Autophagy 6:5, 648-649; July 1, 2010; 2010 Landes Bioscience

AUTOPHAGIC PUNCTUM
648 Autophagy Volume 6 Issue 5
Key words: lysosomal storage diseases,
neurodegeneration, glycosphingolipids,
p62, LAMP-2, Gaucher disease
Submitted: 03/26/10
Revised: 04/12/10
Accepted: 04/14/10
Previously published online:
www.landesbioscience.com/journals/
autophagy/article/12047
*Correspondence to: Gregory A. Grabowski;
Email: greg.grabowski@cchmc.org
Gaucher disease is an inherited autosomal
recessive disease caused by mutations of
acid -glucosidase, a lysosomal hydro-
lase specic for degradation of glucosyl-
ceramide and glucosylsphingosine in the
glycosphingolipid metabolic pathway.
Clinically, Gaucher disease is classied
into three types: type 1 is a visceral dis-
ease, whereas types 2 and 3 are acute and
chronic neuronopathic variants, respec-
tively. In types 2 and 3, the CNS pathology
displays neuronal inclusions and neuron
death. The underlying mechanism(s) by
which the glycosphingolipid storage leads
to this pathology is not fully understood. A
mouse model whose phenotype mimicked
that of the human neuronopathic variants
was generated in our lab. In the brain of
this model, abnormal autophagosomes and
lysosomes implicate autophagy in the neu-
ronal degeneration of Gaucher disease.
The strategy to generate a neuronop-
athic-type Gaucher disease mouse model
is based on the nding that in vitro acid
-glucosidase activity is optimized by,
but does not require, a small lysosomal
protein, saposin C. In addition to its acti-
vation function, saposin C has an anti-
proteolytic protective effect toward acid
-glucosidase. We hypothesized that the
in vivo loss of saposin C in the presence of
a point mutated acid -glucosidase would
potentiate substrate accumulation and the
abnormal phenotype. Crossbreeding of
mice homozygous for the V394L muta-
tion with the saposin C null mice leads to
additional decreases in acid -glucosidase
protein and activity, and the resultant
insufcient degradation of its substrates,
glucosylsphingosine and glucosylceramide;
Impaired autophagosomes and lysosomes
in neuronopathic Gaucher disease
Ying Sun and Gregory A. Grabowski*
The Division of Human Genetics; Cincinnati Childrens Hospital Medical Center and the Department of Pediatrics; University of Cincinnati College
of Medicine; Cincinnati, OH USA
these are signicantly increased in the
brain. Development was nearly normal for
four weeks, but their body weight declined
after 30 days. This is followed by develop-
ment of hind limb weakness and duck-like
walking. Attenuated hippocampal long-
term potentiation correlates with substrate
accumulation. The severe neurological
degeneration leads to death by 48 days.
Electron and light microscopy of brain
sections reveals axonal inclusions contain-
ing undigested material in the spinal cord,
midbrain, brain stem and cerebellum,
whereas the soma of neurons appears nor-
mal. This axonal defect is accompanied
by activation of microglial cells and astro-
cytes, indicating a local proinammatory
reaction. Axonal degeneration can lead to
cell death by blocking retrograde transport
from axons to the soma where the recycled
endosomal membrane can be degraded in
the lysosomes. Apoptotic-mediated cell
death was excluded by negative TUNEL
assays. Immunouorescence staining of
LAMP-2, an endosomal/lysosomal mem-
brane protein, and p62 that participates
in autophagic clearance of ubiquitinated
proteins, shows enhanced signals in the
thalamus, brain stem and basal ganglia.
When autophagy is inhibited, p62 can
aggregate and form inclusion bodies.
The increased p62 in this model (Fig.
1) suggests that in addition to primary
glycosphingolipids accumulation, pro-
tein degradation through the autophagy
pathway is impaired. Autophagy starts
engulfment of cytoplasmic materials by
a phagophore, followed by sequestration
through formation of a double-membrane
vesicle (autophagosome), maturation by
Punctum to: Sun Y, Liou B, Ran H, Skelton MR,
Williams MT, Vorhees CV, Kitatani K, Hannun YA,
Witte DP, Xu YH, Grabowski GA. Neuronopathic
Gaucher disease in the mouse: viable combined
selective saposin C defciency and mutant
glucocerebrosidase (V394L) mice with glucosyl-
sphingosine and glucosylceramide accumulation
and progressive neurological defcits. Hum Mol
Genet 2010; 19:108897; PMID: 20047948; DOI:
10.1093/hmg/ddp580.
www.landesbioscience.com Autophagy 649
AUTOPHAGIC PUNCTUM AUTOPHAGIC PUNCTUM
fusion of the autophagosome with late
endosomes and then with the lysosomes to
form the autolysosome, and ultimate deg-
radation of the autophagosome contents.
Glycosphingolipids in the salvage pathway
are endocytosed into a late endosome, and
then degraded in the lysosome. The accu-
mulation of p62 and LAMP-2 observed
in this model suggests that glucosylcer-
amide and glucosylsphingosine accumula-
tion in lysosomes may disrupt the normal
autophagic fusion process. Cell specicity
of this process is evidenced by the p62 sig-
nal being presented in neurons and astro-
cytes, but not in microglial cells.
Autophagy disturbances have been
observed in several neurodegenerative
lysosomal storage diseases. Blockage of
autophagy is present in multiple sulfatase
deciency and neuronal ceroid lipofusci-
nosis, whereas an induction of autophagy
is observed in Niemann-Pick type C dis-
ease degenerating cerebellar Purkinje cells.
This impairment of autophagy in the soma
differs from that in the neuronopathic
Gaucher disease mouse in which aberrant
autophagy appears to be primarily affected
in axons. Interestingly, autophagy has a
quality control role in protein degradation
for maintaining axonal homeostasis and
its disruption leads to axonal degeneration.
We speculate that the disrupted autophagy
caused by glucosylceramide and glucosyl-
sphingosine storage is causal to this degen-
erative axonal process and is central to the
pathogenesis of neuronopathic Gaucher
disease. Further study will be directed to
determining if autophagy is enhanced or
attenuated. Enhanced autophagy is gen-
erally believed to help remove the mem-
brane structures and serve as protection.
Deciency of autophagy can lead to axonal
dystrophy and degeneration.
Pathological dysregulation of autophagy
is implicated in synaptic dysfunction,
cellular stress, and neuronal cell death
that may be mediated by a rapamycin-
dependent protein signaling process that
regulates synaptic protein synthesis and
maintenance. Impairment of autophagy
at synapses due to substrate accumulation
and inclusions in neurite processes could
affect the long-term potentiation in the
neuronopathic Gaucher disease model.
Figure 1. The neuronopathic Gaucher disease mouse model exhibits a severe neurodegenerative
phenotype and axonal degeneration in the CNS. (A) Electron microscopy of axon in the midbrain
region shows the undigested inclusion materials in the axon. (B) Accumulation of p62 (red) in the
neurons (green, nuclear NeuN staining) of thalamus.