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Whole genome microarray analysis of gene expression in Prader-Willi syndrome.

ABSTRACT: Prader-Willi syndrome (PWS) is caused by loss of function of paternally expressed genes in the 15q11-q13 region and a paucity of data exists on transcriptome variation. To further characterize genetic alterations in this classic obesity syndrome using whole genome microarrays to analyze gene expression, microarray and quantitative RT-PCR analysis were performed using RNA isolated from lymphoblastoid cells from PWS male subjects (four with 15q11-q13 deletion and three with UPD) and three age and cognition matched nonsyndromic comparison males. Of more than 47,000 probes examined in the microarray, 23,383 were detectable and 323 had significantly different expression in the PWS lymphoblastoid cells relative to comparison cells, 14 of which were related to neurodevelopment and function. As expected, there was no evidence of expression of paternally expressed genes from the 15q11-q13 region (e.g., SNRPN) in the PWS cells. Alterations in expression of serotonin receptor genes (e.g., HTR2B) and genes involved in eating behavior and obesity (ADIPOR2, MC2R, HCRT, OXTR) were noted. Other genes of interest with reduced expression in PWS subjects included STAR (a key regulator of steroid synthesis) and SAG (an arrestin family member which desensitizes G-protein-coupled receptors). Quantitative RT-PCR for SAG, OXTR, STAR, HCRT, and HTR2B using RNA isolated from their lymphoblastoid cells and available brain tissue (frontal cortex) from separate individuals with PWS and control subjects and normalized to GAPD gene expression levels validated our microarray gene expression data. Our analysis identified previously unappreciated changes in gene expression which may contribute to the clinical manifestations seen in PWS.

SUBMITTER: Bittel DC

PROVIDER: S-EPMC5467864 | biostudies-literature | 2007 Mar

REPOSITORIES: biostudies-literature

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Whole genome microarray analysis of gene expression in Prader-Willi syndrome.

Bittel Douglas C DC Kibiryeva Nataliya N Sell Susan M SM Strong Theresa V TV Butler Merlin G MG

American journal of medical genetics. Part A 20070301 5

Prader-Willi syndrome (PWS) is caused by loss of function of paternally expressed genes in the 15q11-q13 region and a paucity of data exists on transcriptome variation. To further characterize genetic alterations in this classic obesity syndrome using whole genome microarrays to analyze gene expression, microarray and quantitative RT-PCR analysis were performed using RNA isolated from lymphoblastoid cells from PWS male subjects (four with 15q11-q13 deletion and three with UPD) and three age and ...[more]

PMID: 17236194

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Project description:Prader-Willi syndrome (PWS) is caused by loss of paternally expressed genes in the 15q11-q13 region. To further characterize alterations in gene expression in this classical obesity syndrome we used whole genome microarrays to study a PWS mouse model resulting from a paternally derived imprinting center (IC) deletion (PWS IC deletion). These mice die generally within 2-3 days of life (reflective of failure to thrive in infants with PWS) and therefore, the analysis was performed on RNA extracted from the whole brain of PWS IC deletion mice and normal littermates at less than 24 hr after birth. Of more than 45,000 probes examined, 26,471 (59%) were detected for further analysis, and 69 had a significant change in expression of at least 1.5-fold and a false discovery rate (FDR) of 5%. Eight of the genes with differential expression were imprinted and from the PWS critical region (PWSCR). The three genes with the highest expression in the PWS IC mice were pro-opiomelanocortin (Pomc) and two transcripts of unknown function. Pomc knockout mice have been shown to develop obesity. Therefore, elevated Pomc RNA in PWS IC deletion neonatal mice may be an important genetic factor in the survival of these mice as it may affect eating behavior. Interestingly, Mc5r, a melanocortin receptor known to directly respond to Pomc expression changes, was upregulated as well. Mc5r is known to be involved with thermoregulation which is reportedly abnormal in PWS infants. These observations support a role for Pomc and the network of genes involved in regulating energy homeostasis in the early clinical findings of failure to thrive observed in PWS. Other notable patterns include three previously unstudied transcripts that are expressed only from the paternal allele under regulatory control of the IC and include AK013560, BB3144814, and BB182944 (whose genes are located in the mouse PWSCR on chromosome 7B). As expected, all the known paternally expressed genes from the PWSCR had detection signals below the threshold in the PWS IC deletion mice but were clearly detectable in control littermates. Several of the genes in this study were further examined by quantitative reverse transcription-PCR (RT-PCR) to confirm their expression status. Further analysis of gene expression in these mice may lead to novel pathways affected in PWS. These results, along with other recent reports, suggest that the cumulative effect of modest changes in expression of many genes, especially genes involved in energy metabolism, contribute to the failure to thrive of infants with PWS.

Project description:BACKGROUND:Imprinting disorders (IDs) show overlapping phenotypes, particularly in Silver-Russell syndrome (SRS), Temple syndrome (TS14), and Prader-Willi syndrome (PWS). These three IDs include fetal and postnatal growth failure, feeding difficulty, and muscular hypotonia as major clinical features. However, the mechanism that causes overlapping phenotypes has not been clarified. To investigate the presence or absence of methylation signatures associated with overlapping phenotypes, we performed genome-wide methylation analysis (GWMA). RESULTS:GWMA was carried out on 36 patients with three IDs (SRS [n?=?16], TS14 [n?=?7], PWS [n?=?13]) and 11 child controls using HumanMethylation450 BeadChip including 475,000 CpG sites across the human genome. To reveal an aberrantly methylated region shared by SRS, TS14, and PWS groups, we compared genome-wide methylation data of the three groups with those of control subjects. All the identified regions were known as SRS-, TS14-, and PWS-related imprinting-associated differentially methylated regions (iDMRs), and there was no hypermethylated or hypomethylated region shared by different ID groups. To examine the methylation pattern shared by SRS, TS14, and PWS groups, we performed clustering analysis based on GWMA data. The result focusing on 620 probes at the 62 known iDMRs (except for SRS-, TS14-, and PWS-related iDMRs) classified patients into two categories: (1) category A, grossly normal methylation patterns mainly consisting of SRS group patients; and (2) category B, broad and mild hypermethylation patterns mainly consisting of TS14 and PWS group patients. However, we found no obvious relationship between these methylation patterns and phenotypes of patients. CONCLUSIONS:GWMA in three IDs found no methylation signatures shared by SRS, TS14, and PWS groups. Although clustering analysis showed similar mild hypermethylation patterns in TS14 and PWS groups, further study is needed to clarify the effect of methylation patterns on the overlapping phenotypes.

Project description:PURPOSE:Fragile X syndrome, the most common inherited form of human mental retardation, arises as a consequence of a large expansion of a CGG trinucleotide repeat in 5' untranslated region of the fragile X mental retardation 1 (FMR1) gene located on the X chromosome. Although the FMR1 gene was cloned 15 years ago, the mechanisms that cause fragile X syndrome remain to be elucidated. Multiple studies have identified proteins that potentially interact with FMRP, the product of FMR1, and differentially expressed genes in an Fmr1 knockout mouse. To assess the impact of fragile X syndrome on gene expression in humans and to attempt to identify disturbed genes and gene interactive pathways relevant to fragile X syndrome, we performed gene expression microarray analysis using RNA isolated from lymphoblastoid cells derived from males with fragile X syndrome with and similarly aged control males. METHODS:We used whole genome microarrays consisting of 57,000 probes to analyze global changes to the transcriptome in readily available lymphoblastoid cell lines derived from males with fragile X syndrome and healthy comparison males with normal intelligence. We verified the differential expression of several of these genes with known biological function relevant to fragile X syndrome using quantitative reverse transcription polymerase chain reaction using RNA from lymphoblastoid cells from fragile X syndrome and control males as well as RNA from human brain tissue (frontal cortex) of other affected fragile X syndrome males. RESULTS:We identified more than 90 genes that had significant differences in probe intensity of at least 1.5-fold with a false discovery rate of 5% in cells from males with fragile X syndrome relative to comparison males. The list of 90 differentially expressed genes contained an overrepresentation of genes involved in signaling (e.g., UNC13B [-3.3-fold change in expression in lymphoblasts by quantitative reverse transcription polymerase chain reaction), GABRD [+2.0-fold change] EEF1A2 [+4.3-fold change]), morphogenesis (e.g., MAP1B [-7.5-fold change], ACCN1 [-8.0-fold change]), and neurodevelopment and function (e.g., PPP1R9B [+3.5-fold change], HES1 [+2.8-fold change]). CONCLUSIONS:These genes may represent members of candidate networks disturbed by the loss of FMR1 and consequently fragile X mental retardation protein function, thus lending support for altered fragile X mental retardation protein function resulting in an abnormal transcriptome. Further analyses of the genes, especially those that have been identified in multiple studies, are warranted to develop a more integrated description of the alterations in gene processing that lead to fragile X syndrome.

Dataset Information

Whole genome microarray analysis of gene expression in Prader-Willi syndrome.

Publications

Whole genome microarray analysis of gene expression in Prader-Willi syndrome.

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