1 March 2011. Scientists’ view of amyloid’s role in Alzheimer’s disease has evolved from the view that large amyloid fibrils poison the brain to one primarily blaming soluble, oligomeric forms of Aβ. A similar evolution of thought may be on the horizon for prion diseases. In the February 24 Nature, researchers led by John Collinge at University College London, U.K., show that prion infection and propagation can be separated from toxicity. Using mice that expressed different levels of native prion protein (PrPc), the researchers found that prion infections rose rapidly and plateaued at the same level in all mice. Clinical disease began shortly thereafter in mice with a lot of prion protein, but much later in mice with little protein. The results suggest that prion disease involves two separate phases—an infectious phase and a toxic phase—with toxic prion forms emerging only after the infection has saturated.
In an accompanying Nature commentary, Reed Wickner at the National Institutes of Health, Bethesda, Maryland, wrote, “The authors infer that the toxic, killer PrP species must be distinct from the infectious species. This is unexpected and raises issues central to our understanding of prion diseases.” Lary Walker at Emory University in Atlanta, Georgia, agreed that the paper is conceptually interesting, adding, “We’re going to be thinking about the implications for quite awhile.”
In other prion news, researchers led by Lawrence Goldstein at the University of California in San Diego provide insight into prion transport in the February 18 Cell, identifying kinesin-1 and cytoplasmic dynein as the major motor proteins that tow prion-containing vesicles along axons.
Prion diseases are thought to develop when misfolded proteins infect an animal and catalyze the conversion of the native prion protein into the misfolded form. This mechanism has also captured the interest of AD researchers, since growing evidence suggests that aggregating forms of Aβ can convert harmless versions into toxic entities, thus spreading the disease through the brain (see e.g., ARF related news story on Eisele et al., 2009). Prion diseases involve long incubation periods, as long as 50 years in humans, followed by rapid clinical decline and death. In mice, researchers had noted that mice with more of the normal prion protein had shorter incubation periods than mice with less, but they did not know how the prion infection was progressing during this dormant period.
To answer this question, first author Malin Sandberg used a cell-based assay that measures the concentration of infectious prion particles accurately and rapidly (see Klöhn et al., 2003). Sandberg and colleagues compared brain homogenates from four different mouse strains: animals that lacked prion protein, hemizygous mice that expressed half the normal amount of prion protein, wild-type mice, and transgenic mice with eight times the normal expression level. All animals were inoculated with infectious Rocky Mountain Laboratory prions. At regular intervals, the researchers sacrificed animals and checked the infection level in their brains using the cell-based assay.
Mice without prion expression quickly cleared the infection, in accordance with prior research in the field (see, e.g., Büeler et al., 1993). Unexpectedly, the authors found that prion infection rose rapidly in all other mice, reaching a common plateau level that was independent of the amount of native prion protein in the brains. The infection phase took a little longer in animals with half the normal levels of prion expression, but was virtually the same length in wild-type mice and overexpressors. This suggests that something other than prion concentration limits the reaction, the authors write. It is not clear what sets the maximum level of infection; the authors speculate that there might be an essential cofactor that has a fixed concentration in the brain, or a limited number of sites for misfolded prions to occupy.
After the infections reached the plateau, the time until mice got sick varied inversely with the amount of prion protein the mice expressed. This fits with a model in which the infectious PrPsc particles act as a catalytic surface for production of neurotoxic PrP particles, the authors suggest. The toxic PrP species might be oligomeric, they add. In this model, mice die when poisonous prion particles reach a set toxic threshold that is the same for all mice. The more native prion lying around to serve as raw material, the more quickly the threshold is reached.
In his commentary, Wickner puts forth other possibilities. He points out that Sandberg and colleagues were unable to measure the concentration of toxic prion particles. Therefore, it could be that a higher level of toxic particles accumulates during the long incubation period in hemizygous mice, but these mice have more resistance to the deadly prions. To answer this question will require the development of a cell-based toxicity assay, Wickner suggests.
He also notes that only cells that express PrP on their surface are susceptible to the disease (see, e.g., Brandner et al., 1996). “In this sense, therefore, PrP is a receptor for killing cells,” Wickner wrote. “This would explain a direct relationship between toxicity and PrP expression level.” PrP particles may be assembling into amyloid forms on the cell surface, then getting internalized and clogging up cellular compartments, Wickner speculates.
That fits with previous work from Collinge’s group showing that depleting normal prion protein in mice allows them to recover from a prion infection (see ARF related news story on Mallucci et al., 2007; ARF related news story on Mallucci et al., 2003). These data highlight that the disease cannot take hold without the presence of normal prion protein.
Walker points out that the existence of a two-phase disease process suggests a therapeutic strategy. If physicians could catch the disease during its second, incubation phase, and administer a drug that lowers production of toxic prion forms, it might delay the onset of the disease. Walker said the next step will be to identify the infectious and toxic prion forms and discover how they differ from one another. If other diseases that involve protein aggregation, such as Alzheimer’s disease, also have this two-stage characteristic, the same approach might work for them, Walker speculated, adding that this is a big “if.” For AD, scientists must also gain a better understanding of what different Aβ species do, he said.
Meanwhile, new work from the UC San Diego group opens a window into the intracellular transport of normal prions. Goldstein and colleagues show that kinesin-1 and dynein are the major motor proteins responsible for carting prions around. Kinesin-1 powers anterograde transport toward axon ends, while dynein walks retrogradely, but both motors were always found on prion vesicles regardless of which way the vesicle was moving. The activities of both motor proteins were tightly coupled. This led the authors to suggest that the direction of movement must be controlled by the regulation of motor protein activity, rather than the attachment of specific motor proteins, Goldstein and colleagues found.—Madolyn Bowman Rogers.
Sandberg MK, Al-Doujaily H, Sharps B, Clarke AR, Collinge J. Prion propagation and toxicity in vivo occur in two distinct mechanistic phases. Nature. 2011 Feb 24;470(7335):540-2. Abstract
Wickner RB. Infectivity versus toxicity. Nature. 2011 Feb 24;470(7335):470-1. Abstract
Encalada SE, Szpankowski L, Xia CH, Goldstein LS. Stable kinesin and dynein assemblies drive the axonal transport of mammalian prion protein vesicles. Cell. 2011 Feb 18;144(4):551-65. Abstract