The 2012 Nobel Prize in Physiology and Medicine was jointly awarded to Shinya Yamanaka and John Gurdon for their discovery that mature cells can be reprogrammed to take on a stem-cell-like state. These reprogrammed cells, known as induced pluripotent stem cells (iPSCs), regain the ability to differentiate into a range of cell types in the body.
However, these iPSCs are not totipotent—they are unable to form the placenta. By this definition, even embryonic stem cells (ESCs) are not totipotent. The classification of totipotency is therefore reserved strictly for cells formed during the earliest stages of embryonic development (the zygote and two- to four-cell stages).
“Totipotent stem cells are the most versatile of the stem cell types,” said Wee-Wei Tee, a Principal Investigator at A*STAR's Institute of Molecular and Cell Biology (IMCB). “In this study, we found that we can activate the totipotent state simply by tweaking the gene expression and metabolic programs of ESCs.”
Tee’s team first analyzed the complete gene expression profiles, or transcriptomes, of pre-implantation mouse embryos at various development stages. Using a hierarchical clustering algorithm, the researchers were able to identify stage-specific gene signatures, allowing them to focus their attention on genes expressed at the two-cell stage of embryonic development.
“This unbiased approach led us to uncover, for the first time, a maternal factor called negative elongation factor A (NELFA) that is involved in totipotent gene expression,” Tee explained.
The researchers then overexpressed NELFA in mouse ESCs and observed that genes associated with totipotency were upregulated. They found that the NELFA protein interacted with another protein, Top2a, to activate the transcription factor Dux, which is responsible for increasing the expression of the totipotency gene set.
Further delving into the biological processes altered by NELFA in mouse ESCs, the researchers noticed that glycolysis—the pathway by which cells break down sugar—was suppressed during the two-cell stage of embryonic development. Suspecting that this was a characteristic of totipotent cells, the researchers decided to inhibit glycolysis in ESCs using a chemical that mimics glucose.
They found that the inhibition of glycolysis indeed caused ESCs to revert to a totipotent-like state. Concordantly, NELFA expression was increased in glycolysis-inhibited ESCs. These findings suggest that NELFA is a master regulator of totipotency.
“Discovering this method of inducing totipotency in cells allows us to engineer cells with maximum cell plasticity, thus increasing the potential applications of regenerative medicine,” said Tee.
Moving forward, Tee and colleagues intend to identify non-genetic means, such as the use of small molecules and specific culture conditions, to induce and stably propagate totipotent cells from a variety of cell types.
“Non-transgene approaches are important for cell replacement therapies,” Tee noted.
The A*STAR-affiliated researchers contributing to this research are from the Institute of Molecular and Cell Biology (IMCB), the Bioprocessing Technology Institute (BTI) and the Institute of Medical Biology (IMB).