Abstract
Leukemia Inhibitory Factor (LIF)/Signal transducer and activator of transcription 3 (STAT3) signaling pathway maintains the stemness and pluripotency of mouse embryonic stem cells (mESCs). Detailed knowledge on key intermediates in this pathway as well as any parallel pathways is largely missing. We initiated our study by investigating the effect of small molecule Curcumin on various signalling pathways essential for self-renewal. Curcumin sustained the LIF independent self-renewal of mESCs and induced pluripotent stem cells (miPSCs) in a STAT3 activity dependent manner. Gene expression analysis showed LIF/STAT3 and redox signaling components to be majorly modulated. Amongst ROS genes, expression of Manganese Superoxide Dismutase (MnSOD) specifically relied on STAT3 signaling as evidenced by STAT3 inhibition and reporter assay. The silencing of MnSOD, but not Cu-ZnSOD expression, resulted in the loss of mESC pluripotency in presence of LIF, and the overexpression of MnSOD is sufficient for maintaining the expression of pluripotent genes in the absence of STAT3 signaling. Finally, we demonstrate MnSOD to stabilize the turnover of pluripotent proteins at the post-translational level by modulating proteasomal activity. In conclusion, our findings unravel a novel role of STAT3 mediated MnSOD in the self-renewal of mESCs.
Highlights
Leukemia Inhibitory Factor (LIF)/Signal transducer and activator of transcription 3 (STAT3) signaling pathway maintains the stemness and pluripotency of mouse embryonic stem cells
At passage 1 there was no significant difference between Curcumin treated and untreated cells, during later passages (15th passage), ESCs cultured in absence of LIF lost the expression of pluripotent genes and cells cultured without LIF but in presence of 5 mM Curcumin significantly maintained the expression of pluripotent genes similar to that of mouse embryonic stem cells (mESCs) cultured in presence of LIF (Fig. 1a, Supplementary Fig. 1d)
To rule out the contribution of feeders in the expression of pluripotent genes, mESCs were cultured in the presence of Curcumin and in the absence of LIF and feeders. 5 mM Curcumin treated cells maintained the compact morphology of mESCs similar to the LIF treated cells (Fig. 1d)
Summary
Leukemia Inhibitory Factor (LIF)/Signal transducer and activator of transcription 3 (STAT3) signaling pathway maintains the stemness and pluripotency of mouse embryonic stem cells (mESCs). Many of the signaling pathways mentioned above have been reported to play key role in maintaining self-renewal or differentiation of ESCs23 It is unclear about the role of Curcumin in self-renewal or differentiation of mESCs and the signalling pathways adopted by this small molecule in exerting its effect. To address this question we evaluated the efficacy of Curcumin to maintain mESC cultures in LIF and feeder free culture conditions. By this approach and in combination with gene expression profiling, we demonstrate Curcumin to impart its effect on self-renewal of mESCs by modulating STAT3 mediated MnSOD expression.
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