Courtesy of Cecil Fox and the National Cancer Institute

Differentiation, the stepwise specialization of cells, and transdifferentiation, the apparent switching of one cell type into another, capture much of the stem cell spotlight. But dedifferentiation, the developmental reversal of a cell before it reinvents itself, is an important process, too. This loss of specialization is believed to factor heavily into stem-cell culture techniques and pathological conditions such as cancer. Determining its programming, however, or even definitively identifying the process defies best efforts and flies in the face of Occam's razor: Here, the simplest explanations hardly seem sufficient.

"Dedifferentiation is difficult to look at, but the term is used fairly widely," says Rod Bremner, a senior scientist in the division of cell and molecular biology at the Toronto Western Research Institute. Many times, it seems, dedifferentiation is inferred as the mysterious means to an observable end.


The classic...


Dedifferentiation is easier to observe in unusual situations. Loss of specialization is a defining feature of a cancer cell. Since stem cells are also dedifferentiated, shouldn't errant stem cells beget cancer cells? Irving Weissman, professor of pathology and developmental biology at the Stanford University School of Medicine, calls leukemia "a smoldering stem cell disease," arising in stem cells but wreaking havoc via proliferating progenitors. But again, the obvious explanation might not suffice. "The cell of cancer origin need not be a stem cell, but [one that] became a stem cell by virtue of mutation that confers stem-cell-like properties," says Bremner.

Mutation might also explain why more advanced tumors have fewer specialized cells. "Think of a tumor as a tissue with a balance between three cell types – stem, progenitor, and terminally differentiated – that determines if the tumor is aggressive and rapidly growing, or much less aggressive and fairly well differentiated," explains Bremner. Then another mutation removes some specialization from a cell lineage, bolstering the progenitor cell contingent. "It's not that there is dedifferentiation, but changing of the balance between the proportions of stem cells, progenitors, and differentiated cells," he adds.

Artificial circumstances such as cell culture and somatic cell nuclear transfer (SCNT) evoke dedifferentiation. Arshak Alexanian, an assistant professor in the department of neurosurgery at the Medical College of Wisconsin, showed that glia from the subependymal zone of mouse brains can regress to neural stem cells (NSCs) when exposed to NSCs, losing glia-specific markers and sporting new NSC markers such as nestin and Sox2. Meanwhile, the cells formed neurospheres, which are balls of NSCs. "Cell-cell interaction between multipotent adult NSCs and glial cells can be an important means for plasticity," he says.


© 2004 Nature Publishing Group

An assay screening compounds for the ability to revert myoblasts into progenitor cells turned up reversine, a 2,6-disubstituted purine. (From Nat Biotecnol, 22:833–840, 2004.)

SCNT is the most dramatic demonstration of dedifferentiation, when the nucleus from a differentiated cell recapitulates development, presumably by reactivating its "stemness" genes (see Hot Papers, pg. 24). Recently, two groups produced mice from the nuclei of olfactory neurons. Although an olfactory neuron is about as terminally differentiated as a cell can get, the resulting mice had full smelling capabilities, underscoring the success at reversing cell fate.23 But the apparent nuclear reprogramming is inferred from the end result, the mouse. Again, there might be another explanation. "Reprogramming suggests that a new program is loaded and reloaded, and I'm not sure that's the case," says Mombaerts, one of the principal investigators. "Maybe there is a chaotic constellation of events, and the rare cell that turns on a few genes needed for early development makes it. That might be why SCNT has a success rate of about one percent, no matter what cell type or species is used."


Taking a person's cells back in developmental time to grow spare parts is still in the future, as researchers are just beginning to explore the components of dedifferentiation pathways. Experiments using the cellular slime mold Dictyostelium discoideum and mice allow investigators to take this reductionist approach.

Dictyostelium is a poster child for dedifferentiation, swinging from unicellular to multicellular depending on food availability. Within 24 hours of starving, undifferentiated single cells aggregate into a slug that finds food, halts, telescopes upward, then forms a fruiting body that releases spores, which can dedifferentiate back to the unspecialized, single life. It appears a highly regulated process. "Cells that have differentiated for a short time require less time to dedifferentiate than cells that have differentiated for a longer time," explains Gad Shaulsky, associate professor of molecular and human genetics at the Baylor College of Medicine in Houston.

To investigate how Dictyostelium cells know when to lose their specialization, Shaulsky and colleagues compared transcriptional profiles of fruiting bodies disrupted at different times; they found the same up or down regulation of 122 genes, no matter when the shakedown occurred.4 The pattern of gene expression is not simply the reverse of differentiation. This observation, plus the fact that mutations can derail dedifferentiation, indicates that dedifferentiation is as precisely, if distinctively, genetically controlled as the better-studied differentiation. Yet the researchers identified a key gene active in differentiation and dedifferentiation, histidine kinase dhkA.

Elsewhere, in another organism and through a completely different experimental design, researchers have zeroed in on kinase interactions as part of the mechanistic machinery of dedifferentiation.56 Ding and colleagues used combinatorial chemistry to screen 50,000 synthetic compounds with structures that could bind a kinase. They applied the compounds to mouse myoblasts, which are differentiated cells that merge to form myotubes. (Previous experiments had demonstrated plasticity: Regenerated newt limb extracts could revert myotubes to mesenchymal progenitor cells, which could then reform myotubes.) The assay screened for myoblast cells that, when exposed to a test compound, assumed a progenitor phenotype and then, given appropriate signals, redifferentiated as bone or fat cells. A purine analog, named reversine, fit the bill.

Isolating kinases and their potential partners in signaling pathways adds precision to a line of research founded on multi-molecular leg-of-newt and egg extracts to evoke dedifferentiation. Says Ding, "These extracts are mixtures of many proteins and signaling molecules, and it is hard to find out which one is the master regulator, or which combination of molecules is most effective. We are still far away from what nature does. But we are doing other screens to look for chemicals and genes that would mimic egg extracts. You never know if we will be lucky enough to find them."

Ricki Lewis rlewis@the-scientist.com

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