s laboratory has worked on the NHLH2 gene for 20 years, and has many tools that will allow her laboratory to study how both NHLH2 and PCSK1 mRNAs are differentially expressed in PWS, as compared to control neurons. Her lab will use pluripotent stem cells from PWS patients and controls, as well as the PWS mouse mode, and mouse hypothalamic neurons in cell- and mouse-based studies to identify the molecular genetic mechanism underlying the SNORD116--NHLH2 and PSCK1 interacting pathway. Understanding the basic molecular genetic biology of the SNORD116 locus and its targets will lay the foundation for identifying future clinical targets or therapeutic interventions for PWS patients and their families.

Research Outcomes: Public Summary

Although the smallest human causative deletions for PWS only encompass SNORD116@ (a group of 28 small nucleolar or snoRNAs), SNORD109a, and IPW genes1, there is still a lack of understanding about how deletions in SNORD116@ lead to the complex phenotypes of PWS. Phylogenetic analysis of the SNORD116@ locus in a number of research models (humans, mice, rats, rabbits and primates) concluded that strong homology exists throughout these models; however, there are different numbers of transcripts in each organism, and differences in the “groupings” of these snoRNA transcripts into homology families. In late 2016, Burnett and colleagues proposed that the normal SNORD116@ locus may confer positive regulation on NHLH2 and PCSK1 mRNAs, in that stem cells from PWS patients lacking SNORD116@, and Snord116m+/p- mice expressed low levels of both transcripts. Based on that information, and our 20+-year history in working on the NHLH2 transcription factor, a new FPWR grant was awarded to our laboratory in late November 2017. However, since that time, two other labs were unable to show Nhlh2/NHLH2 and/or Pcsk1/PCSK1 downregulation in Snord116m+/p- mouse and humans, respectively. Our work using cell lines under regular culture conditions, and the Snord116m+/p- mouse also has not yet been able to replicate the initial Burnett et al., 2016 findings. The question remains as to whether a SNORD116: NHLH2 regulatory relationship exists. However, additional support for a SNORD116: NHLH2 molecular relationship lies in the phenotypic similarities between the PWS model mouse, Snord116m+/p- mouse, and the Nhlh2 KO (N2KO) mouse. New data from our laboratory demonstrate additional physical activity/exercise phenotypes in the Snord116m+/p- mice, and in mice with a deletion of both alleles of Snord116. Our laboratory is one of the few with the tools, reagents and expertise to attack the NHLH2:SNORD116 question. The answers we seek will not only help to guide the PWS field on the function of SNORD116, but also contribute to the field of non-coding RNA biology and obesity/metabolism research, possibly leading to new treatment options for many individuals.

Research Outcomes: Publications

Phylogenetic Analysis of the SNORD116. Good, DJ and Kocher, MA.  LocusGenes 2017, 8(12), 358.