Disruption of KCNQ1 prevents methylation of the ICR2 and supports the hypothesis that its transcription is necessary for imprint establishment

Abstract

Beckwith-Wiedemann syndrome (BWS; OMIM #130650) is an imprinting disorder caused by genetic or epigenetic alterations of one or both imprinting control regions on chromosome 11p15.5. Hypomethylation of the centromeric imprinting control region (KCNQ1OT1:TSS-DMR, ICR2) is the most common molecular cause of BWS and is present in about half of the cases. Based on a BWS family with a maternal deletion of the 5' part of KCNQ1 we have recently hypothesised that transcription of KCNQ1 is a prerequisite for the establishment of methylation at the KCNQ1OT1:TSS-DMR in the oocyte. Further evidence for this hypothesis came from a mouse model where methylation failed to be established when a poly(A) truncation cassette was inserted into this locus to prevent transcription through the DMR. Here we report on a family where a balanced translocation disrupts the KCNQ1 gene in intron 9. Maternal inheritance of this translocation is associated with hypomethylation of the KCNQ1OT1:TSS-DMR and BWS. This finding strongly supports our previous hypothesis that transcription of KCNQ1 is required for establishing the maternal methylation imprint at the KCNQ1OT1:TSS-DMR.

Fig. 1

Overview of the imprinted domains on chromosome 11p15.5. Given are the two imprinting control regions (ICRs) on chromosome 11p15.5. The centromeric KCNQ1OT1:TSS-DMR is methylated on the maternal allele (grey square). It regulates the expression of the maternally expressed genes KCNQ1, KCNQ1DN, CDKN1C and SLC22A18 (depicted in red) and the paternally expressed KCNQ1OT1 (blue). The biallelically expressed gene PHLDA2 is shown in black. The telomeric H19/IGF2:IG-DMR is methylated on the paternal allele with the maternally expressed H19 and the paternally expressed IGF2 gene. Tel telomeric, cen centromeric. Modified from Beygo et al. 2016

Fig. 2

a Schematic view of the chromosomal rearrangement and b detail view of the disruption of KCNQ1 and KCNQ1OT1. In part a the chromosomes 13 (purple) and 11 (green, 11p dark green, 11q lighter green) are given in the normal state (outer chromosomes) and with the rearrangement detected in the family (inner chromosomes). The terminal part of 13q is translocated to 11q while the terminal part of 11p is translocated to 13q. Furthermore, the biggest part of the derivative chromosome 11 is inverted (indicated by arrows). No large deletions or duplications associated with the breakpoints have been detected so that the rearrangement is balanced. The breakpoints are localised in 13q21.33, 11p15.5 and 11q22.1 and were confirmed by junction PCRs. The disrupted gene KCNQ1 is indicated as red boxes. Not drawn to scale. Part b shows the detailed view of the maternally expressed KCNQ1 (red) and its antisense transcript, the paternally expressed KCNQ1OT1 (blue). The imprinting control region 2 (ICR2, KCNQ1OT1:TSS-DMR) is given as a grey square. Direction of the transcription is indicated by arrows. From the literature the deletion described in Beygo et al. 2016, the duplication described in Demars et al. 2011 (grey bars) and the region containing the translocation breakpoint described in Kaltenbach et al. 2013 are depicted (gashed grey bar). Tel telomeric, cen centromeric. Modified from Beygo et al. 2016. The translocation of 11p material to chromosome 13q disrupts both transcripts (indicated by zigzag line) and is located in intron nine of KCNQ1. The 5’ part of the gene is translocated to chromosome 13 while the 3’ part including the ICR2 stays on chromosome 11p. The disruption prevents transcription of KCNQ1 through the ICR2. Not drawn to scale. Tel telomeric, cen centromeric. Figure modified from Beygo et al. 2016

Fig. 3

Pedigree of the family including a variant showing the grandparental inheritance of the derivative chromosome 11. The figure depicts the family pedigree with the results of Sanger-sequencing of the breakpoint region 2 showing the grandparental inheritance of the derivative chromosome 11. The grandfather and the grandmother have two normal chromosomes. The grandfather is heterozygous for a private variation (A > G), the grandmother is homozygous A. For the mother and the fetus (III-4) only the derivative chromosome 11 was amplified using primers on each side of the breakpoint (BP2). The derivative chr.11 in the mother shows the G-allele indicating that the rearrangement involves the paternal allele. This is in accordance with her normal phenotype. She passed the derivative chr.11 on to her child (III-4) indicated again by the G-allele