Novel Variant in the ANK2 Membrane-Binding Domain Is Associated With Ankyrin-B Syndrome and Structural Heart Disease in a First Nations Population With a High Rate of Long QT Syndrome

Abstract

Background— Long QT syndrome confers susceptibility to ventricular arrhythmia, predisposing to syncope, seizures, and sudden death. While rare globally, long QT syndrome is ≈15× more common in First Nations of Northern British Columbia largely because of a known mutation in KCNQ1 . However, 2 large multigenerational families were affected, but negative for the known mutation. Methods and Results— Long QT syndrome panel testing was carried out in the index case of each family, and clinical information was collected. Cascade genotyping was performed. Biochemical and myocyte-based assays were performed to evaluate the identified gene variant for loss-of-function activity. Index cases in these 2 families harbored a novel ANK2 c.1937C>T variant (p.S646F). An additional 16 carriers were identified, including 2 with structural heart disease: one with cardiomyopathy resulting in sudden death and the other with congenital heart disease. For all carriers of this variant, the average QTc was 475 ms (±40). Although ankyrin-B p.S646F is appropriately folded and expressed in bacteria, the mutant polypeptide displays reduced expression in cultured H9c2 cells and aberrant localization in primary cardiomyocytes. Furthermore, myocytes expressing ankyrin-B p.S646F lack normal membrane targeting of the ankyrin-binding partner, the Na/Ca exchanger. Thus, ankyrin-B p.S646F is a loss-of-function variant. Conclusions— We identify the first disease-causing ANK2 variant localized to the membrane-binding domain resulting in reduced ankyrin-B expression and abnormal localization. Further study is warranted on the potential association of this variant with structural heart disease given the role of ANK2 in targeting and stabilization of key structural and signaling molecules in cardiac cells.

The evolving role of ankyrin-B in cardiovascular disease

“…The loss-of-function variant was found to prohibit normal membrane targeting of NCX, presumably due to the altered localization of AnkB p.S646F. 38 Ichikawa et al also recently found possibly damaging variants in the MBD in isolated cases of arrhythmia. All other reported disease causing loss-of-function variants occur in the SBD, DD, or C-terminal domain.…”

“…Normal localization of NCX and AnkB was also found to be disrupted in the MBD variant p.S646F, indicating that NCX localization is regulated by two AnkB domains – MBD and RD. 38 ANK2 variants have also been found to cause sinus node disease (SND), and AnkB heterozygous (AnkB +/− ) mice phenocopy human SND patients, presumably due to loss of AnkB-mediated organization and signaling in sinoatrial node (SAN) cells. 34 Another variant in ANK2 that disrupts the AnkB/βII-spectrin interaction (AnkB p.R990Q) causes severe arrhythmia phenotypes.…”

“…65 Furthermore, seizures have been shown to correlate with AnkB-associated cardiac channelopathies. 38, 66, 67 Initially discovered in 1993 and 1995, the 440kDa AnkB (AnkB-440) and 480kDa AnkG (AnkG-480) isoforms, also known as giant ankyrins, are a result of the addition of a single large exon. 68, 69 The role of giant ankyrins in patterning the axon initial segment was demonstrated in Drosophila and was shown to be conserved across Bilatera.…”

“…While this finding is significant in itself, this group also reported the presence of congenital malformations associated with the AnkB p.S646F variant. 38 Interestingly, AnkB −/− mice suffer neonatal lethality 21 , which could be associated with a congenital heart defect. Additionally, these families displayed an incidence of cerebral aneurysms, suggesting a novel vascular role for AnkB.…”

Isolation and characterization of multi-protein complexes enriched in the K-Cl co-transporter 2 from brain plasma membranes

“…The subsequent association of either Sptbn1 (275 kDa), or Itpr1 (310 kDa) would shift the mass to around 800 kDa. Both Sptan1 and Ank2 have a variety of binding domains where interactions with Kcc2 could occur, such as SH3 domains (Sptan1) (61) and the membrane binding domain containing the Ank repeat sequences of Ank2 (62), which has been shown to interact with other ion channels/transporters (63).…”