Lipid Transport between the Endoplasmic Reticulum and Mitochondria

24 Pages·2013·1 MB·English

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Lipid Transport between the Endoplasmic
Reticulum and Mitochondria
VidV.FlisandGu¨ntherDaum
InstituteofBiochemistry,GrazUniversityofTechnology,A-8010Graz,Austria
Correspondence:[email protected]
Mitochondria are partially autonomous organelles that depend on the import of certain
proteinsandlipidstomaintaincellsurvivalandmembraneformation.Althoughphospha-
tidylglycerol,cardiolipin,andphosphatidylethanolaminearesynthesizedbymitochondrial
enzymes,phosphatidylcholine,phosphatidylinositol,phosphatidylserine,andsterolsneed
tobeimportedfromotherorganelles.Theoriginofmostlipidsimportedintomitochondriais
the endoplasmic reticulum, whichrequires interaction of these twosubcellularcompart-
ments.Recently,proteincomplexesthatareinvolvedinmembranecontactbetweenendo-
plasmicreticulumandmitochondriawereidentified,buttheirroleinlipidtransportisstill
unclear. In the present review, we describe components involved in lipid translocation
between the endoplasmic reticulum and mitochondria and discuss functional as well as
regulatoryaspectsthatareimportantforlipidhomeostasis.
Biological membranes are major structural organelles(Futerman2006).Besidestheexport
components of all cell types. They protect ofceramides,theERsuppliesalargeportionof
the cell from external influences, organize the lipids to other organelles, which cannot pro-
interiorindistinctcompartmentsandallowbal- ducetheirownlipidsorhavealimitedcapacity
ancedfluxofcomponents.Besidestheirspecific todoso.Organelleinteractionandtransportof
proteome,organellesexhibituniquelipidcom- lipids require specific carrier proteins, mem-
positions,whichinfluencetheirshape,physical brane contact sites, tethering complexes, and/
properties, and function. Major lipid classes or vesicle flux. These processes are highly im-
foundinbiologicalmembranesarephospholip- portant for the maintenance of cell structure
ids,sterols,andsphingolipids. and survival but are still a matter of dispute.
Themajor“lipidfactory”withinthecellis Mostprominentorganelleinteractionpartners
the endoplasmic reticulum (ER). It is able to aretheERandmitochondria.Asubfractionof
synthesizethebulkofstructuralphospholipids, the ER named mitochondria-associated mem-
sterols,andstoragelipidssuchastriacylglycer- brane (MAM) (Vance 1990) was described to
olsandsterylesters(vanMeeretal.2008).Fur- beinvolvedinlipidtranslocationtomitochon-
thermore, initial steps of ceramide synthesis dria.MAMispartoftheERnetwork,whichwas
occur in the ER providing precursors for the shown to be in contact or close proximity to
formation of complex sphingolipids in other the outer mitochondrial membrane (OMM).
Editors:SusanFerro-Novick,TomA.Rapoport,andRandySchekman
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V.V.FlisandG.Daum
ContactsitesbetweenMAMandmitochondria headgroupandlargehydrophobictail,PEisa
wereassumedtofacilitateexchangeofcompo- typical nonbilayer-forming phospholipid (see
nentsbetweenthetwocompartments.Interest- Fig.1).Thisstructureappearstobeimportant
ingly, MAM harbor a numberof lipid synthe- forsomeperipheralandintegralmembranepro-
sizing enzymes (Gaigg et al. 1994). Recently, teins(vandenBrink-vanderLaanetal.2004).
molecular components governing membrane TogetherwithCL,PEplaysacrucialroleinmain-
contactbetweenthetwoorganelleswereidenti- taining mitochondrial morphology. Loss of
fied(Dolmanetal.2005;Csorda´setal.2006;de bothPEandCLislethalprobablyduetodefects
BritoandScorrano2008;Kornmannetal.2009; inmitochondrialfusion(Joshietal.2012)and/
Friedman et al. 2010; Lavieu et al. 2010), al- orproteindestabilizing(Bogdanovetal.2008b;
though the specific role of these components Osmanetal.2009b).LackofPEsynthesizedin
inlipidtranslocationisnotyetclear. the IMM cannot be fully compensated by PE
importedfromextramitochondrialsites(Birner
etal.2001;Bu¨rgermeisteretal.2004;Joshietal.
MAJORPHOSPHOLIPIDCLASSESOF
2012). However, PE derived from exogenous
MITOCHONDRIA
lyso-PE can restoreboth intra-andextramito-
Cardiolipin (CL) and/or phosphatidylglycerol chondrial PE deficiencies by remodeling pro-
(PG) are considered as mitochondria-specific cesses(RiekhofandVoelker2006).Noteworthy,
phospholipids (Zinseret al. 1991). Both lipids mitochondria are the major supplier of PE to
are synthesized by mitochondria themselves otherorganelles(Voelker1984).
(DavidsonandStanacev1971).Theimportance MitochondrialenzymesofCLandPGsyn-
ofCLforATP-productionhasbeenshownwith thesis,aswellasthemajorPSdecarboxylase,are
differentmembranetypes,e.g.,bacteria,hydro- synthesized on cytosolic ribosomes, imported
genosomes, and mitochondria (Mileykovskaya into mitochondria, and assembled into the
etal.2005;Schlame2008;Acehanetal.2011). IMM (Minskoff and Greenberg 1997; Jiang
Inmitochondria,themajorityofCLislocalized et al. 2000; Nowicki et al. 2005; Nebaueret al.
to the inner membrane (IMM) (Zinser et al. 2007; Choi et al. 2012). Only recently werewe
1991), but substantial amounts were also de- abletodemonstrateexperimentallytheinvolve-
tected in the OMM (Gebert et al. 2009). With ment of components of the mitochondrial
its uncommon, dimeric structure (Fig. 1) CL TOMcomplexandofmitochondrialproteases
together with phosphatidylethanolamine (PE) inimportandprocessingofthemitochondrial
interacts with many mitochondrial proteins PS decarboxylase Psd1p from the yeast (Hor-
(Osmanetal.2011)andstabilizestheirconfor- vathetal.2012).
mation(Joshietal.2012).MutantslackingCL Phosphatidylcholine(PC)hasabighydro-
andPEbiosynthesisaresyntheticlethalinyeast philic head group and a long hydrophobictail
and bacteria (Gohil et al. 2005). In bacteria, (seeFig.1).Itscylindricalshapemakesitaper-
bothlipidsareorganizedinmembraneclusters fect component of bilayer membranes and its
(Matsumoto et al. 2006). Specific interaction roleasastructuralcomponentisessential(van
partnersofCLareproteinsofATPproduction, Meer et al. 2008). Specific functions of PC in
mitochondrial transport systems (Bogdanov mitochondria are not well defined. Similar to
etal.2008a;SchlameandRen2009),andpro- PC, specific functions of phosphatidylinositol
teins requiredformitochondrial structureand (PI) in mitochondria are not known, but its
fusion(Joshietal.2012). essentialroleintotalcellularmetabolismmakes
Another prominent mitochondrial phos- it directly or indirectly indispensible for the
pholipidisPE.Mitochondriaareabletosynthe- maintenance of mitochondria. The bulkhead
sizealargeportionofPEbydecarboxylationof groupofPImakesthisphospholipidaspecific
phosphatidylserine(PS)(Vialetal.1982;Trotter membranebilayercomponent.
etal.1993;Emotoetal.1999;Birneretal.2001; Phosphatidylserine (PS) has also a cylin-
Nerlichetal.2007).Withitssmallhydrophilic dricalshapethatallowsintegrationintobilayer
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LipidTransportbetweentheERandMitochondria
Phosphatidylserine Phosphatidylcholine Phosphatidylinositol Cylindrical
CH3 bilayer
C–O2 N+H3 CH3 N+ CH3 HO HO OH
CH CH2 OH
O O
CH2 CH2 H
*O P O
O O
O
*O P O *O P O
O O CH2 CH CH2
O O
CH2 CH CH2 CH2 CH CH2
O O O O R1 R2
R1 R2 R1 R2
Conical Conical
Cardiolipin Phosphatidylethanolamine
nonbilayer nonbilayer
+
NH3
OH
CH2 CH CH2 CH2
O O CH2
*O P O O P O* O
O O *O P O
CH2 CH CH2 CH2 CH CH2 O
O O O O CH2 CH CH2
R1 R2 R2 R1 O O
R1 R2
Figure1.Majorphospholipidclassesofmitochondria.Chemicalformulasandgeometricalformsofthemajor
mitochondrialphospholipidsareshown.Theshape-structureconceptoflipidscomparestheareaoftheheadgroup
withtheareaoftheiracylchains.Ifthecross-sectionareaoftheheadgroupissimilartothatoftheacylchains,lipids
haveanoverallcylindricalshapeandhaveastrongtendencyofself-assemblyintobilayerphasesofbiological
membranes.AtypicalexampleforsuchgeometryisPC.Ifthecross-sectionareaoftheheadgroupissmallerthan
thatoftheacylchains,lipidshaveaconicalshapeandformstructureswithnegativecurvaturesuchashexagonal
phase.ExamplesofthistypearePEandCL.WhilePC,PS,andPIexhibitcylindricalshapeandself-organizeinto
bilayers,PEandCLwiththeirconicalshapeinducehexagonalphasesanddisturbbilayerarrangement.
membranes(seeFig.1).AlthoughPSispresent Vance1997;vanMeer 2008).OMMandIMM
in low concentrations in organelles of eukary- arestronglydifferentwithrespecttotheirlipid
oticcells(ZinserandDaum1995),itisimpor- equipment. Whereas the IMM is strongly en-
tantasaprecursorforthetwomajorphospho- richedinproteinsandcontainsonly20%lipids
lipids,PEandPC.SupplyofPStomitochondria oftotalmass,theOMMisalipidrichmembrane
isessentialbecauseinmanycellsthemajorityof (ZinserandDaum1995;Dolisetal.1996;Daum
PEisformedbydecarboxylationofPSinmito- andVance1997).AccumulationofCL,PG,and
chondria.ThemetabolicconversionofPStoPE also PE in the IMM appears to be related to
uponimportofPSintomitochondriaprovides functionsmentionedabove.
aconvenientbiochemicalmethodtostudythis
importprocess(Voelker1988,1989b,1991,1992,
SYNTHESISOF
1993;Achleitneretal.1995,1999).
AMINOGLYCEROPHOSPHOLIPIDS
Typically, the lipid composition of mito-
INVOLVESINTERACTIONOFORGANELLES
chondria shows as major components 40%–
44% PC, 27%–34% PE, 1%–3% PS, 5%– The biosynthetic sequence of aminoglycero-
15% PI, and 13%–14% CL depending on the phospholipid formation starts with the syn-
cell type (Zinser and Daum 1995; Daum and thesis of PS in the ER (Fig. 2). The highest
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V.V.FlisandG.Daum
A Endoplasmic reticulum Mitochondria
Pss1p/Cho1p
CDP-DAG PS PS
Serine
Golgi network Psd2p –CO2 –CO2 Psd1p
Eki1p Ect1p Ept1p
Etn PE PE
ATP CTP DAG
SAM Cho2p/Pem1p
Dpl1p
SL
SAM Opi3p/Pem2p
SAM Opi3p/Pem2p
Cki1p Cct1p Cpt1p
Cho PC
ATP CTP DAG
B Mitochondria Endoplasmic reticulum
Ser PSS1 Cho CPT CCT CK
PS PC Cho
PS
DAG CTP ATP
Ser
PSD –CO2 PSS2 PE3M T× SAM CDP-Cho
Etn
EPT ECT EK
PE PE Etn CDP-Etn
DAG CTP ATP
C PL PL CDP-Etn/Cho
Methylation pathway
Decarboxylation pathway
Land’s Acyl-CoA:lysophospholipid
PLA2 cycle acyltransferase
Fatty acid Acyl-CoA
Lyso-PL
Figure2.Biosyntheticpathwaysofaminoglycerophospholipids.(A)Biosynthesisofaminoglycerophospholipids
inyeast.FormationofPSisaccomplishedintheERandcatalyzedinaCDP-DAG-dependentreactionbythePS
synthase,Pss1p.DecarboxylationofPSyieldingPEoccursinmitochondria(viaPsd1p)andaGolgi/vacuolar
compartment (viaPsd2p).Three-stepmethylationofPEintheERcatalyzedbyCho2p/Pem1pandOpi3p/
Pem2p leadsto formation ofPC with S-adenosine methionine (SAM)as methyldonor. PC andPE can be
formedalsobytheCDP-ethanolamineandCDP-cholinebranchesoftheso-calledKennedypathwaymakinguse
ofexogenousorendogenouscholine(Cho)andethanolamine(Etn),respectively.SL,sphingolipids.(B)Amino-
glycerophospholipidbiosynthesisinmammaliancells.ThemajormechanismofPSproductioninmammalsis
baseexchangewithPC(PSS1)andPE(PSS2)assubstrates.AmajorrouteofPEandPCproductionaretheCDP-
ethanolamineandCDP-cholinepathways.Inmammaliancells,PEcanalsobeproducedbydecarboxylationof
PS.Inhepatocytes,asinglemethyltransferasecatalyzesmethylationofPEtoPC.(C)TheLand’scycledescribesa
sequenceofdeacylationandreacylation.PEandPCareconvertedtolyso-PEandlyso-PCandviceversa.
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LipidTransportbetweentheERandMitochondria
concentration of PS synthesizing enzymes has pathway cannot fully complement for the mi-
been detected in the MAM (mitochondria-as- tochondrial requirement of PE in cells deleted
sociatedmembrane)(Kuchleretal.1986;Vance of both PSD1 and PSD2 (Trotter and Voelker
1990).Interestingly,mammaliancellsandyeast 1995). Ethanolamine phosphate, which is an
havedifferentpathwaystoproducePS.Inmam- intermediate in this biosynthetic sequence,
malian cells, two phosphatidylserine synthases canalsobederivedfromsphingolipiddegrada-
(PSS-1andPSS-2)producePSbybaseexchange tion providing a link between sphingolipid
attheheadgroupofPEorPC(Vance2008)ina and PE metabolism (Mao et al. 1997; Saba
Ca2þ-dependentreaction(Fig.2B).Thisrelease etal.1997).Finally,yeastharborsenzymesthat
of Ca2þ into the lumen of the ER is energy catalyze acylation of lyso-PE (Riekhof and
dependent. In yeast, formation of PS is cata- Voelker2006; Riekhof etal. 2007b; Deng etal.
lyzedbythegeneproductofPSS1/CHO1(Letts 2010).ThistypeofreactionnamedLand’scycle
etal.1983),whichrequiresCDP-diacylglycerol includes deacylation/reacylation of phospho-
(CDP-DAG)andserine(NikawaandYamashita lipids(Lands1958)andappearstobeimportant
1981) as substrates and depends on Mg2þ or forremodelingprocesses.Thereasonfortheex-
Mn2þ(Fig.2A).Cellularenergyisrequiredfor istenceofoverlappingpathwaysisstillamatter
the formation of CDP-DAG. In plants, both ofdispute,althoughevidencefordistinctpools
pathwaysdescribedaboveareactiveforPSpro- of PE has been presented (Bu¨rgermeisteret al.
duction(Delhaizeetal.1999;Manoharanetal. 2004).
2000;Ronteinetal.2003).Inalltypesofcells, Mammalian cells have the capacity to syn-
PS synthesized in the ER is exported to other thesizetheirentirePEthroughthePSDpathway
organelles, including mitochondria where it in mitochondria (Voelker and Frazier 1986),
servesasasubstrateforPEsynthesis. but in the presenceof ethanolamine the CDP-
PE is the second most abundant lipid of ethanolamine pathway can fulfill most of the
eukaryotic cells. It can be synthesized by four extramitochondrial requirements for this lipid
different pathways: (i) the CDP-ethanolamine (Vance 2008). The Land’s cycle is important
pathway(alsonamedKennedypathway)(Kan- for remodeling phospholipids that have been
ferandKennedy1964),(ii)decarboxylationof formedbyotherpathways.Asanexample,for-
PStoPE,(iii)baseexchangebetweendifferent mation of PE from lyso-PE can be accom-
phospholipids, and (iv) acylation of lyso-PE plishedbyLPEAT2(acyl-CoA:lysophosphatidyl-
(see Fig. 2). Yeast has two PS decarboxylases ethanolamine acyltransferase 2) in the brain
(PSD) with overlapping functions (Trotter et cells. Ethanolamine phospholipids are major
al. 1993). Psd1p is localized to mitochondria constituents of the myelin sheath increasing
(Trotteretal.1993),andPsd2phasbeenfound the signal transmission speed along the axons
inaGolgi/vacuolarcompartment (Trotterand (Caoetal.2008).Inplants,remodelingofPEto
Voelker 1995). Psd1p is a component of the lyso-PE was reported to be involved in plant
IMM/intermembrane space and produces the growthpromotionandleafsenescence(Cowan
majorityoftotalcellularandmitochondrialPE 2009;Hongetal.2009).Intheparasites,Trypa-
(Trotteretal.1993;Birneretal.2001). nosoma brucei and Plasmodium berghei, PE is
YeastcanalsoproducePEthroughtheCDP- exclusively produced through the CDP-etha-
ethanolamine branch of the Kennedy pathway nolaminebranchoftheKennedypathway(Ser-
(KennedyandWeiss 1956).TheCDP-ethanol- ricchioandBu¨tikofer2011).
amine pathway incorporates externally added PC is the most abundant phospholipid in
orendogenousethanolaminethroughstepwise eukaryoticcells(vanMeeretal.2008).Because
phosphorylation, activation with CTP, and at- mitochondria lack PC synthesizing enzymes,
tachment to diacylglycerol (DAG). In the final thisphospholipidhastobeimportedfromthe
step,phosphoethanolamineistransferredfrom ER. PC can be produced via three pathways:
CDP-ethanolamine to a DAG acceptor form- (i)theCDP-cholinebranchoftheKennedypath-
ing PE. However, PE synthesized through this way(GibelliniandSmith2010),(ii)methylation
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V.V.FlisandG.Daum
ofPE(SundlerandAkesson1975;Lietal.2005), Cds1p,eitherintheER/MAMorinmitochon-
and(iii)theLand’scycle,inwhichLPCAT(lyso- dria. Then, conversion of CDP-DAG to phos-
phosphocholine acyltransferase) produces PC phatidylglycerolphosphate(PGP)catalyzedby
from lyso-PC and fatty acids (Riekhof et al. Pgs1p occurs (Chang et al. 1998a). PGP is de-
2007a; Hishikawa et al. 2008; Shindou et al. phosphorylated to PG in a reaction that is ac-
2009) (see Fig. 2). In most mammalian cells, complished by the PGP phosphatase Gep4p
themajorityofPCisformedthroughtheCDP- (Osman et al. 2010). In the final step of CL
choline pathway (Kennedy 1956; Gibellini and synthesis,aphosphatidate(PA)moietyistrans-
Smith 2010; Hermansson et al. 2011), and ferredfromCDP-DAGtothehydroxylgroupin
only inhepatocytes PEmethylation isthepre- the headgroup of PG. Thecleavage of the py-
dominantpathway(SundlerandAkesson1975; rophosphategroupprovidesthechemicalener-
Li et al. 2005). Reactions of the CDP-cho- gy for the latter reaction. In yeast (Jiang et al.
line pathway are similar to the CDP-ethanol- 1997; Chang et al. 1998b; Tuller et al. 1998),
aminepathwayandcatalyzedbycholinekinase, Arabidopsis(Katayamaetal.2004;Nowickietal.
phosphocholinecytidyltransferase,andcholine 2005),andhumancells(Chenetal.2006;Hout-
phosphotransferase.Inthefinalstep,phospho- kooperetal.2006;Luetal.2006),CLsynthase
cholineistransferredfromCDP-cholinetoDAG waslocalizedtomitochondria.Importantly,CL
formingPC.Inyeast (Boumannetal.2004)as undergoes remodeling processes (Joshi et al.
wellasinmammals(HenneberryandMcMaster 2009; Schlame and Ren 2009) with so-called
1999), enzymes of the both branches of the tafazzins(Malhotraetal.2009)involved.Tafaz-
Kennedy pathway have overlapping substrate zinsarephospholipidtransacylasesthattransfer
specificities. acyl groups from phospholipids, preferentially
The second pathway of PC production in PC, to monolysocardiolipin (MLCL). The re-
mammalian cells (Van Pilsum and Carlson versereactionofthismodificationisthetransfer
1970) and yeast (Kodaki and Yamashita 1987) ofanacylgroupfromCLtolyso-PC.Thereac-
isPEmethylation.Inthissequence,S-adenosyl- tion does not require activation of fatty acids
methionine(SAM) serves as methyl donor for butoccursinalysophospholipid–phospholip-
threestepsofmethyltransferasereactions.Inthe idcomplexbydeprotonationandnucleophilic
firststep,PEismethylatedtomonomethyl-PE attackontheesterbondoftheacyldonor(Xu
and then further converted to dimethyl-PE in etal.2006).
the second step. The biosynthetic sequence is PA and CDP-DAG are important interme-
completedbyathirdstepofmethylationyield- diatesnotonlyforPG/CLsynthesisbutalsofor
ing the final product PC. In yeast, the gene the formation of PI and PS (Athenstaedt and
products of CHO2/PEM1 and OPI3/PEM2 Daum 1999). In the yeast, PA is synthesized
are involved (Kodaki and Yamashita 1987), from glycerol-3-phosphate and/or dihydroxy-
whereashepatocytesharboronlyoneN-meth- acetonephosphate(DHAP)intheERandlipid
yltransferase, which catalyzes all three steps of dropletsbytwostepsofacylation.Thefirstacet-
PE methylation (Sundler and Akesson 1975; ylationreactionwithDHAPasasubstrateleads
Ridgwayetal.1989).Thelowereukaryote,Try- to acyl-DHAP, which is then reduced by the
panosoma brucei, lacks genes coding for PE 1-acyl-DHAP reductase Ayr1p in an NADPH-
N-methyltransferases(Gibellinietal.2009). dependent reaction to lyso-PA. Alternatively,
acylation of glycerol-3-phosphate leads to for-
mationoflyso-PA.Inafinalacylationstep,lyso-
CARDIOLIPINAND
PAisconvertedtoPA(Athenstaedtetal.1999a;
PHOSPHATIDYLGLYCEROLSYNTHESIS
SorgerandDaum2003).PAcanalsobegener-
INMITOCHONDRIA
ated by phospholipase D (Osman et al. 2011)
SynthesisofPGandCLinmitochondriaoccurs in mitochondria. In plants, PA synthesis oc-
byamultistepreactionsequence(Schlameetal. curs in plastids, mitochondria, and micro-
1993). First, PA is converted to CDP-DAG by somes. Dephosphorylation of PA is catalyzed
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LipidTransportbetweentheERandMitochondria
by mammalian lipin and the yeast ortholog cellshavebeenperformed(ButlerandThomp-
Pah1p(CarmanandHan2006,2009;Brindley son 1975; Voelker 1985, 1989a,b, 1990, 1991;
etal.2009;HarrisandFinck2011),whichwere Vance1990; Achleitneret al. 1995; Shiao et al.
recently shown to be pacemakers and switch 1995;Emotoetal.1999;Kugeetal.2001;Schu-
pointsinlipidmetabolism.Dephosphorylation macheretal.2002;WuandVoelker2004;Car-
ofPAisacrucialstep,becausetheproductofthe rasco et al. 2006; Riekhof and Voelker 2006;
dephosphorylation reaction, DAG, becomes Kornmannetal.2009;Nguyenetal.2012;Padi-
substrate for triacylglycerol synthases and thus lla-Lo´pezetal.2012;Tamuraetal.2012).These
promotes biogenesis of lipid droplets (Adeyo studies showed that in mammalian cells PS
etal.2011).PA,whichisnotdephosphorylated, transport from MAM to the OMM requires
ispreferentiallyconvertedtoCDP-DAG,which ATP. Further import of PS to the IMM yields
isfurtherusedforthesynthesisofphospholip- PE (Voelker and Frazier 1986; Voelker 1990,
ids. CDP-DAG synthase activity in yeast, as 1991;Shiaoetal.1995).
well as in mammals, was localized to the ER Experiments with isolated organelles pro-
and mitochondria (Kuchler et al. 1986; Kelley vided more detailed information about the
andCarman1987;Chenetal.2006). processofphospholipidimportintomitochon-
dria.Reconstitutionoftransportsystemsusing
isolatedmitochondria andmicrosomes/MAM
PHOSPHATIDYLINOSITOLSYNTHESIS
(Kuchleretal.1986;Simbenietal.1990;Vance
PIisproducedintheERandhastobeimported 1990;Achleitneretal.1999;Emotoetal.1999)
intomitochondria.PIissynthesizedfromCDP- revealedthatuptakeofPSbymitochondriade-
DAGandinositol(Gardockietal.2005).Inthe pended on the PS concentration in the do-
yeast,deletionoftheonlyphosphatidylinositol nor membranes (ER, Golgi) (Wu and Voelker
synthasegenePIS1islethal(Nikawaetal.1987). 2004).AlthoughATPwasrequiredfortransport
Furthermore,PIservesasprecursorforcellsig- ofPSinintactandpermeabilizedcells(seebe-
naling molecules such as phosphatidylinosi- low),nosuchobservationwasmadeinthere-
tolphosphates(PIP),phosphatidylinositolbis- constituted in vitro system. It was argued that
phosphates (PIP2) and phosphatidylinositol closeappositionofdonor(ER/MAM)andac-
triphosphates (PIP3) and for the biosynthesis ceptor (mitochondria) membranes was suffi-
ofGPIanchors(SerricchioandBu¨tikofer2011). cient for transport,butthatATPwasprobably
requiredtoprovideappropriateconditionsfor
transport of this phospholipid in intact cells.
TRANSPORTOFPHOSPHOLIPIDSBETWEEN
Experiments making use of the coisolation of
THEENDOPLASMICRETICULUMAND
MAM with mitochondria suggested that the
MITOCHONDRIA
contact of these two cellular fractions is fairly
AsthebiosyntheticsequenceofPS-PE-PCsyn- tight(Vance1990;Achleitneretal.1999).Alto-
thesis occurs in different subcellularcompart- gether,itwasconcludedthatmembranecontact
ments (see above), the amounts of intermedi- betweenERandmitochondriaisimportantfor
ates and products are indication and measure lipid translocation between these two organ-
for transport between organelles. Therefore, elles.
interorganelle translocation of aminoglycero- Experimentsperformedwithpermeabilized
phospholipidscanbeinvestigatedbyfollowing cellsprovidedausefulalternativetotheexper-
the incorporation of serine into PS in the ER, imentsdescribedabove(Limetal.1986;Voelker
decarboxylation of PS to PE in the mitochon- 1992;Achleitneretal.1995;Kugeetal.2001;Wu
dria, and methylation of PE to PC upon the and Voelker 2001). Permeabilized cells are ob-
returnofPEtotheERwithoutisolationofor- tained by mild chemical or mechanical treat-
ganelles.Makinguseofthisexperimentalstrat- ment of whole cells, resulting in the internal
egy, phospholipid transport studies with in- organelle structures remaining largely intact,
tactcells,isolatedorganelles,andpermeabilized while access to the interior is possible for
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V.V.FlisandG.Daum
compoundsthatcannotenteranintactcell.In fectssubstratespecificityoftheubiquitinligase
permeabilizedyeastcells(Achleitneretal.1999) (Chandrasekaran et al. 2006; Yan and Xiong
[3H]serine is incorporated into PS in the ER. 2010).OnesubstratefortheE3ubiquitinligase
Transport of [3H]PS to mitochondria yields isthetranscriptionfactorMet4p,whichisinac-
[3H]PE formed by mitochondrial Psd1p. Use tivated upon ubiquitination. It was suggested
of psd2Dmutants and depletion of ethanol- thatMet30pmediatesubiquitinationofcertain
amine from the medium allows attribution of targetproteinsandmayplayaroleeitherinER-
theformed[3H]PEtotransportof[3H]PSfrom mitochondria recognition,inthe inhibition of
the ER. Transport of [3H]PE from mitochon- this docking process, or in regulation of tran-
driatotheERresultsinformationof[3H]PCas scriptionofafactorinvolvedinlipidtransport.
a measure for this transport route (Achleitner Thesecondcomponentidentifiedinthescreen
etal.1995,1999;Shiaoetal.1998). was the phosphatidylinositol 4-kinase Stt4p,
Theadvantageofpermeabilizedcellsisthat which affects transport of PS to the site of
reactions can be manipulated by addition of Psd2p-driven conversion to PE (Trotter et al.
chemicals such as divalent ions, energy block- 1998).ThemodeofactionofStt4ponorganelle
ers, or cytoskeleton inhibiting reagents (Eilers interactionand/orPStranslocationremainsto
et al. 1989) or by removing the cytosol. This beelucidated.ItwassuggestedthattheC2do-
feature allowed additional characterization of mainofPsd2p,whichbindsCa2þ,interactswith
lipid transport between ER and mitochondria proteinsorlipidsandrecognizesanioniclipids
withmammalian(Voelker1990)andyeastcells such as PS and polyphosphoinositides (Choi
(Achleitneretal.1995).TransportofPStomi- etal.2005)andmaybethebridgeforPStrans-
tochondriawasshowntobedependentonATP location. Finally, the gene product of PDR17/
inmammaliancells(Voelker1989a)butnotin SFH4wasfoundtoaffectPStransporttoPsd2p.
yeast(Achleitneretal.1999).However,acertain Reconstitution assays with permeabilized cells
amount of ATP must be present in both cell and isolated organelles showed that Pdr17p
types (Shiao et al. 1995) because upstream re- mustbepresentontheacceptormembranefor
actionsoflipidprecursorformationareenergy transfer of PS to Psd2p (van den Hazel et al.
consuming.Itwasalsoshownthatongoingsyn- 1999;Wuetal.2000).Themechanismofaction
thesisofPSisnotrequiredfortranslocationto ofPdr17pisstillunknown.
mitochondria because preformed PS was effi- Reconstitution of PS synthesis and trans-
ciently used as a substrate for mitochondrial portinpermeabilizedmammaliancellsidenti-
PSdecarboxylation.Interestingly,whenexoge- fiedtwoothercomponentsthataffectPStrans-
nous mitochondria were added to permeabi- port from the ER/MAM to mitochondria
lized cells, they failed to serve as acceptors for (Emoto et al. 1999; Kuge et al. 2001). One of
PSimport (Voelker1993).Thisresultsuggests these proteins is S100B (NP_006236), an EF-
that pre-existing, stable associations between hand domain-Ca2þ-binding protein. It is not
mitochondriaandER/MAMplayanimportant known whether this protein participates in
roleinthetransportprocess. transport or promotes stability and/or assem-
In a yeast genetic screen, Voelker and co- blyofinteractionsofER/MAMandmitochon-
workerssearchedforcomponentsthatinfluence dria.Thesecondcomponentwasonlyindirectly
the translocation of PSfrom ERtothe sites of identified in the mammalian cell line R41,
decarboxylation in mitochondria (via Psd1p) whichhasadefectinPStransportbetweenthe
(Schumacher et al. 2002) and Golgi/vacuoles OMMandIMM.
(viaPsd2p)(Trotteretal.1998;WuandVoelker Studies with isolated organelles treated
2001, 2004). The first gene product identified with protease (Shiao et al. 1998; Achleitner
was Met30p, which influenced the import of et al. 1999) or dinitrophenol, which alters the
PS into mitochondria. MET30 encodes a sub- distance between the ER and mitochondria
unit of a multicomponent E3 ubiquitin ligase (Hoviusetal.1992),suggestedtheparticipation
(Aghajanetal.2010;Ounietal.2010) andaf- of organelle surface proteins in PS transport.
8 CitethisarticleasColdSpringHarbPerspectBiol2013;5:a013235
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LipidTransportbetweentheERandMitochondria
Electron microscopy supported this view by heritance (Frederick et al. 2004, 2008) and
demonstrating close contact zones between mostlikelyconnectsERMEStotheactincyto-
ER/MAM and mitochondria, which might skeleton (Kornmann et al. 2011; Michel and
serveasbridgesforlipidtranslocation(Csorda´s Kornmann 2012). Itwas assumed that Gem1p
et al. 2006). Furthermore, the involvement of shuttlesbetweenafreeandERMESboundform
acetylated microtubules in ER-mitochondria (seeFig.3),althoughthemolecularmechanism
dynamics was demonstrated, a mechanism by ofGem1premains unknown.Physicalinterac-
which membrane contact can be established tion ofPsd1pwithERMESorGem1pwasnot
and/or maintained (Friedman et al. 2010). demonstrated.Nevertheless,itwasreportedthat
Moreover, mitochondrial division sites pro- a gem1Dpsd2Ddpl1Dyeast mutant grown on a
ducedbyDnm1pandDrp1pwerefoundtobe nonfermentablecarbonsourcewasdefectivein
physically connected with the ER (Friedman its PC synthesis (Kornmann et al. 2011), most
etal.2011). likelybyimpairedPEandPStransport.Asimilar
ToshedmorelightontheroleofER-mito- growth phenotypewas reported before for the
chondriacontact,geneticandsyntheticgenetic gem1Dsingledeletionstraingrownonsynthetic
interactionsanalysismainlywiththeyeast,Sac- glycerol media (Frederick et al. 2008). GEM1
charomyces cerevisiae, were performed (Birner also shows strong synthetic lethality with
etal.2003;Kornmannetal.2009;Osmanetal. GEP4,encodingaPGPphosphatase(Kornmann
2009a). Such genetic screens were designed et al. 2011). Gem1p is well conserved in the
inawaythatdefectsintwoormoreinteracting eukaryotic kingdom, suggesting that Miro
gene products become lethal. Recently, Korn- GTPases are general components of ER-mito-
mann et al. (2009) identified components of chondriaconnections.Miro-1,themammalian
ER-mitochondriacontactinyeastatthemolec- homologofGem1p,interactswithmitofusin1
ularlevel.Theyshowedthatinwild-typecells,a and 2, the human homologs of yeast Fzo1p.
proteincomplextetherstheER/MAMandmi- Mfn-2wassuggestedtotethermitochondriato
tochondria.Mutationsinthisproteincomplex ERandbeinginvolvedinmitochondrialmove-
causedslowgrowthorwerelethalforthecells, mentalongaxons(deBritoandScorrano2008;
butcouldbesuppressedbyanartificialtether- Miskoetal.2010).Mostrecently,however,the
ingconstructcalledChiMERA,whichcarrieda roleofERMESandGem1pintheimportofPS
GFP molecule. When staining the mitochon- intomitochondriaanditsconversiontoPEwas
driawithmito-trackerandexpressingtheChi- challenged(Nguyenetal.2012).Itwasargued
MERA, association was visualized confirming that, despite their genetic and physical inter-
that ChiMERA acted as a bridge between the action, ERMES and Gem1p function in dis-
ER and mitochondria. The authentic complex tinctpathways,andtheabsenceofERMESand
wasnamedERMES(ERmitochondriaencoun- Gem1phadonlylittleeffectonPSimportinto
tered structures) and contains the OMM pro- mitochondriaandconversiontoPE.Theeffect
teins,Mdm10p,Mdm12p,andMdm34p,which on PC formation as described by Kornmann
interactwiththeERmembraneanchoredpro- et al. (2009) was regarded as minor. Nguyen
teinMmm1p.IntheabsenceofERMES,theCL et al. (2012) argued that changes in the lipid
level and the PS/PC conversion rates were de- profileofcellslackingERMESweresideeffects
creased,suggestingtheinvolvementofERMES ofdefectsinmitochondrialmorphology.
in phospholipids transport between ER and Recently, another mitochondrial complex
mitochondria(Fig.3).Moreover,geneticinter- namedMitOS,MICOS,orMINOSwasidenti-
actionsbetweenPSD1,GEM1,andtheERMES fied (Harner et al. 2011; Hoppins et al. 2011;
complex were shown (Kornmann et al. 2009). vonderMalsburgetal.2011).ThisIMM-asso-
TheCa2þMiro(mitochondriarho-like)GTPase ciated complex functions in cristae formation
Gem1p is a regulatory component of ERMES andmorphology(Rabletal.2009).Itwaspro-
(Kornmann et al. 2011; Stroud et al. 2011), posed (van der Laan et al. 2012) that MINOS
maintains mitochondrial morphology and in- forms the central core of a large and complex
CitethisarticleasColdSpringHarbPerspectBiol2013;5:a013235 9
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V.V.FlisandG.Daum
MLCL
TAZ1 CL
CLD1
CL Nonbilayer P-lipids
CRD1
Bilayer P-lipids
PG
PHB1/PHB2
PA CDP-DAG
PGP
PGS1 GEP4
MDM35
CDS1 PSD1 IMM
UPS1/UPS2
YME1 ATP23
PA CDP-DAG PC PE PS UPS2 UPS1
MDM34 MDM10 OMM
2
M1 GEM1 m
MD MMM1 10–40 n
CDS1 ER/MAM
PA CDP-DAG PEM2 PC PEM1PE PS ERMES
Figure 3. Componentsinvolvedintranslocationofphospholipidstoandwithinmitochondriaofyeast.The
directionsoflipidtranslocationareindicatedbyarrows.WhilePSandPCareimportedintomitochondria,alarge
portionPEisexported.UponimportthemajorityofPSisconvertedtoPEbyPsd1p.DecarboxylationofPStoPE
inmitochondriaandmethylationofPEtoPCintheERaresufficientforsupplyingcellswithPEandPCeveninthe
absenceofexternalethanolamineorcholine.SimilartoPSandPC,PAandCDP-DAGalsohavetobeimported
intomitochondriawheretheyarerequiredforCLsynthesis.ThepathwayofCLsynthesisintheIMMstartswith
CDP-DAGandiscompletedduringseveralstepsbyamultienzymecascade.AsmallportionofCLisexportedto
theOMM.TheERMEScomplexcontaininggeneproductsofMDM10,MDM34,MDM12,andMMM1tethersER
andmitochondria.Inmammaliancells,Mfn1/2seemtohaveasimilarfunction.ERMESassociateswiththe
GTPaseGEM1,cyclingbetweenERMESboundandfreeform.UPS1/UPS2interactwithMdm35pandregulate
themitochondriallevelsofPEandCL.UPS1andUPS2aredegradedbyATP23andYEM1,respectively.Prohibitin
ringlikestructuresmadefromtwoproteinsPHB1/2contributetotheformationofPEandCLclustersinthe
IMM.Foradetaileddescriptionofbiosyntheticandtranslocationprocessesandcomponentsinvolved,seetext.
organizing system named ERMIONE, which et al. 2009a; Tamura et al. 2009, 2012; Potting
includes the ERMES complex, the prohibitin et al. 2010). However, direct involvement of
ringlike structures, the TOM and TIM com- MINOS in lipid transport and/or assembly
plexes, and Mdm31/32 proteins required for into mitochondrial membrane has not been
mtDNA maintenance. MINOSwas also found demonstrated.
tointeractwiththeproteinVDACandwiththe
fusionproteinUgo1(vanderLaanetal.2012).
LINKBETWEENCARDIOLIPINFORMATION
BothERMESandMINOSaregeneticallylinked
ANDINTRAMITOCHONDRIALTRANSPORT
to the prohibitin ring complexes of the IMM
OFAMINOGLYCEROPHOSPHOLIPIDS
that are integrated into the lipid network me-
tabolism(Psd1,Ups1/Ups2)(Birneretal.2003; A link between CL and PE assembly in mito-
Gohiletal.2005;Kornmannetal.2009;Osman chondrial membranes, PE synthesis by Psd1p,
10 CitethisarticleasColdSpringHarbPerspectBiol2013;5:a013235