209 Rho signaling-directed regulation of YAP/TAZ in parthenogenetic stem cells

Abstract

Mammalian oocyte maturation is characterised by asymmetric meiotic division that is regulated by specific cytoskeleton organisation. Similarly, during early embryonic divisions, one of the most important steps is the establishment of polarity that allows cells to adopt distinct developmental fates. All of these events are driven by dynamic changes in actin filaments. It has been demonstrated recently that the Rho signalling pathway plays a key role in the organisation and rearrangement of actin-containing structures, regulating cell polarity and migration. In addition, beside its effect on cell cytoskeleton, Rho directly interacts with the Hippo pathway, influencing both embryonic cell proliferation and differentiation. Because both Rho and Hippo are expressed by the oocyte and maternally inherited (Zhang et al. 2014 Cell Cycle 13, 3390-3403, https://doi.org/10.4161/15384101.2014.952967; Menchero et al. 2017 Dev. Dyn. 246, 245-261, https://doi.org/10.1002/dvdy.24471), we investigated their regulation in parthenogenetic embryonic stem cells (ParthESC) that possess exclusively maternal genetic material, and compared the results with biparental ESCs. Previous results obtained by whole-transcriptome analysis revealed the presence of several differentially expressed genes involved in the Rho pathway and showed no differences for most of the Hippo signalling genes. To better elucidate the molecular mechanisms involved, in the present study, we dissected the expression pattern of the Rho and Hippo regulatory genes in human biparental ESCs and ParthESC. Experiments were performed on 4 biparental ESC and 4 ParthESC lines using cells between passages 5 to 25. The results showed significantly increased transcription of the Rho GTPase family genes (RHOA, RHOB, and RHOC) in ParthESC compared with biparental ESCs. Consistent with this, 12 of 17 Rho activators were significantly upregulated, whereas 8 of 11 Rho inhibitors were significantly decreased in ParthESC. Furthermore, monoparental cells displayed significantly higher expression levels of YAP and TAZ, whereas the upstream genes involved in the Hippo pathway (LATS1/2, MOB1, MST1/2, NF2) were comparable in the two cell types. Interestingly, a significantly higher total YAP protein content was detected in ParthESC, whereas the quantity of the phosphorylated form was comparable in the two cell types. This accounts for the observed upregulation of Rho genes, which stimulate the assembly of contractile actin stress fibres, inhibiting LATS1/2 phosphorylation and preventing subsequent phosphorylation of YAP/TAZ (Yu and Guan 2013 Genes Dev. 27, 355-371; https://doi.org/10.1101/gad.210773.112). Altogether, our results suggest that the Rho pathway may regulate YAP/TAZ behaviour via a LATS/MST/NF2-independent process in ParthESC, similarly to a previous report in oocytes (Posfai and Rossant 2016 Cell Res. 26, 393-394; https://doi.org/10.1038/cr.2016). Although further clarifications are needed, we hypothesise that the regulatory mechanisms detected in ParthESC may be related to their strictly maternal origin, with a possible impact on their plasticity and potency.

This study was supported by Carraresi Foundation. Authors are members of the COST Actions CA16119.