PWS/AS MS-MLPA Confirms Maternal Origin of 15q11.2 Microduplication

Abstract

The proximal region of the long arm of chromosome 15q11.2-q13 is associated with various neurodevelopmental disorders, including Prader-Willi (PWS) and Angelman (AS) syndromes, autism, and other developmental abnormalities resulting from deletions and duplications. In addition, this region encompasses imprinted genes that cause PWS or AS, depending on the parent-of-origin. This imprinting allows for diagnosis of PWS or AS based on methylation status using methylation sensitive (MS) multiplex ligation dependent probe amplification (MLPA). Maternally derived microduplications at 15q11.2-q13 have been associated with autism and other neuropsychiatric disorders. Multiple methods have been used to determine the parent-of-origin for 15q11.2-q13 microdeletions and microduplications. In the present study, a four-year-old nondysmorphic female patient with developmental delay was found to have a de novo ~5 Mb duplication within 15q11.2 by oligonucleotide genomic array. In order to determine the significance of this microduplication to the clinical phenotype, the parent-of-origin needed to be identified. The PWS/AS MS-MLPA assay is generally used to distinguish between deletion and uniparental disomy (UPD) of 15q11.2-q13, resulting in either PWS or AS. However, our study shows that PWS/AS MS-MLPA can also efficiently distinguish the parental origin of duplications of 15q11.2-q13.

Figure 1

Chromosome 15 ideogram showing breakpoints BP1–BP5 in 15q11.2-q13 and the PWS/AS common deletion regions (green). Genes located in each breakpoint interval are shown below (blue = paternal expression/maternal imprint; purple = maternal expression/paternal imprint; black = biallelic expression/no imprint). ^*APBA2 is a nonimprinted gene utilized in MS-MLPA (see Figure 2).

Figure 2

MS-MLPA analysis for determination of 15q11.2 duplication. x-axes represent fragment size in bps and y-axes represent probe peak ratios. (a) MLPA copy number peak ratio: red = PWS/AS region probes; green = non-PWS/AS region probes. 1 = TUBGCP5; 2 = APBA2; 3 = NIPA1; X (2 copies) and Y (0 copies) chromosome in female patient. (b) MS-MLPA digestion of five HhaI methylation sensitive probes: 4 = SNRPN intron 1; 5 = SNRPN exon 1; 6 = SNRPN intron 1; 7 = SNRPN promoter; 8 = NDN; ∗ = HhaI digestion controls.