In the early embryo of vertebrates, totipotent cells have the potential to differentiate and give rise to cells that function in specific tissues, ultimately forming an entire organism, including the extra-embryonic tissues, such as the placenta. This process of cell specification is controlled by the interplay of endogenous and exogenous factors (see page 713). At the blastocyst stage of the early embryo, the cells of the inner cell mass (from which embryonic stem (ES) cell lines are derived1,2
) are pluripotent: they are able to form each of the three germ layers — the endoderm, ectoderm and mesoderm. Eventually, cells that are committed to each of these germ layers specialize to give rise to the tissues of the adult body, such as the brain, intestine or cardiac muscle. These differentiated adult cells generally do not switch fates; for example, hepatocytes do not spontaneously become cardiomyocytes.
Several classic studies, however, suggested that ‘committed’ cells of the embryo are ‘plastic’, because the fate of these cells can change when they are explanted and exposed to a different microenvironment. In one of these studies, cells from the imaginal discs of Drosophila melanogaster
pupae were serially transplanted into the abdomen of an adult fly, and ‘transdetermination’ was observed: cells that were originally destined to form genital structures gave rise to leg or head structures and, eventually, on subsequent transplantations, to wings3,4
. Although such switches in cell fate occurred at a low frequency, these experiments by Hadorn3
provided evidence that explanted cells are surprisingly plastic. In another elegant study5
, cells were transplanted from quails to chickens: these cells were sufficiently similar to be able to participate in normal development on transplantation but were histologically distinct, enabling them to be tracked. Using this property, Le Lievre and Le Douarin5
showed that explanted neural crest cells adopt new fates (bone, cartilage and connective tissue) that are dictated by their new cellular neighbourhood and not their original location in the avian embryo. One caveat of these findings is that the fate of single cells could not be followed. But, as early as the mid-1960s, such embryonic cell transplantation experiments suggested that the generally stable state of a specialized cell was plastic and could be altered in response to the extracellular environment.
It was long thought that when a cell differentiates, it loses chromosomes or permanently inactivates genes that it no longer needs. Why would a specialized cell maintain the potential to reactivate genes typical of another cell type? This would seem to be a risky mechanism, given the possibility that genes could be inappropriately activated. Yet three approaches to nuclear reprogramming — nuclear transfer, cell fusion and transcription-factor transduction (described in detail below) — have shown conclusively in a defined specialized cell type (that is, in a cell that has been carefully determined to be differentiated) that cell fate can be reversed, returning the cell to an embryonic state (). These three experimental models therefore provide evidence that, with few exceptions (such as homologous recombination in lymphocytes), highly specialized somatic cells retain all of the genetic information that is needed for them to revert to ES cells and that the genes of the somatic cells have not been permanently inactivated. In addition, the three approaches show that, although the differentiated state of a cell is generally stable, cellular ‘memory’ is dynamically controlled and subject to changes induced by perturbations in the stoichiometry of the transcriptional regulators present in the cell at any given time.
Three approaches to nuclear reprogramming to pluripotency
Recent studies show that pluripotent stem cells with properties similar to ES cells (called iPS cells) can be induced readily from differentiated somatic cells. This finding has led to great excitement regarding the potential of these cells for improving the understanding and treatment of disease and has highlighted the need for a better mechanistic understanding of the reprogramming process. Insight is needed into which regulators are required for iPS cells to be reliably and efficiently generated and induced to differentiate towards the specialized cell fate of interest. To achieve this goal, all three approaches to nuclear reprogramming need to be enlisted.
This Review provides a historical perspective on the key findings () that led to the discovery of cellular plasticity, discussing studies using each of the three approaches to nuclear reprogramming in turn. It also indicates the questions that must be answered before nuclear reprogramming can fulfil its potential in medical applications.
Timeline of discoveries in nuclear reprogramming