More support for what might be called the axonal "traffic jam" hypothesis for the pathogenesis of AD - from Larry Goldstein's lab. This paper should be read in conjunction with Orly Lazarov et al., J Neurosci March 2, 2005, which integrates work from Sam Sisodia's lab and five other labs and which provides evidence against that hypothesis. It would be nice if experiments could sharply differentiate between axonal transport peripherally and centrally. One would expect fierce traffic jams in peripheral axons, but AD patients do not appear to be particularly susceptible to peripheral neuropathy. (Peripheral neuropathy is very common in older people and is a sadly neglected research topic.)

View all comments by George M. Martin

This interesting paper shows that our perception of AD physiopathology is getting more complex, but more realistic. We are far away from the simple explanation of the amyloid cascade hypothesis (ACH). To summarize, neurodegeneration is associated with a defect of the axonal transport: key players involved are the microtubules stabilized by tau proteins, the motor proteins that transport the cargo- vesicle along microtubules, and especially kinesin-I, and APP as well as PS1 in the cargo-vesicles.

One big surprise is that the axonal transport defect generated by reducing the genetic dosage of kinesin increases Ab42 secretion and deposition. This sequence of events is the opposite of the ACH.

To conclude, kinesin-I is likely to be an additional risk factor of AD. But behind the paper, even if bypassed, is the role of tau to stabilize and control axonal transport. Cause and effects have still to be untangled in AD.

References:
Among the references related to this approach, I recommend also the papers of Beyreuther on the fast axonal transport of APP and those of the Mandelkow's related to kinesin, tau, APP and the axonal transport (J Cell Biol. 2002 Mar 18;156(6):1051-63 and other related papers)

View all comments by Andre Delacourte

This is an important study that may usher in a new perspective on the normal function of APP and clarify its role in AD pathogenesis.

View all comments by John Trojanowski

This paper underscores the importance of impaired axonal transport and motor neuron deficits induced by familial mutations in PS1. We agree with the notion that the problem in AD is intraneuronal mistrafficking of different axonal proteins, and the results presented may explain some pathological features we have previously observed. We have studied two bigenic AD mouse models with abundant intraneuronal Aβ accumulation, which correlated well with the observed neuron loss, and axonal degeneration in brain and spinal cord. We agree with Lazarov et al. that these defects are likely induced by a different trafficking of APP due to expression of mutant PS1. In both models, the APP751SL/PS1M146L (Schmitz et al., 2004), and the APP/PS1KI (APP751SL and knock-in of PS1M233T and PS1L235P) (Casas et al., 2004) mouse model, we have shown that neuronal dysfunction is plaque-independent (Wirths et al., 2006a; Wirths et al., 2006b).

The APP/PS1KI mouse model is especially interesting, because 50 percent of CA1 neurons are lost at 10 months of age (Casas et al., 2004). These mice also...  Read more

This paper underscores the importance of impaired axonal transport and motor neuron deficits induced by familial mutations in PS1. We agree with the notion that the problem in AD is intraneuronal mistrafficking of different axonal proteins, and the results presented may explain some pathological features we have previously observed. We have studied two bigenic AD mouse models with abundant intraneuronal Aβ accumulation, which correlated well with the observed neuron loss, and axonal degeneration in brain and spinal cord. We agree with Lazarov et al. that these defects are likely induced by a different trafficking of APP due to expression of mutant PS1. In both models, the APP751SL/PS1M146L (Schmitz et al., 2004), and the APP/PS1KI (APP751SL and knock-in of PS1M233T and PS1L235P) (Casas et al., 2004) mouse model, we have shown that neuronal dysfunction is plaque-independent (Wirths et al., 2006a; Wirths et al., 2006b).

The APP/PS1KI mouse model is especially interesting, because 50 percent of CA1 neurons are lost at 10 months of age (Casas et al., 2004). These mice also exhibit early and robust brain and spinal cord axonal degeneration, as shown by the occurrence of axonal spheroids, together with a reduced ability to perform motor performance tasks, including balance beam, string suspension, or rotarod. Working memory deficits were also evident at that time point (6 months of age) (Wirths et al., 2007). In good agreement with Lazarov et al. we have found evidence for increased levels of phosphorylated proteins (Tau [pS199] and APP [Thr668]) in degenerating axons inducing a loss of trophic support which might explain the robust age-dependent axonal degeneration in APP/PS1KI mice.

References:
Casas C, Sergeant N, Itier JM, Blanchard V, Wirths O, van der Kolk N, Vingtdeux V, van de Steeg E, Ret G, Canton T, Drobecq H, Clark A, Bonici B, Delacourte A, Benavides J, Schmitz C, Tremp G, Bayer TA, Benoit P, Pradier L. Massive CA1/2 neuronal loss with intraneuronal and N-terminal truncated Abeta42 accumulation in a novel Alzheimer transgenic model. Am J Pathol. 2004 Oct;165(4):1289-300. Abstract

Schmitz C, Rutten BP, Pielen A, Schafer S, Wirths O, Tremp G, Czech C, Blanchard V, Multhaup G, Rezaie P, Korr H, Steinbusch HW, Pradier L, Bayer TA. Hippocampal neuron loss exceeds amyloid plaque load in a transgenic mouse model of Alzheimer's disease. Am J Pathol. 2004 Apr;164(4):1495-502. Abstract

Wirths O, Weis J, Kayed R, Saido TC, Bayer TA. Age-dependent axonal degeneration in an Alzheimer mouse model. Neurobiol Aging. 2006 Sep 8; [Epub ahead of print] Abstract

Wirths O, Weis J, Szczygielski J, Multhaup G, Bayer TA. Axonopathy in an APP/PS1 transgenic mouse model of Alzheimer's disease. Acta Neuropathol (Berl). 2006 Apr;111(4):312-9. Epub 2006 Mar 7. Abstract

Wirths O, Breyhan H, Schafer S, Roth C, Bayer TA. Deficits in working memory and motor performance in the APP/PS1ki mouse model for Alzheimer's disease. Neurobiol Aging. 2007 Jan 8; [Epub ahead of print] Abstract

View all comments by Thomas Bayer
View all comments by Oliver Wirths

Reply by Aidong Yuan and Ralph Nixon to comments
Our study was an in vivo test of the hypothesis that moderate overexpression of tau directly impairs axonal transport. We expected our in vivo results to support in vitro data, but the surprising absence of a significant effect was clear and is compatible with existing data. The thoughtful comments in this forum highlight important general issues for consideration in future investigations in this area. As discussed in our report, we agree with the view expressed by Fred Van Leuven that our findings do not exclude the possibility that pathological states of tau might directly or indirectly alter axonal transport. Going forward, however, the task of defining any meaningful direct connection between pathological tau and transport disruption will require that primary effects on transport be distinguished from indirect effects that are secondary to neurodegeneration, which inevitably disrupts transport. In the studies of mice overexpressing tau 4R cited by Van Leuven (1-4), the inference that axonal transport may be...  Read more

Reply by Aidong Yuan and Ralph Nixon to comments
Our study was an in vivo test of the hypothesis that moderate overexpression of tau directly impairs axonal transport. We expected our in vivo results to support in vitro data, but the surprising absence of a significant effect was clear and is compatible with existing data. The thoughtful comments in this forum highlight important general issues for consideration in future investigations in this area. As discussed in our report, we agree with the view expressed by Fred Van Leuven that our findings do not exclude the possibility that pathological states of tau might directly or indirectly alter axonal transport. Going forward, however, the task of defining any meaningful direct connection between pathological tau and transport disruption will require that primary effects on transport be distinguished from indirect effects that are secondary to neurodegeneration, which inevitably disrupts transport. In the studies of mice overexpressing tau 4R cited by Van Leuven (1-4), the inference that axonal transport may be disrupted is based on neuropathological detection of focal organelle accumulations in the degenerating axons of tg mice. While the conclusion may well be correct, evidence from the only one of these studies that investigated axonal transport directly (3) suggests that effects on transport in this case are more likely secondary to neurodegeneration than related to a primary effect of tau 4R on transport mechanisms.

In these time-lapse transport analyses of fluorescent dextran-loaded vesicles in DRG axons of tau 4R tg mice, “the calculated mean of absolute velocities of fluorescent vesicles moving >0.1 μm/min reveals that both retrograde and anterograde vesicle transport did not differ significantly between transgenic and control mice” (3). Also, in vivo tracking of fluorescent vesicle numbers after intranerve injection of fluorescent dextran in live mice showed that tau 4R and wt mice differed at 1 hour, but not at 2 hours or 3 hours (3). Therefore, in the one context where axonal transport was measured directly, overexpression of tau 4R had no or equivocal effects on transport velocities. This observation is consistent with our data and with the Alzforum comment of Akihiko Takashima that “if tau overexpression induces impairment of axonal transport, tau tg mice must show neuronal dysfunction in the entire brain from a young age on” and not only after 20 months of age, when aging-related factors might combine with tau cytotoxicity to cause axonal degeneration and secondary transport failure. Rescue of this pathology by GSK-3β overexpression in tau 4R mice (4) may well be directly related to tau hyperphosphorylation, although neuroprotection could also involve reversal of transport-independent cytotoxic effects of tau.

We agree with Virginia Lee and John Trojanowski that our results are compatible with findings from their labs. The overexpression of the shortest isoform of tau at levels that were higher than those in our study was associated with neurofibrillary degeneration and reduced fast transport rates measured directly in vivo in side-by-side comparisons with appropriate control groups. Our discussion of these studies was not a misread of the two papers we cited (5-6). It instead drew attention to a comparison specifically of the untreated (or sham-treated) tau tg mice across the two studies showing that the range of fast transport rates for these tg mice overlapped with the rate measured for wt mice. Regardless of the precise extent of transport impairment, this model features significant neurofibrillary degeneration unlike the model we studied, as they pointed out. If high tau levels have direct effects on transport in this model, it would need to be shown in the absence of indirect effects on transport arising from tau-mediated neurodegeneration.

The Alzforum comments and data from in vivo and in vitro studies emphasize that the degree of tau overexpression is critical to any observed effects on axonal transport. Transport impairments have been associated so far only with very high tau expression, although Fred Van Leuven points out that the emergence of pathological effects at any level of tau overexpression may be influenced by additional vulnerabilities of the animal contributed by genotype and phenotype. Finally, any observed effect of tau at altered expression levels will need to be evaluated in the light of disease relevance, namely, clear evidence that tau levels are comparably altered in Alzheimer disease or other tauopathies. Existing evidence on this issue is not so clear.

References:
1. Spittaels K, Van den Haute C, Van Dorpe J, Bruynseels K, Vandezande K, Laenen I, Geerts H, Mercken M, Sciot R, Van Lommel A, Loos R, Van Leuven F. Prominent axonopathy in the brain and spinal cord of transgenic mice overexpressing four-repeat human tau protein. Am J Pathol. 1999 Dec 1;155(6):2153-65. Abstract

2. Spittaels K, Van den Haute C, Van Dorpe J, Geerts H, Mercken M, Bruynseels K, Lasrado R, Vandezande K, Laenen I, Boon T, Van Lint J, Vandenheede J, Moechars D, Loos R, Van Leuven F. Glycogen synthase kinase-3beta phosphorylates protein tau and rescues the axonopathy in the central nervous system of human four-repeat tau transgenic mice. J Biol Chem. 2000 Dec 29;275(52):41340-9. Abstract

3. Künzi V, Glatzel M, Nakano MY, Greber UF, Van Leuven F, Aguzzi A. Unhampered prion neuroinvasion despite impaired fast axonal transport in transgenic mice overexpressing four-repeat tau. J Neurosci. 2002 Sep 1;22(17):7471-7. Abstract

4. Schindowski K, Bretteville A, Leroy K, Bégard S, Brion JP, Hamdane M, Buée L. Alzheimer's disease-like tau neuropathology leads to memory deficits and loss of functional synapses in a novel mutated tau transgenic mouse without any motor deficits. Am J Pathol. 2006 Aug 1;169(2):599-616. Abstract

5. Zhang B, Maiti A, Shively S, Lakhani F, McDonald-Jones G, Bruce J, Lee EB, Xie SX, Joyce S, Li C, Toleikis PM, Lee VM, Trojanowski JQ. Microtubule-binding drugs offset tau sequestration by stabilizing microtubules and reversing fast axonal transport deficits in a tauopathy model. Proc Natl Acad Sci U S A. 2005 Jan 4;102(1):227-31. Abstract

6. Ishihara T, Hong M, Zhang B, Nakagawa Y, Lee MK, Trojanowski JQ, Lee VM. Age-dependent emergence and progression of a tauopathy in transgenic mice overexpressing the shortest human tau isoform. Neuron. 1999 Nov 1;24(3):751-62. Abstract

View all comments by Ralph Nixon
View all comments by Aidong Yuan

This interesting paper uses classical assays to measure slow transport and some markers for fast axonal transport. It sees no differences in gross rates of axonal transport in the absence of tau, or upon tau overexpression. Thus, this work differs significantly from observations made by the Mandelkow lab looking at the effects of tau expression on axonal transport and organelle localization.

The reasons for the apparent discrepancies between these observations remain to be determined. One possibility is the nature of the cargos under investigation, as the paper by Yuan et al. is focusing primarily on cargos undergoing slow transport along the axon. An alternate possibility is the relatively insensitive nature of the transport assay used by Yuan et al. For example, previous work using this approach in the SOD1 model for familial ALS did not reveal significant defects in anterograde transport until relatively late in disease (Zhang et al., 1997; Williamson and Cleveland, 1999), whereas...  Read more

This interesting paper uses classical assays to measure slow transport and some markers for fast axonal transport. It sees no differences in gross rates of axonal transport in the absence of tau, or upon tau overexpression. Thus, this work differs significantly from observations made by the Mandelkow lab looking at the effects of tau expression on axonal transport and organelle localization.

The reasons for the apparent discrepancies between these observations remain to be determined. One possibility is the nature of the cargos under investigation, as the paper by Yuan et al. is focusing primarily on cargos undergoing slow transport along the axon. An alternate possibility is the relatively insensitive nature of the transport assay used by Yuan et al. For example, previous work using this approach in the SOD1 model for familial ALS did not reveal significant defects in anterograde transport until relatively late in disease (Zhang et al., 1997; Williamson and Cleveland, 1999), whereas live cell assays for vesicular transport that we have done on neurons from the SOD1 mouse (Perlson et al., submitted) do reveal significant changes in transport velocities. Alternatively, isoform expression may affect the observations, as we and others have seen pronounced differences in the effects of tau isoforms on microtubule motors at the single molecule level.

In our recent work on the differential effects of tau on kinesin and dynein (Dixit et al., 2008), the motility of individual kinesin motors was strongly affected when the motors encountered patches of tau bound along the microtubule. These encounters most frequently resulted in either pausing of the motor or detachment of the kinesin from the microtubule. It is possible that for larger cargos undergoing longer-distance transport, these molecular-level changes are damped out. This is an interesting question that needs to be investigated further with high-resolution assays. Alternatively, as Yuan et al. suggest, there may be compensatory mechanisms operating in vivo that mitigate the effects seen at the molecular level.

Thus, their argument is a good one: we need to keep examining in vivo biology in concert with higher-resolution studies that offer more mechanistic insights.

View all comments by Erika Holzbaur

These two reports from Scott Brady’s and Larry Goldstein’s laboratories are highly significant because they extend the concept that neurodegenerative disease is caused by impaired axonal transport, beyond more common disorders like Alzheimer's, to also include triplet-repeat diseases. The implication is that multiple neurodegenerative diseases may share a similar mechanism. This notion was proposed nearly 20 years ago by Carlton Gajdusek, but many years went by before sufficient technical advances occurred in AD research to provide circumstantial and experimental data supporting this view. Traction in this area began with the demonstration that tau (a microtubule binding protein) was the building block of AD neurofibrillary tangles (NFTs). Also helpful was the resolution of the controversy over the role of NFT formation in AD in 1991 by studies showing that abnormally phosphorylated CNS tau proteins (PHFtau) form the paired helical filaments in AD NFTs, and that excessive phosphorylation of PHFtau reduced its...  Read more

These two reports from Scott Brady’s and Larry Goldstein’s laboratories are highly significant because they extend the concept that neurodegenerative disease is caused by impaired axonal transport, beyond more common disorders like Alzheimer's, to also include triplet-repeat diseases. The implication is that multiple neurodegenerative diseases may share a similar mechanism. This notion was proposed nearly 20 years ago by Carlton Gajdusek, but many years went by before sufficient technical advances occurred in AD research to provide circumstantial and experimental data supporting this view. Traction in this area began with the demonstration that tau (a microtubule binding protein) was the building block of AD neurofibrillary tangles (NFTs). Also helpful was the resolution of the controversy over the role of NFT formation in AD in 1991 by studies showing that abnormally phosphorylated CNS tau proteins (PHFtau) form the paired helical filaments in AD NFTs, and that excessive phosphorylation of PHFtau reduced its ability to bind microtubules (MTs) and stabilize them in order to support axonal transport. For a detailed review of this research area, see Lee et al., 2001.

Thus, years before it was discovered that loss of tau function was the consequence of tau gene mutations in hereditary tauopathies, such as frontotemporal dementia with parkinsonism linked to chromosome 17 or FTDP-17, it was already appreciated that wild-type tau, when altered by hyperphosphorylaton in AD, sustained a loss of function that might impair axonal transport and so lead to a neurodegenerative disease. This prompted the hypothesis that the generation of PHFtau depletes neurons of tau able to bind microtubules, thereby leading to brain degeneration in AD. This model predicted that: 1) the conversion of tau into PHFtau disrupts MT-based transport as well as perhaps physically “blocking” transport due to accumulations of PHFs in neurons and their processes, and 2) the failure of neurons to export proteins from the cell body to distal processes and to retrieve substances (e.g., trophic factors) internalized at axon terminals compromises neuronal viability. It was proposed that these events would culminate in neuronal dysfunction and degeneration leading to the onset/progression of AD. Remarkably, nearly all of the predictions of this disease model of tau pathology in AD and related tauopathies were validated in the last four years through studies of tau-transgenic mice. Some of these provided experimental proof that neurodegeneration caused by tau aggregation was linked to axonal transport failure (Isihara et al., 1999). Indeed, a consensus in favor of this notion appears to be building and a whole issue of (Neuromolecular Medicine) was dedicated to this topic last year.

It will be important to confirm and extend the findings described in these two studies, which differ in some details. Both papers conclude that impaired axonal transport plays a significant role in mechanisms underlying neurodegeneration. Significantly, the views proposed in these papers complement and extend the earlier concept of a loss of function and impairment of axonal transport when tau is altered in AD, FTDP-17, and other tauopathies. Specifically, the authors of both of these Neuron papers propose that polyQ species acquire a toxic gain of function that disrupts axonal transport. By adding a toxic gain of function in disease proteins to the more well-documented loss of normal function (as in hyperphosphorylated tau), and linking these abnormalities to impaired axonal transport, these two studies open up bold new avenues for advancing insights into mechanisms of neurodegenerative disease. All of this could have substantial implications for the discovery of new and better therapies for AD and other less common neurodegenerative diseases such as Huntington’s, FTDP-17, other tauopathies, and related disorders.

View all comments by John Trojanowski

Building on their earlier provocative findings linking APP function to fast axonal transport, Stokin and colleagues, in this latest report, reinforce several important themes that are emerging from recent studies. First, significant neuronal pathobiology, especially evidence of altered vesicular trafficking, can be detected very early in Alzheimer disease (AD), before classical Alzheimer neuropathology appears. Second, these early disturbances at least partly stem from a behavior of APP or one of its processed forms; however, the issue of whether Aβ generation is an effect rather than the cause of this pathophysiology needs to be considered seriously. Finally, beyond its implications for Aβ generation, the defective vesicular transport observed in this study, and early endosomal-lysosomal dysfunction seen in other studies, are in their own right very likely to impair synapse function and axon/dendrite maintenance (Nixon, 2005). The new studies by the Goldstein group will hopefully encourage further exploration of these research themes, which are relatively understudied....  Read more

Building on their earlier provocative findings linking APP function to fast axonal transport, Stokin and colleagues, in this latest report, reinforce several important themes that are emerging from recent studies. First, significant neuronal pathobiology, especially evidence of altered vesicular trafficking, can be detected very early in Alzheimer disease (AD), before classical Alzheimer neuropathology appears. Second, these early disturbances at least partly stem from a behavior of APP or one of its processed forms; however, the issue of whether Aβ generation is an effect rather than the cause of this pathophysiology needs to be considered seriously. Finally, beyond its implications for Aβ generation, the defective vesicular transport observed in this study, and early endosomal-lysosomal dysfunction seen in other studies, are in their own right very likely to impair synapse function and axon/dendrite maintenance (Nixon, 2005). The new studies by the Goldstein group will hopefully encourage further exploration of these research themes, which are relatively understudied.

The report provides evidence for an early failure of anterograde axonal transport in AD and implicates the transport motor, kinesin-1, as one route to this failure. This could nicely explain an initial report suggesting that KLC1 polymorphisms may influence risk for AD. A more generalized defect of vesicular transport in AD could also be envisioned. A dysfunctional microtubule "track," possibly involving tau, or an altered vesicular cargo, perhaps involving post-translationally modified APP, would be expected to impair not only anterograde axonal transport, but also retrograde traffic in dendrites, where dystrophy and accumulation of vesicular cargoes is more profoundly affected than in axons. The accumulating vesicles in dystrophic neurites in the Alzheimer brain include many of lysosomal origin, as initially pointed out by Robert Terry and colleagues. At the same time, many, if not most, correspond to autophagic vacuoles, which are early and late compartments of macroautophagy, a pathway for the turnover of organelles and long-lived proteins (Nixon et al. 2005). Interestingly, autophagic vacuoles are enriched in γ-secretase activity and contain Aβ in addition to the necessary components to generate Aβ (Yu et al., 2004). In the Stokin et al. study, a proportion of the vesicles accumulating in pathologic axons of the mouse model appear to have the distinctive double limiting-membrane morphology of early autophagic vacuoles, suggesting one possible source for the extra Aβ in these mice. Endosomes, another site of amyloidogenic APP processing, are known to be abundant anterograde vesicular cargoes in axons, so it will be interesting in future studies to sort out the relative contributions of these different vesicular compartments to the Aβ effect.

References:
Nixon RA. Endosome function and dysfunction in Alzheimer's disease and other neurodegenerative diseases. Neurobiol Aging. 2005 Mar;26(3):373-82. Abstract

Nixon RA; Wegiel J, Kumar A, Yu WH, Peterhoff C, Cataldo A, Cuervo AM. Extensive Involvement of Autophagy in Alzheimer Disease: An Immuno-Electron Microscopy Study. Journal of Neuropathology and Experimental Neurology: 2005 Feb; 64(2):113-122.

Yu WH, Kumar A, Peterhoff C, Shapiro Kulkane L, Uchiyama Y, Lamb BT, Cuervo AM, Nixon RA. Autophagic vacuoles are enriched in APP-secretase activities: Implications for Aβ peptide over-production and localization in Alzheimer’s disease. International Journal of Biochemistry and Cell Biology 2004; 36:2531-2540. Abstract

See also ARF related conference report.

View all comments by Ralph Nixon

The paper by Stokin et al is most remarkable and very convincing. Reducing axonal transport enhanced axonopathy, increased intracellular Aβ levels and extracellular deposition. Stimulation of APP cleavage may be the consequence of enhanced presence of APP-containing vesicles in axonal and/or somatodendritic compartments due to mistrafficking. Increased intraneuronal Aβ accumulation as a consequence has been earlier shown to trigger neuronal death in APP/PS1 mouse models. Impaired axonal transport may be the result of age-dependent processes leading to axonal deafferentiation and loss of synaptic contacts.

In my opinion, this is a milestone paper, because it shows that intraneuronal deficits, like axonopathy, are observed prior to plaque induction. It provides further evidence for a central role of intraneuronal Aβ accumulation in the pathological processes of Alzheimer disease.

View all comments by Thomas Bayer

This paper by Stokin et al. from the lab of Larry Goldstein has some interesting and important findings. I think the finding that APPsw transgenics having half the dose of kinesin-1 have increased Aβ deposition and pathology strongly argues that normal axonal transport is involved in the development of Aβ-related pathologies in AD. This is important, as it suggests that augmentation of this function or factors that prevent axonopathy may be protective against AD.

The finding that there are neuritic swellings in very young APP transgenic mice is interesting, but whether this is relevant to AD is unclear. First, these swellings are smaller and different in appearance than the neuritic dystrophy around amyloid deposits. Second, and more importantly, the APP transgenic mice being studied overexpress mutant APP many-fold. Humans with AD of any type do not overexpress mutant APP (except in Down syndrome, in which there is APP overexpression but at a much lower level than in these mice). The overexpression of human APP increases human Aβ (required for Aβ...  Read more

This paper by Stokin et al. from the lab of Larry Goldstein has some interesting and important findings. I think the finding that APPsw transgenics having half the dose of kinesin-1 have increased Aβ deposition and pathology strongly argues that normal axonal transport is involved in the development of Aβ-related pathologies in AD. This is important, as it suggests that augmentation of this function or factors that prevent axonopathy may be protective against AD.

The finding that there are neuritic swellings in very young APP transgenic mice is interesting, but whether this is relevant to AD is unclear. First, these swellings are smaller and different in appearance than the neuritic dystrophy around amyloid deposits. Second, and more importantly, the APP transgenic mice being studied overexpress mutant APP many-fold. Humans with AD of any type do not overexpress mutant APP (except in Down syndrome, in which there is APP overexpression but at a much lower level than in these mice). The overexpression of human APP increases human Aβ (required for Aβ aggregation in mice), but also may be resulting in other biological effects of mutant APP overexpression.

It is possible that the neuritic changes described in the young APPsw mice are secondary to increased soluble Aβ. It is also possible that they are due to APPsw overexpression. Appropriate controls to sort this out might be overexpression of APPsw with the Aβ region changed in sequence or determining whether pharmacological or other inhibition of Aβ blocks the early neuritic changes. While the neuritic swellings seen in young APPsw mice are interesting and may have relevance to AD, I think this remains unclear at this point.

View all comments by David Holtzman

Kinesin molecular motor protein is involved axonal transport along microtubules. Tau protein is a major constituent of mircrotubules and thus disruption of tau (hyperphophorylation as an example) or any other part of microtubules have been shown to interfere with anterograde transport and retrograde transport. In the case of AD the research seems to point more towards APP buildup as a result of neuronal structure degradation. A drastic reduction of kinesin is merely a symptom and not directly causal of APP and amyloid beta. Presenilin mutations that affect the enzyme's activity in cutting APP are shown in a wide variety of axonal dysfucntion in AD patients.

View all comments by Jacob Mack

Aluminum could be a co-factor in the findings of Stokin and collegues. Aluminum was found to inhibit neurofilament assembly, cytoskeletal incorporation, and axonal transport by Shea et al, 1997. Deloncle et al, 2001 found that aluminum L-glutamate causes massive mitochondrial swelling in the hippocampus of younger laboratory rats that mimics similar effects of the aging process in older animals. Stokin et al. found mitochondria in the axons. Aluminum is known to interfere with ATP and is linked with neurofibrillary degeneration. Bioaccumulation of aluminum in the human brain over the lifespan exposes the aging brain to potentially significant dosages.

References:
T.B. Shea, E. Wheeler and C. Jung, Aluminum inhibits neurofilament assembly, cytoskeletal incorporation and axonal transport. Dynamic nature of aluminum-induced perikaryl neuro-filament accumulations as revealed by subunit turnover, Mol Chem Neuropathol 32(1-3)1997, 17-39

R. Deloncle, F. Huguet, B. Fernandez, N. Quellard, P. Babin and O. Guillard, Ultrastructural study of rat hippocampus after chronic adminstration of aluminum L-glutamate: an acceleration of the aging process, Exp Gerontol 36(2) 2001, 231-44

View all comments by Erik Jansson

This excellent study clearly demonstrates that axonal damage occurs long before amyloid deposition in both early stage AD and an APP mouse model. Furthermore, the authors demonstrate that reduced expression of the motor protein KCL-1 increases both the production and deposition of Aβ. However, it is unclear which comes first, the generation of soluble toxic Aβ species and then disruption of axonal transport, or disruption of transport leading to increased Aβ production and subsequent generation of toxic assemblies. A clear understanding of the pathogenic sequence is essential for the rational development of therapies and thus the temporal relationship between axonopathy and soluble Aβ species demands further investigation. Specifically, in light of the finding that anti-Aβ antibodies can lead to the clearance of early hyperphosphorylated forms of tau, it would be worthwhile determining if either passive or active immunization can rescue the pre-amyloid axonopathy.

View all comments by Dominic Walsh

The study of the biology of APP and its proteolytic products, although pioneered in the early 1990s by Eddie Koo, Joseph Buxbaum, Sam Sisodia, and others, has nevertheless remained mostly out of the limelight until the last few years. The present study from Elaine Bearer’s laboratory now illuminates part of a picture that has been taking shape in the last few years suggesting that APP is likely involved in the modulation of synaptic activity in adults (Priller et al., 2006; Yang et al., 2005; Seabrook et al., 1999), in synapse formation and function (Wang et al., 2005), and in neuronal migration and adhesion during development (Herms et al., 2004).

APP is a synaptic protein that is anterogradely transported to terminals. A few years ago Kamal et al. suggested that the C-terminus of APP could serve as a receptor for kinesin (  Read more

The study of the biology of APP and its proteolytic products, although pioneered in the early 1990s by Eddie Koo, Joseph Buxbaum, Sam Sisodia, and others, has nevertheless remained mostly out of the limelight until the last few years. The present study from Elaine Bearer’s laboratory now illuminates part of a picture that has been taking shape in the last few years suggesting that APP is likely involved in the modulation of synaptic activity in adults (Priller et al., 2006; Yang et al., 2005; Seabrook et al., 1999), in synapse formation and function (Wang et al., 2005), and in neuronal migration and adhesion during development (Herms et al., 2004).

APP is a synaptic protein that is anterogradely transported to terminals. A few years ago Kamal et al. suggested that the C-terminus of APP could serve as a receptor for kinesin (Kamal et al., 2000), but this observation was subsequently questioned by Lazarov et al. (Lazarov et al., 2005). The present study by Satpute-Krishnan et al. provides strong evidence that the C-terminus of APP may indeed contain sequences sufficient for its association with axonal transport components. The careful experiments addressed this question using a fairly well-defined system, the squid giant axon, and the investigators’ observations indicate that the C-terminal domain of APP, either through a direct interaction with kinesin or indirectly via scaffolding proteins such as JIPs, participates in fast anterograde axonal transport. Quoting their discussion, “The robust motility of C99 beads in the intact axon argues for a physiological role of APP in recruitment of anterograde transport machinery inside cells.” It certainly does, and it comes as no surprise. Although the study by Satpute-Krishnan et al. does not answer the question of whether the interaction of APP with kinesin is or is not direct, it significantly adds to the rapidly growing evidence suggesting a crucial role of the C-terminus of APP (and possibly its family members APLP1 and 2) in neuronal biology, possibly at synaptic sites.

The remarkable conservation of the C-terminal sequences of APP across phyla suggests conservation of function. Supporting this idea, the phenotypes of APP/APLP2 double and APP/APLP1/APLP2 triple knockouts and those of two prominent APP-interacting proteins (X11 and the Fe65 family) involve alterations in neuronal function, synaptic formation, function, and regulation (Wang et al., 2005; Ho et al., 2003; Yang et al., 2005; Priller et al., 2006), and in the case of the Fe65/FE65L double and APP/APLP1/APLP2 triple knockouts, result in cortical dysplasias and heterotopias (Herms et al., 2004, Guenette et al., 2006). Interestingly, it was recently shown that transgenic expression of AICD in combination with Fe65 causes alterations in signaling (Ryan and Pimplikar, 2005) and activation of proteins involved in growth cone collapse and axonal guidance.

Why is this important? Most of all, because a significant component of amyloid-β toxicity requires multimerization of APP and cleavage of its C-terminus at Asp664 (Lu et al., 2003; Lu et al., 2003; Shaked et al., 2006). This cleavage not only releases a toxic peptide, but also removes the sequences required for the formation of a multiplicity of protein complexes at APP’s cytoplasmic domain, and as Satpute-Krishnan now suggest, for fast axonal transport. Consistent with what may be an important role of the extreme C-terminal sequences of APP in transducing amyloid-β toxicity, we recently showed that stabilization of APP’s cytoplasmic tail by mutation of the Asp664 cleavage site had a dramatic effect in the development of AD-like deficits in transgenic mice (Galvan et al., 2006)—even in the presence of abundant amyloid-β. With this in mind, the question arises as to whether cleavage at Asp664 while in transit towards synaptic sites would, as expected, prevent delivery of the molecule to its destination—and if the hypothesis of Kamal et al. is correct, whether it would affect the delivery of any subset of associated axonal transport vesicles. Thus, a population of Asp664-intact (transport-competent) and Asp664-cleaved (transport-incompetent) APP molecules may exist. Satpute-Krishnan et al. estimate that 3,000 copies of APP may be associated with each motile bead in their system; although in this study they don’t address the question of what is the minimal number of APP molecules required for transport, it is conceivable that transport-incompetent (Asp664-cleaved) APP molecules may be “carried along” in vesicles containing a sufficient number of transport-competent (Asp664-intact) APP. Cleavage of APP at Asp664 would thus affect not only the transport-competence (and thus the rate of delivery) of APP to neuronal terminals, but since the motifs required for the interaction of APP with a variety of cellular functions reside downstream of Asp664, it would also affect the overall signaling ability of populations of APP molecules at their destination at synaptic sites.

View all comments by Veronica Galvan

In this paper, Yuan et al. report elegant studies of axonal transport in vivo using tau transgenic and tau knockout mice that overexpress human tau isoforms or completely lack tau expression, respectively. These studies sought to elucidate the consequences of too much tau or a complete lack of tau on axonal transport in living mice. This is a most welcome study by the Nixon lab, which has made important contributions to the understanding of axonal transport dynamics for over 2 decades. This study makes increasingly clear that there is a critical need for more studies of this kind to understand how perturbations in tau expression levels or tau pathologies are linked to axonal transport failure and tau-mediated neurodegeneration in Alzheimer disease (AD) and related tauopathies. Indeed, there is growing evidence that failed axonal transport might be the underlying basis for several neurodegenerative diseases in addition to tauopathies (8). It is especially important and timely to undertake in vivo axonal transport studies using the classic Lasek paradigm for measuring rates of...  Read more

In this paper, Yuan et al. report elegant studies of axonal transport in vivo using tau transgenic and tau knockout mice that overexpress human tau isoforms or completely lack tau expression, respectively. These studies sought to elucidate the consequences of too much tau or a complete lack of tau on axonal transport in living mice. This is a most welcome study by the Nixon lab, which has made important contributions to the understanding of axonal transport dynamics for over 2 decades. This study makes increasingly clear that there is a critical need for more studies of this kind to understand how perturbations in tau expression levels or tau pathologies are linked to axonal transport failure and tau-mediated neurodegeneration in Alzheimer disease (AD) and related tauopathies. Indeed, there is growing evidence that failed axonal transport might be the underlying basis for several neurodegenerative diseases in addition to tauopathies (8). It is especially important and timely to undertake in vivo axonal transport studies using the classic Lasek paradigm for measuring rates of axonal transport in living animals (8). This was done in the present study by Yuan et al., as well as in previous studies we conducted on tau transgenic mice (4,13).

Cell culture studies to examine this issue are being reported, but it is not always clear how well these model systems recapitulate what occurs in living animals. This point is driven home by the data reported here because the results on tau overexpression do not support the cell culture reports by Stamer et al. (10) suggesting that excess tau in the absence of fibrillary tau inclusions “clogs” microtubules (MTs) and impedes axonal transport. The in vivo data reported by Yuan et al. do not confirm or support these prior in vitro studies. It will be important to understand the basis of these discrepant findings in cell culture versus animal model systems. However, it also is noteworthy that Yuan et al. point out that there is no unequivocal evidence that tau is overexpressed in AD or any other known human tauopathy.

At first blush, the data reported by Yuan et al. also appear to differ from studies we have reported on tau transgenic mice that overexpress the smallest human tau isoform to perturb the 3R-to-4R tau ratio in these mice. However, there are significant differences between the transgenic mice in our studies (T44 line) and those studies in the report by Yuan (8c line). Most significantly, the 8c mice overexpress human tau isoforms but do not develop neurofibrillary tau pathology, as do our T44 transgenic mice (4,5,14). Thus, human tau overexpression that results in the development of neurofibrillary tau pathology can model authentic human neurodegenerative tauopathies, whereas there is no clear human counterpart, disease or otherwise, of tau overexpression alone. To understand the significance of this, some background on tau pathology in AD and related tauopathies is important.

Briefly, as reviewed recently (2), most early insights into AD and other tauopathies came from studies of AD. At the same time, a substantial amount of data has come more recently from studies of non-AD tauopathies, and it has shown that tau pathology is the critical underlying abnormality that links AD and these disorders to a shared mechanism of neurodegeneration. (View Slide reprinted from Journal of Alzheimer Disease)

For example, when tau becomes hyperphosphorylated or, as a result of other mechanisms, disengages from MTs, higher concentrations of cytosolic tau lead to tau fibrillization and the formation amyloid-like paired helical filaments that aggregate to form neurofibrillary tangles (NFTs). Thus, as a consequence of this tau amyloidosis in the CNS, normal tau proteins will be sequestered. This depletes the levels of normal tau in affected CNS neurons. As a result, this leaves less normal tau available to stabilize MTs, and, when MTs are destabilized, this compromises intraneuronal transport leading to neurodegeneration. Most elements of this tau-mediated neurodegeneration hypothesis in AD and related tauopathies were demonstrated to occur in experimental animals, when Ishihara et al. showed for the first time that the development of fibrillary tau pathology was linked to MT loss, impaired fast axonal transport (FAT) using the Lasek et al. paradigm, and neurodegeneration (4).

This study and subsequent studies by our group indicated that the T44 line recapitulates features of AD tau pathology. However, the overall phenotype of the T44 mice is most similar to Guam amyotrophic lateral sclerosis/Parkinson’s dementia complex or ALS/PDC (5,11). Thus, the T44 mice do produce a phenotype that recapitulates features of authentic human neurodegenerative tauopathies.

Yuan et al. misread the Zhang et al. paper (13) when they infer on page 1686-1687 that the Zhang et al. study of the T44 mice contradicts the data in Ishihara et al. on these same mice. The faster rate of FAT reported in Zhang et al. was the result of a treatment intervention with an MT-stabilizing drug, i.e., taxol. The use of taxol to treat the T44 mice was designed to functionally replace tau and stabilize MTs to offset the sequestration of tau into inclusions in the T44 mice. Specifically, Zhang et al. used the same Lasek et al. paradigm described in Ishihara et al. (and also by Yuan et al.). Zhang et al. confirmed that sham-treated T44 tau-transgenic mice had impaired FAT and the other phenotypic features described by Ishihara et al. (4). By contrast, taxol treatment corrected the FAT deficit as well as the motor impairment in these mice, and this was associated with increased numbers of MTs relative to sham-treated T44 mice (14). Thus, taxol made up for the loss of tau function that resulted from tau sequestration in fibrillary tau inclusions in the T44 line. While there are several problems with taxol for use as therapy for AD and related tauopathies, work is in progress to develop other MT-stabilizing compounds that could go on to become potential disease-modifying therapies for tauopathy patients (1,12).

These and other studies of transgenic mouse models of tauopathies have increased efforts to develop tau-focused interventions for AD and related tauopathies. Some interventions are directed at abrogating/reversing tau fibrillization or tau hyperphosphorylation; others are designed to stabilize MTs by compensating for the sequestration of tau in NFTs (9). However, while it may be desirable to suppress mutant forms of tau in hereditary tauopathies or in tauopathies with an abnormal ratio of 3R versus 4R tau isoforms (6), reducing tau levels in AD (7), especially to a degree that compromises MT stability, is likely to have long-term deleterious effects as exemplified by the studies cited above (4,13). For example, Hirokowa’s laboratory reported that tau knockout mice develop cognitive and motor abnormalities with age, thereby signifying that reducing tau levels may have negative consequences (3). However, there are few studies examining the behavioral and functional consequences of reducing CNS tau levels over the life span. The present study makes it clear that far more research is needed on this topic, as well as on the role of axonal transport in animal models of tauopathies in order to identify the optimal targets for tau-focused drug discovery for AD and related tauopathies.

References:
1. Ballatore C, Hyde E, Deiches RF, Lee VM, Trojanowski JQ, Huryn D, Smith AB. Paclitaxel C-10 carbamates: potential candidates for the treatment of neurodegenerative tauopathies. Bioorg Med Chem Lett. 2007 Jul 1;17(13):3642-6. Abstract

2. Ballatore C, Lee VM, Trojanowski JQ. Tau-mediated neurodegeneration in Alzheimer's disease and related disorders. Nat Rev Neurosci. 2007 Sep 1;8(9):663-72. Abstract

3. Ikegami S, Harada A, Hirokawa N. Muscle weakness, hyperactivity, and impairment in fear conditioning in tau-deficient mice. Neurosci Lett. 2000 Feb 4;279(3):129-32. Abstract

4. Ishihara T, Hong M, Zhang B, Nakagawa Y, Lee MK, Trojanowski JQ, Lee VM. Age-dependent emergence and progression of a tauopathy in transgenic mice overexpressing the shortest human tau isoform. Neuron. 1999 Nov 1;24(3):751-62. Abstract

5. Ishihara T, Zhang B, Higuchi M, Yoshiyama Y, Trojanowski JQ, Lee VM. Age-dependent induction of congophilic neurofibrillary tau inclusions in tau transgenic mice. Am J Pathol. 2001 Feb 1;158(2):555-62. Abstract

6. Miller VM, Xia H, Marrs GL, Gouvion CM, Lee G, Davidson BL, Paulson HL. Allele-specific silencing of dominant disease genes. Proc Natl Acad Sci U S A. 2003 Jun 10;100(12):7195-200. Abstract

7. Roberson ED, Scearce-Levie K, Palop JJ, Yan F, Cheng IH, Wu T, Gerstein H, Yu GQ, Mucke L. Reducing endogenous tau ameliorates amyloid beta-induced deficits in an Alzheimer's disease mouse model. Science. 2007 May 4;316(5825):750-4. Abstract

8. Roy S, Zhang B, Lee VM, Trojanowski JQ. Axonal transport defects: a common theme in neurodegenerative diseases. Acta Neuropathol. 2005 Jan 1;109(1):5-13. Abstract

9. Skovronsky DM, Lee VM, Trojanowski JQ. Neurodegenerative diseases: new concepts of pathogenesis and their therapeutic implications. Annu Rev Pathol. 2006 Jan 1;1():151-70. Abstract

10. Stamer K, Vogel R, Thies E, Mandelkow E, Mandelkow EM. Tau blocks traffic of organelles, neurofilaments, and APP vesicles in neurons and enhances oxidative stress. J Cell Biol. 2002 Mar 18;156(6):1051-63. Abstract

11. Trojanowski JQ, Ishihara T, Higuchi M, Yoshiyama Y, Hong M, Zhang B, Forman MS, Zhukareva V, Lee VM. Amyotrophic lateral sclerosis/parkinsonism dementia complex: transgenic mice provide insights into mechanisms underlying a common tauopathy in an ethnic minority on Guam. Exp Neurol. 2002 Jul 1;176(1):1-11. Abstract

12. Trojanowski JQ, Smith AB, Huryn D, Lee VM. Microtubule-stabilising drugs for therapy of Alzheimer's disease and other neurodegenerative disorders with axonal transport impairments. Expert Opin Pharmacother. 2005 May 1;6(5):683-6. Abstract

13. Zhang B, Maiti A, Shively S, Lakhani F, McDonald-Jones G, Bruce J, Lee EB, Xie SX, Joyce S, Li C, Toleikis PM, Lee VM, Trojanowski JQ. Microtubule-binding drugs offset tau sequestration by stabilizing microtubules and reversing fast axonal transport deficits in a tauopathy model. Proc Natl Acad Sci U S A. 2005 Jan 4;102(1):227-31. Abstract

View all comments by Virginia Lee
View all comments by John Trojanowski

In this paper, Randy Nixon’s group first demonstrated in vivo that axonal transport rates are not significantly affected by tau deletion or overexpression in mouse brain. The results are highly convincing.

In in vitro studies, the Mandelkows’ group and Hirokawa’s group have suggested that tau overexpression inhibits anterograde transport in cultured cells and neurons. Recently, Holzbaur’s group indicated that when kinesin motor protein encountered tau patches on microtubules, composed of 10 tau molecules, it detached from microtubules (Dixit et al., 2008). However, monomeric tau levels 20-fold above physiological concentration did not affect axonal transport in squid axon (Morfini et al., 2007). Taken together, aggregated tau on microtubules, but not monomeric tau, may induce inhibition of axonal transport.

Ishihara and colleagues showed that expressing the shortest human tau fivefold to 10-fold over endogenous tau inhibited axonal transport (  Read more

In this paper, Randy Nixon’s group first demonstrated in vivo that axonal transport rates are not significantly affected by tau deletion or overexpression in mouse brain. The results are highly convincing.

In in vitro studies, the Mandelkows’ group and Hirokawa’s group have suggested that tau overexpression inhibits anterograde transport in cultured cells and neurons. Recently, Holzbaur’s group indicated that when kinesin motor protein encountered tau patches on microtubules, composed of 10 tau molecules, it detached from microtubules (Dixit et al., 2008). However, monomeric tau levels 20-fold above physiological concentration did not affect axonal transport in squid axon (Morfini et al., 2007). Taken together, aggregated tau on microtubules, but not monomeric tau, may induce inhibition of axonal transport.

Ishihara and colleagues showed that expressing the shortest human tau fivefold to 10-fold over endogenous tau inhibited axonal transport (Ishihara et al., 1999), although Nixon’s group reports no inhibition of axonal transport in fourfold overexpression of human tau. Therefore, it is possible that the effect of tau on axonal transport in vivo may be dependent on the level of tau overexpression.

Even if we accept that tau overexpression induces axonal transport, and that it may be a cause of neuronal dysfunction or synapse loss in tauopathy, the question still remains how tau impairs neuronal function in a specific brain region in tauopathy. If tau overexpression induces impairment of axonal transport, tau Tg mice must show neuronal dysfunction in the entire brain from a young age on. We recently showed that human wild-type tau (the longest form), expressed about threefold over endogenous levels, induces neural dysfunction in entorhinal cortex at old age (more than 20 months), accompanied by synapse loss and accumulation of hyperphosphorylated tau resulting in a memory deficit, while adult mice (>12 months old) are no different from non-Tg (Kimura et al., 2007). The level of tau on microtubules shows no significant difference between adult and aged mice. Therefore, our results suggest that hyperphosphorylation of tau at old age itself, rather than tau-induced impairment of axonal transport, may be a cause of neuronal dysfunction in the entorhinal cortex, which shows the earliest pathological change in AD.

View all comments by Akihiko Takashima

The picture is more complicated than the title of Yuan et al. would lead us to believe. Our group has generated many tau transgenic mice strains, and at least Tau-4R mice have impaired axonal transport (Spittaels et al., 1999; Künzi et al., 2002), which, moreover, can be rescued by GSK-3β (Spittaels et al., 2000).

Whether or not axonal transport is impaired depends not only on expression levels, as our Tau-4R mice expressed only about twofold over endogenous mouse tau, and we did not observe aggregates of tau.

Other factors must play a role, from the actual tau isoform and promoter used, up to integration site effects. The latter is illustrated by the "selection" of tau mutant mice (Schindowsky et al., 2006). Other, as yet unknown factors play a role, based on heterogeneity of phenotype, gender differences, variability in response to treatments, etc.

There is clearly more to tau and transport than currently meets the eye (just as is the case with APP).

References:
Spittaels K, Van den Haute C, Van Dorpe J, Bruynseels K, Vandezande K, Laenen I, Geerts H, Mercken M, Sciot R, Van Lommel A, Loos R, Van Leuven F. Prominent axonopathy in the brain and spinal cord of transgenic mice overexpressing four-repeat human tau protein. Am J Pathol. 1999 Dec 1;155(6):2153-65. Abstract

Spittaels K, Van den Haute C, Van Dorpe J, Geerts H, Mercken M, Bruynseels K, Lasrado R, Vandezande K, Laenen I, Boon T, Van Lint J, Vandenheede J, Moechars D, Loos R, Van Leuven F. Glycogen synthase kinase-3beta phosphorylates protein tau and rescues the axonopathy in the central nervous system of human four-repeat tau transgenic mice. J Biol Chem. 2000 Dec 29;275(52):41340-9. Abstract

Künzi V, Glatzel M, Nakano MY, Greber UF, Van Leuven F, Aguzzi A. Unhampered prion neuroinvasion despite impaired fast axonal transport in transgenic mice overexpressing four-repeat tau. J Neurosci. 2002 Sep 1;22(17):7471-7. Abstract

Schindowski K, Bretteville A, Leroy K, Bégard S, Brion JP, Hamdane M, Buée L. Alzheimer's disease-like tau neuropathology leads to memory deficits and loss of functional synapses in a novel mutated tau transgenic mouse without any motor deficits. Am J Pathol. 2006 Aug 1;169(2):599-616. Abstract

View all comments by Fred Van Leuven

These papers represent exciting work describing a new genetic mutation associated with familial ALS. The results further highlight the importance for RNA processing in at least familial forms of motor neuron disease. Much work remains to determine the exact mechanisms by which FUS modulates motor neuron survival. It may be related to that of TDP-43. However, the lack of cytoplasmic aggregation of TDP-43, and rare ubiquitin inclusions in the patients with FUS mutations, suggest the mechanisms may be distinct. It is interesting that FUS protein did not accumulate in the cytoplasm of motor neurons in sporadic ALS patients, again suggestive that the pathogenic mechanisms of mutant FUS-induced motor neuron degeneration may be distinct from that in sporadic ALS.

View all comments by Robert Bowser

These studies raise interesting questions about whether one problem in ALS and perhaps other neurodegenerative diseases is that RNA trafficking proteins fail to properly deliver RNAs to dendritic spines. The paper by Kwiatkowski et al. reports evidence that wild-type FUS and TDP-43 may be involved in transporting RNA into dendrites, where it mediates local protein synthesis that can be stimulated by neural activity. The clumping of the mutant form described by both new papers could therefore perturb the transport of RNA. Local protein synthesis in dendrites plays a major role in the activity-dependent modulation of synaptic strength. Changes in synaptic activity have been recently reported in the mouse model of SOD1 mutation (van Zundert et al., 2008), so it will be worthwhile to examine this issue in the FUS mice that will certainly be developed by these investigators.

View all comments by Eric Frank

This is an extremely exiting story in the understanding of ALS pathogenesis. It actually it dates back to 1998—with the first description of mRNA processing errors in sporadic ALS (Lin et al., 1998), which, interestingly, was made not in the SOD1 mouse model. At the same time, the spinal muscular atrophy gene was discovered. SMA is not unlike a childhood ALS, though predominately lower motor neurons are affected in that disease. The SMA gene defect is involved in RNA metabolism. So for the next 10 years, the SMA field has investigated the pathobiology of the defective protein. At the time it made the link between sporadic ALS and the SMA story intriguing. But there was no clear genetic link (or cause for the changes in sporadic ALS).

Feed forward to 2008, when Chris Shaw and others found a true genetic defect in RNA metabolism-based protein TDP-43. (Of course more work needs to be done on that.) And now another gene by the Shaw group, and now verified by the group in Boston, does set a string of targets that all focus on RNA...  Read more

This is an extremely exiting story in the understanding of ALS pathogenesis. It actually it dates back to 1998—with the first description of mRNA processing errors in sporadic ALS (Lin et al., 1998), which, interestingly, was made not in the SOD1 mouse model. At the same time, the spinal muscular atrophy gene was discovered. SMA is not unlike a childhood ALS, though predominately lower motor neurons are affected in that disease. The SMA gene defect is involved in RNA metabolism. So for the next 10 years, the SMA field has investigated the pathobiology of the defective protein. At the time it made the link between sporadic ALS and the SMA story intriguing. But there was no clear genetic link (or cause for the changes in sporadic ALS).

Feed forward to 2008, when Chris Shaw and others found a true genetic defect in RNA metabolism-based protein TDP-43. (Of course more work needs to be done on that.) And now another gene by the Shaw group, and now verified by the group in Boston, does set a string of targets that all focus on RNA metabolism and (lower) motor neurons.

By the way, all these cases appear to predominately involve a lower motor neuron form of ALS. The hint from genetics does suggest more of a loss of function rather than gain, but cell biology will ultimately sort that out. We certainly await the generation of mouse or fly models, which are now well underway for TDP-43. However, this may be a particularly difficult target for specific, non-toxic drug therapy.

View all comments by Jeffrey D. Rothstein

These back-to-back papers on the identification of FUS (fused in sarcoma) gene as a new genetic component of ALS open a new era of research and direct our attention to mRNA biology with respect to disease. After the first identification of mRNA processing errors in ALS patients (Lin, Bristol et al., 1998), the discovery of TDP-43 (Neumann, Sampathu et al., 2006) and now the FUS gene clearly indicate the importance of mRNA management in neurodegenerative diseases. Defects in RNA transcription, splicing, and trafficking may be the reason for cell-type-specific degeneration of motor neurons in ALS. Motor neurons both in the cortex and spinal cord are very large excitatory neurons that extend long axons to their targets and require high levels of energy and protein integrity for survival and function. Defects in transcriptional mechanisms may result in splicing defects, which could give rise to formation of non-functional proteins that would deplete the pool of required proteins for cellular function, and these non-functional proteins may form aggregates that are toxic to neurons. In...  Read more

These back-to-back papers on the identification of FUS (fused in sarcoma) gene as a new genetic component of ALS open a new era of research and direct our attention to mRNA biology with respect to disease. After the first identification of mRNA processing errors in ALS patients (Lin, Bristol et al., 1998), the discovery of TDP-43 (Neumann, Sampathu et al., 2006) and now the FUS gene clearly indicate the importance of mRNA management in neurodegenerative diseases. Defects in RNA transcription, splicing, and trafficking may be the reason for cell-type-specific degeneration of motor neurons in ALS. Motor neurons both in the cortex and spinal cord are very large excitatory neurons that extend long axons to their targets and require high levels of energy and protein integrity for survival and function. Defects in transcriptional mechanisms may result in splicing defects, which could give rise to formation of non-functional proteins that would deplete the pool of required proteins for cellular function, and these non-functional proteins may form aggregates that are toxic to neurons. In addition, defects in the trafficking of mRNA may lead to depletion of key proteins that are in high demand locally for motor neuron function. But if FUS has a general function in mRNA transcription, splicing, and trafficking, why do mutations in this gene cause ALS and not other neurodegenerative diseases? What makes motor neurons more vulnerable in the presence of defective FUS? It could be true that in motor neurons FUS controls the transcription of a distinct set of mRNA that is expressed in a cell-type-specific manner in motor neurons, or that FUS controls the production of a key protein that is highly required in motor neurons when compared to other cell-types, and thus motor neurons may become vulnerable first. FUS seems to be the tip of the iceberg. Finding effectors, binding partners including mRNA, may lead to the identification of key components of both familial and sporadic ALS. More work is on the way!

References:

Kneussel M. Dynamic regulation of GABA(A) receptors at synaptic sites. Brain Res Brain Res Rev. 2002 Jun ;39(1):74-83. Abstract

Lin CL, Bristol LA, Jin L, Dykes-Hoberg M, Crawford T, Clawson L, Rothstein JD. Aberrant RNA processing in a neurodegenerative disease: the cause for absent EAAT2, a glutamate transporter, in amyotrophic lateral sclerosis. Neuron. 1998 Mar;20(3):589-602. Abstract

Neumann M, Sampathu DM, Kwong LK, Truax AC, Micsenyi MC, Chou TT, Bruce J, Schuck T, Grossman M, Clark CM, McCluskey LF, Miller BL, Masliah E, Mackenzie IR, Feldman H, Feiden W, Kretzschmar HA, Trojanowski JQ, Lee VM. Ubiquitinated TDP-43 in frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Science. 2006 Oct 6;314(5796):130-3. Abstract

Vance C, Rogelj B, Hortobágyi T, De Vos KJ, Nishimura AL, Sreedharan J, Hu X, Smith B, Ruddy D, Wright P, Ganesalingam J, Williams KL, Tripathi V, Al-Saraj S, Al-Chalabi A, Leigh PN, Blair IP, Nicholson G, de Belleroche J, Gallo JM, Miller CC, Shaw CE. Mutations in FUS, an RNA processing protein, cause familial amyotrophic lateral sclerosis type 6. Science. 2009 Feb 27;323(5918):1208-11. Abstract

View all comments by P. Hande Ozdinler

The study by Pigino et al. study elegantly highlights a possible mechanism by which Aβ oligomers can influence axonal transport. Though the validity of intracellular Aβ is debatable in the context of human AD pathology, Pigino et al. convincingly show that in a simple model-system of axonal transport, nanomolar levels of Aβ can influence transport; they also provide convincing evidence for the involvement of a specific signaling cascade in this process. The paper is a must-read!

View all comments by Subhojit Roy

We would like to comment on the interesting results of the recent study by Morfini et al. (1). Kinesin-1, a major microtubule motor that transports cargo in the plus-end direction of microtubules, is a heterotetramer consisting of two microtubule-binding, motor polypeptides (the heavy chains; KHCs) and two cargo-binding polypeptides (the light chains; KLCs). Being a soluble, cytoplasmic protein, kinesin-1 needs to bind the cargo in order to transport it. Therefore, recruitment of kinesin-1 to the cargo vesicle, and its release from it, are important regulatory steps of axonal transport. About 10 years ago, Scott Brady’s laboratory identified the first mechanism leading to the release of kinesin-1 from vesicles. According to this model, kinesin-1 is released through the action of the chaperone HSC70, and is nucleotide-dependent and NEM-sensitive (2). One year later, work from Larry Goldstein’s laboratory suggested that the premature release of kinesin-1 from cargo vesicles in neurons could impair fast axonal transport and lead to neuronal pathology and disease (3). Although the...  Read more

We would like to comment on the interesting results of the recent study by Morfini et al. (1). Kinesin-1, a major microtubule motor that transports cargo in the plus-end direction of microtubules, is a heterotetramer consisting of two microtubule-binding, motor polypeptides (the heavy chains; KHCs) and two cargo-binding polypeptides (the light chains; KLCs). Being a soluble, cytoplasmic protein, kinesin-1 needs to bind the cargo in order to transport it. Therefore, recruitment of kinesin-1 to the cargo vesicle, and its release from it, are important regulatory steps of axonal transport. About 10 years ago, Scott Brady’s laboratory identified the first mechanism leading to the release of kinesin-1 from vesicles. According to this model, kinesin-1 is released through the action of the chaperone HSC70, and is nucleotide-dependent and NEM-sensitive (2). One year later, work from Larry Goldstein’s laboratory suggested that the premature release of kinesin-1 from cargo vesicles in neurons could impair fast axonal transport and lead to neuronal pathology and disease (3). Although the mechanisms for the release of kinesin-1 from its vesicular cargos were incompletely understood at that time, the general idea that a premature release of the motor from its cargo could be at the core of the pathology in neurodegenerative diseases turned out to be correct, and generated an increased interest for research in this direction. Thus, work from the Brady and Busciglio laboratories identified at least two pathways for release of kinesin-1 from vesicles and halt of transport, which are likely to be factors leading to the axonal pathology and synaptic failure in Alzheimer’s disease (AD) (4-6).

Both pathways lead to phosphorylation of the KLCs, followed by detachment of kinesin-1 from the cargo, and impairment of vesicle transport. They are initiated by the addition of soluble Aβ oligomers, or expression of FAD-linked presenilin 1 variants, which trigger aberrant activation of casein kinase 2 or glycogen synthase 3β, which phosphorylate the KLCs. Why the phosphorylated kinesin-1 is released from vesicles is still not fully understood.

Along with AD, kinesin-1 is a target for abnormal phosphorylation in other neurodegenerative diseases, such as spinal and bulbar muscular atrophy (SBMA) and Huntington’s disease, as revealed by the studies from the Brady laboratory, including the work featured here (1, 7). However, in this case, the phosphorylation targets the KHCs, and the activated kinase that performs the phosphorylation is the cJun-N-terminal kinease (JNK). The phosphorylation of the KHCs leads to inhibition of binding of kinesin-1 to microtubules. As a result, the kinesin-1-cargo complex is released from the microtubules, and the transport is halted. These studies showed that the abnormal activation of JNK is triggered by the pathogenic, polyglutaminated, mutant proteins characteristic for polyglutamine (polyQ) expansion diseases: polyQ-androgen receptor in SBMA) (7), and polyQ-huntingtin in Huntingon’s disease(1). As the study by Morfini et al. (1) showed, polyQ-huntingtin activates JNK3, a neuron-specific JNK, that in turn phosphorylates KHC at a serine residue critical for the microtubule-binding function of kinesin-1. While in this case JNK3 is aberrantly activated by a disease factor, it is likely that, under normal conditions, the JNK-3 pathway contributes to the regulation of axonal transport.

Interestingly, in the squid axon system used in these studies, polyQ-huntingtin inhibits, not only the anterograde (kinesin-driven), but also the retrograde (cytoplasmic dynein-driven) fast axonal transport (1). It is not clear whether this inhibition of transport in both directions is due to the fact that kinesin-1 and cytoplasmic dynein interact and coordinate each other’s function (8), or is caused by a direct effect on the dynein machinery. Other studies showed that huntingtin regulates dynein-mediated vesicle transport, and can interact with both dynein and its accessory complex, dynactin (9, 10); however, the assays used by Morfini et al. (1) did not detect an interaction of huntingtin with dynein.

Certainly, other mechanisms, besides the release of the kinesin motor from the cargo or the microtubules, could contribute to the pathogenic processes in these neurodegenerative diseases. Other potentially damaging pathways that target the intracellular transport by affecting the cytoskeleton or the supply of ATP (by disrupting mitochondrial function) have been described (reviewed in (11)). Also, the activation of the kinases is likely to lead to the abnormal phosphorylation of other protein targets as well, with detrimental consequences for the function of neurons via mechanisms that may not involve abnormal axonal transport. For now, a picture emerges where the release of kinesin-1 from either cargo or microtubules, followed by impairment of axonal transport, becomes an important component of the pathogenic process in many neurodegenerative diseases. Therefore, it is the time to think of possibilities to correct the deficiencies, or to find means to enhance the disease-inflicted axonal transport.

References:
1. Morfini GA, You YM, Pollema SL, Kaminska A, Liu K, Yoshioka K, Björkblom B, Coffey ET, Bagnato C, Han D, Huang CF, Banker G, Pigino G, Brady ST. Pathogenic huntingtin inhibits fast axonal transport by activating JNK3 and phosphorylating kinesin. Nat Neurosci. 2009 Jul;12(7):864-71. Abstract

2. Tsai MY, Morfini G, Szebenyi G, Brady ST. Release of kinesin from vesicles by hsc70 and regulation of fast axonal transport. Mol Biol Cell. 2000 Jun;11(6):2161-73. Abstract

3. Kamal A, Almenar-Queralt A, LeBlanc JF, Roberts EA, Goldstein LS. Kinesin-mediated axonal transport of a membrane compartment containing beta-secretase and presenilin-1 requires APP. Nature. 2001 Dec 6;414(6864):643-8. Abstract

4. Morfini G, Szebenyi G, Elluru R, Ratner N, Brady ST. Glycogen synthase kinase 3 phosphorylates kinesin light chains and negatively regulates kinesin-based motility. EMBO J. 2002 Feb 1;21(3):281-93. Abstract

5. Pigino G, Morfini G, Atagi Y, Deshpande A, Yu C, Jungbauer L, Ladu M, Busciglio J, Brady S. Disruption of fast axonal transport is a pathogenic mechanism for intraneuronal amyloid beta. Proc Natl Acad Sci U S A. 2009 Apr 7;106(14):5907-12. Abstract

6. Pigino G, Morfini G, Pelsman A, Mattson MP, Brady ST, Busciglio J. Alzheimer's presenilin 1 mutations impair kinesin-based axonal transport. J Neurosci. 2003 Jun 1;23(11):4499-508. Abstract

7. Morfini G, Pigino G, Szebenyi G, You Y, Pollema S, Brady ST. JNK mediates pathogenic effects of polyglutamine-expanded androgen receptor on fast axonal transport. Nat Neurosci. 2006 Jul;9(7):907-16. Abstract

8. Ligon LA, Tokito M, Finklestein JM, Grossman FE, Holzbaur EL. A direct interaction between cytoplasmic dynein and kinesin I may coordinate motor activity. J Biol Chem. 2004 Apr 30;279(18):19201-8. Abstract

9. Caviston JP, Ross JL, Antony SM, Tokito M, Holzbaur EL. Huntingtin facilitates dynein/dynactin-mediated vesicle transport. Proc Natl Acad Sci U S A. 2007 Jun 12;104(24):10045-50. Abstract

10. Zala D, Colin E, Rangone H, Liot G, Humbert S, Saudou F. Phosphorylation of mutant huntingtin at S421 restores anterograde and retrograde transport in neurons. Hum Mol Genet. 2008 Dec 15;17(24):3837-46. Abstract

11. De Vos KJ, Grierson AJ, Ackerley S, Miller CC. Role of axonal transport in neurodegenerative diseases. Annu Rev Neurosci. 2008;31:151-73. Abstract

View all comments by Zoia Muresan
View all comments by Virgil Muresan

This may be a naive question, but if amyloid deposition in the brain is a critical factor in AD-related behavioral sequelae, why is it so difficult to induce a behavioral modification of statistical relevance following Aβ vaccination, since reports show a strong amyloid plaque clearance effect?

View all comments by Elliott Mufson

I think another model for this kind of study (after parabiotics and vampires) could be pregnant mice. The placental barrier between mother and fetus highly leaky, allowing the passage of, for instance, maternal antibodies (mainly IgG). It seems to me that there is a general observation that the maternal organism appears 'rejuvenated' during pregnancy.

View all comments by Ivan Goussakov

It's of interest that mRNA levels of the calcineurin inhibitor, DSCR1, are also much higher in AD brain (1). The recent study be Lee and colleagues finds that DSCR1 interacts with Tollip and positively modulates IL-1R signalling (2). Tollip is an IRAK-1 inhibitor. This would seem to suggest problems with TLR2/TLR4 signalling in AD. This is supported by the Landreth study finding that CD14 and TLR2 and TLR4 bind Aβ to stimulate microglial activation (3). The KEGG link is below for the TOLL RECEPTOR signaling pathway (4).

References:
1. Ermak G, Morgan TE, Davies KJ. Chronic overexpression of the calcineurin inhibitory gene DSCR1 (Adapt78) is associated with Alzheimer's disease. J Biol Chem. 2001 Oct 19;276(42):38787-94. Abstract

2. Lee JY, Lee HJ, Lee EJ, Jang SH, Kim H, Yoon JH, Chung KC. Down syndrome candidate region-1 protein interacts with Tollip and positively modulates interleukin-1 receptor-mediated signaling. Biochim Biophys Acta. 2009 Dec;1790(12):1673-80. Abstract

3. Reed-Geaghan EG, Savage JC, Hise AG, Landreth GE. CD14 and toll-like receptors 2 and 4 are required for fibrillar A{beta}-stimulated microglial activation. J Neurosci. 2009 Sep 23;29(38):11982-92. Abstract

4. Toll-like receptor signaling pathway—Homo sapiens (human)

View all comments by Mary Reid