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Cited2 in cardiac development: an inside and outside job

Publication . Santos, João Miguel Almeida; Bragança, José; Futschik, Matthias E.

Despite the remarkable knowledge acquired in the formation of the heart during embryonic development and the molecular mechanisms involved in heart function and physiology, there is no efficient way to prevent adult heart disease and congenital heart disease (CHD). The transcriptional modulator Cited2 is required for normal embryogenesis of mice and humans, particularly for heart development. Indeed, mouse lacking Cited2 alleles die in utero displaying many cardiovascular defects, and mutations in human CITED2 have long been associated with CHD. However, the exact role and the molecular mechanisms involving Cited2 during these processes are largely unknown. Using mouse Embryonic Stem Cells (ESC) as a model system, we have established that the depletion of Cited2 at the onset of differentiation resulted in a decline of ESC ability to generate cardiac cells. These cardiogenic defects in Cited2-depleted cells were rescued by treatment with a recombinant CITED2 protein. To further investigate the mechanisms caused by the loss of Cited2 in pluripotency and differentiation, we compared the gene expression profiles of control cells and Cited2-depleted cells upon differentiation. We determined that loss of Cited2 expression delays the expression of early mesoderm transcription factors and cardiopoietic factors. We found that the secretome of Cited2 overexpressing ESC is enough to restore the emergence of beating colonies in Cited2 depleted cells, upon differentiation. We identified WNT5a and WNT11 as two of the proteins enriched in the Conditioned Medium and crucial for rescuing cardiomyocyte differentiation defects caused by Cited2 depletion. Our results point that Cited2 is a co-transcriptional activator of Wnt5a and Wnt11 and that both proteins can restore cardiogenesis in Cited2-depleted cells. Additionally, using zebrafish as a model system, we demonstrated that WNT5a and WNT11 also rescued the development defects caused by Cited2 depletion in vivo. Collectively, our results show that WNT5a and WNT11 rescue cardiogenic defects caused by Cited2 depletion both in vitro, as well as in vivo.

2019-07-18Doctoral thesis

Open access