INTRODUCTION
Neurodegenerative disorders include a number of diseases caused by
progressive loss of neurons. A hallmark of most neurodegenerative
diseases is aberrant accumulation and aggregation of misfolded proteins
inside neurons, including mutant huntingtin (mHTT) aggregates in
Huntington’s disease (HD) (1 ), mutant ataxin-3 in spinocerebellar
ataxia type 3/Machado-Joseph disease (SCA3/MJD) (2 ), Amyloid-beta
(Aβ or Abeta) in Alzheimer’s disease (which also has extracellular
aggregates) (3 ), and α-synuclein in Parkinson’s disease
(4 ). HD is an appealing model for pathological study as well as
for drug discovery for degenerative diseases because of its simplicity
in genetics and its relatively high prevalence (5 ). It is an
autosomal dominant neurodegenerative disorder caused by toxic aggregates
of mutant HTT proteins (mHTT) in the brain (6 ). HTT exon1
contains a polyglutamine (polyQ) tract that consists of 6–35 glutamine
residues in normal neurons. Mutant HTT proteins with expansions greater
than 35 glutamine residues induce protein misfolding and are more prone
to aggregation, leading to onset of HD. The ability of neuron to target
these aggregates for degradation is insufficient, causing proteins
accumulation, aggregation and protein dyshomeostasis, a common thread in
neurodegenerative diseases (7 ). mHTT abundance and proteostasis
are indicators of neurodegeneration and disease severity (8 ).
Thus, induction of degradation of disease-causing proteins should
prevent aggregate accumulation and neuronal cell death, representing a
general therapeutic strategy for HD and other neurodegenerative
disorders.
Valosin-containing protein (VCP/p97) is a central and important
regulator of proteostasis, which has been implicated in degenerative
diseases (9). Chaperone VCP protein (p97 in mouse, TER94 in Drosophila
melanogaster, and CDC48 in S. cerevisiae) belongs to the AAA protein
family ATPase (10 ). VCP is composed of N terminal domain,
followed by tandem D1 and D2 ATPase domain joined by a linker region.
VCP forms a hexametric ring in cylindrical structure with the N-terminal
domain devoted to allow VCP interaction with adaptor or substrate
proteins. VCP regulates degradation of damaged proteins and organelles
in proteasome and autophagic pathway (11,12,13 ) and has been
linked both directly and indirectly to neurodegenerative disorders. In
HD disease as well as other polyQ-related diseases, endogenous VCP
co-localizes with the polyQ containing aggregates in cultured
polyQ-expressing neuronal cells (14 ). Colocalization of VCP with
disease-causing aggregates has been reported in not only Huntington
disease but also Machado–Joseph disease (MJD) and Parkinson disease
(15,16 ). Furthermore, Missense mutations in the VCP gene has been
implicated in pathogenesis of human degenerative disorders including
IBMPFD (inclusion body myopathy associated with Paget disease of bone
and frontotemporal dementia), and rare cases of familial amyotrophic
lateral sclerosis (ALS) (9 ). These results have emphasized the
importance of VCP in the pathology of human neurodegenerative disorders
and raised the possibility of targeting VCP to treat HD as well as other
neurodegenerative disorders.
One of the crucial mechanisms for clearance of the disease-causing mHTT
protein in HD is macroautophagy (hereafter autophagy). Autophagy is a
highly conserved cellular process which engulfs cytosolic components for
degradation, including damaged organelles, misfolded proteins and
aggregates, to maintain cytosolic homeostasis (17 ). It mediates
the degradation of large disease proteins or aggregates that cause
neurodegenerative diseases, which are mostly inaccessible to proteasome
(18 ). In HD, the abnormal accumulation of mHTT proteins arises
from insufficiency of the autophagy machinery (19). Strategies that
induce moderate autophagy upregulation are believed to be of therapeutic
benefit in treatment of HD as well as other neurodegenerative diseases
(20,21 ). Microtubule-associated protein 1A/1B-light chain 3
(LC3), the core autophagy component in mammalian cells, plays a pivotal
role in the autophagy-lysosome pathway, regulating both autophagosome
formation and cargo recognition (22 ). LC3 has been demonstrated
to recognize protein aggregates for autophagy removal (23 ). It
has been reported that LC3 colocalizes with mHTT aggregates and
facilitated mHTT protein degradation by autophagy (24, 25 ). VCP
could interacts with LC3 (26) and has also been reported to co-localize
with mHTT (14 ), indicating potential involvement of VCP in
autophagy-mediated mHTT degradation. However, the role of VCP in
autophagy-mediated mHTT clearance remains undefined
To explore the potential treatment of HD by small molecules targeting
VCP, we established a high-throughput screening assay using VCP’s ATPase
activity as readout and identified gossypol acetate as a novel modulator
of VCP. Gossypol acetate acted in a gain-of-function way to induce
mHTT-VCP-LC3 complex formation, enabling autophagy-mediated degradation
of endogenous mHTT. Gossypol acetate reduced mHTT levels, alleviated
neuronal toxicity in HD cells and improved behaviors in Drosophila and
mouse HD models, making gossypol a promising lead for the development of
new HD therapy.