Identification and Functional Investigation of SOX4 as a Novel Gene Underpinning Familial Atrial Fibrillation

Figures

• 

  Figure 1. Pedigree suffering from atrial fibrillation. The pedigree was arbitrarily termed as Family AF-01. “+” signifies a member harboring the discovered heterogeneous SOX4 mutation; “–” represents a non-carrier of the discovered SOX4 mutation.

• 

  Figure 2. A representative electrocardiogram recorded from the proband (II-7) of Family AF-01. The standard 12-lead electrocardiogram illustrates atrial fibrillation.

• 

  Figure 3. A novel SOX4 mutation predisposing to atrial fibrillation. (A) Sequencing electropherograms exhibiting the heterogeneous SOX4 mutation identified in the index patient with atrial fibrillation (mutant) along with its wild-type control detected in an unaffected subject (wild type). An arrow pinpoints where the mutation occurs. (B) Schematic drawings describing the main structural domains of SOX4. TAD: transcriptional activation domain; HMG: high mobility group.

• 

  Figure 4. A novel SOX4 mutation contributing to atrial fibrillation. (A) Sequencing electropherograms displaying the heterogeneous SOX4 mutation detected in one of the cohort patients with atrial fibrillation (mutant) and its wild-type control detected in a control individual (wild type). An arrow directs where the mutation occurs. (B) Schematic diagrams illustrating the main structural domains of SOX4. HMG: high mobility group; TAD: transcriptional activation domain.

• 

  Figure 5. A representative electrocardiogram from the patient harboring the SOX4 c.290G>A mutation. The electrocardiogram documents atrial fibrillation.

• 

  Figure 6. Functional failure of Gln71*- or Trp97*-mutant SOX4. In COS-7 cells cultivated in vitro, dual-reporter measurement of the transactivation of the GJA1/Cx43 promoter-driven firefly luciferase by wild-type SOX4 or Gln71*-mutant SOX4 (Gln71*) or Trp97*-mutant SOX4 (Trp97*), separately or in combination, unveiled that the Gln71* mutant (A) or Trp97* mutant (B) failed to transactivate GJA1/Cx43. Here “a” and “c” mark p < 0.001, and “b” and “d” indicate p < 0.01, when compared with wild-type SOX4 (400 ng).

• 

  Figure 7. No Synergistic Transactivation of SCN5A Between TBX5 and Gln71*- or Trp97*-Mutant SOX4. In cultured HEK293 cells, dual-reporter gene assessment of the synergistic transactivation of the SCN5A promoter by TBX5 and SOX4 demonstrated that the synergy was disrupted by the Gln71* (A) or Trp97* (B) mutation. Herein “e”, “f” and “g” denote p < 0.005, and “h” means p < 0.001 when compared with the corresponding wild-type counterparts.

• 

  Figure 8. Multiple SOX4-binding sites located within the promoter of GJA1/Cx43. Red color was used to mark the SOX4-binding sites; green color: the first exon mRNA; underlined bases in bold: primer sequences.

• 

  Figure 9. Multiple SOX4- and TBX5-binding sites mapped within the promoter of SCN5A. Red color was used to mark the SOX4-binding sites; blue color: the TBX5-binding sites; green color: the first exon mRNA; underlined bases in bold: primer sequences.