Non-coding RNAs associated with Prader–Willi syndrome regulate transcription of neurodevelopmental genes in human induced pluripotent stem cells

Abstract

Mutations and aberrant gene expression during cellular differentiation lead to neurodevelopmental disorders, such as Prader-Willi syndrome (PWS) which results from the deletion of an imprinted locus on paternally inherited chromosome 15. We analysed chromatin-associated RNA in human induced pluripotent cells (iPSCs) upon depletion of hybrid small nucleolar long non-coding RNAs (sno-lncRNAs) and 5’ snoRNA capped and polyadenylated long non-coding RNAs (SPA-lncRNAs) transcribed from the locus deleted in PWS. We found that rapid ablation of these lncRNAs affects transcription of specific gene classes. Downregulated genes contribute to neurodevelopment and neuronal maintenance while genes that are upregulated are predominantly involved in the negative regulation of cellular metabolism and apoptotic processes. Our data reveal the importance of SPA-lncRNAs and sno-lncRNAs in controlling gene expression in iPSCs and provide a platform for synthetic experimental approaches in PWS studies. We conclude that ncRNAs transcribed from the PWS locus are critical regulators of a transcriptional signature, which is important for neuronal differentiation and development.

Roles of SNORD115 and SNORD116 ncRNA clusters in neuronal differentiation

“…This indicated that transcription and splicing of the SNHG14 primary transcript is essentially unaltered during differentiation. We detected previously reported sno-lncRNAs, long ncRNAs containing two SNORD116 snoRNAs joined by a linker (Sledziowska et al ., 2023; Wu et al ., 2016; Yin et al ., 2012). During differentiation, accumulation of sno-lncRNA 1-3 decreased, sno-lncRNA 4 was unchanged, while sno-lncRNA 5 was not detected.…”

“…This confirmed that transcripts around SNORD115 markedly increased during differentiation, while those around SNORD116 and at sites further 5’ were essentially unchanged. Extended snoRNA-related RNAs SPA1 and sno-lncRNAs 1-4 (Sledziowska et al ., 2023; Wu et al ., 2016; Yin et al ., 2012) were readily detected (Fig. 2B), but showed only modest changes during differentiation.…”

“…SNHG14 generates a very long non-protein coding RNA (lncRNA) with a predicted primary transcript around 600 Kb in length including 145 annotated introns (Runte et al , 2001). It is processed into multiple ncRNAs; including mature snoRNAs, extended snoRNA-related ncRNA species (SPA-lncRNAs and sno-lncRNAs), and alternatively spliced versions of the SNHG14 exons ((Sledziowska et al , 2023; Yin et al , 2012) reviewed in (Ariyanfar & Good, 2022)). Multiple, non-identical versions of SNORD116, are encoded by 29 tandem introns of SNHG14 and excised following splicing.…”

Identifying key underlying regulatory networks and predicting targets of orphan C/D boxSNORD116snoRNAs in Prader-Willi syndrome

“…While it has long been understood that perturbations of the chr15q11-13 region cause PWS, it is unclear if the genes included in the deletions are directly related to PWS phenotypes, if genes regulated by them are to blame, or if it is some combination of these effects. Multiple studies have atempted to address this issue by characterizing gene expression in postmortem PWS brain tissues and neurons differentiated from PWS patient-derived pluripotent stem cell lines to identify genes dysregulated in this disorder (Bochukova et al, 2018; Falaleeva et al, 2015; Huang et al, 2021; Sledziowska et al, 2023; Victor et al, 2021). While these studies indicate gene expression is indeed dysregulated in PWS patient samples, our analysis here showed few genes had consistent dysregulation across a subset of these studies (Supplemental Figure 1A).…”

“…Since the function of SNORD116 thus far has remained elusive, much effort has recently been expended to identify gene expression paterns that are dysregulated in PWS. Several studies have compared gene expression between tissue or cell lines derived from PWS patients and those from unrelated controls (Bochukova et al, 2018; Falaleeva et al, 2015; Huang et al, 2021; Sledziowska et al, 2023; Victor et al, 2021). While each of these studies identified numerous genes with distinct expression paterns in the PWS context, a coherent set of consistently dysregulated disease relevant genes has not been identified.…”