Overexpression of proteins of interest by mRNA delivery in human induced pluripotent stem cell-derived cardiomyocytes

Human induced pluripotent stem cells-derived cardiomyocytes (hiPSC-CMs) are widely used systems for studying heart development, disease modelling and testing treatments. They are known examples of hard-to-transfect cells. In these cases, mRNA delivery improved the overexpression of a protein of interest with reduced toxicity. The aim of this work is to establish a protocol for in vitro transcription to obtain a synthetic mRNA encoding a protein of interest that will be overexpressed in hiPSC-CMs. We selected QKI-5 as an example of a target gene. It is the most expressed isoform of the Quaking (QKI) gene, which is essential for the maturation of hiPSC-CMs and is known to be downregulated in doxorubicin-induced cardiotoxicity. We successfully generated mRNA by in vitro transcription, ensuring high quality and amount for small-scale transfection experiments. The next step will be to overexpress QKI-5 in iPSC-CMs. Overall, although some optimization is still required for complete and stable mRNA production, this workflow can be applied to virtually any other gene of interest in the future.