This finding sheds light on a puzzling behavior. For some differentiating stem cells, the first step leads not to a final decision but to a new choice. In response to the initial chemical signal, these cells take on the genetic signatures of two different cell types. It often requires a second signal for them to commit to a single cellular identity.
In the Aug. 25 2006 issue of Cell, the researchers, working with hematopoietic stem cells, which give rise to the many types of blood cells, show how "pioneer transcription factors" trigger the first step, pushing these stem cells towards this mixed lineage, midway between two related cell types -- in this case between a macrophage and a neutrophil.
Then one of two rival "secondary factors" activates the genes that lead to one cell type and shuts down the genes that lead to the alternative.
Understanding the circuitry that controls these decisions is central to learning how different kinds of stem cells develop. It provides insights into how to transform stem cells into therapeutically useful cells and suggests possible new treatments for leukemias, in which a persistent mixed lineage seems to drive cancerous proliferation.
Although the researchers worked only with blood-forming stem cells, they suspect that the same basic regulatory principles govern cell type determination in other tissues such as skin, brain and intestine.
"We see elements of this framework of primary and secondary cell-fate determinants throughout the hematopoietic system," said study author Harinder Singh, the Louis Block Professor of Molecular Genetics & Cell Biology and a Howard Hughes Medical Institute Investigator at the University of Chicago, "and we suspect such networks also regulate cell fate in other systems."
This finding "represents a significant advance in understanding the molecular mechanisms that regulate [stem cell] development," according to a commentary in the journal by Dale Muzzy of Harvard University and Alexander van Oudenaarden of the Massachusetts Institute of Technology. "Irreversible resolution of lineage priming appears to be a common feature of blood cell differentiation and may in fact be a general feature of other developmental processes."
"Understanding the genetic circuitry that orchestrates development of each specialized cell type," Singh added, "should enable us to manipulate it for our own purposes."
The researchers focused on how hematopoietic stem cells developed into one of two types of white blood cells: macrophages or neutrophils. Macrophages are the long-lived garbage disposals of the immune system, indiscriminately engulfing and digesting cellular debris and pathogens. The shorter-lived neutrophils are the immune system's vultures, flocking to the site of an infection to target and ingest invading organisms.
Although both cell types come from cells known as myeloid progenitors, each type relies on its own set of functionally active genes to carry out its particular role in fighting infection. A major scientific puzzle has been how and why immature hematopoietic stem cells initially express genes that are characteristic of both cell lineages.
Until recently, however, there was no experimental system that researchers could manipulate to solve this puzzle. A few years ago, Singh and colleagues identified a transcription factor called PU.1 that acts as the primary signal, a central genetic switch to initiate development of myeloid progenitor cells. Other researchers identified a rival transcription factor, C/EBPƒÑ.
Cells from mice bred in Singh¡¦s lab to lack PU.1 allowed the researchers to manipulate the cells' decision-making machinery by introducing different amounts of PU.1. When the researchers introduced low concentration of PU.1, they found that the cells activated both macrophage and neutrophil genes.
"This showed for the first time how this mixed lineage pattern is set up," Singh said.
When they increased the concentration of PU.1, however, the cells quickly passed through a transitory mixed lineage state and produced new regulatory proteins that activated macrophage genes and repressed neutrophil genes. Higher levels of C/EBPƒÑ tipped the balance the other way.
The researchers were then able to identify both sets of antagonistic secondary regulators. Egr-1 and Egr-2 activate macrophage and repress neutrophil genes. Gif-1 is required to turn on neutrophil genes and repress macrophage counterparts.
Such counteracting repression circuitry may be the key to understanding stem cell regulation in general, Singh said. "We think that if this property of mixed lineage transcriptional priming is shared amongst different kinds of stem cells, then resolving these mixed-lineage states will invariably involve counteracting repressors."
In collaboration with colleague Aaron Dinner, the research team also formulated a mathematical model that depicts the regulatory network governing progenitor cell development. This model, he said, could have important implications for the therapeutic use of stem cells to rejuvenate damaged tissues.
Understanding of leukemias also could be aided by insight into this regulatory circuitry, Singh said. Many leukemias exhibit mixed-lineage patterns of gene expression, for example of both macrophage and lymphocyte genes.
"It may be that these cells are stuck in a progenitor-like state," said Singh. "If you could induce them to resolve that state ¡X to differentiate into one or the other cell type ¡X they would cease to be tumorigenic."