Recessive TAF1A mutations reveal ribosomopathy in siblings with end-stage pediatric dilated cardiomyopathy

Abstract

Non-ischemic dilated cardiomyopathy (DCM) has been recognized as a heritable disorder for over 25 years, yet clinical genetic testing is non-diagnostic in >50% of patients, underscoring the ongoing need for DCM gene discovery. Here, whole exome sequencing uncovered a novel molecular basis for idiopathic end-stage heart failure in two sisters who underwent cardiac transplantation at three years of age. Compound heterozygous recessive mutations in TAF1A, encoding an RNA polymerase I complex protein, were associated with marked fibrosis of explanted hearts and gene-specific nucleolar segregation defects in cardiomyocytes, indicative of impaired ribosomal RNA synthesis. Knockout of the homologous gene in zebrafish recapitulated a heart failure phenotype with pericardial edema, decreased ventricular systolic function, and embryonic mortality. These findings expand the clinical spectrum of ribosomopathies to include pediatric DCM.

Figure 1

Identification of recessive TAF1A mutations in a sibling pair with DCM. (A) Family pedigree. Square, male; circle, female; solid, affected; open, unaffected; black font, age at screening echocardiography; red font, age at diagnosis; arrow, proband. (B) An iterative filtering scheme of whole exome sequencing variant calls identified a single candidate gene, TAF1A. (C) Sanger sequencing verified compound heterozygous missense mutations. (D) Conservation of L84, G341, and surrounding residues.

Figure 2

TAF1A-associated pathological and ultrastructural alterations. (A) Cross section of explanted heart from a normal male control, age 6, and the affected sibling pair, both 3 years of age at time of cardiac transplantation. Increased myocardial mass and extensive fibrosis was seen in both siblings. (B–D) Representative photomicrographs of Masson’s trichrome stained left ventricular myocardial tissue from a normal male control, age 13, a female DCM control, age 9, and the proband. Replacement fibrosis was observed, unique to the affected sibling pair. Scale bars, 200 µm (40X magnification). (E–G) Hematoxylin and eosin staining revealed marked cardiomyocyte hypertrophy in both siblings. Scale bars, 20 µm (400X magnification). (H–J) Transmission electron microscopy revealed nucleolar segregation that was unique to the affected sibling pair. Scale bars, 400 nm (30,000X magnification). (K), Nucleolar segregation quantification. In total, 60 nucleoli/group were examined (30 nucleoli/individual) and classified as segregated or non-segregated based on morphological appearance.

Figure 3

taf1a^−/− zebrafish recapitulate a heart failure phenotype. (A) Lateral view of WT and taf1a^−/− embryos at 6, 7, and 8 days post-fertilization, demonstrating pericardial edema that was absent in WT fish. (B) taf1a^−/− embryos exhibited a significant reduction in ventricular fractional shortening compared to WT (n = 5 per group, P<0.05). No difference was observed between taf1a^+/− and WT embryos (data not shown). (C) Kaplan-Meier survival curve demonstrates complete lethality in taf1a^−/− fish by 11 days post-fertilization (WT = 65, taf1a^−/− =64; P < 0.001). (D) Ribosome biogenesis. Ribosomal DNA is transcribed within the nucleolar compartment of the nucleus. Pre-ribosomal RNA is processed into mature ribosomal RNA, and then assembled into ribosomal subunits before being exported to the cytoplasm for protein translation. In cardiomyocytes, nascent proteins are continuously integrated into the existing sarcomere. Loss-of-function of TAF1A and impaired rRNA synthesis could result in an imbalance between protein synthesis and degradation.