Since discovering it as a gene for frontotemporal lobar degeneration (ARF related news story on Baker et al., 2006 and Cruts et al., 2006), scientists have puzzled over what progranulin does in the central nervous system, and how inadequate levels of this protein lead to neurodegeneration. A new study in C. elegans roundworms suggests an intriguing possibility: Progranulin slows clearance of sick or dying cells. In this scenario, phagocytes—cells that engulf other cells and debris—become dangerously ravenous without progranulin, gobbling sick cells that might otherwise have recovered. This could explain how a non-lethal insult can result in massive neuron loss, Cynthia Kenyon, University of California, San Francisco, and colleagues propose in a paper published online February 28 in the Proceedings of the National Academy of Sciences U.S.A.

Progranulin functions in embryogenesis, inflammation, and wound healing, and some tumor cells produce a lot of the protein (reviewed in Bateman and Bennett, 2009). Mutations in the progranulin gene may also be linked to Alzheimer’s disease (see ARF related news story). In cultured neurons and neuronal cell lines, the glycoprotein seems to act as a trophic factor, promoting neurite outgrowth and cell survival (Van Damme et al., 2008; Ryan et al., 2009). Hence, it “made perfect sense” to think that neurons lacking progranulin would get sick and die, first author Aimee Kao said in an interview with ARF.

Yet several mouse models of progranulin deficiency fail to show the expected neuron loss (Kayasuga et al., 2007; Yin et al., 2010; Yin et al., 2010). “But they did have angry, active microglia,” Kao said. And since neurons that die in neurodegenerative disease are known to do so in a controlled way, the UCSF researchers wondered whether progranulin might have a role in apoptosis, or programmed cell death.

The C. elegans nematode seemed the ideal system for addressing this question. The lineage of every one of its 302 neurons has been mapped out, such that researchers know exactly which cells die during development, as well as precisely when—down to the minute. Plus, since the worms are transparent, “you can really watch cells in the process of dying,” Kao said.

The researchers approached Shohei Mitani at Tokyo Women’s Medical University, Japan, for his progranulin-null strain. These mutant worms look fine and have a normal lifespan. However, using microscopy to visualize dying cells during development, the scientists found the opposite of what they expected. In the progranulin-null embryos, “we thought we’d see more programmed cell death,” Kao said. “We ended up seeing less.” At any given point in time, the mutant embryos had 20 percent fewer apoptotic “corpses,” compared to wild-type controls.

With help from coauthor Juan Cabello at the Center for Biomedical Research of La Rioja, Spain, the researchers took a closer look, using time-lapse microscopy to track individual cells minute by minute across several hours of development. They found that, on average, “the amount of time it took for a progranulin-deficient cell to die was about half the time it took for wild-type cells,” Kao said.

As for possible reasons why, the scientists envisioned several situations. It could be that cells in the progranulin-deficient embryos “are dying faster,” Kao suggested, “or that the engulfing cells are coming along more quickly, to help the dying cells die.” To tease out those possibilities, she isolated peritoneal macrophages from progranulin-knockout mice made by UCSF collaborator Robert Farese (see ARF related conference story), and used in vitro assays to measure how many microscopic latex beads, or apoptotic wild-type thymocytes, they could scarf in a 90-minute period. In both tests, the progranulin-knockout macrophages out-gobbled the macrophages from wild-type littermates.

All told, the data “suggest a model where progranulin acts normally as a factor to slow [removal of apoptotic cells],” Kao said. “And if you don’t have progranulin, this whole process occurs more quickly.”

In this model, when wild-type levels of progranulin are present, a cell has enough time to repair itself and survive a “sublethal insult,” the authors wrote. However, in conditions of progranulin deficiency, “the rate at which an injured cell is recognized and/or engulfed by phagocytic cells is accelerated, and the damaged cell has less time to recover.” Such changes in cell death kinetics could increase susceptibility to neurodegeneration and, in turn, facilitate disease progression, they wrote.

Benjamin Wolozin of Boston University in Massachusetts found this “a provocative and interesting study that definitely moves the field forward.” The authors “convincingly demonstrate a role for progranulin in phagocytosis,” he wrote in an e-mail to ARF. However, Wolozin suggested that “jumping from there to proposing that this is a cause of frontotemporal dementia is too large of a leap.” (See full comment below.) Kao and colleagues note that their explanation for how progranulin deficiency might lead to neurodegeneration “remains speculative.” In addition, they acknowledge that these changes in the kinetics of apoptosis would have been hard to see in mammals.

Chris Link, University of Colorado, Boulder, said the study could have been strengthened by measuring phagocytic activity of progranulin-deficient brain microglia, instead of peritoneal macrophages, which “are influenced by lots of things, such as whether you had infection or not.” In addition, since the same corpse removal pathways are used for both apoptotic and necrotic cell death in worms (Chung et al., 2000), “it would be nice to know if the progranulin mutation also increased the rate of corpse removal of necrotic neurons,” he noted. The current study only examined apoptotic cells.

On a broader level, Link questioned whether it is even useful to consider how a role in apoptosis could explain the contribution of progranulin deficiency to neurodegenerative disease. “There’s little evidence in my mind that any neurodegenerative condition is specifically due to neuronal loss,” Link told ARF. “There is lots of evidence that before you get neuron loss, you get synapse damage and behavioral effects. In neurodegenerative disease, you have apoptosis, but I think that is appropriate because the cells are already damaged. So what you need to understand is why they got damaged in the first place, not why they’re dying.”

To determine whether the observed effects of progranulin deficiency on cell death kinetics carry through to mammalian models, Kao said she plans to collaborate with multiphoton microscopy experts to track individual neurons in the brains of progranulin-deficient mice “to look at whether they live or die, and see what happens to the microglia around them.”—Esther Landhuis

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Comments on News and Primary Papers

  1. This is a very interesting study by Cynthia Kenyon's team. Their work in nematodes convincingly provides clear demonstration of a role for progranulin in phagocytosis, and confirms it by abrogation of the effect of progranulin deletion by deletion of other genes in the phagocytic pathway. The work with mouse macrophages provides useful translation to the mammalian context. A weakness in the C. elegans study is the inability to fully genetically complement the deletion of progranulin by expressing the progranulin gene transgenically. Kenyon's team suggests that this is likely due to differences in gene expression between endogenous and expressed genes. This explanation is probably correct; however, it is also possible that the incomplete complementation results from mutations at other sites in the nematode genome that might impact on phagocytosis.

    The team's discovery that progranulin insufficiency increases phagocytosis is a surprise, because we have come to associate neurodegeneration with reduced catabolic activity, rather than an increase in degradative processes. Kenyon proposes that the increased phagocytosis that might be associated with frontotemporal dementia (FTD) is the cause of the illness. I think this hypothesis is premature because progranulin is expressed in many cell types and exerts many different actions. The role in phagocytosis is quite believable and probably true, but jumping from there to proposing that this is the cause of FTD is too large of a leap. However, the proposal does suggest experiments that are relatively straightforward to test, such as the conditional knockout of progranulin gene in the macrophage lineage. Regardless, this is a provocative and interesting study that definitely moves the field forward.

    View all comments by Benjamin Wolozin

References

News Citations

  1. Birds of a Feather…Mutations in Tau Gene Neighbor Progranulin Cause FTD
  2. Genetics of FTD: New Gene, PGRN Variety, and a Bit of FUS
  3. San Diego: Progranulin, Wnt, and Frizzled, Frazzle Neurons in FTD

Paper Citations

  1. . Mutations in progranulin cause tau-negative frontotemporal dementia linked to chromosome 17. Nature. 2006 Aug 24;442(7105):916-9. PubMed.
  2. . Null mutations in progranulin cause ubiquitin-positive frontotemporal dementia linked to chromosome 17q21. Nature. 2006 Aug 24;442(7105):920-4. PubMed.
  3. . The granulin gene family: from cancer to dementia. Bioessays. 2009 Nov;31(11):1245-54. PubMed.
  4. . Progranulin functions as a neurotrophic factor to regulate neurite outgrowth and enhance neuronal survival. J Cell Biol. 2008 Apr 7;181(1):37-41. PubMed.
  5. . Progranulin is expressed within motor neurons and promotes neuronal cell survival. BMC Neurosci. 2009;10:130. PubMed.
  6. . Alteration of behavioural phenotype in mice by targeted disruption of the progranulin gene. Behav Brain Res. 2007 Dec 28;185(2):110-8. PubMed.
  7. . Exaggerated inflammation, impaired host defense, and neuropathology in progranulin-deficient mice. J Exp Med. 2010 Jan 18;207(1):117-28. PubMed.
  8. . Behavioral deficits and progressive neuropathology in progranulin-deficient mice: a mouse model of frontotemporal dementia. FASEB J. 2010 Dec;24(12):4639-47. PubMed.
  9. . A common set of engulfment genes mediates removal of both apoptotic and necrotic cell corpses in C. elegans. Nat Cell Biol. 2000 Dec;2(12):931-7. PubMed.

Further Reading

Papers

  1. . Progranulin: normal function and role in neurodegeneration. J Neurochem. 2008 Jan;104(2):287-97. PubMed.

Primary Papers

  1. . A neurodegenerative disease mutation that accelerates the clearance of apoptotic cells. Proc Natl Acad Sci U S A. 2011 Mar 15;108(11):4441-6. PubMed.