Does XEN Cell Epigenetic Memory Influence Reprogramming Outcomes?

Undergraduate #12
Discipline: Biological Sciences
Subcategory: Cell and Molecular Biology
Session: 3
Room: Harding

Victor Gipson II - University of Georgia
Co-Author(s): Tayler Murphy, Michigan State University, East Lansing, MI; Dr. Amy Ralston, Michigan State University, East Lansing, MI

In 2006, Takahashi and Yamanaka published a paper detailing their success in deriving pluripotent stem cells from a non-embryonic source which were called induced pluripotent stem cells (iPS cells). In 2016, Parenti found that a new type of stem cell, dubbed induced extraembryonic endoderm cells (iXEN cells), were produced with iPS cells when reprogramming. The ratio between these cells is 1:3 iPS:iXEN. iXEN cells mimic naturally occurring extraembryonic endoderm cells (XEN cells) from the embryo. XEN and iXEN cells are multipotent. They can become yolk sac and visceral endoderm cell types. The goal is to determine why some cells become iPS cells and others become iXEN. This will help us understand what controls cell fate during development or reprogramming. My hypothesis is epigenetic memory influences reprogramming outcomes. If the hypothesis is correct, reprogramming XEN cells will produce more iXEN cells than iPS cells. I will derive mouse XEN cells with the rtTa Tet-on OSKM gene which makes the reprogramming factors inducible through doxycycline treatment. After reprogramming, I will utilize immunofluorescence and qPCR analyses to determine if XEN cells are primed to become iXEN cells due to epigenetic memory. We were not successful in deriving XEN cells from the embryos possibly because we tried deriving XEN cells too early. To address this, embryos will be outgrown longer before extraction.

Funder Acknowledgement(s): Michigan State University: The Graduate School

Faculty Advisor: Dr. Amy Ralston, aralston@msu.edu

Role: I did the embryo extraction from mice uterine horns and then their transfer to culture media, the culture of the embryos, extraembryonic endoderm (XEN) Cell Derivation Protocol, XEN cell culture, imaging of embryos, imaging of XEN cells, media making, reprogramming of mouse embryonic fibroblasts (MEFs), embryonic stem (ES) cell culture and imaging, genotyping of mice, etc.