synapse complexity swartz foundation banbury meeting, cshl 15 th april 2009 seth grant
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Synapse Complexity
Swartz FoundationBanbury Meeting, CSHL
15th April 2009
Seth Grant
Genes to Cognition programwww.genes2cognition.org
Wellcome Trust Sanger Institute, Cambridge UK
MASC
PSDMASC
Post-Synaptic Proteome
MASC MAGUK Associated Signaling ComplexPSD Post Synaptic Density------------------------------------------------------------------------------------------PSP Post Synaptic Proteome (total set of postsynaptic proteins)
Presynaptic terminal
Postsynaptic terminal
NR
2
PSD-95fyn
NMDA receptor
Genetic evidence for postsynaptic complexes
Grant, et al, Science. 258, 1903-10. 1992Migaud et al, Nature, 396; 433-439. 1998Sprengel et al, Cell 92, 279-89. 1998
Learning impairmentsPlasticity impairments
MAGUK proteins
PSD95
NR
Husi et al. Nature Neuroscience, 3 (7), 661-669. 2000.Husi & Grant. J. Neurochem, 77, 281-291. 2001Collins et al, J. Neurochem. 2005Fernandez et al, submitted
Proteomic characterisation of complexes
• 2.5 Mda
• 77 proteins (2000)• 186 (2005)• 158 (2009)• 118 core (2009)
Migaud et al, Nature, 396; 433-439. 1998; Husi et al, Nature Neuroscience, 2000
synaptic strength gene expressionmRNA turnoverProtein turnover
BehavioursCognition & plasticity
NRC / MASC
2-3 MDa186 proteins
47 genes in human disease48 synaptic plasticity42 rodent behaviour
Complexity
PSDMASC
PSD 700 -1500 proteins
MASC 77 - 185 proteins
MASC occupies ~10 % of the PSP
- a core subset- same classes of proteins- ideal model of PSP- multiple isolation methods available
Grant. (2006) Biochemical Society Transactions. 34, 59-63. 2006
Post Synaptic Density 1124ER:microsomes 491Splicesome 311NRC/MASC 186Nucleolus 147Peroxisomes 181Mitochondria 179Phagosomes 140Golgi 81Choroplasts 81Lysosomes 27Exosomes 21
Is there some logic to this complexity ?
Pocklington, et al, Molecular Systems Biol 2006.
Molecular Networks: modular protein interaction networks
upstream
downstream
Upstream and Downstream modules
A useful tool for handling complexity
Human cortex PSD: 1462 protein network
Alex Bayes, Mark Collins, Louie Van De Lagemaat, Ian Whittle, Jyoti Choudhary
The origin and evolution of synapses and the brain
Emes et al, Nature Neuroscience 11, 799 (2008)
6 myr
75 myr
600 myr
1000 myr
Tree of life
6 myr
75 myr
600 myr
1000 myr
chordates
metazoans
eukaryotes
Origin of the brain
The first neurons and first brains
protosynapse
combinations
expansion
Emes et al, Nature Neuroscience 11, 799 (2008)
570 genes: 186 NRC/MASC; 570 PSD
Number of orthologues compared to mouse
Comparative genomics
Synapse orthologues in single cell eukaryotes control response to environment
Yeast behaviours:- Environmental responses
- - stress- - nutrients- - pH
143 PSD proteins
Origin of the brain
synapse first modelneuron first model
6 myr
75 myr
600 myr
1000 myr
chordates
metazoans
eukaryotes
Origin of the brain
protosynapse
Unicellulareukaryotes
Invertebrates Vertebrates
Evolutionary elaboration and expansion of the protosynapse
upstream
downstream
upstream
downstream
Y yeastI invertebrateV vertebrates
Method: Proportion of each functional class with earliest identifiable orthologue
Origins of functional classes
Evolution of ‘learning molecules’
• neurotransmitter receptors• second messengers• protein synthesis
GO terms
Y yeastW wormF flyZ zebrafishC chickenM mouseH human
DLG
NRC / MASC
NR
1NR
2
Increased combinations of complexes in vertebratesfrom expansion in paralogues
Invertebrates Vertebrates
Proteins
NR1 1 1NR2 1 4DLG 1 4
Complexes1 16
Theoretically possible MASC combinations in mammals >1030
Protosynapse‘first synapse’
Synapse evolution
Protosynapse‘first synapse’ first neurons
Synapse evolution
Protosynapse‘first synapse’ first neurons
Synapse evolution
Brain size vs Synapse complexity
6 myr
75 myr
600 myr
1000 myr
complex s
ynapse
s big brains
Big synapse proteomes evolved before big brains
How are complex synapses used in complex brains ?
Examine synapse proteome composition in different in classes of neurons in mouse
Chris Anderson, Cathy Vickers, Andrew Pocklington
anatomical expression level profiling
>150 MASC/PSD proteins in 22 regions of mouse brain
variation in levels & overlap
Measured:
mRNA
• in situ
• microarray
protein
• immunohistochem
• western
Combinations of synapse proteome define brain regions, neuron types
Expression barcode
MASC
0 4 4 3 1
1 4 3 2 3
2 3 2 2 1
anatomicalvariation
phylogeny
recent
ancient
upstream
downstream
Expression barcode
MASC
0 4 4 3 1
1 4 3 2 3
2 3 2 2 1
anatomicalvariation
phylogeny
recent
ancient
upstream
downstream
Protosynapse has most conserved & uniform expression pattern
Evolutionary expansions gave combinations used to generate anatomical diversity
Combinations
Signaling networks in the PSP
a complex combinatorial signalling machine
NMDA receptor activation with a synaptic plasticity protocol
how many substrates change?
Marcelo Coba, Andrew Pocklington, Mark Collins, Jyoti Choudhary (Science Signalling 2009)
NMDA stimulation (long term depression, LTD) > 200 phosphorylation sites.
> 120 proteins
PSD
9 of 21 kinases tested
NMDA receptor activation with a synaptic plasticity protocol
how many kinases change?
Marcelo Coba
Combinatorial outputs
10 phosphorylation sites on 4 proteins
Stimulus:
No changeIncrease phos
Decrease phos
Marcelo Coba
Combinatorial outputs
10 phosphorylation sites on 4 proteins
Stimulus:
No changeIncrease phos
Decrease phos
Combinatorial outputs
10 phosphorylation sites on 4 proteins
Stimulus:
No changeIncrease phos
Decrease phos
Combinatorial outputs
10 phosphorylation sites on 4 proteins
Stimulus:
No changeIncrease phos
Decrease phos
sites states
GluR1 3 8 (23)
10 proteins 100 1030 (2100)
PSP 104 1060
Combinatorial outputs = states
Are any two synapses the same?Can a synapse ever be in the same state twice?What are the physiological constraints?
kinases
substrates
Postsynaptic kinase-substrate network - mapping 743 phosphorylation events
Marcelo Coba, Andrew Pocklington
Building the network
defining minimal units
Number of kinases with n substrates
0123456789
0-5 5-10 10-15 15-20 20-25 25-30 30-35 35-40n substrate
num
ber o
f kin
ases
How many substrates for a kinase?
20.5 ( 8.3) protein substrates phosphorylated by each kinase
k1
PPPkinasedivergence
Number of sites with n kinases
01020304050607080
1 2 3 4 5 6 8 9 10 12n kinase
num
ber o
f site
s
How many sites were phosphorylated by more than one kinase?
65% (129) sites phosphorylated by multiple kinases
kinaseconvergence
k1 k2 kn
P
• redundance / robustness• multiple upstream receptors
Hubs – highly connected
SitesProteins
coincidence detectors
NMDA receptor
mGluR receptor
Dopaminereceptor
Combinations of sites
Functional orchestration
EPMBEHAVIOUR
OF NOE FC RR
Synapse evolution and expansion model of synapse diversity and behaviour
-20 0 20 40 6050
100
150
200
250
AcknowledgementsProteomicsMarcelo CobaAlex Bayes
Bilal Malik
Mark CollinsJyoti Choudhary
ExpressionChristopher AndersonCathy Vickers
InformaticsAndrew PocklingtonJ. Douglas Armstrong
Mike CroningRichard Emes
Support
Wellcome TrustMedical Research CouncilEuropean Union
www.genes2cognition.orgwww.g2conline.org
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