Abstract Body (Do not enter title and authors here): Background: In regenerative hearts, robust cardiomyocyte (CM) proliferation was observed in mice that underwent myocardial infarction on postnatal (P) day 1 (MI_P1) and pigs that underwent P1 apical resection followed by P28 myocardial infarction (AR_P1MI_P28). Regulators promoting CM proliferation were discovered by analyzing transcriptomic data generated from these models. Most of these regulators supported mRNA production of cell-cycle machinery; yet the mRNA required translation into functional protein and required the activity of RNA-binding proteins (RBPs). The relationship between RBP and the cell cycle in CM is an open question.
Hypothesis: The RBP expressions strongly correlate with the CM cell cycle in mice, pigs, and humans.
Approach: We performed a meta-analysis to study the relationship between RBP expression and CM proliferation. In mouse and pig CM proliferation, the single-nuclei RNA-sequencing (snRNAseq) data from regenerating mouse (Gene Expression Omnibus GSE130699) and pig (GSE185289) hearts were reanalyzed via an Autoencoder with a focus on RBP expression. We also analyzed bulk RNA-sequencing, via DeSeq2 software, from two human-induced pluripotent stem cell-derived (hiPSC) cardiomyocyte (hiPSC-CM) cell lines (GSE289839); the first cell line was harvested sixteen days (hiPSC-CM-D16) after differentiation, when the cells still actively proliferated; the second cell line, hiPSC-CM-D140, was harvested when the cells ceased cell cycle. Then, the RBPs associated with mouse, pig, and hiPSC-CM were compared across species. Protein expressions of RBPs, which were consistently upregulated in proliferating CM among the three species, were examined in pig CMs via immunohistological analysis.
Result: Proliferating CM was retrieved by RBP-focused analysis, where five proliferation markers AURKB, MKI67, INCENP, CDCA8, and BIRC5 co-localized. Twenty-one RBPs were found to be consistently upregulated in proliferating mouse, pig, and hiPSC-derived CMs. Histological analysis showed a significant increase of DHX9 and PTBP3 in regenerative (AR_P1MI_P28P35), compared to non-regenerative (MI_P28P35) pig hearts.
Conclusion: The results demonstrated a strong relationship between RBP expression and CM proliferation.