Using long-read sequencing to detect and subtype a case with Temple syndrome

Abstract

Temple syndrome is an imprinting disorder resulting from abnormal genomic or epigenomic aberrations of chromosome 14 including maternal uniparental disomy (matUPD), paternal deletion of 14q32, or aberrant methylation of the imprinting control regions at 14q32. Understanding the underlying molecular mechanism is essential to understanding the recurrence risk and physical effects. Currently, diagnosis requires the detection of aberrant methylation and copy number loss via methylation-sensitive assays such as methylation-specific multiplex ligation-dependent probe amplification, and short tandem repeat analysis to detect matUPD and the presence of epimutation. Therefore, a one-step approach that can detect aberrant methylation and underlying genetic mechanisms would be of high clinical value. Here we use nanopore sequencing to delineate the molecular diagnosis of a case with Temple syndrome. We demonstrate the application of nanopore sequencing to detect aberrant methylation and underlying genetic mechanisms simultaneously in this case, thus providing a proof of concept for a one-step approach for molecular diagnosis of this disorder.

Keywords: DNA Methylation; Epigenomics; Genetics, Medical; Genomics; Nanopore Sequencing.

Figure 1. Patient with Temple syndrome with mixed maternal UPD. On the uppermost part, an ideogram shows regions of chromosome 14. Below this, the CN plot demonstrates a normal CN across Chr14, while the BAF plot shows a proximal centromeric heterozygosity and homozygosity from 14q12 to the telomere of Chr14. On the bottom from left to right, IGV screenshots of nanopore reads showing the methylation status at known maternally methylated RANBP20P, approximate breakpoint region within PRKD1, paternally methylated IG-DMR and MEG3, and maternally methylated MEG8 imprinted regions. Identifying exact breakpoints was not possible as the reads spanning the breakpoints were in a difficult-to-map region. However, the presence of two haplotypes followed by the loss of allelic phasing as a result of the loss of heterozygous variants is present in the figure. In the nanopore reads, red indicates methylated CpGs and blue indicates unmethylated CpGs. BAF, B allele frequency; Chr14, chromosome 14; CN, copy number; IGV, Integrative Genomics Viewer; UPD, uniparental disomy. The gap with no data points in the CN plot is the region ignored by the CN caller, Delly, based on the required GRCh38 mappability map file.