Background and Hypothesis: Loss of S100A1 expression in heart failure contributes to adverse cardiomyocyte (CM) remodeling via an unknown molecular mechanism. Conversely, the antihypertrophic effect of viral-based restoration of S100A1 expression in diseased hearts is not understood yet. We hypothesized that S100A1 may directly interfere with mRNA translation in CMs.

Methods and Results: An unbiased mass spectrometry/affinity purification approach using human recombinant S100A1 protein as a bait and murine S100A1^-/- knock-out CM homogenates as prey identified the translation initiation factors EIF4G, EIF2b, RPS6 and PABP as targets for Ca²⁺-bound (holo) S100A1 protein, amongst other translational regulators. Computational modeling revealed top-scoring binding modes for holo-S100A1’s C-terminus in close proximity to RPS6’s ser-235/236 phosphorylation site harboring domain, and PLA detected S100A1/RPS6 complexes in neonatal ventricular cardiomyocytes (NVCMs). siRNA silencing of S100A1 in NVCMs caused a significant increase in cell size (+44%*) and fetal gene expression (e.g. MYH7 4.2-fold*) with a prompt enhancement of the de-novo protein synthesis rate (+89%*), as assessed by IF, RT-PCR and puromycin incorp. up to 48h (n=3-5, *P<0.05 vs contr). Although phospho-specific IB revealed an increase in RPS6-ser235/236 phosphorylation (+38%, n=10, P<0.05) due to S100A1 knock-down, the activity of the upstream Akt/mTORC2 pathway remained unaltered. A nascent proteome analysis unveiled the significant increase and decrease in the de-novo translation of 554 and 17 proteins (FC>1.5), respectively, in S100A1-silenced NVCMs vs contr. with e.g. MYH7, ALC-1, ACTN2 or Nppb in the top 10 newly synthesized proteins. Heart muscle contraction and metabolism-related GO-terms (e.g. myofibril assembly or FA oxidation and OXPHOS) characterized the top 100 upregulated proteins.

Conclusion: This is the first study providing evidence for an impact of S100A1 on the mRNA translation machinery in CMs by inhibiting translation of contractile and mitochondrial protein programs required for hypertrophic growth. S100A1 may mechanistically interfere with molecular factors of the translation initiation machinery, which warrants further investigation.