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Axonal transport, synapse developmentand mental retardation
Yong Q. Zhang
Institute of Genetics and Developmental Biology Chinese Academy of Sciences
Mental retardation(Intellectual Disability, Cognitive Disorder)
a generalized disorder, characterized by significantly impaired cognitive functioning and deficits in two or more adaptive behaviors with onset before the age of 18. 1-3% of general population is mentally retarded
from wikipedia.com
82 MR genes on the X-chrom.
Chiurazzi et al., EJHG, 2008
Syndromic MR: black Non-syndromic MR: *greyMR with neuromuscular defects: +grey
CUL4B
Chiurazzi et al., EJHG, 2008
Molecular Functions of MR Genes
1
2
3
35
✓
Transcriptional/translational Cell-cycle-related Ros/Rho/PSD95
Nature 2011
Neuronal functions of MR-related genes
1, Neurogenesis (microcephaly)
2, Neuronal migration (Lissencephaly)
3, Synapse formation and plasticity
Neuronal functions of MR-related genes
1, Neurogenesis (microcephaly)
2, Neuronal migration (Lissencephaly)
3, Synapse formation and plasticity
Synapse dysgenesis in MR patients
Purpura, Science, 1974Dendritic spines
Mutations in ACSL4 are associated with non-syndromic X-linked mental retardation
Meloni et al., Nat Genet, 2002Longo et al., J Med Genet, 2003
aa
Splicing mutant resulting in truncation
ACSL: Acyl-CoA Synthetase Long-chain (C12-20)
ACSL in Lipid Synthesis and ATP Production
Fatty acid
Acyl-CoA
Coenzyme A
Glycerol-3-P
Diacylglycerol (DAG)
Triacylglycerol (TAG)
PC, PE, PS
Lipid droplet
MembraneComponents
Fatty acid
Acyl-CoA
Coenzyme A
β-oxidation
ATP
ACSL
ACSL
Endoplasmic Reticulum Mitochondria
Lipid biosynthesis Fatty acid degradation
Phosphatidic acid (PA)
Coleman et al., Annu Rev Nutri, 2000
The role of ACSL4 in neurodevelopment?
The mechanism of how the disease develops?
ACSL Family
ACSL1, 5, 6 ACSL3, 4
{subfamilies
no mouse models
50% identity and 67% similarity
Zhaohui Wang and colleagues, Hum Mol Genet, 2009
ACSL Family: 1, 3, 4, 5, and 6
ACSL1, 5, 6 ACSL3, 4 (fly dAcsl)
Larval Neuromuscular Junctions
NMJ synapses
Muscles
Large, simple and accessible
Griffth and Budnik, 2006
Nerves and NMJ synapses
Axonal accumulation of synaptic vesicle protein CSP in dAcsl mutants
Wild type
dAcsl
Ubi. rescue
Khc, DhcWT
Hurd and Saxton, Genetics, 1996Martin et al, Mol Cell Biol, 1999
A Control
B1 dAcslKO/05847
B2
OK6>Syt-eGFP ChAT
B3
B2’
B3'
B1’
A’
OK6>Syt-eGFP
ChAT
merge
20 μm
Distally-Biased Axonal Aggregates of SVs
ChAT: choline acetyltransferase
Motor Neu
Sens. Neu
Anterior Posterior
Sensory NeuronMotor Neuron
SV protein
Dynein
Active zone
t-SNARE
Mitochondria
Cell adhesion
Accumulation of selective axonal cargos
Retrograde cargos accumulated in dAcsl mutants
MVB:Multiple vesicle body
PLB:Prelysosomal body
Lb:Lamellated body
.
..
..
..
..
.
..
Late endosome/lysosome
Autophago-some
MVBs
Retrograde cargos accumulated in mutants
Lysosome+autophagosome Multiple vesicle body
dAcsl mutations lead to specific accumulation of retrograde cargoes
Synaptic vesiclesAutophagosomeLysosome
Immunostaining and EM analysis
Why did the axonal aggregates of retrograde cargos form in dAcsl mutants?
Live imaging of axonal transport of GFP-tagged synaptic vesicles
Flux
Velocity
Processivity
Retrograde transport of synaptic vesicles was impaired in dAcsl mutants
Axonal transport of mitochondria was normal in dAcsl mutants
FR
WT dfmr1 NOE WT dfmr1 NOE
Tim
e p
erce
nta
ge
*
**
*
*
AT RT
Stop
RR
E
10 m
CA B
time
Anterograde Retrograde
50 s
C,A, B,
AT RT
***
***
Flu
x (m
ito.
nu
mb
er/m
in)
*
*
WT
dfmr1
NOE
D
WT dfmr1 dfmr1 overexpres.
Yao et al., Hum Mol Genet, 2011
dfmr1 regulates axonal transport of mitochondria
Role of axonal transport in neurodegenerative diseasesDe Vos et al., Annu Rev Neurosci, 2008
Axonal transport and neurodegenerative diseaseChevalier-Larsen and HolzbaurBiochim Biophys Acta, 2006
SV transport defects in dAcsl mutants
Mitochondria transport defects in dfmr1 mutants
Accumulation of retrograde cargoesand transport defects
?
Defects in synaptic development and function
Larval Neuromuscular Junctions
NMJ synapses
MusclesLarge, simple and accessible
Griffth and Budnik, 2006
Nerves and NMJ synapses
dAcsl mutants show atrophic synaptic terminals
Neuronal but not muscular rescue by human ACSL4
dAcsl is required for NMJ growth and stability
Pre- and post-synaptic components concomitantly reduced in dystrophic NMJs
No increased apoptosis
Futsch/MAP1B active zone protein Postsynaptic scaffold
CNS
7 6
NMJ
Nerve stimulation
Vm
I Postsynaptic current (EJC)
Nerve
Will atrophic NMJs show defective transmission?
mEJP: miniature EJPs
Spontaneous release of single SVs
EJP:Excitatory Junction Potentials
dAcsl mutations impaired neurotransmissionEJP: Excitatory Junction Potentials
mEJP: miniature excitatory junction potentials
dAcsl mutations impaired neurotransmission
Neuronal but not muscular rescue by human ACSL4
Axonal jam and retrograde transport defects
NMJ synapse atrophy
Reduced neurotransmission
Can these phenotypes be corrected by induced expression?
Drug-induced tissue specific expression
Osterwalder et al., PNAS, 2001
Gal4
Aggregates were rescued by induced expression
Together with NMJ rescue
dAcsl mutants show distally-biased axonal aggregates and impaired retrograde transport of synaptic vesicles
dAcsl is required for synaptic growth, stability
and neurotransmission
Summary
Liu et al., J Neurosci., 2011
SVSVSVSVSVSV
AcknowledgmentsZhihua Liu, PhD
Yan Huang
Zhaohui Wang (PI)
Yi Zhang (mutants & Ab)
Di Chen (mutants)
Zhihua Liu
Yan Huang
Grants:
National Science Foundation of China
Chinese Academy of Sciences
The Ministry of Science and Technology
Acyl-CoA is central to diverse processes
K294E
Q419R
W685@
R570S
P375L
V594D
dACSL-8 dACSL-1 mutations in patients
Conservation is not restricted