Search

Background: Prader-Willi syndrome (PWS) is a rare genetic disorder resulting from the lack of expression of the PWS region (locus q11-q13) on the paternally derived chromosome 15, as a result of a type I or II paternal deletion (50%), maternal uniparental disomy (43%), imprinting defect (4%) or translocation (

The eutherian-specific SNORD116 family of repeated box C/D snoRNA genes is suspected to play a major role in the Prader–Willi syndrome (PWS), yet its molecular function remains poorly understood. Here, we combined phylogenetic and molecular analyses to identify candidate RNA targets. Based on the analysis of several eutherian orthologs, we found evidence of extensive birth-and-death and conversion events during SNORD116 gene history. However, the consequences for phylogenetic conservation were heterogeneous along the gene sequence. The standard snoRNA elements necessary for RNA stability and association with dedicated core proteins were the most conserved, in agreement with the hypothesis that SNORD116 generate genuine snoRNAs. In addition, one of the two antisense elements typically involved in RNA target recognition was largely dominated by a unique sequence present in at least one subset of gene paralogs in most species, likely the result of a selective effect. In agreement with a functional role, this ASE exhibited a hybridization capacity with putative mRNA targets that was strongly conserved in eutherians. Moreover, transient downregulation experiments in human cells showed that Snord116 controls the expression and splicing levels of these mRNAs. The functions of two of them, diacylglycerol kinase kappa and Neuroligin 3, extend the description of the molecular bases of PWS and reveal unexpected molecular links with the Fragile X syndrome and autism spectrum disorders.

Background Prader–Willi syndrome is a rare genetic neurodevelopmental disorder caused by a paternal deficiency of maternally imprinted gene expression located in the chromosome 15q11–q13 region. Previous studies have demonstrated that several classes of neurodevelopmental disorders can be attributed to either over- or under-expression of specific genes that may lead to impairments in neuronal generation, differentiation, maturation and growth. Epigenetic changes that modify gene expression have been highlighted in these disorders. One recent study focused on epigenetic analysis and compared patients with PWS with patients with other imprinting disorders. No study, however, has yet focused on epigenetics in patients with PWS specifically by comparing the mutations associated with this syndrome. Objective This study investigated the epigenetic modifications in patients with PWS and patients with PWS-related disorders caused by inactivation of two genes of the PWS chromosomal region, SNORD116 and MAGEL2. Our approach also aimed to compare the epigenetic modifications in PWS and PWS-related disorders. Methods We compared genome-wide methylation analysis (GWAS) in seven blood samples from patients with PWS phenotype (five with deletions of the PWS locus, one with a microdeletion of SNORD116 and one with a frameshift mutation of MAGEL2 presenting with Schaaf–Yang syndrome), as well as two control patients. Controls were infants that had been studied for suspicion of genetic diseases that was not confirmed by the genetic analysis and the clinical follow-up. Results The analysis identified 29,234 differentially methylated cytosines, corresponding to 5,308 differentially methylated regions (DMRs), which matched with 2,280 genes. The DMRs in patients with PWS were associated with neurodevelopmental pathways, endocrine dysfunction and social and addictive processes consistent with the key features of the PWS phenotype. In addition, the separate analysis for the SNORD116 and MAGEL2 deletions revealed that the DMRs associated with the SNORD116 microdeletion were found in genes implicated in metabolic pathways and nervous system development, whereas MAGEL2 mutations mostly concerned genes involved in macromolecule biosynthesis. Conclusion The PWS is associated with epigenetic modifications with differences in SNORD116 and MAGEL2 mutations, which seem to be relevant to the different associated phenotypes.

Prader–Willi syndrome (PWS) is a complex neurodevelopmental disorder caused by the deletion or inactivation of paternally expressed imprinted genes at the chromosomal region 15q11–q13. The PWS-critical region (PWScr) harbors tandemly repeated non-protein coding IPW-A exons hosting the intronic SNORD116 snoRNA gene array that is predominantly expressed in brain. Paternal deletion of PWScr is associated with key PWS symptoms in humans and growth retardation in mice (PWScr model). Dysregulation of the hypothalamic–pituitary axis (HPA) is thought to be causally involved in the PWS phenotype. Here we performed a comprehensive reverse transcription quantitative PCR (RT-qPCR) analysis across nine different brain regions of wild-type (WT) and PWScr mice to identify stably expressed reference genes. Four methods (Delta Ct, BestKeeper, Normfinder and Genorm) were applied to rank 11 selected reference gene candidates according to their expression stability. The resulting panel consists of the top three most stably expressed genes suitable for gene-expression profiling and comparative transcriptome analysis of WT and/or PWScr mouse brain regions. Using these reference genes, we revealed significant differences in the expression patterns of Igfbp7, Nlgn3 and three HPA associated genes: Pcsk1,Pcsk2 and Nhlh2 across investigated brain regions of wild-type and PWScr mice. Our results raise a reasonable doubt on the involvement of the Snord116 in posttranscriptional regulation of Nlgn3 and Nhlh2 genes. We provide a valuable tool for expression analysis of specific genes across different areas of the mouse brain and for comparative investigation of PWScr mouse models to discover and verify different regulatory pathways affecting this complex disorder.

Prader-Willi Syndrome (PWS) is a genetic disease that results in abnormal hormone levels, developmental delay, intellectual disability, hypogonadism, and excessive appetite. The disease is caused by a de novo genetic deletion in chromosome 15. While many of the deleted genes have been identified, there is little known about their molecular function. There is evidence that a cluster of non-coding RNA genes in the deleted region known as the SNORD116 genes may be the most critical genes deleted in Prader-Willi Syndrome. It is unknown what the SNORD116 genes do at the molecular level, but recent evidence suggests they regulate the expression of other genes involved in the neuroendocrine system. Specifically, the SNORD116 gene is implicated in regulation of NHLH2, a transcription factor gene which plays a key role in development, hormonal regulation, and body weight. In this study we identify phylogenetically conserved regions of SNORD116 and predict interactions with its potential downstream RNA targets. We show that mouse Snord116 post-transcriptionally increases Nhlh2 RNA levels dependent on its 3'UTR and protects it from degradation within 45 minutes of its transcription. Additionally, a single nucleotide variant within Nhlh2 at the predicted Snord116 interaction site may disrupt Snord116's protective effect. This is the first observation of a molecular mechanism for Snord116, identifying its role in RNA stability, and leads us closer to understanding Prader-Willi Syndrome and finding a possible treatment. However, Snord116 in vitro knockdown or paternally inherited in vivo deletion fail to detect differential expression of Nhlh2, likely due to missing the key timepoint of Snord116 regulatory effects on Nhlh2 RNA soon after its transcriptional stimulation, and dependent on leptin signals. Furthermore, the hypothalamic mRNA expression profile of PWS mouse models fed a nutraceutical dietary supplement of conjugated linoleic acid reveals minimal overall changes, while the effect of diet may be stronger than genotype and potentially changes gene expression of metabolic molecular pathways. Doctor of Philosophy Prader-Willi Syndrome is a genetic disease that results in abnormal hormone levels, slow development, intellectual disability, gonad deficiency, and excessive appetite. The disease is caused by a genetic deletion in chromosome 15 that is almost always a spontaneous mutation not inherited from the parents. While many of the deleted genes have been identified, there is little known about what their molecular function is. There is evidence that a cluster of genes in the deleted region known as the SNORD116 genes may be the most critical genes deleted in Prader-Willi Syndrome. It is unknown what the SNORD116 genes do at the molecular level, but recent evidence suggests that it regulates other genes involved in the hormone system. Specifically, the SNORD116 gene is implicated to regulate the levels of NHLH2, a gene which plays a key role in development, hormonal regulation, and body weight. In this study we identify key regions of SNORD116 and predict interactions with its potential downstream targets. We show that SNORD116 increases NHLH2 levels and slows its degradation at the RNA transcript level. This is the first observation of a molecular mechanism for SNORD116 and leads us closer to understanding Prader-Willi Syndrome and finding a possible treatment. However, other mouse models of Snord116 deletion fail to find differences in Nhlh2. This is likely due to missing a brief key timepoint and hormonal signal when Nhlh2 is most subject to Snord116's effects. Furthermore, PWS mouse models fed a supplement intended for weight loss leads to mild overall gene expression changes in the hypothalamus, a brain region that regulates many hormonal signals including appetite and energy balance. The effect of diet may be stronger than genotype in this brain region, with diet potentially changing the activity of metabolic molecular pathways.

The human genome contains more than two hundred C/D box small nucleolar RNAs (snoRNAs) genes. C/D snoRNAs associate with a set of four core proteins to catalyze 2ʹ-O-methylations of ribosomal RNAs (rRNAs) and small nuclear RNAs (snRNAs). Interestingly, over the past decade, studies done mostly on mammalian models suggested that snoRNPs are involved in unrelated cellular processes. The human genome encodes numerous orphan C/D snoRNAs that do not possess obvious complementarities with canonical target RNAs. Also, several lines of evidence pointed to the possibility that snoRNAs associate with other proteins than snoRNP core proteins and may affect cell metabolism and proliferation. Finally, specific snoRNAs are deregulated in pathologies such as the Prader-Willi syndrome or in several cancers. In front of this initially unsuspected complex picture, we emit the hypothesis that the formation, the fate, and the functions of snoRNPs are regulated by unappreciated protein factors, in physiological and pathological contexts. Thus, our main objectives were to study new molecular mechanisms involved in C/D box snoRNPs functions using a combination of several complementary approaches, based on the study of RNA:protein and RNA:RNA interactions. We identified that the orphan snoRNA SNORD116 possesses a complementarity antisense element with several mRNAs sequences and we observed that the depletion of SNORD116 by ASO affects their steady state levels. We searched for new snoRNP protein partners by a combination of co-immunoprecipitation (IP) and mass spectrometry (MS) analyses. We identified that GNL3/Nucleostemin, a GTP-binding protein interacts with protein complexes involved in C/D snoRNP biogenesis. We also identified that the RNA binding protein ILF3 interacts with mature snoRNPs. Depletion of ILF3 by siRNAs does not affect snoRNA steady state levels and modestly modulates their association with the ribosome. Analysis of rRNAs 2ʹ-O-methylations will be crucial to determine whether ILF3 affects snoRNPs catalytic activities. Moreover, data obtained from IP-MS and CLIP (crosslinking immunoprecipitation)-microarray suggested that ILF3 impacts the association of snoRNPs with the splicing machinery which leads to a change in the splicing pattern of several cellular mRNAs., Le génome humain contient plus de deux cents gènes de petits ARN nucléolaires (snoARN) à boîtes C/D. Les snoARN C/D s'associent avec un jeu de quatre protéines cœurs pour catalyser les 2ʹ-O-méthylations des ARN ribosomiques (ARNr) et des petits ARN nucléaires (snARN). De manière intéressante, au cours de la dernière décennie, des études effectuées principalement sur des modèles de mammifères ont suggéré que les snoRNP sont impliquées dans d’autres processus cellulaires. Le génome humain contient de nombreux snoARN C/D orphelins qui ne possèdent pas de complémentarités évidentes avec les ARN cibles canoniques. En outre, plusieurs évidences ont souligné la possibilité que les snoARN s'associent avec de nouveaux partenaires protéiques et pourraient affecter le métabolisme et la prolifération des cellules. Enfin, certains snoARN sont spécifiquement dérégulés dans des pathologies comme le syndrome de Prader-Willi ou dans plusieurs cancers. Face à la complexité émergente des données issues de la littérature, nous émettons l'hypothèse que la formation, le devenir et les fonctions des snoRNP sont régulés par des facteurs protéiques non identifiés, dans des contextes physiologiques et pathologiques. Ainsi, nos principaux objectifs étaient d'étudier de nouveaux mécanismes moléculaires impliqués dans les fonctions cellulaires des snoRNP à boîtes C/D en utilisant une combinaison de plusieurs approches complémentaires, basée sur l'étude d’interactions ARN:protéine et ARN:ARN. Nous avons identifié que le snoARN orphelin SNORD116 possède des éléments antisens complémentaires de séquences contenues dans plusieurs ARNm et nous avons observé que la déplétion de SNORD116 par des oligonucléotide antisens (ASO) affecte leur taux à l’état d'équilibre. Nous avons recherché de nouveaux partenaires protéiques des snoRNP en utilisant une combinaison d'analyses par co-immunoprécipitation (IP) et spectrométrie de masse (MS). Nous avons identifié que GNL3/Nucléostemine, une protéine de liaison au GTP, interagit avec les complexes protéiques impliqués dans la biogenèse des snoRNP à boîtes C/D. Nous avons également observé que la protéine de fixation aux ARN, ILF3, interagit avec des snoRNP matures. La réduction du taux d'ILF3 par l’utilisation de siARN n'affecte pas le taux à l’équilibre des snoARN et module modestement leur association avec le ribosome. L'analyse des 2ʹ-O-méthylations portées par ARNr nous permettra de déterminer si ILF3 affecte les activités catalytiques des snoRNP. De plus, les données obtenues à partir d'IP-MS et de CLIP (crosslinking-immunoprecipitation)-microarray suggèrent que ILF3 impacte l'association des snoRNP avec des protéines de la machinerie d’épissage, ce qui a pour conséquence d’altérer le profil d’épissage de certains ARNm cellulaires.

We report a 17-year-old boy who met most of the major Prader−Willi syndrome (PWS) diagnostic criteria, including infantile hypotonia and poor feeding followed by hyperphagia, early-onset morbid obesity, delayed development, and characteristic facial features. However, unlike many children with PWS, he had spontaneous onset of puberty and reached a tall adult stature without growth hormone replacement therapy. A phenotype-driven genetic analysis using exome sequencing identified a heterozygous microdeletion of 71 kb in size at chr15:25,296,613-25,367,633, genome build hg 19. This deletion does not affect the SNURF-SNRPN locus, but results in the loss of several of the PWS-associated non-coding RNA species, including the SNORD116 cluster. We compared with six previous reports of patients with PWS who carried small atypical deletions encompassing the snoRNA SNORD116 cluster. These patients share similar core symptoms of PWS while displaying some atypical features, suggesting that other genes in the region may make lesser phenotypic contributions. Altogether, these rare cases provide convincing evidence that loss of the paternal copy of the SNORD116 snoRNA is sufficient to cause most of the major clinical features of PWS.

Prader–Willi Syndrome (PWS) is a human genetic condition that affects up to 1 in 10,000 live births. Affected infants present with hypotonia and developmental delay. Hyperphagia and increasing body weight follow unless drastic calorie restriction is initiated. Recently, our laboratory showed that one of the genes in the deleted locus causative for PWS, Snord116, maintains increased expression of hypothalamic Nhlh2, a basic helix–loop–helix transcription factor. We have previously also shown that obese mice with a deletion of Nhlh2 respond to a conjugated linoleic acid (CLA) diet with weight and fat loss. In this study, we investigated whether mice with a paternal deletion of Snord116 (Snord116m+/p−) would respond similarly. We found that while Snord116m+/p− mice and mice with a deletion of both Snord116 alleles were not significantly obese on a high-fat diet, they did lose body weight and fat on a high-fat/CLA diet, suggesting that the genotype did not interfere with CLA actions. There were no changes in food intake or metabolic rate, and only moderate differences in exercise performance. RNA-seq and microbiome analyses identified hypothalamic mRNAs, and differentially populated gut bacteria, that support future mechanistic analyses. CLA may be useful as a food additive to reduce obesity in humans with PWS. Published version

Summary Transcriptional analysis of brain tissue from people with molecularly defined causes of obesity may highlight disease mechanisms and therapeutic targets. We performed RNA sequencing of hypothalamus from individuals with Prader-Willi syndrome (PWS), a genetic obesity syndrome characterized by severe hyperphagia. We found that upregulated genes overlap with the transcriptome of mouse Agrp neurons that signal hunger, while downregulated genes overlap with the expression profile of Pomc neurons activated by feeding. Downregulated genes are expressed mainly in neuronal cells and contribute to neurogenesis, neurotransmitter release, and synaptic plasticity, while upregulated, predominantly microglial genes are involved in inflammatory responses. This transcriptional signature may be mediated by reduced brain-derived neurotrophic factor expression. Additionally, we implicate disruption of alternative splicing as a potential molecular mechanism underlying neuronal dysfunction in PWS. Transcriptomic analysis of the human hypothalamus may identify neural mechanisms involved in energy homeostasis and potential therapeutic targets for weight loss., Graphical Abstract, Highlights • Overlap between genes expressed in human PWS hypothalamus and mouse Agrp neurons • Downregulated genes are involved in neuronal development • SNORD116 deletion reduces neural development and survival in cells • Alternative splicing is disturbed in PWS, Prader-Willi syndrome (PWS) is a genetic obesity syndrome. Bochukova et al. report gene expression changes in the hypothalamus of people with PWS that support neurodegeneration and neuroinflammation as key processes involved in this condition.

// Elena Vendramini 1, 2, 3 , Marco Giordan 1 , Emanuela Giarin 1 , Barbara Michielotto 1 , Grazia Fazio 4 , Gianni Cazzaniga 4 , Andrea Biondi 4 , Daniela Silvestri 4 , Maria Grazia Valsecchi 5 , Martina U. Muckenthaler 6 , Andreas E. Kulozik 6 , Valter Gattei 7 , Shai Izraeli 2, 3 , Giuseppe Basso 1 and Geertruy te Kronnie 1 1 Department of Women’s and Children’s Health, University of Padova, Padova, Italy 2 Edmond and Lily Safra Children’s Hospital, Sheba Medical Center, Ramat Gan, Israel 3 Tel Aviv University, Tel Aviv, Israel 4 Centro Ricerca Tettamanti, Clinica Pediatrica, University of Milano-Bicocca, Monza, Italy 5 School of Medicine and Surgery, University of Milano-Bicocca, Milano, Italy 6 Department of Pediatric Oncology Hematology, University of Heidelberg, Heidelberg, Germany 7 Clinical and Experimental Onco-Hematology Unit, Centro di Riferimento Oncologico, I.R.C.C.S., Aviano (PN), Italy Correspondence to: Elena Vendramini, email: elena.vendramini@gmail.com Keywords: B cell precursor acute lymphoblastic leukemia, ERG aberrations, noncoding RNAs, miR-125, SNORD116 Received: June 29, 2016 Accepted: March 04, 2017 Published: March 21, 2017 ABSTRACT ERG-related leukemia is a B cell precursor acute lymphoblastic leukemia (BCP ALL) subtype characterized by aberrant expression of DUX4 and ERG transcription factors, and highly recurrent ERG intragenic deletions. ERG-related patients have remarkably favorable outcome despite a high incidence of inauspicious IKZF1 aberrations. We describe clinical and genomic features of the ERG-related cases in an unselected cohort of B-other BCP ALL pediatric patients enrolled in the AIEOP ALL 2000 therapeutic protocol. We report a small noncoding RNA signature specific of ERG-related group, with up-regulation of miR-125b-2 cluster on chromosome 21 and several snoRNAs in the Prader-Willi locus at 15q11.2, including the orphan SNORD116 cluster.

Prader-Willi syndrome (PWS) is the most common genetic obesity syndrome, with associated learning difficulties, neuroendocrine deficits, and behavioural and psychiatric problems. As the life expectancy of individuals with PWS increases, there is concern that alterations in brain structure associated with the syndrome, as a direct result of absent expression of PWS genes, and its metabolic complications and hormonal deficits, might cause early onset of physiological and brain aging. In this study, a machine learning approach was used to predict brain age based on grey matter (GM) and white matter (WM) maps derived from structural neuroimaging data using T1-weighted magnetic resonance imaging (MRI) scans. Brain-predicted age difference (brain-PAD) scores, calculated as the difference between chronological age and brain-predicted age, are designed to reflect deviations from healthy brain aging, with higher brain-PAD scores indicating premature aging. Two separate adult cohorts underwent brain-predicted age calculation. The main cohort consisted of adults with PWS (n = 20; age mean 23.1 years, range 19.8–27.7; 70.0% male; body mass index (BMI) mean 30.1 kg/m2, 21.5–47.7; n = 19 paternal chromosome 15q11–13 deletion) and age- and sex-matched controls (n = 40; age 22.9 years, 19.6–29.0; 65.0% male; BMI 24.1 kg/m2, 19.2–34.2) adults (BMI PWS vs. control P = .002). Brain-PAD was significantly greater in PWS than controls (effect size mean ± SEM +7.24 ± 2.20 years [95% CI 2.83, 11.63], P = .002). Brain-PAD remained significantly greater in PWS than controls when restricting analysis to a sub-cohort matched for BMI consisting of n = 15 with PWS with BMI range 21.5–33.7 kg/m2, and n = 29 controls with BMI 21.7–34.2 kg/m2 (effect size +5.51 ± 2.56 years [95% CI 3.44, 10.38], P = .037). In the PWS group, brain-PAD scores were not associated with intelligence quotient (IQ), use of hormonal and psychotropic medications, nor severity of repetitive or disruptive behaviours. A 24.5 year old man (BMI 36.9 kg/m2) with PWS from a SNORD116 microdeletion also had increased brain PAD of 12.87 years, compared to 0.84 ± 6.52 years in a second control adult cohort (n = 95; age mean 34.0 years, range 19.9–55.5; 38.9% male; BMI 28.7 kg/m2, 19.1–43.1). This increase in brain-PAD in adults with PWS indicates abnormal brain structure that may reflect premature brain aging or abnormal brain development. The similar finding in a rare patient with a SNORD116 microdeletion implicates a potential causative role for this PWS region gene cluster in the structural brain abnormalities associated primarily with the syndrome and/or its complications. Further longitudinal neuroimaging studies are needed to clarify the natural history of this increase in brain age in PWS, its relationship with obesity, and whether similar findings are seen in those with PWS from maternal uniparental disomy., Highlights • Prader-Willi syndrome is the commonest genetic obesity syndrome. • 19 of the 20 adults with PWS had a paternal deletion in chromosome 15q11–13. • Adults with PWS had increased brain age using structural MRI with machine learning. • This was independent of their higher BMI, and use of growth and sex hormones. • Another man with a SNORD116 microdeletion also had an increased brain age.

Prader-Willi syndrome (PWS) is a neurogenetic multifactorial disorder caused by the deletion or inactivation of paternally imprinted genes on human chromosome 15q11-q13. The affected homologous locus is on mouse chromosome 7C. The positional conservation and organization of genes including the imprinting pattern between mice and men implies similar physiological functions of this locus. Therefore, considerable efforts to recreate the pathogenesis of PWS have been accomplished in mouse models. We provide a summary of different mouse models that were generated for the analysis of PWS and discuss their impact on our current understanding of corresponding genes, their putative functions and the pathogenesis of PWS. Murine models of PWS unveiled the contribution of each affected gene to this multi-facetted disease, and also enabled the establishment of the minimal critical genomic region (PWScr) responsible for core symptoms, highlighting the importance of non-protein coding genes in the PWS locus. Although the underlying disease-causing mechanisms of PWS remain widely unresolved and existing mouse models do not fully capture the entire spectrum of the human PWS disorder, continuous improvements of genetically engineered mouse models have proven to be very powerful and valuable tools in PWS research.

Prader-Willi syndrome (PWS) is an imprinted neurodevelopmental disease caused by a loss of paternal genes on chromosome 15q11-q13. It is characterized by cognitive impairments, developmental delay, sleep abnormalities, and hyperphagia often leading to obesity. Clinical research has shown that a lack of expression of SNORD116, a paternally expressed imprinted gene cluster that encodes multiple copies of a small nucleolar RNA (snoRNA) in both humans and mice, is most likely responsible for many PWS symptoms seen in humans. The majority of previous research using PWS preclinical models focused on characterization of the hyperphagic and metabolic phenotypes. However, a crucial understudied clinical phenotype is cognitive impairments and thus we investigated the learning and memory abilities using a model of PWS, with a heterozygous deletion in Snord116. We utilized the novel object recognition task, which doesn’t require external motivation, or exhaustive swim training. Automated findings were further confirmed with manual scoring by a highly trained blinded investigator. We discovered deficits in Snord116 +/– mutant mice in the novel object recognition, location memory and tone cue fear conditioning assays when compared to age-, sex- matched, littermate control Snord116 +/+ mice. Further, we confirmed that despite physical neo-natal developmental delays, Snord116 +/− mice had normal exploratory and motor abilities. These results show that the Snord116 + /– deletion murine model is a valuable preclinical model for investigating learning and memory impairments in individuals with PWS without common confounding phenotypes.

Study objectives Sleep-wake disturbances are often reported in Prader-Willi syndrome (PWS), a rare neurodevelopmental syndrome that is associated with paternally-expressed genomic imprinting defects within the human chromosome region 15q11-13. One of the candidate genes, prevalently expressed in the brain, is the small nucleolar ribonucleic acid-116 (SNORD116). Here we conducted a translational study into the sleep abnormalities of PWS, testing the hypothesis that SNORD116 is responsible for sleep defects that characterize the syndrome. Methods We studied sleep in mutant mice that carry a deletion of Snord116 at the orthologous locus (mouse chromosome 7) of the human PWS critical region (PWScr). In particular, we assessed EEG and temperature profiles, across 24-h, in PWScr (m+/p-) heterozygous mutants compared to wild-type littermates. High-resolution magnetic resonance imaging (MRI) was performed to explore morphoanatomical differences according to the genotype. Moreover, we complemented the mouse work by presenting two patients with a diagnosis of PWS and characterized by atypical small deletions of SNORD116. We compared the individual EEG parameters of patients with healthy subjects and with a cohort of obese subjects. Results By studying the mouse mutant line PWScr(m+/p-), we observed specific rapid eye movement (REM) sleep alterations including abnormal electroencephalograph (EEG) theta waves. Remarkably, we observed identical sleep/EEG defects in the two PWS cases. We report brain morphological abnormalities that are associated with the EEG alterations. In particular, mouse mutants have a bilateral reduction of the gray matter volume in the ventral hippocampus and in the septum areas, which are pivotal structures for maintaining theta rhythms throughout the brain. In PWScr(m+/p-) mice we also observed increased body temperature that is coherent with REM sleep alterations in mice and human patients. Conclusions Our study indicates that paternally expressed Snord116 is involved in the 24-h regulation of sleep physiological measures, suggesting that it is a candidate gene for the sleep disturbances that most individuals with PWS experience.

Prader-Willi syndrome (PWS) is the most common genetic obesity syndrome, with associated learning difficulties, neuroendocrine deficits, and behavioural and psychiatric problems. As the life expectancy of individuals with PWS increases, there is concern that alterations in brain structure associated with the syndrome, as a direct result of absent expression of PWS genes, and its metabolic complications and hormonal deficits, might cause early onset of physiological and brain aging. In this study, a machine learning approach was used to predict brain age based on grey matter (GM) and white matter (WM) maps derived from structural neuroimaging data using T1-weighted magnetic resonance imaging (MRI) scans. Brain-predicted age difference (brain-PAD) scores, calculated as the difference between chronological age and brain-predicted age, are designed to reflect deviations from healthy brain aging, with higher brain-PAD scores indicating premature aging. Two separate adult cohorts underwent brain-predicted age calculation. The main cohort consisted of adults with PWS (n = 20; age mean 23.1 years, range 19.8-27.7; 70.0% male; body mass index (BMI) mean 30.1 kg/m2, 21.5-47.7; n = 19 paternal chromosome 15q11-13 deletion) and age- and sex-matched controls (n = 40; age 22.9 years, 19.6-29.0; 65.0% male; BMI 24.1 kg/m2, 19.2-34.2) adults (BMI PWS vs. control P = .002). Brain-PAD was significantly greater in PWS than controls (effect size mean ± SEM +7.24 ± 2.20 years [95% CI 2.83, 11.63], P = .002). Brain-PAD remained significantly greater in PWS than controls when restricting analysis to a sub-cohort matched for BMI consisting of n = 15 with PWS with BMI range 21.5-33.7 kg/m2, and n = 29 controls with BMI 21.7-34.2 kg/m2 (effect size +5.51 ± 2.56 years [95% CI 3.44, 10.38], P = .037). In the PWS group, brain-PAD scores were not associated with intelligence quotient (IQ), use of hormonal and psychotropic medications, nor severity of repetitive or disruptive behaviours. A 24.5 year old man (BMI 36.9 kg/m2) with PWS from a SNORD116 microdeletion also had increased brain PAD of 12.87 years, compared to 0.84 ± 6.52 years in a second control adult cohort (n = 95; age mean 34.0 years, range 19.9-55.5; 38.9% male; BMI 28.7 kg/m2, 19.1-43.1). This increase in brain-PAD in adults with PWS indicates abnormal brain structure that may reflect premature brain aging or abnormal brain development. The similar finding in a rare patient with a SNORD116 microdeletion implicates a potential causative role for this PWS region gene cluster in the structural brain abnormalities associated primarily with the syndrome and/or its complications. Further longitudinal neuroimaging studies are needed to clarify the natural history of this increase in brain age in PWS, its relationship with obesity, and whether similar findings are seen in those with PWS from maternal uniparental disomy.