Localized translation and sarcomere maintenance requires ribosomal protein SA in mice
Rami Haddad, Omer Sadeh, Tamar Ziv, Itai Erlich, Lilac Haimovich-Caspi, Ariel Shemesh, Jolanda van der Velden, Izhak Kehat
Rami Haddad, Omer Sadeh, Tamar Ziv, Itai Erlich, Lilac Haimovich-Caspi, Ariel Shemesh, Jolanda van der Velden, Izhak Kehat
View: Text | PDF
Research Article Cardiology

Localized translation and sarcomere maintenance requires ribosomal protein SA in mice

Abstract

Cardiomyocyte sarcomeres contain localized ribosomes, but the factors responsible for their localization and the significance of localized translation are unknown. Using proximity labeling, we identified ribosomal protein SA (RPSA) as a Z-line protein. In cultured cardiomyocytes, the loss of RPSA led to impaired local protein translation and reduced sarcomere integrity. By employing CAS9-expressing mice, along with adeno-associated viruses expressing CRE recombinase and single-guide RNAs targeting Rpsa, we knocked out Rpsa in vivo and observed mislocalization of ribosomes and diminished local translation. These genetic mosaic mice with Rpsa knockout in a subset of cardiomyocytes developed dilated cardiomyopathy, featuring atrophy of RPSA-deficient cardiomyocytes, compensatory hypertrophy of unaffected cardiomyocytes, left ventricular dilation, and impaired contractile function. We demonstrated that RPSA C-terminal domain is sufficient for localization to the Z-lines and that if the microtubule network is disrupted RPSA loses its sarcomeric localization. These findings highlight RPSA as a ribosomal factor essential for ribosome localization to the Z-line, facilitating local translation and sarcomere maintenance.

Authors

Rami Haddad, Omer Sadeh, Tamar Ziv, Itai Erlich, Lilac Haimovich-Caspi, Ariel Shemesh, Jolanda van der Velden, Izhak Kehat

×

Figure 2

Proximity labeling with Cypher-BioID2 identifies Z-line proteins and components of the translational machinery.

Options: View larger image (or click on image) Download as PowerPoint
Proximity labeling with Cypher-BioID2 identifies Z-line proteins and com...
(A) Volcano plot of proteins identified by mass spectrometry showing the log2 fold change in protein abundance with Cypher-BioID2 versus BioID2 alone and the t test –log10 (P value) of the difference significance. Dashed horizontal lines show the ≥2 and ≤2 cutoffs for fold change, and the vertical dashed bar shows the P < 0.05 cutoff for significance. Proteins highlighted in red are those with a fold change ≥2 and P < 0.05 that were used for further pathway enrichment analysis. Other proteins were denoted as nonsignificant and are shown in gray. Several proteins identified as significant are highlighted. (B) Pathway enrichment analysis of BioID2 significantly enriched proteins (fold change ≥2 and P < 0.05) showing fold enrichment of GO terms associated with cytoskeletal elements of the sarcomere and with ribosomes and protein translation. The number of proteins identified for each term is shown as a circle, and the –log10 of the false discovery rate (FDR) is color coded. (C) Unsupervised clustering of enriched GO terms showing identified proteins were predominantly associated with two groups — cytoskeletal sarcomere proteins or ribosomal components. The enrichment FDR of specific GO terms is shown. (D and E) Representative immunofluorescence images (D) and line scan (E, white line) analysis of isolated adult rat ventricular cardiomyocytes stained for α-actinin (red), RPSA (green), and DAPI (blue), showing localization of endogenous RPSA to both sides of the Z-line identified with α-actinin. Scale bar: 10 μm.