TRPC3 is required for the survival, pluripotency and neural differentiation of mouse embryonic stem cells (mESCs)
Transient receptor potential canonical subfamily member 3 (TRPC3) is known to be important for neural development and the formation of neuronal networks. Here, we investigated the role of TRPC3 in undifferentiated mouse embryonic stem cells (mESCs) and during the differentiation of mESCs into neurons. CRISPR/Cas9-mediated knockout (KO) of TRPC3 induced apoptosis and the disruption of mitochondrial membrane potential both in undifferentiated mESCs and in those undergoing neural differentiation. In addition, TRPC3 KO impaired the pluripotency of mESCs. TRPC3 KO also dramatically repressed the neural differentiation of mESCs by inhibiting the expression of markers for neural progenitors, neurons, astrocytes and oligodendrocytes. Taken together, our new data demonstrate an important function of TRPC3 with regards to the survival, pluripotency and neural differentiation of mESCs.
Keywordstransient receptor potential canonical subfamily member 3 (TRPC3) mouse embryonic stem cells (mESCs) neuron differentiation CRISPR/Cas9 pluripotency apoptosis mitochondrial membrane potential
We thank Prof. Austin Smith (Cambridge Stem Cell Institute, Cambridge, UK), for kindly providing us with the 46C cells. We also thank Ms. Mandy Chan (HKUST, Hong Kong) for her technical support and Prof. Jacques Haiech (University of Strasbourg, France) for his helpful comments about the project. This work was supported by the Hong Kong Research Grants Council (RGC) General Research Fund awards (662113, 16101714, 16100115), the ANR/RGC joint research scheme award (AHKUST601/ 13), the Hong Kong Theme-based Research Scheme award (T13-706/11-1) and the Hong Kong Innovation and Technology Commission (ITCPD/17-9).
- Becker, E.B.E., Oliver, P.L., Glitsch, M.D., Banks, G.T., Achilli, F., Hardy, A., Nolan, P.M., Fisher, E.M.C., and Davies, K.E. (2009). A point mutation in TRPC3 causes abnormal Purkinje cell development and cerebellar ataxia in moonwalker mice. Proc Natl Acad Sci USA 106, 6706–6711.CrossRefPubMedPubMedCentralGoogle Scholar
- Fusco, F.R., Martorana, A., Giampà, C., De March, Z., Vacca, F., Tozzi, A., Longone, P., Piccirilli, S., Paolucci, S., Sancesario, G., Mercuri, N.B., Bernardi, G. (2004). Cellular localization of TRPC3 channel in rat brain: preferential distribution to oligodendrocytes. Neurosci Lett 365, 137–142.CrossRefPubMedGoogle Scholar
- Hartmann, J., Dragicevic, E., Adelsberger, H., Henning, H.A., Sumser, M., Abramowitz, J., Blum, R., Dietrich, A., Freichel, M., Flockerzi, V., Birnbaumer, L., Konnerth, A. (2008). TRPC3 channels are required for synaptic transmission and motor coordination. Neuron 59, 392–398.CrossRefPubMedPubMedCentralGoogle Scholar
- Louhivuori, L.M., Jansson, L., Turunen, P.M., Jäntti, M.H., Nordström, T., Louhivuori, V., and Åkerman, K.E. (2015). Transient receptor potential channels and their role in modulating radial glial-neuronal interaction: a signaling pathway involving mGluR5. Stem Cells Dev 24, 701–713.CrossRefPubMedGoogle Scholar
- Nagasaka, R., Matsumoto, M., Okada, M., Sasaki, H., Kanie, K., Kii, H., Uozumi, T., Kiyota, Y., Honda, H., and Kato, R. (2017). Visualization of morphological categories of colonies for monitoring of effect on induced pluripotent stem cell culture status. Reg Ther 6, 41–51.Google Scholar