The mutant protein that causes Huntington disease blocks axonal traffic by putting the molecular equivalent of the wheel-locking “boot” on motor proteins, according to a paper published online June 14 in Nature Neuroscience. Principal investigators Gerardo Morfini and Scott Brady, both of the University of Illinois in Chicago, and colleagues teased out the specifics of huntingtin’s attack on kinesin-1, showing that mutant huntingtin activates cJun N-terminal kinase 3 (JNK3) to phosphorylate the motor protein and prevent it from binding to the microtubule tracks along the axon. The work suggests that JNK3, or the upstream kinases that likely activate it, could be drug targets for Huntington disease.

“This mutant protein looks like it tweaks one particular signaling pathway, and then all hell breaks loose,” said Peter Hollenbeck of Purdue University in West Lafayette, Indiana, who was not involved in the current research. The pathway is currently incomplete with the cascade between huntingtin and JNK3 remaining a bit of a black box.

Neurons depend on reliable fast axonal transport to supply the synapses at the end of long axons, making transport a vulnerable point for nerve cells. Scientists already knew that the pathogenic proteins in several neurodegenerative diseases activate kinases to gum up axonal transport. Huntingtin has been repeatedly implicated in vesicular trafficking (for review, see Caviston and Holzbaur, 2009), and Brady’s group and others have shown that in Alzheimer disease, Aβ inhibits transport via casein kinase 2 (see ARF related news story; Pigino et al., 2009; Moreno et al., 2009).

The huntingtin gene causes disease when it contains too many CAG trinucleotide repeats, causing an extra long succession of glutamine (polyQ expansion) in the protein. Morfini and Brady suspected JNK might interact with huntingtin because they had already shown that another polyglutamine-expanded protein, the androgen receptor, causes spinal and bulbar muscular atrophy via JNK’s action on fast axonal transport (see ARF related news story; Morfini et al., 2006). JNK phosphorylates the heavy chain of kinesin, inhibiting microtubule binding.

The researchers found that polyQ-expanded huntingtin (polyQ-Htt) inhibited both retrograde and anterograde transport in isolated squid axons, whereas the wild-type protein did not. When JNK inhibitors were included, the effect of polyQ-expanded huntingtin was blocked, suggesting JNK carries out huntingtin’s dirty work. In mouse neuroblastoma cells, polyQ-Htt transfection increased the levels of phosphorylated, active JNK.

There are three isoforms of JNK, and using inhibitors that affect them differentially, the scientists determined that JNK3 was the most important for huntingtin’s effect on transport. Many kinases come in more than one isoform, and researchers should consider that fact, Brady said: “They’re not interchangeable.” The authors used mass spectrometry to map the residue that JNK3 phosphorylates, serine-176 in the kinesin heavy chain.

As powerful as the squid assay is, the important question is what happens in mammalian neurons, said Erika Holzbaur of the University of Pennsylvania in Philadelphia, who was not part of the current research. Morfini and colleagues transfected cultured hippocampal cells with GFP-tagged kinesin heavy chain constructs that included wild-type serine-176, a glutamate at position 176 (to mimic constant phosphorylation), or an alanine at 176 (an construct that cannot be phosphorylated). To evaluate transport, they observed how much GFP-tagged kinesin reached the axon tips. The S176E mutant allowed approximately 55 percent of kinesin to travel to the tips, while the other constructs permitted approximately 75 percent or more to reach the final destination. However, the error bars were large. “It’s not day and night,” Holzbaur said, and she suggested that JNK3 serine-176 may contribute to kinesin transport, but is an insufficient explanation on its own.

The paper offers a tantalizing explanation as to why Huntington’s affects only the nervous system. Although huntingtin expression is ubiquitous, JNK3 expression is limited to the brain and testes. Brady suspects that huntingtin, and JNK3, are involved in normal regulation of kinesin-based transport, but that the poly-Q expanded huntingtin throws the system off balance. “It’s when you get activation at too high a level, or in the wrong subcellular compartment, that you get into trouble,” Brady said. The researchers are currently working on defining the pathway between huntingtin and JNK3 activation; any member of that cascade could be a potential new target for HD therapies.—Amber Dance


  1. 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.


    . Pathogenic huntingtin inhibits fast axonal transport by activating JNK3 and phosphorylating kinesin. Nat Neurosci. 2009 Jul;12(7):864-71. PubMed.

    . Release of kinesin from vesicles by hsc70 and regulation of fast axonal transport. Mol Biol Cell. 2000 Jun;11(6):2161-73. PubMed.

    . Kinesin-mediated axonal transport of a membrane compartment containing beta-secretase and presenilin-1 requires APP. Nature. 2001 Dec 6;414(6864):643-8. PubMed.

    . Glycogen synthase kinase 3 phosphorylates kinesin light chains and negatively regulates kinesin-based motility. EMBO J. 2002 Feb 1;21(3):281-93. PubMed.

    . 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. PubMed.

    . Alzheimer's presenilin 1 mutations impair kinesin-based axonal transport. J Neurosci. 2003 Jun 1;23(11):4499-508. PubMed.

    . JNK mediates pathogenic effects of polyglutamine-expanded androgen receptor on fast axonal transport. Nat Neurosci. 2006 Jul;9(7):907-16. PubMed.

    . A direct interaction between cytoplasmic dynein and kinesin I may coordinate motor activity. J Biol Chem. 2004 Apr 30;279(18):19201-8. PubMed.

    . Huntingtin facilitates dynein/dynactin-mediated vesicle transport. Proc Natl Acad Sci U S A. 2007 Jun 12;104(24):10045-50. PubMed.

    . Phosphorylation of mutant huntingtin at S421 restores anterograde and retrograde transport in neurons. Hum Mol Genet. 2008 Dec 15;17(24):3837-46. PubMed.

    . Role of axonal transport in neurodegenerative diseases. Annu Rev Neurosci. 2008;31:151-73. PubMed.

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News Citations

  1. The Many Misdeeds of Aβ—Seizures and Axonal Transport Interference
  2. JNK Clogs Axonal Transport

Paper Citations

  1. . 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. PubMed.
  2. . Synaptic transmission block by presynaptic injection of oligomeric amyloid beta. Proc Natl Acad Sci U S A. 2009 Apr 7;106(14):5901-6. PubMed.
  3. . JNK mediates pathogenic effects of polyglutamine-expanded androgen receptor on fast axonal transport. Nat Neurosci. 2006 Jul;9(7):907-16. PubMed.

Further Reading


  1. . Phosphorylated Tau interacts with c-Jun N-terminal kinase-interacting protein 1 (JIP1) in Alzheimer disease. J Biol Chem. 2009 Jul 31;284(31):20909-16. PubMed.
  2. . Examination of potential mechanisms of amyloid-induced defects in neuronal transport. Neurobiol Dis. 2009 Oct;36(1):11-25. PubMed.
  3. . Rhes, a striatal specific protein, mediates mutant-huntingtin cytotoxicity. Science. 2009 Jun 5;324(5932):1327-30. PubMed.
  4. . Huntingtin facilitates dynein/dynactin-mediated vesicle transport. Proc Natl Acad Sci U S A. 2007 Jun 12;104(24):10045-50. PubMed.

Primary Papers

  1. . Pathogenic huntingtin inhibits fast axonal transport by activating JNK3 and phosphorylating kinesin. Nat Neurosci. 2009 Jul;12(7):864-71. PubMed.