Kinesin motor proteins regulate mitosis and anterograde cargo
transport as exemplified by fast axonal transport (FAT) in neurons.
Neurons depend on kinesins for cell cycle regulation, especially
the assembly and function of the mitotic spindle, a macromolecular
structure composed primarily of microtubules (MTs) that undergo
cycles of polymerization and depolymerization to properly segregate
duplicate chromosomes into separate daughter cells. In addition,
the kinesin motors use MTs to transport cargo such as proteins,
lipids, and mitochondria to the axon and axon terminal, an
essential process for normal neuron development, growth, and
communication as the cell body is the site of synthesis1.
In neurodegenerative diseases such as Alzheimer’s disease (AD),
cell cycle defects (e.g., chromosome mis-segregation, abnormal
mitotic spindle structure/function, aneuploidy) and impaired FAT
are pathophysiological hallmarks of AD, the most common form of
dementia2,3. Indeed, FAT deficits and axonal swellings occur before
the classic neuropathological signs of dementia (plaques and
tangles), leading some researchers to posit that transport deficits
are an early sign of neuron vulnerability to neurodegeneration4-8.
This newsletter focuses on the role Eg5 (KSP, kinesin spindle
protein, KIF11, kinesin-5) and kinesin-1 have in AD-associated cell
cycle and FAT defects, respectively. Eg5 is a kinesin motor that
interacts with MTs and is essential for mitotic spindle formation
and function. Kinesin-1 is the prototypical kinesin that mediates
FAT. For a discussion of the role of the dynein motor in
neurodegeneration, see the 2014 January/February newsletter.
Motors and Neurodegeneration: Cell Cycle Defects
The neuropathological hallmarks of AD are beta amyloid (Ab)
plaques and hyperphosphorylated tau neurofibrillary tangles. Ab is
produced by enzymatic cleavage of amyloid precursor protein (APP)
by beta-secretase 1 (BACE1) and the presenilin (PS)-containing
gamma-secretase complex2,3. In AD-associated cell cycle defects,
both Ab and tau inhibit Eg5 activity and its interaction with MTs,
which negatively affects mitotic spindle structure and function9-11
(Fig. 1). A dysfunctional spindle results in the mis-segregation of
chromosomes, aneuploidy/hyperploidy, and cell death9-11. Tau's
inhibition of Eg5 requires excess levels of MT-bound tau10. Neurons
displaying aneuploidy/hyperploidy are increased in preclinical AD
and selectively die as AD progresses. Indeed, 90% of cell death in
autopsied AD brains is comprised of hyperploid neurons12.
Eg5’s role in AD is not limited to cell cycle defects as mature,
post-mitotic neurons express this protein. Eg5 mediates Ab-induced
inhibition of long-term potentiation and loss of hippocampal
spines/synapses13,14, at least partly through a reduction in cell
surface trafficking of NMDA and nerve growth factor/p75 neutrophin
receptors13.
Kinesin Motor Proteins and Neurodegeneration v
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JUNE2016
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Kinesin Motor Proteins and Neurodegeneration Related
Publications
Research Tools
Fig. 1. Kinesin motor Eg5 binding to MTs, essential for proper
mitotic spindle structure and function, is inhibited by Ab and
tau.
Fig. 2. Kinesin-1-mediated anterograde FAT is inhibited by
GSK-3b-mediated phosphorylation of kinesin-1 light chains which
causes dissociation of kinesin-1 and its cargo. GSK-3b can be
activated by PS1 and/or PP1 with the latter activated by
hyper-phosphorylated tau filaments.
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Anterograde Tau
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Motors and Neurodegeneration: Axonal Transport Defects
All 4 neurodegeneration-related proteins (APP, Ab, PS, and tau)
inhibit kinesin-1-mediated FAT. For APP and Ab disruption of
FAT15-17, various mechanisms have been reported, ranging from actin
aggregation and dynamic changes in actin polymerization16 to
activation of casein kinase 2, which initiates a sequential cascade
of kinesin-1 light chain phosphorylation and release of cargo from
kinesin-117. Interestingly, APP undergoes anterograde FAT18 via
binding with kinesin-1 light chains as part of an axon membrane
compartment which also contains BACE1 and PS119-21. In this way,
APP could be cleaved into Ab during and/or after kinesin-1-mediated
FAT21. However, this finding has been refuted by others22.
Both tau and mutant PS1 (M146V) regulate kinesin-1-mediated FAT
via dephosphorylation-stimulated activation of GSK-3b, resulting in
GSK-3b-mediated phosphorylation of kinesin-1 light chains, which in
turn induce separation of kinesin-1 and its cargo5 (Fig. 2). As a
tau kinase, GSK-3b can mediate formation of pathological,
hyper-phosphorylated tau, which dissociates from MTs and causes MT
depolymerization, followed by formation of tau filaments and
eventually neurofibrillary tangles2,3. The amino terminus of either
tau filaments or C-terminal-truncated, unbound monomers (C-terminus
contains MT binding domains) inhibits kinesin-dependent FAT. Here,
GSK-3b activity is increased via protein phosphatase 1
(PP1)-mediated dephosphorylation-induced activation of GSK-3b23
(Fig. 2). These studies have led researchers to suggest that FAT is
vulnerable to disruption early in the pathophysiology of AD2-5;
indeed, axonal/transport defects occur long before amyloid
deposition4. Furthermore, depletion of kinesin-1 produces similar
transport deficits while also increasing levels of Ab peptide
levels and deposits4.
Conclusion
Normal neuron physiology relies on kinesin motors for a variety
of processes, including proper mitosis and FAT. Impaired kinesin
function has a profound and wide-spread effect on neuron health as
kinesin motor dysfunctions are not only observed in AD, but
Huntington's disease24, upper and lower motor neuron diseases, and
Charcot-Marie-Tooth peripheral neuropathy2,3. To assist in studying
how kinesin and dynein motor proteins regulate neuron health and
survival, Cytoskeleton, Inc. offers purified kinesin motor proteins
and dynein protein, along with kits to measure MT-induced motor
ATPase activity.
ReferencesContinued from Page 1
www.cytoskeleton.com
MOTOR PROTEIN PRODUCTS
1. Bass P.W. 1998. The role of motor proteins in establishing
the microtubule arrays of axons and dendrites. J. Chem. Neuroanat.
14, 175-180.
2. Millecamps S. and Julien J.-P. 2013. Axonal transport
deficits and neurodegenerative diseases. Nat. Rev. Neurosci. 14,
161-176.
3. Morfini G.A. et al. 2009. Minisymposium: Axonal transport
defects in neurodegenerative diseases. J. Neurosci. 29,
12776-12786.
4. Stokin G.B. et al. 2005. Axonopathy and transport deficits
early in the pathogenesis of Alzheimer’s disease. Science. 307,
1282-1288.
5. Pigino G. et al. 2003. Alzheimer’s presenilin 1 mutations
impair kinesin-based axonal transport. J. Neurosci. 23,
4499-4508.
6. Lazarov O. et al. 2007. Impairments in fast axonal transport
and motor neuron deficits in transgenic mice expressing familial
Alzheimer’s disease-linked mutant presenilin 1. J. Neurosci. 27,
7011-7020.
7. Ishihara T. et al. 1999. Age-dependent emergence and
progression of a tauopathy in transgenic mice overexpressing the
shortest human tau isoform. Neuron. 24, 751-762.
8. Zhang B. et al. 2004. Retarded axonal transport of R406W
mutant tau in transgenic mice with a neurodegenerative tauopathy.
J. Neurosci. 24, 4657-4667.
9. Borysov S.I. et al. 2011. Alzheimer Ab disrupts the mitotic
spindle and directly inhibits mitotic microtubule motors. Cell
Cycle. 10, 1397-1410.
10. Bouge A.-L. and Parmentier M.-L. 2016. Tau excess impairs
mitosis and kinesin-5 function, leading to aneuploidy and cell
death. Dis. Model. Mech. 9, 307-319.
11. Rossi G. et al. 2008. A new function of
microtubule-associated protein tau: involvement in chromosome
stability. Cell Cycle. 7, 1788-1794.
12. Arendt T. et al. 2010. Selective cell death of hyperploid
neurons in Alzheimer’s disease. Am J. Pathol. 177, 15-20.
13. Ari C. et al. 2014. Alzheimer Ab inhibition of eg5/kin5
reduces neurotrophin/transmitter receptor function. Neurobiol.
Aging. 35, 1839-1849.
14. Freund R.K. et al. 2016. Inhibition of the motor protein
Eg5/Kinesin-5 in amyloid b-mediated impairment of hippocampal
long-term potentiation and dendritic spine loss. Mol. Pharmacol.
89, 552-559.
15. Gunawardena S. and Goldstein L.S.B. 2001. Disruption of
axonal transport and neuronal viability by amyloid precursor
protein mutations in Drosophila. Neuron. 32, 389-401.
16. Hiruma H. et al. Glutamate and amyloid b-protein rapidly
inhibit fast axonal transport in cultured rat hippocampal neurons
by different mechanisms. J. Neurosci. 23, 8967-8977.
17. Pigino G. et al. 2009. Disruption of fast axonal transport
is a pathogenic mechanism for intraneuronal amyloid beta. Proc.
Natl. Acad. Sci. USA. 106, 5907-5912.
18. Koo E.H. et al. 1990. Precursor of amyloid protein in
Alzheimer disease undergoes fast anterograde axonal transport.
Proc. Natl. Acad. Sci. USA. 87, 1561-1565.
19. Ferreira A. et al. 1993. Intraneuronal compartments if the
amyloid precursor protein. J. Neurosci. 13, 3112-3123.
20. Kamal A. et al. 2000. Axonal transport of amyloid precursor
protein is mediated by direct binding to the kinesin light chain
subunit of kinesin-I. Neuron. 28, 449-459.
21. Kamal A. et al. 2001. Kinesin-mediated axonal transport of a
membrane compartment containing b-secretase and presenilin-1
requires APP. Nature. 414, 643-648.
22. Lazarov O. et al. 2005. Axonal transport, amyloid precursor
protein, kinesin-1, and the processing apparatus: Revisited. J.
Neurosci. 25, 2386-2395.
23. LaPointe N.E. et al. 2009. The amino terminus of tau
inhibits kinesin-dependent axonal transport: Implications for
filament toxicity. J. Neurosci. Res. 87, 440-451.
24. Morfini G.A. et al. 2009. Pathogenic Huntingtin inhibits
fast axonal transport by activating JNK3 and phosphorylating
kinesin. Nat. Neurosci. 12, 864-871.
Kits and Assays
Kinesin & Dynein Proteins
Microtubules
Product Source Purity Cat. # Amount
CENP-E Motor Domain Protein H. sapiens >85% CP01-ACP01-XL2 x
25 µg1 x 1 mg
Chromokinesin Motor Domain Protein H. sapiens >85% CR01-A 2 x
25 µg
Dynein (cytoplasmic) Porcine brain >80% CS-DN01 1 x 50 µg
Eg5 Motor Domain Protein H. sapiens >85% EG01-AEG01-B2 x 25
µg10 x 25 µg
KIFC3 Motor Domain Protein H. sapiens >85% KC01-A 2 x 25
µg
KIF3C Motor Domain Protein H. sapiens >85% KF01-A 2 x 25
µg
KIF7 motor domain H. sapiens >85% CS-KF51 1 x 100 µg
Kinesin Heavy Chain Motor Domain Protein H. sapiens >85%
KR01-AKR01-XL
2 x 25 µg1 x 1 mg
MCAK Motor Domain Protein H. sapiens >85% MK01-A 2 x 25
µg
MKLP1 Motor Domain Protein H. sapiens >85% MP01-AMP01-XL2 x
25 µg1 x 1 mg
MKLP2 Motor Domain Protein H. sapiens >85% CS-MP05 1 x 50
µg
Product Cat. # AmountATPase ELIPA™ (enzyme-linked,
colorimetric)Kinetic quantitation of ATP hydrolysis (Kcat 0.05 to
>1.0) BK051 96 assays
CytoPhos™ Phosphate Assay (endpoint assay)Colorimetric assay for
ATPases & GTPases (Kcat 0.01 to >1.0) BK054 1000 assays
GTPase ELIPA™ (enzyme-linked, colorimetric)Kinetic quantitation
of GTP hydrolysis (Kcat 0.05 to >1.0) BK052 96 assays
Kinesin ELIPA™ Biochem KitFor real time kinetic and Vmax kinesin
ATPase measurements BK060 96 assays
Kinesin ATPase Endpoint AssayFor endpoint measurement of kinesin
ATPase activity BK053 1000 assays
Product Cat. # AmountMicrotubules, Pre-formed, lyophilized,
porcine source, substrate for kinesin ATPase assays
MT002-AMT002-XL
4 x 500 µg1 x 10 mg