By flipping a genetic switch, HHMI researchers can control a white blood cell's destiny.
By increasing or decreasing levels of a specific protein, researchers can control the developmental destiny of white blood cells.
The finding represents the first evidence that mammalian immune system cells respond to such a dual control mechanism. The discovery raises the possibility that scientists might be able to use the developmental switch to manipulate the immune system to produce specific disease-fighting cells. Furthermore, throwing the switch by using drugs or gene therapy might jump-start the proper development of immature blood cells whose maturation is stalled in certain cancers.
We believe that development of different lineages of blood cells is triggered by a combination of particular sets of transcription factors and their levels relative to one another.
In an article in the May 26, 2000, issue of Science, Howard Hughes Medical Institute investigator Harinder Singh and his colleague Rodney DeKoter report that the concentration of the transcription factor PU.1 in progenitor immune cells determines whether those cells become macrophages or B cells.
Transcription factors are proteins that activate genes to transcribe their information into messenger RNA. Macrophages are white blood cells that attack bacteria and other invaders by engulfing and digesting them; while B cells generate antibodies that tag pathogens bearing foreign proteins, marking them for attack by other immune cells.
In previous studies, Singh and his colleagues at the University of Chicago showed that knocking out the PU.1 gene in mice eliminated the ability of such mice to make white blood cells. The scientists found that PU.1 switched on genes specific to progenitors of macrophages or B cells that were necessary for their normal differentiation into mature cells.
"This left us with a puzzle," said Singh. "If this transcription factor is needed for development of different cell types in the immune system—each of which expresses very different sets of genes—how could a single factor participate in regulating these very diverse programs of gene expression?"
In exploring PU.1's role in development, the researchers created a method for isolating mutant mouse progenitor blood cells that lacked PU.1. Once they had harvested the PU.1-deficient cells, they used a viral vector to shuttle the PU.1 genes back into the cells.
"We found that these cells that were otherwise blocked from developing any further recovered their ability to differentiate properly into macrophages or B cells when they received the gene for PU.1," said Singh.
A striking result of this experiment, said Singh, was that the resulting macrophages expressed high concentrations of the PU.1 protein, while the B cells expressed low concentrations.
"It was quite a surprise," said Singh. "Although there had been some reports that PU.1 levels were different in macrophages and B cells, no one had ascribed any functional significance to that difference."
To explore the functional role of PU.1 levels in immune cell differentiation, the researchers next raised the level of PU.1 in normal progenitor cells. They found that higher PU.1 levels effectively caused the cells to become macrophages, rather than B cells.
And in a third set of experiments, the scientists inserted a truncated form of the PU.1 protein into cells that lacked PU.1 and found that this "weakened" transcription factor efficiently rescued the development of B cells, but not of macrophages.
"Thus, we have shown that a lower concentration or a lower activity state translates into one developmental outcome for these cells, turning them into B cells," said Singh. "And conversely, higher concentrations of the regulator produce macrophages.
"The idea that differing concentrations of a transcription factor can control the development of different cell types from progenitors has been an attractive and commonly held view among developmental biologists," said Singh. "It has been exceedingly difficult, however, to show that this principle is actually valid in any given organism."
Singh noted that, while researchers had shown that developing fruit flies use graded levels of certain transcription factors to control the generation of different embryonic cell types, "there hasn't been strong evidence for this mechanism in mammalian systems. Thus, we are providing good evidence that concentration-based control exists in the immune system, and of course, in mammals in general."
Singh emphasized that despite PU.1's importance, it is certainly not the lone determinant of immune cell differentiation.
"We believe that development of different lineages of blood cells is triggered by a combination of particular sets of transcription factors and their levels relative to one another," he said.
Understanding how PU.1 controls immune cell development could have important medical implications, he noted.
"Once we understand the regulatory circuitry that controls immune cell development, we could exploit that knowledge to develop treatments that direct progenitor cells to differentiate into particular cell types for therapeutic purposes," he said.
"Also, many cancers of the immune system involve interruption of blood cell differentiation. It's possible that drugs or gene therapy could be developed to increase PU.1 activity to unblock differentiation of these cancer cells and make them become benign."