Bi-allelic missense disease-causing variants in RPL3L associate neonatal dilated cardiomyopathy with muscle-specific ribosome biogenesis

Abstract

Dilated cardiomyopathy (DCM) belongs to the most frequent forms of cardiomyopathy mainly characterized by cardiac dilatation and reduced systolic function. Although most cases of DCM are classified as sporadic, 20-30% of cases show a heritable pattern. Familial forms of DCM are genetically heterogeneous, and mutations in several genes have been identified that most commonly play a role in cytoskeleton and sarcomere-associated processes. Still, a large number of familial cases remain unsolved. Here, we report five individuals from three independent families who presented with severe dilated cardiomyopathy during the neonatal period. Using whole-exome sequencing (WES), we identified causative, compound heterozygous missense variants in RPL3L (ribosomal protein L3-like) in all the affected individuals. The identified variants co-segregated with the disease in each of the three families and were absent or very rare in the human population, in line with an autosomal recessive inheritance pattern. They are located within the conserved RPL3 domain of the protein and were classified as deleterious by several in silico prediction software applications. RPL3L is one of the four non-canonical riboprotein genes and it encodes the 60S ribosomal protein L3-like protein that is highly expressed only in cardiac and skeletal muscle. Three-dimensional homology modeling and in silico analysis of the affected residues in RPL3L indicate that the identified changes specifically alter the interaction of RPL3L with the RNA components of the 60S ribosomal subunit and thus destabilize its binding to the 60S subunit. In conclusion, we report that bi-allelic pathogenic variants in RPL3L are causative of an early-onset, severe neonatal form of dilated cardiomyopathy, and we show for the first time that cytoplasmic ribosomal proteins are involved in the pathogenesis of non-syndromic cardiomyopathies.

Fig. 1

Pedigrees, genetic and clinical characterization of three families with DCM carrying bi-allelic variants in RPL3L. a Pedigrees of families 1–3, genotypes and electropherograms of the identified RPL3L variants. All affected siblings carry compound heterozygous variants in RPL3L, while all parents were heterozygous carriers of one of the identified variants. Non-affected siblings were either homozygous for the wild-type allele (individual III.2, family 1) or heterozygous carrier of only one identified RPL3L variant (individual IV.2, family 2; individual III.1, family 3). n/a, DNA sample not available. b Echocardiogram in parasternal short-axis view of individuals III.1 (upper panel) and III.3 (lower panel) of family 1 including measurements and z-scores. Note end-diastolic dilation of the right ventricle and muscular hypotrophy of the septal and posterior wall. Asterisk and bold letter indicate pathological values. RVAW, right ventricular anterior wall. RVedD right ventricular end-diastolic diameter. IVSed intraventricular septum end-diastolic. LVedD left ventricular end-diastolic dimension. LVPWDd left ventricular posterior wall dimension diastole. c Pathological evaluation of the explant heart tissue of individual IV.3 (family 2). H & E staining of the myocardium revealed myocytolysis (green arrows) and fibrotic regions (black arrows)

Fig. 2

Molecular characterization of the identified RPL3L variants. a Schematic RPL3L protein structure and the localization of the identified variants. All missense variants are located within the conserved RPL3 domain of the protein. b Amino acid sequence alignment of RPL3L residues that are altered in the affected individuals including surrounding residues across different species. c Three-dimensional homology model of the RPL3L protein. RPL3L is shown in worm representation (yellow). Affected amino acid residues are labeled in black, shown in sphere representation and colored according to atom type (blue = nitrogen, white = carbon, red = oxygen). Portions of the 60S RNA in proximity to altered residues (R343, R161 and G27) are shown in ball-and-stick representation and colored according to atom type