Normal prion protein finally has a job.
A few unsettled questions notwithstanding, there is broad consensus in the scientific community about what misfolded prion protein does. It appears to be an infectious agent and the culprit in transmitting spongiform encephalopathies - degenerative brain disorders that include mad cow disease in cows, scrapie in sheep and Creutzfeldt-Jacob disease (CJD) in humans (see BioWorld Today, Nov 10, 2005.)
But what the normally folded prion protein does has been more of a mystery until now. Despite being widely expressed on many different cell types, both during development and in adulthood, prion protein knockouts do not have much of a phenotype.
"There are lots of hints and little agreement" on what happens to prion protein knockouts, Andrew Steele told BioWorld Today. Steele is a graduate student in Susan Lindquist’s lab at the Whitehead Institute for Biomedical Research and a co-author on two papers that have appeared in the Proceedings of the National Academy of Sciences. In the papers, Steele and his colleagues at the Whitehead Institute and Harvard Medical School in Cambridge, Mass., and Boston, respectively, show that prion protein plays a role in self-renewal and proliferation of blood-forming stem cells and neural precursors.
The realization that prion protein might have a role in cellular self-renewal was "fortuitous," Steele said, discovered in the course of microarray studies on blood-forming stem cells done by postdoctoral fellow Cheng Cheng Zhang, at Whitehead Institute. "By studying the normal function of prion protein, we hope to decipher its role in disease," he added.
To study prion protein’s role in the blood system, the researchers performed bone marrow transplantations of cells that did not express prion protein together with wild-type bone marrow stem cells. They found that after a single transplantation, prion protein knockout stem cells replicated about as well as their wild-type counterparts.
However, when the scientists upped the ante by performing serial transplantations, prion knockout cells did not hold up well under pressure. In serial transplants, mice are irradiated, receive bone marrow and are allowed to recover. Once they have recovered, they serve as bone marrow donors for another group in a secondary transplant. This process can be repeated for tertiary transplant; unsurprisingly, it takes robust cells to survive such multiple transplant procedures.
When tertiary transplantations were performed, only roughly 20 percent of cells in peripheral blood samples were descended from prionless cells four months after transplantation, showing that the cells lacking prion protein did not replicate as well as wild-type cells. In noncompetitive transplants where each mouse received either purely wild-type or purely prion knockout cells, both groups had full survival after a primary transplant, but the survival prospects diverged sharply by the secondary transplant, and by the tertiary transplant. None of the animals transplanted with prion knockout cells survived, whereas roughly half of the animals who received wild-type cells did.
The authors concluded that blood-forming stem cells lacking prion protein "have impaired self-renewal capabilities," which may not be critical under normal conditions but becomes apparent in times of stress. The experiments were published in the Feb. 7 issue of PNAS.
In the second paper, which also appeared in PNAS and now is available via early online publication, the scientists turned their attention to neural precursor cells.
Steele avoided calling the neural cells described in the paper stem cells. "I wish we were sure they were stem cells," he said, "but we did not do the experiments that would let us say that." Technically speaking, a true stem cell gives rise to another stem cell as part of its cell division, while precursor cells divide to produce two differentiated cells.
The scientists found that prion protein was expressed in increasing amounts as cells took the multistep trip from neural precursor to differentiated neuron. In mature brain cells, prion protein was expressed in high levels in neurons, but not in support cells known as glia.
When they compared knockouts, wild-type animals and overexpressors of prion protein, the levels of prion protein correlated with how rapidly cells went from precursor to a differentiated state. However, while prion protein lends a helping hand in adult neurogenesis, it does not seem to be absolutely necessary. Though they were slower about it, the precursor cells of prion knockout mice ultimately also differentiated into neurons, and prion knockouts did not have fewer neurons or obvious structural defects.
The obvious next question is whether prion protein’s role in cellular self-renewal is a general one. Steele and his colleagues plan on investigating other stem cell types.
They are planning to investigate prion protein’s role in lung stem cells, which have been isolated and can be unambiguously identified. That is an advantage they have over blood-forming stem cells and neural stem cells; despite the fact that new cells are clearly formed in adults in both the blood and the brain, which cells do the job is still somewhat murky. "No one really knows what they are," Steele said, whereas "lung stem cells have been identified."