complex lipids and neudegenerative diseases - mondino.it · 16/03/2018 6 hydrogen bonds at the...
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Sandro SonninoDepartment of Medical Biotechnology and Translational Medicine
Study Center for the Functional Biochemistry of Brain LipidsUniversity of Milan - Italy
Complex lipids and neudegenerative diseases
Complex lipids are amphiphilic components of cell membranes
In neurons:Glycerophospholipids: 60% of total membrane lipidsCholesterol: 30% of total membrane lipidsGlycosphingolipids: 10% of total membrane lipids
neutral glycosphingolipids: 10%acidic glycosphingolipids (gangliosides): 90%
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HHO
O
O
NHH
R
2S,3R,4E
Configuration of the glycolipid lipid moiety
HHO
O
O
OHOH
HO
OO
OH
OO
OH
O
NHH
OH
NHAc
O
O
OH
OH
-OOC
H
H
HHO
OOH
HO OHO
O
O
OHOH
OH
HO
OH
OO
OOH
OO
OH
O
NHHNHAc
OH
NHAc
O
O
OH
OHO
OH
OH
OH
NHAc
NHAcHO
HO
O
O
HO
OH
-OOC
-OOC
-OOC
AcO
GM3
O-Ac-GT1b
Gangliosides
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J. Biochem. (1984) 96: 1943-46
Chloroform/methanol/0.2% aqueous CaCl2, 50:42:11, first runPropan-1-ol/17M NH4OH/water, 6:2:1, second run
Two dimensional TLC of the ganglioside mixture from adulthuman temporal cortex.
Separation of the ganglioside mixture bycombining normalphase- with reversed phase HPLC
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Minimum energy conformation of the head group of phosphatidylcholine (PC) and of GM1 ganglioside.
PC GM1
Brocca,P., Berthault,P., Sonnino,S. (1998). Biophysical J. 74, 309.
Acquotti,D, Poppe,L., Dabrowski,J., von der Lieth G.W., Sonnino,S., Tettamanti,G. (1990) J.Am.Chem.Soc. 112, 7772.
Volumes occupied by phosphatidylcholine and ganglioside GM1 hydrophilic heads at the membrane surface
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GLYCEROPHOSPHOLIPIDS
SPHINGOMYELIN
CHOLESTEROL
GANGLIOSIDES
PC
PE/PS
GEOMETRICAL PROPERTIES OF SOME AMPHIPHILES
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HYDROGEN BONDS AT THE WATER/LIPID INTERFACE OF SPHI NGO-AND GLYCEROLIPID AGGREGATES
OR
O
O
H
O
O
ROH HN
HHO
O
RHXR
R
O
RXH
RXH
Membrane Lipid Domains: Known as Lipid Rafts
Concerning glycosphingolipids: the big oligosaccharidechain, the existence in solution of severaloligosaccharide conformers, the trans configuration ofthe double bond of sphingosine, the network of hydrogenbonds involving the amide linkage and the absence ofdouble bonds inside the acyl chain, are the driving forcesfor the glycosphingolipid phase separation.All this reduces the fluidity of this membrane portionallowing sorting of cholesterol and DPPC.These membrane domains result in an enrichment ofglsphingolipids, cholesterol and DPPC with respect ofthe total glycerophospholipids. In addition they contain afew proteins but mainly involved in cell signaling.
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Changes of the membrane curvature following oligosaccharide contentand structure.
Transmission electron micrographs of clustered polysialogangliosides at the outer surface of fish brainneurons, using gold labeled anti-polysialoganglioside antibody. Accumulation of binding sites is shown forneuronal cell bodies (A), nerve fibers (B) and synaptic terminals (C). Note that polysialogangliosides areclustered in neuronal cells and nerve fibers, and accumulated/clustered at the neuronal junction sites,where Ca2+-dependent ATPase, ion channels, and ion carrier are also clustered.
A B C
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Changes in ganglioside expression during mouse brain development
J Lipid Res. 2009 50(Suppl): S440–S445
Sphingolipid metabolic pathways
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Glycosyltransferase properties
DNA�RNA�proteins�Golgi enzymes
Recognizing sites for the acceptor and for the donor
Partially sequentially distributed in different cisternae of Golgi
Ganglioside biosynthetic pathways
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sialyltransferase: Preti A. et al. (1980) J. Neurochem 35, 281
sphingomyelin sinthase II: Huitema K. et al. (2004) EMBO J 23, 33
Cell surface associated enzymes involved in glycosphingolipid biosynthesis
sialidase Neu3: Monti E. et al. (2000) Biochem. J. 1, 343
β-hexosaminidase: Mencarelli S. et al. (2005) FEBS Lett. 579, 5501
β-glucosidase GBA1: Aureli M. et al. (2009) FEBS Lett. 583, 2469
sphingomyelinases: Levade T. et al. (1999) BBA 1438, 1
β-glalactosidase: Aureli M. et al. (2009) FEBS Lett. 583, 2469
sialyltransferase SAT2: Crespo PM et al. (2010) J Biol Chem 285, 29179
β-glucosidase GBA2: van Weely et al. (1993) BBA 1181, 157
β-N-acetylgalattosaminyltransferase: Crespo PM et al. (2010) J Biol Chem 285, 29179
Glycosylation/deglycosylation cycles, directly triggered at the cell surface,
might be very important mechanism responsible for rapid and possibly
transient changes of the plasma membrane structure and curvature in
response to specific signals.
Changes of the membrane curvature following oligosaccharide changes.
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O
OH
OOH
HO
OH
O
OH OH
NHAc
O
OO
OHO
OH
O
-OOC
OH
HO
O
OH
O
OH
HOOH
OCer
AcHN
O
-OOC
OH
HO
HO
OHAcHN
Lactonization of GD1b under acidic conditions
H+
O
OH
O
OH
HO
OH
O
OH OH
NHAc
O
OO
OHO
OH
O
-OOC
HOHO
O
OH
HOOH
OCer
AcHNO
O
O O
HOAcHN
HO
OH
HO
Na+, H+ antiporter
Na+
GD1b-lactone
PK
C
-
+
+
PIP2
IP3
DAG
+
+Extracellular signal
Ganglioside-protein specific interactions
unknown protein
H+
GD1b
+
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J Neurochem, 57, 1207-1221
Modulation of the membrane kinase activity by GD1b and GD1b-lactone
Sphingolipid metabolic pathways
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Reorganization of secondary lysosom
membrane enriched of cholesterol
membrane enriched of bis(monoacylglycero)phosphoric acid
1-enzyme: exoglycohydrolase very specific for the disaccharide linkage
2-cofactor: proteic activator
1 + 2a b c d
a
b c d
Lysosomal glycosphingolipid degradation
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Sphingolipidosis
Several severe diseases due to the impairment of one of the sequential
steps of (glyco)sphingolipid lysosomalcatabolism with consequent
accumulation of a specific sphingolipid
Gal-GalNAc-Gal-Glc-CerNeu5Ac
- GalNAc-Gal-Glc-Cer Neu5Ac
-
Gal
β-galactosidase
Gal-Glc-Cer Neu5Ac
-
GalNAc
β-hexosaminidase
GM2 activator protein
Gal-Glc-Cer
Glc-Cer Cer
Neu5Ac
Gal
α-sialidaseGM2 activator protein
β-galactosidase
Gaucher disease
β-glucocerebrosidase
saposin activator
saposin activator
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Glc-Cer Cer β-glucocerebrosidase
Gaucher disease/GBA1 associated Parkinson disease
Reduced or absent GBA1 activity
β-Glucosidase activity in GD fibroblasts
nmol
es/(
106
cells
*h)
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Glucosylceramide accumulation in the brain of a Gaucher disease mouse model
mild inflammation, no neurodegenerative symptomes
strong inflammation, neurodegenerative symptomes
Hum Mol Genet. 2014 23:843-54
Lysosome
Lysosomal accumulationof undegradedmolecules
1
TFEBP
TFEB nucleartranslocation
TFEBP
Nucleus
TFEB
Lysosomalimpairment
LysosomalbiogenesisSecondary
storage of uncatabolizedglycosphingolipids
Accumulationof impairedlysosomes
Fusion Lysosomes-PM
Plasma membrane (PM)
Complex glycosphingolipidsand their catabolic enzymes
Alteration of PM glycosphingolipidcomposition
Cell damage
2
3
4
5
6
78
9
Lysosomal hydrolase
Glycosphingolipids
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Parkinson’s disease
Parkinson's disease (PD), the most prevalent movementdisorder of our time, can occur as an inherited disorder or morecommonly in sporadic form of indefinite etiology.Degeneration of dopaminergic (DA) neurons of the substantianigra (SN).A well-recognized feature is loss of pigmented DA neurons inthe SN that project to the striatum, resulting in progressive DAdeficit and malfunctioning of the neural circuits within the basalganglia that regulate movement. Lewy bodies that containaggregated α-synuclein (aSyn) together with ubiquitin withinthe cytosol and neurites are an additional hallmark
Brains showing GM1 low content, such asParkinson's disease brains, show elevation ofaggregated forms of α-Syn.
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O
H
O
OH
HO
OH
O
OH OH
NHAc
OO
O
OH
O
OH
O
-OOC
HO
HO
OH
OH
O
OH
HO
OH
O
H HN
HO H
O
AcHN
O
OO
OH OH
NHAc
OO
O
OH
O
OH
O
-OOC
HO
HO
OH
OH
O
OH
HO
OH
O
H HN
HO H
O
AcHN
H
UDP-GalGalactosyltransferase, B4galnt1
GM2
GM1
Genetically engineered mice (B4galnt1−/−) devoid of GM1 areconsidered to develop parkinsonism, acquiring characteristicsymptoms of this disorder, including motor impairment,depletion of striatal dopamine, selective loss of tyrosinehydroxylase-expressing neurons, and accumulate alpha-synuclein in nigral neurons.This seems to be due to a failure in signaling efficiency by thecatecholaminergic growth factor, GDNF.
Experimental increase of the GM1 content reduces theaggregated forms of α-Syn in B4galnt1−/+ animal models andimprove several animal behaviors.
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Deficiency of ganglioside GM1 correlates with Parki nson's disease in mice
Intraperitoneal injection into B4galnt1−/+ mice of a GM1derivative, GM1 containing an acetyl group as acyl chainand capable to cross the blood brain barrier, reduces theaggregated forms of α-Syn in B4galnt1−/+ animal modelsand improve several animal behaviors.
O
H
O
OH
HO
OH
O
OH OH
NHAc
OO
O
OH
O
OH
O
-OOC
HO
HO
OH
OH
O
OH
HO
OH
O
H HN
HO T
O
AcHN
O
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WT
0 1 2 3 40
5
10
15
Weeks
time
(sec
)
Saline
0 1 2 3 40
10
20
30
40
50
Weeks
time
(sec
)
Liga4
0 1 2 3 40
10
20
30
40
Weeks
time
(sec
)
OligoGM1
0 1 2 3 40
10
20
30
40
50
Weeks
time
(sec
)
Irritant removal test
0 1 2 3 40
10
20
30
40
50 Saline
Weeks
Tim
e (s
ec)
0 1 2 3 40
100
200
300
400 WT
Weeks
Tim
e (s
ec)
0 1 2 3 40
100
200
300
400
500 OligoGM1
Weeks
Tim
e (s
ec)
0 1 2 3 40
100
200
300
400 Liga4
Weeks
Tim
e (s
ec)
Grip test
GM1-acetyl GM1-acetyl