Title:
Studies on mouse peri-implantation development using developmental mutant and stem cells
Speaker:
Kuniya Abe, Ph.D.
Mammalian Genome Dynamics, RIKEN BioResource Center, Tsukuba, Ibaraki, Japan
Abstract:
In early mammalian embryos, there exist undifferentiated cells possessing ability to become various kinds of differentiated cells. Inner cell mass of preimplantation blastocyst and epiblast of postimplantation embryos are such pluripotential cells. In order to understand factors/ genes essential for growth and differentiation of these pluripotential cells, we have been using mouse developmental mutants and stem cells derived from these pluripotent cells. tw5 is one of such developmental mutants mapped in mouse t-complex region on chromosome 17. tw5 homozygotes die around embryonic day 6.5, showing specific defects in epiblast. Our genetic study has narrowed down the tw5 critical region to 750-kb, and identified a candidate gene,Vps52, and confirmed its authenticity by various means. Vps52 is the mouse homolog of yeast VPS52 involved in the retrograde trafficking of endosomes. Its function in the development of multicellular organisms has been totally unknown. Our data strongly suggest that via cell–cell interactions, Vps52 acts in extraembryonic tissues, most likely in the visceral endoderm, to support the developmental transition of pluripotent cells and growth.
Pluripotent cells can be cultured in appropriate conditions and propagated indefinitely in vitro, and termed pluripotent stem cells (PSC). There are at least two kinds of PSCs in mammals, i.e. naïve and primed PSCs. Mouse embryonic stem cells (mESCs) are naïve PSC, isolated from pre-implantation embryos. Mouse epiblast stem cells (EpiSCs) are derived from epiblast of post-implantation embryos and classified as primed PSC. Interestingly, EpiSCs share many characteristics with human PSCs such as human ES cells (hESC) and human induced pluripotent stem cells (hiPSC). Because of enormous advantages of mice as an experimental system, EpiSCs should serve as a useful model for studying primed states of pluripotency. However, studies using the EpiSCs are still limited relative to mouse ESCs, partly due to technical difficulties in derivation and maintenance of EpiSCs.
Here we have devised a simple yet highly efficient protocol for EpiSC derivation and maintenance of homogenous, high-quality EpiSCs using inhibitor of Wnt signaling. Using this method, EpiSCs can be readily derived from any mouse strains tested. The EpiSCs derived by this protocol maintain homogenous, undifferentiated status, yet retain high differentiation potential. Unlike EpiSCs established by the original protocol, the new EpiSC lines requires the continued presence of the WNT inhibitor, suggesting intrinsic differences from the EpiSCs made by the original method. This new version of EpiSCs will provide clues to understand nature of primed states of mammalian pluripotent cells and open up a new avenue for stem cell applications.
References:
Sugimoto, M., Kondo, M., Hirose, M., Suzuki, M., Mekada, K., Abe, T., Kiyonari, H., Ogura, A., Takagi, N., Artzt, K. and Abe, K. (2012). Molecular identification of tw5: Vps52 promotes pluripotential cell differentiation through cell-cell interactions. Cell Reports 2, 1363-1374.
Sugimoto, M., Kondo, M., Koga, Y., Shiura, H., Ikeda, R., Hirose, M., Ogura, A., Murakami, A., Yoshiki, A., Chuva de Sousa Lopes, S., Abe, K. (2015). A simple and robust method for establishing homogeneous mouse epiblast stem cell lines by Wnt inhibition. Stem Cell Reports 4, 744-757.
