PAR-1, a homologue of microtubule-affinity regulating kinase (MARK), kicks off the phosphorylation events that culminate in cell death in a fly model of tauopathy, according to a report in today’s issue of Cell by Bingwei Lu and colleagues.

The list of enzymes that can regulate tau phosphorylation in the test tube is quite long—in addition to MARK/PAR-1, there are GSK-3β, MAP kinase, CDK2 and 5 (see ARF related news story), PKA, CaMKII, and protein phosphatases 1, 2A, 2B, and 2C. GSK-3 and CDK5 are most likely involved in vivo, too. But which one is the ringleader of the neurodegenerative process?

In the current study, first author Isao Nishimura of the Rockefeller University in New York City, with Lu and Yufeng Yang, who are now at Stanford University in Palo Alto, California, take advantage of the ease with which researchers can manipulate Drosophila genetics. They studied the role the fly’s MARK homologue PAR-1 (see ARF related news story) plays in tau-related degeneration of photoreceptor cells. The researchers first established that overexpression of PAR-1 in photoreceptor neurons led to degeneration of these cells in a dose-dependent manner. This effect was mediated by an endogenous fly tau homologue. PAR-1 overexpression also increased neurodegeneration in flies transfected with either wild-type or mutant (4 repeat with R406W mutation) human tau.

These experiments did not produce obvious neurofibrillary tangles (NFTs). Conversely, George Jackson and colleagues have found that coexpression of GSK-3 with human tau did lead to evidence of NFTs in a similar photoreceptor model (see ARF related news story).

The researchers next sought to answer the question of what happens to tau if there is no PAR-1 present. Since embryos lacking PAR-1 don't survive, the researchers created tissue clones that lacked PAR-1 but contained mutant human tau. TUNEL assays revealed a threefold drop in apoptosis in cells lacking PAR-1, accompanied by a drop in tau phosphorylation at the S262 and S356 residues. These two sites are frequented by MARK and other tau phosphorylators, and the researchers confirmed their importance by showing that point mutations in S262 and S356 abolished the toxicity of mutant h-tau in photoreceptors.

Yet more interesting was the finding that these mutations in the PAR-1/MARK binding site also abolished phosphorylation at other sites implicated in AD, including those recognized by the antibodies AT100 (pT212 and pS214) and AT8 (pS202 and pT205), but not the AT270 site (S199 and S202). The suggestion that phosphorylation at S262 and/or S356 is necessary for phosphorylation at these other sites was confirmed in neurons without PAR-1.

What does this mean for other phosphorylation suspects? The authors found that elevation of GSK-3 and CDK5, which have affinity for these inhibited sites, raised phosphorylation at these residues, an effect that was compounded by overexpression of PAR-1. Thus, PAR-1 phosphorylation seems to be a prerequisite for GSK-3- and CDK5-mediated phosphorylation.

In an editorial in the same issue of Cell, Mark Fortini of the National Cancer Institute in Frederick, Maryland, suggests an emerging picture "in which PAR-1 initiates a temporally ordered series of tau phosphorylations, with the early PAR-1 phosphorylation step generating soluble non-aggregating tau forms that are converted into hyperphosphorylated, aggregation-prone tau through downstream phosphorylations performed by CDK5, GSK-3β and perhaps additional kinases."

Speculating on tau's role in AD and other tauopathies, the authors make special mention of the possibility that hyperphosphorylation of tau could be involved in the early synaptic dysfunction postulated in AD (see also Mandelkow portion of ARF related news story). They suggest several candidate mechanisms for such an effect, including a direct effect of hyperphosphorylated tau at the synapse, or an effect via disrupted microtubule dynamics.—Hakon Heimer.

References:
Nishimura I, Yang Y, Lu B. PAR-1 kinase plays an initiator role in a temporally ordered phosphorylation process that confers tau toxicity in Drosophila. Cell. 2004 Mar 5;116:671-82. Abstract

Fortini ME. PAR-1 for the course of neurodegeneration. Cell. 2004 Mar 5;116(5):631-2. Abstract

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  1. This paper describes an intriguing Drosophila model of tau phosphorylation causing tau neurotoxicity. So far, therapeutic approaches to tau pathology in AD did not progress beyond the preclinical stage and were mainly directed at the inhibition of the CDK5 and GSK3 kinases. However, the MARK pathway may offer more promising targets. We and others have recently shown that MARKs are activated by LKB1/Par-4 [1,2]. This may represent a neurotoxic signal which is not specific for AD pathology, since it was just shown that both LKB1 and MARK4 become rapidly upregulated in a murine stroke model [3].

    Since confirmation of the Drosophila model by mouse knockouts may be difficult due to the presence of four MARK genes—whereas flies possess only a single gene—we may need to await the development of specific MARK inhibitors, and see whether these are able to inhibit P-tau (and Aβ-?) induced neuronal cell death.

    References:
    1. Brajenovic M, Joberty G, Kuster B, Bouwmeester T, Drewes G. Comprehensive proteomic analysis of human Par protein complexes reveals an interconnected protein network. J Biol Chem. 2003 Dec 15 [Epub ahead of print] Abstract

    2. Lizcano JM, Goransson O, Toth R, Deak M, Morrice NA, Boudeau J, Hawley SA, Udd L, Makela TP, Hardie DG, Alessi DR. LKB1 is a master kinase that activates 13 kinases of the AMPK subfamily, including MARK/PAR-1. EMBO J. 2004 Feb 25;23(4):833-43. Abstract

    3. Schneider A. et al. Identification of regulated genes during permanent focal cerebral ischaemia: characterization of the protein kinase 9b5/MARKL1/MARK4. J Neurochem. 2004 Mar 1; 88 (5)1114-26.

  2. This excellent paper draws renewed attention to the (other) central problem in neurodegeneration in general and AD in particular: How does the tau pathology originate? This essentially boils down to the question of what is the initial kinase, i.e., the kinase that triggers the phosphorylation that eventually results in hyperphosphorylation of tau and instigates the deadly cascade ending in paired helical filaments, neurofibrillary tangles and cell death. In that respect, tau is definitely the prime suspect and candidate "executer" of neurons in many neurodegenerative disorders, including AD. The pathological definition of AD as "plaques + tangles" does not allow or permit the AD field to escape this problem, despite the fact that amyloid attracts 10 times (my wild guess) more attention than tau.

    Through the work of the Mandelkow lab and many others, the functions of MARK kinase have been defined in some detail, in terms of phosphorylating tau and other MAPs, and in terms of neurite outgrowth and polarization. What was missing was a definite link to pathology, and that is provided by this paper. The authors define PAR1 kinase as responsible for phosphorylation of serines 262 and 356 in tau, thereby causing cells to die. Many studies have indirectly implicated MARK, GSK-3, PKA, and CaMKII in phosphorylating these sites, but no animal study is yet available to validate these kinases as the physiological kinase for these sites.

    So, are "S262 and S356" going to be magical for tau pathology as "β and γ" are for amyloid pathology? At the least, these serine residues are located in the region of tau that matters most, i.e., the microtubule-binding domain, which, incidentally, is also the region that is littered with mutations giving rise to the family of tauopathies known as FTD, or frontotemporal dementia! Antibody 12E8 specifically detects pS262 and pS356 in the MTBD of tau (Seubert et al., 1995), and it is definitely going to be popular and in demand among tauists and perhaps baptists.

    As always, many questions and some caveats remain. For one, the authors use not wild-type human tau, but the FTD mutant tau-R406W. This might explain why the final outcome is cell death but no tau aggregates in whatever form. Moreover, the choice of this mutant might have been fortuitous, since it is the most C-terminal of all known clinical mutations, closely positioned to the AD2/PHF1 epitope that is important in binding to MT and in tau aggregation (Spittaels et al., 2000; Vandebroek et al., 2004,). Further along the path to understand it all remains the question why no mice have yet been produced (or reported) with overexpression or deficiency of MARK? Those who have such mice available should come forward and inform the community what and how and where …

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References

News Citations

  1. Aiding and Abetting, Hyperactive CDK5 Gives Mouse Tangles
  2. New Tau Kinase Makes Its MARK
  3. Wingless Pathway Helps Tauopathy Take Off
  4. New Orleans: Symposium Probes Why Synapses Are Suffering

Paper Citations

  1. . PAR-1 kinase plays an initiator role in a temporally ordered phosphorylation process that confers tau toxicity in Drosophila. Cell. 2004 Mar 5;116(5):671-82. PubMed.
  2. . PAR-1 for the course of neurodegeneration. Cell. 2004 Mar 5;116(5):631-2. PubMed.

Further Reading

Papers

  1. . MARKK, a Ste20-like kinase, activates the polarity-inducing kinase MARK/PAR-1. EMBO J. 2003 Oct 1;22(19):5090-101. PubMed.
  2. . Protein kinase MARK/PAR-1 is required for neurite outgrowth and establishment of neuronal polarity. Mol Biol Cell. 2002 Nov;13(11):4013-28. PubMed.
  3. . MARK, a novel family of protein kinases that phosphorylate microtubule-associated proteins and trigger microtubule disruption. Cell. 1997 Apr 18;89(2):297-308. PubMed.
  4. . PAR-1 kinase plays an initiator role in a temporally ordered phosphorylation process that confers tau toxicity in Drosophila. Cell. 2004 Mar 5;116(5):671-82. PubMed.
  5. . Axonal neuregulin-1 regulates myelin sheath thickness. Science. 2004 Apr 30;304(5671):700-3. PubMed.
  6. . PAR-1 for the course of neurodegeneration. Cell. 2004 Mar 5;116(5):631-2. PubMed.

News

  1. Aiding and Abetting, Hyperactive CDK5 Gives Mouse Tangles
  2. New Orleans: Symposium Probes Why Synapses Are Suffering
  3. New Tau Kinase Makes Its MARK

Primary Papers

  1. . PAR-1 kinase plays an initiator role in a temporally ordered phosphorylation process that confers tau toxicity in Drosophila. Cell. 2004 Mar 5;116(5):671-82. PubMed.
  2. . PAR-1 for the course of neurodegeneration. Cell. 2004 Mar 5;116(5):631-2. PubMed.
  3. . Axonal neuregulin-1 regulates myelin sheath thickness. Science. 2004 Apr 30;304(5671):700-3. PubMed.