Abstract
BackgroundDuplications of the chromosome 15q11-q13.1 region are associated with an estimated 1 to 3% of all autism cases, making this copy number variation (CNV) one of the most frequent chromosome abnormalities associated with autism spectrum disorder (ASD). Several genes located within the 15q11-q13.1 duplication region including ubiquitin protein ligase E3A (UBE3A), the gene disrupted in Angelman syndrome (AS), are involved in neural function and may play important roles in the neurobehavioral phenotypes associated with chromosome 15q11-q13.1 duplication (Dup15q) syndrome.MethodsWe have generated induced pluripotent stem cell (iPSC) lines from five different individuals containing CNVs of 15q11-q13.1. The iPSC lines were differentiated into mature, functional neurons. Gene expression across the 15q11-q13.1 locus was compared among the five iPSC lines and corresponding iPSC-derived neurons using quantitative reverse transcription PCR (qRT-PCR). Genome-wide gene expression was compared between neurons derived from three iPSC lines using mRNA-Seq.ResultsAnalysis of 15q11-q13.1 gene expression in neurons derived from Dup15q iPSCs reveals that gene copy number does not consistently predict expression levels in cells with interstitial duplications of 15q11-q13.1. mRNA-Seq experiments show that there is substantial overlap in the genes differentially expressed between 15q11-q13.1 deletion and duplication neurons, Finally, we demonstrate that UBE3A transcripts can be pharmacologically rescued to normal levels in iPSC-derived neurons with a 15q11-q13.1 duplication.ConclusionsChromatin structure may influence gene expression across the 15q11-q13.1 region in neurons. Genome-wide analyses suggest that common neuronal pathways may be disrupted in both the Angelman and Dup15q syndromes. These data demonstrate that our disease-specific stem cell models provide a new tool to decipher the underlying cellular and genetic disease mechanisms of ASD and may also offer a pathway to novel therapeutic intervention in Dup15q syndrome.
Highlights
Duplications of the chromosome 15q11-q13.1 region are associated with an estimated 1 to 3% of all autism cases, making this copy number variation (CNV) one of the most frequent chromosome abnormalities associated with autism spectrum disorder (ASD)
Generation of induced pluripotent stem cell (iPSC) from Dup15q patients iPSC lines were established using fibroblasts from one idic (15) individual, one individual with a paternally-inherited duplication of chromosome 15q11-q13.1, and one individual who was mosaic for a maternally-inherited interstitial duplication of chromosome 15q11-q13.1 (Figure 1a). iPSC lines were established from a cord blood sample from one individual with idic(15) (Figure 1a). iPSCs were generated using either retroviral, lentiviral, or episomal vectors encoding POU class 5 homeobox 1 (OCT4), SRY-box 2 (SOX2), Kruppel-like factor 4 (KLF4), v-myc avian myelocytomatosis viral oncogene homolog (MYC), and lin28 homolog A (LIN28) [see Additional file 1: Table S1] [26,27,28]
Reprogrammed colonies were initially identified morphologically, and were subsequently validated using quantitative reverse transcription PCR (qRT-PCR) and/or immunocytochemistry to verify the expression of the pluripotency markers, NANOG, stage specific embryonic antigen 4 (SSEA4), and TRA1-60 (Figure 1b-d) and pluripotency genes [see Additional file 2: Figure S1A]. iPSCs were shown to have the expected karyotypes of 46, XX; 46,XX.ish dup(15)(q11.2q11.2)(SNRPN++); or 47,XX,+idic(15).ish15q12 small nuclear ribonucleoprotein polypeptide N (SNRPN) x 4, 15qter X2 (Figure 1e)
Summary
Duplications of the chromosome 15q11-q13.1 region are associated with an estimated 1 to 3% of all autism cases, making this copy number variation (CNV) one of the most frequent chromosome abnormalities associated with autism spectrum disorder (ASD). One of the most frequent chromosome anomalies associated with autism is the duplication of chromosome 15q11-q13.1 [1,2,3,4,5,6,7,8]. The parent-of-origin is an important factor for chromosome 15q11-q13.1 duplication (Dup15q) syndrome because the chromosome 15q11-q13.1 region is subject to genomic imprinting, which is an epigenetic process that results in monoallelic gene expression. The 15q11-q13.1 duplications that lead to autism are most frequently of maternal origin. Interstitial duplications (int dup(15)) result in tandem copies of maternal 15q11-q13.1 lying in a head-to-head orientation on the same chromosome arm. Isodicentric chromosome 15 (idic(15)) duplications result in two additional copies of maternal 15q11-q13.1 which are flanked by two centromeres on a supernumerary chromosome. Individuals with idic(15), who have 4 copies of 15q11-q13.1, are more severely affected than those with int dup(15), who have 3 copies [9]
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