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Endoplasmic reticulum: one continuous network compartmentalized by extrinsic cuesTim Levine1 and Catherine Rabouille2

The endoplasmic reticulum (ER) is an extensive three-

dimensional network that stretches from the inner nuclear

envelope to the cell cortex with a single, continuous membrane

and a single, continuous lumen. Yet the ER contains

specialized regions that carry out unique functions. The

question that immediately arises is how the ER can be

compartmentalized if it is continuous, and the answer to this is

that cellular landmarks with unique sub-cellular distributions

impose non-uniformity on the ER from outside, creating

structural and functional sub-domains of the ER.

Addresses1Division of Cell Biology, Institute of Ophthalmology, 11–43 Bath St,

London EC1V 9EL, UK2The Cell Microscopy Centre, Department of Cell Biology and Institute of Biomembranes, University Medical Centre Utrecht, AZU Rm G02.525,

Heidelberglaan 100, 3584 CX Utrecht, The Netherlands

Corresponding author: Rabouille, Catherine ([email protected])

Current Opinion in Cell Biology 2005, 17:362–368

This review comes from a themed issue onMembranes and organelles

Edited by Vivek Malhotra and Mike Yaffe

Available online 21st June 2005

0955-0674/$ – see front matter

# 2005 Elsevier Ltd. All rights reserved.

DOI 10.1016/j.ceb.2005.06.005

IntroductionThe endoplasmic reticulum (ER) is ubiquitous and cells

contain just a single copy. Yet specialized sub-compart-ments of the ER carrying out unique functions can be

distinguished morphologically: the rough ER (RER),

which is studded with ribosomes, and is typically involved

in coupling protein synthesis to protein translocation

into the ER lumen through the translocon; the transi-

tional ER (tER) sites, which are specialized cup-shapedER cisternae from which newly synthesized proteins exit

via COPII vesicles en route to the remainder of the

secretory pathway [1]; and the smooth ER, which is

dedicated to calcium storage and metabolic pathways

involved in drug handling and lipid and steroid synthesis

(reviewed in [2]).

The question that immediately arises is how the ER can

be compartmentalized if it is continuous, and the answer

to this is that cellular landmarks with unique sub-cellular

distributions impose non-uniformity on the ER from

outside, creating ER sub-domains. In some cases, these

extrinsic cues organising the ER are associated withunique structural types of the ER. A clear example of

this is the nuclear envelope, which is organized by

the nuclear material via lamins and lamin receptors.

The notion of extrinsic organization may also apply to

the RER, which is fixed to regions containing ribosomes

[3]. Furthermore, functional heterogeneities within ER

sub-compartments that are not obviously structurally

distinct can also be created by extrinsic cues. Below,

we will discuss some of the recent interesting discoveries

about issues surrounding the nature of the extrinsic

organizer of the ER.

ER positioning imposed by the cytoskeletonPolarity and structure are imparted to cells by the cytos-

keleton, which allows placement of proteins and orga-

nelles in a characteristic relationship to each other.

Therefore, a large degree of the extrinsic organization

of the pan-cellular ER derives from the cytoskeleton. In

many animal cells, the ER is particularly associated with

the microtubules. In the past, ER tubules were found to

be transported on cytoplasmic microtubules predomi-

nantly (but not exclusively) by kinesins, which pull theER from the centre of the cell toward the cell cortex (i.e.

they are plus-end-directed motors). Recently, ER–micro-

tubule associations have been shown to be independentof motors [4,5]. ER–microtubule interactions without a

motor may be important in highly differentiated cells

such as neurons, where large-scale relocation of the ER is

not constitutively required.

The actin cytoskeleton is the major cytoskeletal partner

of the ER in yeast and plant cells, but also plays a role in

animal cells [6]. For instance, the rearrangements of ER

architecture in C. elegans during early embryogenesis have

recently been shown to depend on actin [7]. Therefore, it

is likely that both microtubules and actin control different

aspects of ER positioning and structure.

Organization of tER sitesLittle is known about how tER sites form and maintain

their seemingly random distribution throughout the cell’s

ubiquitous ER. One approach to understanding tERbiogenesis has been to identify the landmark proteins

that recruit the remainder of the tER machinery, much of

this work being carried out in yeast. The COPII subunit

Sec12 was considered to be the tER landmark, until the

COPII subunits Sec23 and Sar1 were shown to concen-

trate into tER sites even after its depletion [8]. Another

candidate is Sec16, a component of the tER sites in yeast,

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fly and human. Depletion or mutation of Sec16 comple-

tely disorganizes tER sites, with failure to recruit Sec23

(Ben Glick, personal communication; and C Rabouille,

unpublished). The final candidate to consider is the Golgi

matrix protein p115, since in Drosophila S2 cells depletion

of the p115 homologue causes an alteration in the orga-nization of the tER sites [9].

Although the biogenesis of the tER is unclear, it is clear

that tER sites are relatively stable structures, in the sense

that they do not move long distances, although they

undergo fission and fusion in both yeast and mammalian

cells [10,11]. Like the rest of the ER, tER sites may be

spatially arranged along microtubules, and a unique rela-

tionship has recently been discovered between Sec23 and

dynactin [12]. It also seems that the tER sites are stablestructures in the sense that they are not generated in

response to protein synthesis and transport. This has been

addressed by live cell imaging using Sec23, VSVG (pro-

tein G from vesicular stomatitis virus) [13], and procolla-

gen [14] tagged with respective fluorescent tags: Sec23

spots did not appear in response to cargo transport in this

system and did not fade away afterwards.

ER non-uniformities derived from localizedmRNA Localization of mRNA is a clear example of how an

extrinsic cue can impose functional non-uniformity onthe RER and tER without being associated with any

obvious structural features. Many mRNAs exhibit a

restricted localization within the cell [15], leading to

restricted protein synthesis, and recent work has shown

that they form particles associated with microtubules[16,17,18]. This is a key mechanism in the establish-

ment of asymmetric protein distribution, f or instance in

cell polarity (reviewed in [16] and [19]). Localized

mRNA codes not only for cytoplasmic proteins, but also

for transmembrane and secreted proteins. For instance,

out of the 22 mRNAs localized to the bud-tip in yeast, at

least five are membrane-associated [20]. The localizedmRNAs encoding transmembrane and secreted proteins

will create heterogeneity within the RER by localized

translation, and heterogeneity among tER sites by loca-

lized transport.

mRNA localization on ER subdomainsTargeting of mRNA to the RER was first discovered in

Xenopus oocytes [21,22]. Different mRNAs can bind

directly to different sub-domains of the RER. For exam-

ple, two mRNAs for membrane proteins are specificallylocalized to different parts of the ER in rice seeds

[23,24]. One mRNA codes for prolamine, which forms

inclusions derived from the ER. The other mRNA codes

for glutein, which is initially translocated into the ER, but

is not found in prolamine inclusions to be transported via

the Golgi apparatus to the vacuole, where it too forms

inclusions. The separate localizations of the mRNAs

minimize mixing of the two proteins and allow the for-

mation of separate inclusions.

Multiple tER–Golgi secretory units

Can localization of mRNAs mediate polarised deposition

or secretion of the resulting protein in just one segment of a cell? This is possible in any cell type that has multiple

tER–Golgi units. This type of exocytic pathway organi-

zation was first described in the yeast Pichia, where each

tER site is close to a Golgi mini-stack, thus forming a

tER–Golgi unit [25]. Similar arrangement of the tER and

Golgi is found in Drosophila cells [9] and Drosophila

oocytes, which are very large cells containing 1000

tER–Golgi units [26], providing a unique system to

study localized protein synthesis.

The Drosophila oocyte is highly polarized. In particular,

secretion of the protein Gurken occurs only at the dorsal–

anterior corner [27]. Gurken is synthesized as a trans-

membrane protein. A cargo-specific co-factor called Cor-

nichon then escorts it rapidly from the ER via the tER to

the Golgi apparatus, where Gurken’s bioactive lumenal

domain is cleaved. Polarized secretion is achieved by

localizing gurken mRNA to the dorsal–anterior corner

of the oocyte, but how does this ensure local secretion?

Firstly, local synthesis leads to local transport because

Cornichon ensures rapid ER transit, greatly limiting

diffusion in the ER, which is continuous here as it is inall cells. Secondly, it appears that each tER–Golgi unit

is able to secrete independently: just 10–15 out of

the 1000 units transport Gurken protein, limiting the

distribution of secreted Gurken (Figure 1b) [26].

This pre-translational mechanism is a previouslyunknown way of creating polarised deposition of trans-

membrane and/or secreted proteins, complementary to

post-translational sorting in the trans-Golgi network

(TGN) [28], and may apply to other secreted proteins

in Drosophila [29].

In mammalian cells, the localisation of mRNAs encodingtransmembrane or secreted proteins has not been system-

atically studied. One reason for this is that past work has

focused in particular on model cargo proteins such as

VSVG being secreted by model cells, often fibroblasts (for

example HeLa cells), where the Golgi apparatus is a

single-copy juxtanuclear ribbon in which all the mini-stacks are linked together [30]. In one cell type, however,

Golgi outposts have been described. These are found in

dendrites of rat neurons, which also contain associated

RER and tER sites [31], and these dendrites are able tosustain local synthesis and transport of transmembrane

proteins, such as the AMPA receptor, independently of

the cell body [32,33]. This is likely to be the result of

translation of localized mRNA (although the AMPA

mRNA has not been formally localized) followed by

transport in these Golgi outposts equivalent to what

occurs in tER–Golgi units. Similar localized translation

Endoplasmic reticulum: one continuous network Levine and Rabouille 363

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and transport is suggested to also take place in axons

[34,35,36].

An additional question yet to be addressed is the extent to

which tER sites are equivalent. That they can function

independently from one another is shown by the local

transport of Gurken protein in Drosophila oocytes

(Figure 1b). However, it is not known whether all the

peripheral tER sites of a HeLa cell are active in the

transport of generic transport markers and it is pos-

sible that these sites are in fact used for the synthesis

of localized, as yet unidentified mRNAs.

364 Membranes and organelles

Figure 1

N

P

mRNA

G tER

site

M

PM

ER

N

Exocyst Lipid transfer protein tSNAREs

Translocon VAP vSNARE

PM

ER

(a)

(b) (c)

(d)

ER heterogeneity is created by extrinsic cues. (a) A diagrammatic representation of the interactions between the ER (green) and different extrinsic

cues including localized mRNAs near peripheral tER sites, the plasma membrane, mitochondria (M) and a phagosome (P). Microtubules are

drawn in purple. G, Golgi apparatus. (b) A single stage-9 Drosophila oocyte surrounded by follicle cells. Within the oocyte, Gurken protein (red)

is restricted to the dorsal/anterior corner near the nucleus (N) in dots representing the tER–Golgi units. The ER in all cells is green. (Courtesy of Bram Herpers, Department of Cell Biology, UMC Utrecht, The Netherlands.) (c) Diagram of three protein complexes potentially involved in

ER–plasma membrane contact [44–47]. (d) Electron micrograph of a mitochondrion wrapped in ER showing zones of apposition (arrows), where

ER depleted of ribosomes comes to within 30nm of the mitochondrion. (Courtesy of Ann de Maziere, Department of Cell Biology, UMC Utrecht,

The Netherlands.) Scale bar represents 200nm.


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ER non-uniformities derived from othermembrane-bound organellesIn addition to the well-known subdomains of the ER (the

RER, smooth ER and tER sites), we address here ER

non-uniformities that are induced by other organelles.

These sub-domains of the ER have no well-describedmorphological features apart from the proximity of other

compartments. The ER is intimately associated with

many other organelles, including the TGN [37], endo-

somes [38], chloroplasts [39] and peroxisomes [40], as well

as with the plasma membrane and the mitochondria,

which we will discuss in greater detail below. The mem-

brane contact sites between the ER and these other

compartments form specialized sub-domains of each

organelle that engage in communication of information

and material across narrow cytoplasmic gaps.

ER and plasma membrane

Cortical ER is closely apposed to a portion of plasma

membrane in all cell types. This proximity has been

supposed to underlie the ability of lipids to traf fic bidir-

ectionally between the ER and the plasma membrane

independently of the secretory pathway [41,42]. Only

in yeast has the portion of the ER attached to the plasma

membrane been purified. In comparison to generic ER it

is enriched in lipid synthases, suggesting that it is adapted

to supply lipids [43].

The physical nature of the ER–plasma-membrane link is

still not certain, but three different bridging complexes

have been characterized recently (Figure 1c). First, in

mammalian cells and yeast, the translocon of RER inter-

acts directly with plasma-membrane-bound exocyst(reviewed in [44]). Further evidence for the significance

of this interaction comes from the recruitment of cortical

ER to the new bud in yeast by the exocyst subunit Sec3

[45]. Second, a conserved ER protein called VAP (VAMP-

associated protein, where VAMP is the v-SNARE vesicle-

associated membrane protein) forms bridging complexes

with peripheral membrane proteins on the plasma mem-brane, as well as other sites in the cell [46]. Third, when

phagocytes engulf large bodies, the plasma membrane

must grow rapidly and under these extreme circum-

stances extra membrane is provided by the ER. This

step is mediated by an ER v-SNARE homologous to

Sec22, the only ER v-SNARE that pairs up with plasmamembrane t-SNAREs [47], implicating direct SNARE-

mediated fusion of the ER with the plasma membrane.

Similarly, stimulation of human lymphocytes induces

contact between the exofacial leaflet of the plasma mem-

brane and the lumenal leaflet of the ER, suggesting the

presence of transient continuity between the ER and the

plasma membrane in these cells [48].

In addition to these ER–plasma-membrane bridges,

there is evidence for membrane traf fic between these

two compartments that bypasses the Golgi apparatus.

The most intriguing data comes from yeast, in which

the polytopic plasma membrane protein Ist2 appears to

traf fic from the cortical ER of the bud-tip (where its

mRNA is specifically localized) to the adjacent plasmamembrane [49]. Presuming that Ist2 is translocated into

the ER, this work suggests that traf fic of Ist2 is indepen-dent of the classical exocytic pathway, as it is unaffected

by a variety of mutations in the secretory pathway

[49,50]. The alternative is that Ist2 might undergo

one of the previously described forms of non-classical

protein secretion (i.e. direct insertion into the plasma

membrane after translation in the cytosol), which hitherto

have mainly been restricted to small secreted cytokines

[51]. The suggestion that direct traf fic occurs between

the ER and the plasma membrane is supported by the

finding that other blocks in the yeast secretory pathwayonly moderately affect the plasma membrane deposition

of the bulk of integral plasma membrane proteins [52].

While it is possible that ER–plasma-membrane contact

sustains a novel traf ficking pathway, it is important to

substantiate this by identifying the molecular machinery

involved, for example by determining whether Ist2 traf fic

requires the SNAREs that pair the ER and the plasma

membrane [47].

ER and mitochondria

The ER and mitochondria are intimately associated witheach other. They are physically linked together in many

cell types, and they co-regulate some highly important

functions, including apoptosis and traf fic of lipids and

Ca2+ [53].

Non-vesicular traffic and the physical link between

the ER and the mitochondria

The specific ER compartment that interacts with the

mitochondria is called the mitochondria-associated ER

membranes (MAM). Very little is known about MAM,

except that it is enriched in many lipid synthases com-

pared to generic ER, and is de-enriched in ribosomes [54].The enrichment of lipid synthases is compatible with one

of the clearest functions of MAM, which is to participate

in lipid traf fic to and from mitochondria [55]. Similarly,

ER sites of Ca2+ release channels appear to be preferen-

tially localized near to mitochondria, implying that MAM

may be specialized for calcium traf fic from the ER to themitochondria [56].

Despite the absence of membrane fusion between the

ER and mitochondria, an emerging theme is that someproteins are shared between these organelles, including

some integral membrane proteins. For these membrane

proteins, post-translational insertion into the mitochon-

dria tends to occur if co-translational insertion into the ER

is inef ficient [57–59]. Importantly, any protein shared

between the ER and the mitochondria that can form

head-to-head dimers is a good candidate for a bridging



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component at the zone of apposition (arrows in

Figure 1d). One proposed candidate is the voltage-depen-

dent anion channel (VDAC or ‘porin’) [60], and this may

be an important beginning in mapping the bridging

proteins between MAM and mitochondria, none of which

has been positively identified.

Apoptosis

The ER and mitochondria both control important aspects

of apoptosis, and the pathways involving these two orga-

nelles are highly interdependent. For example, the pro-

apoptotic protein BAP31 resides on the ER and enhances

the pro-apoptotic Ca2+ signal that passes from the ER to

the mitochondria after BAP31 is cleaved by the apoptosis

initiator caspase-8 [61]. Mitochondria are involved in this

cleavage, since caspase 8 is localized on mitochondria andacts in trans to cleave BAP31 on the ER [62]. Thus,

intimate contact between the ER and mitochondria is

important for programmed cell death.

One aspect of this inter-dependence is that the ER and

mitochondria share many apoptotic regulators. This

includes PACS-2, which binds to the proapoptotic

BH3-domain-only protein Bid [63]. In resting cells,

phosphorylated Bid is found both in the cytoplasm and

to some extent on the ER. Following an apoptotic sti-

mulus, Bid is dephosphorylated and then binds to PACS-

2 on mitochondria. Bid is then activated by caspase-8cleavage, forming truncated Bid (tBid), which in turn

induces release of cytochrome c . Interestingly, PACS-2

responds to apoptotic stimuli by translocating from the

ER to mitochondria, and the structure of MAM itself

depends on PACS-2 for its normal integrity [63

],although this may turn out to be a secondary effect of

PACS-2’s role in altering apoptotic pathways. Despite

these exciting findings, we still know very little about how

the portion of ER that is MAM is organized extrinsically

by mitochondria.

ConclusionsThe ER maintains variety in unity. For a long time, the

ER was considered to act in a unified manner, in keeping

with the unitary nature of the rest of the secretory path-

way. However, we now know that localized mRNAs allow

the sub-division of the classical secretory pathway into

separate compartments. In addition, the ER has specificrelationships with many non-ER cellular components,

inducing multiple sub-domains in the ER [64]. For the

sake of brevity, we have only discussed two of these in

detail, the plasma membrane and the mitochondria. Themolecular bridges between the ER and other organelles

are almost entirely undescribed, except in a single case

(the nuclear–vacuole junction in yeast) [65]. In any event,

the idea that extrinsic cues organize the ER leads to a

further question: what provides these cues with their

positional information? The simple answer must be that

the cytoskeleton contains most of the information needed

to organize the different cellular components and their

inter-relationships. In some cases, the molecular mechan-

isms that set up these relationships are now becoming

understood, such as the pathways underpinning the

movement and anchoring of Gurken mRNA [17,18].

However, in most cases of ER sub-compartmentalizationonly a small fraction of the overall mechanism is known at

the molecular level, making this an interesting field to

work in.

AcknowledgementsWe wish to thank Vangelis Kondylis, Judith Klumperman and ChrisLoewen for critically reading this manuscript. We thank Adrian Oprinsfor Figure 1a. Figure 1b and 1d are courtesy of Bram Herpers and Annde Maziere (Utrecht, The Netherlands), respectively. We apologize toour colleagues whose work we have not been able to include because of space constraints.

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This article, together with [34] and [36], exemplifies the interesting andcontroversial possibility that synthesis and transport of transmembraneproteins occurs not only in neuronal cell bodies but also in axons (as indendrites, see [31,32,33] ). Conopressin receptor, for which the mRNA is injected [36], the plasma membrane receptor EphA2, for which themRNA is also injected [34], and Syntaxin [35] are all shown to betranslated and inserted in the plasma membrane independently fromthe cell body.

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37. Ladinsky MS, Mastronarde DN, McIntosh JR, Howell KE,Staehelin LA: Golgi structure in three dimensions: functionalinsights from the normal rat kidney cell . J Cell Biol 1999,144:1135-1149.

38. Haj FG, Verveer PJ, Squire A, Neel BG, Bastiaens PI: Imagingsites of receptor dephosphorylation by PTP1B on thesurface of the endoplasmic reticulum. Science 2002,295:1708-1711.

39. XuC, Fan J,RiekhofW, Froehlich JE, Benning C: A permease-likeproteininvolvedin ER to thylakoid lipid transfer in Arabidopsis.EMBO J 2003, 22:2370-2379.

40. Geuze HJ, Murk JL, Stroobants AK, Griffith JM, Kleijmeer MJ,Koster AJ, Verkleij AJ, Distel B, Tabak HF: Involvement of theendoplasmic reticulum in peroxisome formation. Mol Biol Cell 2003, 14:2900-2907.

41.

Li Y, Prinz WA: ATP-binding cassette (ABC) transportersmediate nonvesicular, raft-modulated sterol movement fromthe plasma membrane to the endoplasmic reticulum . J Biol Chem 2004, 279:45226-45234.



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The traf fic of sterol in yeast is independent of the classical secretorypathway, even though the sterol studied here is cholesterol rather thanendogenous ergosterol. Also, as found in the study by Baumann et al.[42], the propensity for sterol to escape the ER is correlated with itsability to form complexes.

42.

Baumann N, Sullivan D, Ohvo-Rekila ¨ H, Simonot C, Pottekat A,Klaassen Z, Beh C, Menon A: Transport of newly synthesized

sterol to the sterol-enriched plasma membrane occurs vianon-vesicular equilibration. Biochemistry 2005, in press.The kinetics of sterol traf fic to the plasma membrane in yeast are notaffected by blocking all SNARE-mediated traf fic, suggesting that steroltraf fics by a non-vesicular route. Furthermore, sphingolipidsin the plasmamembrane act as a sink for sterol, as together these lipids form high-af finity complexes, so that the total sterol concentration in the plasmamembrane is far higher in than in the ER, although the concentration of free sterol is the same in both compartments.

43. Pichler H, Gaigg B, Hrastnik C, Achleitner G, Kohlwein SD,Zellnig G, Perktold A, Daum G: A subfraction of the yeastendoplasmic reticulum associates withthe plasma membraneand has a high capacity to synthesize lipids . Eur J Biochem2001, 268:2351-2361.

44. Guo W, Novick P: The exocyst meets the translocon: aregulatory circuit for secretion and protein synthesis?Trends Cell Biol 2004, 14:61-63.

45. Wiederkehr A, Du Y, Pypaert M, Ferro-Novick S, Novick P: Sec3pis needed for the spatial regulation of secretion and for theinheritance of the cortical endoplasmic reticulum. Mol Biol Cell 2003, 14:4770-4782.

46. LoewenCJ, Roy A,Levine TP: A conserved ER targeting motif inthree familiesof lipid binding proteins and in Opi1p binds VAP.EMBO J 2003, 22:2025-2035.

47. Becker T, Volchuk A, Rothman JE: Differential use ofendoplasmic reticulum membrane for phagocytosis in J774macrophages. Proc Natl Acad Sci USA 2005, 102:4022-4026.

48. Dadsetan S, Shishkin V, Fomina AF: Intracellular Ca2+ releasetriggers translocation of membrane marker FM1-43 from theextracellular leaflet of the plasma membrane intoendoplasmic reticulum in T lymphocytes. J Biol Chem 2005, inpress.

49.

Juschke C, Ferring D, Jansen RP, Seedorf M: A novel transportpathway for a yeast plasma membrane protein encoded by alocalized mRNA . Curr Biol 2004, 14:406-411.

Transport of the integral membrane protein Ist2 to the plasma membraneis unaffected by various sec mutations inactivating the classical secretorypathway at different points, and appears to rely instead on direct trans-port from the ER to the plasma membrane. In addition, Ist2 mRNA is oneof very few mRNAs localized to the bud-tip that code for a membraneprotein, although the relationship between this and the means of exo-cytosis has not been examined.

50. Juschke C, Wachter A, Schwappach B, Seedorf M: SEC18/NSF-independent, protein-sorting pathway from the yeast corticalER to the plasma membrane. J Cell Biol 2005, in press.

51.

Nickel W: The mystery of nonclassical protein secretion: acurrent view on cargo proteins and potential export routes.Eur J Biochem 2003, 270:2109-2119.

This review describes protein insertion in the plasma membrane that isindependent of the ER. This differs from the novel membrane traf fickingpathway suggested for Ist2 from the ER to the plasma membrane.

52. Schnabl M, Daum G, Pichler H: Multiple lipid transport pathwaysto the plasma membrane in yeast. Biochim Biophys Acta 2005,1687:130-140.

53. Levine T: Short-range intracellular trafficking of smallmolecules across endoplasmic reticulum junctions.Trends Cell Biol 2004, 14:483-490.

54. Wang HJ, Guay G, Pogan L, Sauve R, Nabi IR: Calcium regulatesthe association between mitochondria and a smoothsubdomain of the endoplasmic reticulum. J Cell Biol 2000,150:1489-1498.

55. Vance JE: Phospholipid synthesis in a membrane fractionassociated with mitochondria. J Biol Chem 1990,265:7248-7256.

56.

Filippin L, Magalhaes PJ, Di Benedetto G, Colella M, Pozzan T:Stable interactions between mitochondria and endoplasmicreticulum allow rapid accumulation of calcium in asubpopulation of mitochondria. J Biol Chem 2003,278:39224-39234.

Ithasbeenknownforsometime thatCa2+ released from theER canreachhigh local concentrations at mitochondrial Ca2+ uptake pumps, ratherthan being released into the cytoplasm at large. Here, it is shown thatindividual Ca2+ release channels remain physically linked to a specificmitochondrion over reasonably long periods of time, indicating that ER-to-mitochondrion Ca2+ traf fic occurs at stable structures similar to MAM.

57. Colombo S, Longhi R, Alcaro S, Ortuso F, Sprocati T, Flora A,Borgese N: N-myristoylation determines dual targeting ofmammalian NADH-Cytochrome b(5) reductase to ER andmitochondrial outer membranes by a mechanism of kineticpartitioning. J Cell Biol 2005, 168:735-745.

58. Miyazaki E, Kida Y, MiharaK, SakaguchiM: Switching the sorting

mode of membrane proteins from co-translational ERtargeting to post-translational mitochondrial import. Mol Biol Cell 2005, in press.

59. van Herpen RE, Oude Ophuis RJ, Wijers M, Bennink MB,van de Loo FA, Fransen J, Wieringa B, Wansink DG: Divergentmitochondrial and endoplasmic reticulum association ofDMPK splice isoforms depends on unique sequencearrangements in tail anchors. Mol Cell Biol 2005,25:1402-1414.

60. Shoshan-Barmatz V, Zalk R, Gincel D, Vardi N: Subcellularlocalization of VDAC in mitochondria and ER in thecerebellum. Biochim Biophys Acta 2004, 1657:105-114.

61. Breckenridge DG, Stojanovic M, Marcellus RC, Shore GC:Caspase cleavage product of BAP31 induces mitochondrialfission through endoplasmic reticulum calcium signals,enhancing cytochrome c release to the cytosol. J Cell Biol 2003, 160:1115-1127.

62. Chandra D, Choy G, Deng X, Bhatia B, Daniel P, Tang DG: Association of active caspase 8 with the mitochondrialmembrane duringapoptosis: potential roles in cleaving BAP31and caspase 3 and mediating mitochondrion–endoplasmic-reticulum cross talk in etoposide-induced cell death. Mol Cell Biol 2004, 24:6592-6607.

63.

Simmen T, Aslan JE, Blagoveshchenskaya AD, Thomas L,Wan L, Xiang Y, Feliciangeli SF, Hung CH, Crump CM, Thomas G:PACS-2 controls endoplasmic reticulum-mitochondriacommunication and Bid-mediated apoptosis. EMBO J 2005,24:717-729.

PACS-2, like PACS-1, is shown to be a cargo sorting protein, but unlikePACS-1, which targets cargo within the late secretory pathway, PACS-2targets Bid to mitochondria. Furthermore, PACS-2 is among the firstproteins identified that are essential for normal architecture of ER–mitochondrial-membrane contact sites.

64. StaehelinLA: The plantER: a dynamic organellecomposed of alarge number of discrete functional domains. Plant J 1997,11:1151-1165.

65. Pan X,RobertsP, ChenY, KvamE, ShulgaN, Huang K,Lemmon S,Goldfarb DS: Nucleus-vacuole junctions in Saccharomycescerevisiae are formed through the direct interaction of Vac8pwith Nvj1p. Mol Biol Cell 2000, 11:2445-2457.



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