Transcription factors (TFs), through their gene regulation activity, are known to be powerful regulators of organogenesis, including the heart. IRX5 TF belongs to the Iroquois homeobox (IRX) family and studies in mice have shown that it is expressed from the early stages of cardiac development throughout adulthood. Irx5 has also been shown to play a crucial role in establishing the ventricular repolarization gradient through the modulation of certain cardiac genes: Kcnd2 in mice and SCN5A in humans. However, these studies are gene-candidate based, therefore the global target gene-set of IRX5 and its protein partners in human cardiomyocytes is still to be identified.

The aim of this project is to identify all the IRX5 target genes and its protein partners in human cardiomyocytes, and to study its functions during cardiomyocyte differentiation.

In order to perform studies in a human context, we used cardiomyocytes derived from induced pluripotent stem cells (iPS-CMs), generated from patients with cardiac defects linked to IRX5 loss of function mutations and healthy individuals. High-throughput transcriptomic experiments as well as immunoprecipitation and luciferase analyses were performed to obtain a global understanding of IRX5 functions and partners.

We have shown that IRX5 can interact with major cardiac TFs such as GATA4, TBX5 and NKX2-5. More precisely, IRX5 interacts with them as a multi-protein complex to modulate the expression of target genes. We demonstrated the central role of a conserved region of IRX5 in these interactions: its homeodomain. We have also identified misregulated genes between control and IRX5^Mut iPS-CMs, associated to the cardiac contraction and fibrosis pathways.

IRX5 is an important actor of the regulation of human cardiac functions, and these regulations seem to require the establishment of a multiprotein complex with other major cardiac TFs.