Inhibition of protein kinase D by CID755673 promotes maintenance of the pluripotency of embryonic stem cells

The identification of novel mechanisms to maintain embryonic stem cell (ESC) pluripotency is of crucial importance, because the currently used culture conditions are not suitable for ESCs from all species. In this study, we show that the protein kinase D (PKD) inhibitor CID755673 (CID) is able to maintain the undifferentiated state of mouse ESCs in combination with the mitogen-activated protein kinase kinase (MEK) inhibitor. The expression levels of PKD members, including PKD1, PKD2 and PKD3, were low in mouse ESCs but significantly increased under differentiation conditions. Therefore, depletion of three PKD genes was able to phenocopy PKD inhibition. Mechanistically, PKD inhibition activated PI3K/AKT signaling by increasing the level of AKT phosphorylation, and the addition of a PI3K/AKT signaling pathway inhibitor partially reduced the cellular response to PKD inhibition. Importantly, the self-renewal-promoting effect of CID was maintained in human ESCs. Simultaneous knockdown of the three human PKD isoforms enabled short-term self-renewal in human ESCs, whereas PI3K/AKT signaling inhibition eliminated this self-renewal ability downstream of the PKD inhibitor. These findings expand our understanding of the gene regulatory network of ESC pluripotency.