PRRX1 Loss‐of‐Function Mutations Underlying Familial Atrial Fibrillation

Abstract

Background Atrial fibrillation (AF) is the most common form of clinical cardiac dysrhythmia responsible for thromboembolic cerebral stroke, congestive heart failure, and death. Aggregating evidence highlights the strong genetic basis of AF. Nevertheless, AF is of pronounced genetic heterogeneity, and in an overwhelming majority of patients, the genetic determinants underpinning AF remain elusive. Methods and Results By genome‐wide screening with polymorphic microsatellite markers and linkage analysis in a 4‐generation Chinese family affected with autosomal‐dominant AF, a novel locus for AF was mapped to chromosome 1q24.2–q25.1, a 3.20‐cM (≈4.19 Mbp) interval between markers D1S2851 and D1S218, with the greatest 2‐point logarithm of odds score of 4.8165 for the marker D1S452 at recombination fraction=0.00. Whole‐exome sequencing and bioinformatics analyses showed that within the mapping region, only the mutation in the paired related homeobox 1 ( PRRX1 ) gene, NM_022716.4:c.319C>T;(p.Gln107*), cosegregated with AF in the family. In addition, sequencing analyses of PRRX1 in another cohort of 225 unrelated patients with AF revealed a new mutation, NM_022716.4:c.437G>T; (p.Arg146Ile), in a patient. The 2 mutations were absent in 908 control subjects. Biological analyses in HeLa cells demonstrated that the 2 mutants had significantly diminished transactivation on the target genes ISL1 and SHOX2 and markedly decreased ability to bind the promoters of ISL1 and SHOX2 (2 genes causally linked to AF), although with normal intracellular distribution. Conclusions This study first indicates that PRRX1 loss‐of‐function mutations predispose to AF, which provides novel insight into the molecular pathogenesis underpinning AF, implying potential implications for precisive prophylaxis and management of AF.

A novel PRRX1 loss-of-function variation contributing to familial atrial fibrillation and congenital patent ductus arteriosus

“…Notably, in PRRX1 -knockout mice, cardiovascular developmental malformations occurred, encompassing awkward curvature and abnormal positioning of the aortic arch, an aberrant retro-esophageal right subclavian artery as well as a misdirected and elongated ductus arteriosus, highlighting the crucial role of PRRX1 in the proper development of vessels and perivascular matrices ( Bergwerff et al , 2000 ). Moreover, a recent study has demonstrated that PRRX1 physically binds to the promoters of SHOX2 and ISL1 and transcriptionally activates the expression of SHOX2 and ISL1 ( Guo et al , 2021 ), two key downstream target genes responsible for the normal development of the heart, especially for its pacing and conducting system ( Blaschke et al , 2007 ; Liang et al , 2015 ; Vedantham et al , 2015 ; Galang et al , 2020 ) and variations in both SHOX2 and ISL1 have been causally linked to AF and CHD ( Blaschke et al , 2007 ; Hoffmann et al , 2016 , 2019 ; Li et al , 2018 ; Ma et al , 2019 ; Wang et al , 2019 ; Wu et al , 2020 ). In the present research, a new PRRX1 loss-of-function variation was discovered to lead to AF and PDA.…”

“…The recombinant eukaryotic expression plasmid PRRX1 -pcDNA3.1 expressing human wild-type PRRX1 was constructed as previously described ( Guo et al , 2021 ). The Glu125 * -mutant PRRX1 -pcDNA3.1 was generated by site-targeted mutagenesis utilizing a complimentary pair of primers (forward primer: 5'-GATGCTTTTGTGCGATAAGACCTTGCCCGCC-3'; reverse primer: 5'-GGCGGGCAAGGTCTTATCGCACAAAAGCATC-3') and a site-directed mutagenesis kit (Stratagene, USA) following the manufacturer’s instructions.…”

“…The Glu125 * -mutant PRRX1 -pcDNA3.1 underwent selection by DpnI (NEB, Hitchin, UK) and was confirmed by sequencing analysis. The SHOX2 -luciferase ( SHOX2 -luc) and ISL1 -luciferase ( ISL1 -luc) reporter plasmids, which both express firefly luciferase, were created as described elsewhere ( Guo et al , 2021 ).…”

“…Hela cells were grown in a 12-well plate at an initial density of 1×10 5 cells per well 24 h before transient transfection. Cells were transfected with various amounts of expression plasmids as described previously ( Guo et al , 2021 ). The plasmid pGL4.75 (Promega, USA) expressing renilla luciferase was co-transfected as an internal control to normalize transfection efficiency.…”

“…Up to now, in addition to the association of ~140 genetic loci with increased predisposition to AF revealed by genome-wide association studies ( Kim et al , 2021 ), rare variations in over 50 distinct genes have been discovered to contribute to AF, amidst which the majority encode cardiac potassium ion channels, sodium channels, gap junction channels, calcium channels, signaling molecules, structural proteins and transcription factors ( Choi et al , 2020 ; Ghazizadeh et al , 2020 ; Hansen et al , 2020 ; Huang et al , 2020 ; Jiang et al , 2020 ; Ragab et al , 2020 ; Roselli et al , 2020 ; van Ouwerkerk et al , 2020 ; Wu et al , 2020 ; Yang et al , 2020 ; Chalazan et al , 2021 ; Lazarte et al , 2021 ; Li et al , 2021a , b ; Ziki et al , 2021 ). Interestingly, multiple variations in or near the PRRX1 gene, has recently been associated with an enhanced susceptibility to AF in humans ( Tucker et al , 2017 ; Guo et al , 2021 ; Wu et al , 2021 ). However, due to pronounced genetical heterogeneity, the genetic determinants underlying AF remain largely elusive.…”