The Plant ER–Golgi Interface

# 2008 The Authors

Journal compilation # 2008 Blackwell Munksgaard

doi: 10.1111/j.1600-0854.2008.00773.xTraffic 2008; 9: 1571–1580Blackwell Munksgaard

Review

The Plant ER–Golgi Interface

Chris Hawes*, Anne Osterrieder, Eric Hummel

and Imogen Sparkes

School of Life Sciences, Oxford Brookes University,Headington, Oxford OX3 0BP, UK*Corresponding author: Chris Hawes,[email protected]

The interface between the endoplasmic reticulum (ER) and

the Golgi apparatus is a critical junction in the secretory

pathway mediating the transport of both soluble and

membrane cargo between the two organelles. Such trans-

port can be bidirectional and is mediated by coated mem-

branes. In this review, we consider the organization and

dynamics of this interface in plant cells, the putative struc-

ture ofwhich has caused somecontroversy in the literature,

andwespeculateon the stagesofGolgi biogenesis fromthe

ER and the role of theGolgi and ERon each other’smotility.

Key words: COPII, endoplasmic reticulum, ER exit sites,

Golgi apparatus, myosins, SNAREs

Received 23 April 2008, revised and accepted for publica-

tion 28 May 2008, uncorrected manuscript published

online 30 May 2008, published online 20 June 2008

The plant Golgi apparatus is characterized by numerous

individual cisternal stacks that appear more or less randomly

distributedthroughoutthecytoplasm.Whilesuchadistribution

isdifferent to that commonlydescribed formammaliancells, it

ismoreakin to thatof insectssuchasDrosophila (1). However,

in many plant cell types, it is apparent that Golgi stacks are

motile and closely associatedwith the endoplasmic reticulum

(ER) (2), while in others stacks can exist isolated from the ER

(3). In this review, we consider the unique nature of the plant

ER–Golgi interface and speculate as to how this is organized

in a system that can be continually motile.

Organization of the Plant ER–Golgi Interface

Anterograde protein transport from the ER to the Golgi

takes place at specialized ER exit sites (ERES) and is

mediated by the Sar1p guanosine triphosphatase (GTPase)

and its exchange factor Sec12 plus the coat protein (COP)II

coat comprising the heterodimeric Sec23/Sec24 and

Sec13/31 complexes (4,5). Homologues of those proteins

have been identified in plants (6–12), and the COPII

machinery appears to be conserved in plants as over-

expression of Sec12 and expression of Sar1 mutants

disrupt anterograde protein transport (13–15).

What are the COPII Carriers?

COPI vesicles have been identified (12,16), yet the nature

of COPII-coated carriers remains elusive. Although COPII

proteins have been located (11,17–19) (Figure 1A–C and

Table 1), to date, there is little hard evidence for the

existence of COPII vesicles in plants. Such vesicles have

been described in meristematic cells in high-pressure-

frozen freeze-substituted material and in unicellular algae

(20), but the specificity of the Sar1 antibody used to

identify COPII components was not demonstrated. Bud-

ding profiles on the ER have been reported in BY2 cells,

but immunolabelling failed to identify COPII components

(21).

The possibility of membrane connections between the ER

and the Golgi, be they permanent or transitory, tubular or

direct, has been debated in several recent reviews and has

been described in many electron micrographs (3,22–24).

It does however need to be emphasized that the canonical

view that all transport between ER and Golgi must be

mediated by COPII vesicles is being challenged (reviewed

in 4). Indeed, much of the early evidence of such vesicles

came from in vitro reconstituted yeast systems (25), which

may not reflect the in vivo state. In mammalian cells, it has

now been suggested that the vesicular tubular carriers

between the ER and the Golgi may be generated by the

fusion of COPII vesicles or from tubules or specialized

domains at the ERES (26). Such ER-to-Golgi carriers pro-

duced directly from the ER may be COPII dependent in

formation but may not involve COPII vesicles (27). Thus,

there is no a priori reason to assume that the COPII

vesicles must be the carriers across what must be a very

small divide in the plant ER–Golgi interface in many tissues

such as leaves and hypocotyls. However, the question

then needs to be answered as to what is the situation in

cells where the Golgi bodies appear separate from the ER,

such as the isodiametric less vacuolated cells in meris-

tems? Is there a long-range transport of carriers from ER to

Golgi or do the Golgi bodies dock onto ERES, retrieve

cargo and then separate from the ER (28)?

Proteins Acting at the Plant ER–Golgi Interface

Nomatter what the physical nature of the ER-to-Golgi vector

might be, transport between the two organelles requires, at

some stage, fusion between the membranes of donor and

acceptor compartments. A number of ER and Golgi SNARE

proteins have been described for Arabidopsis (29), and four

of these, Sec 22, Memb11, Bet 11 and Sed5 (Syp 31), have

been suggested to play a role at the ER–Golgi interface (30).

Thewhole transport processbetween the twoorganellesalso

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Hawes et al.

appears to be regulated by the plant homologue of the Rab1

GTPase (RabD2A) (31).TheArabidopsis homologues of these

regulatory proteins are summarized in Table 1 together

with other key proteins that are implicated to act at the plant

ER–Golgi interface. The putative functions and importance

of COPII isoforms and coat-related proteins, Golgi matrix

proteins and movement proteins will be discussed in more

detail in the following sections of this review.

What is the Function of COPII Isoforms?

Plants possess genes encoding for multiple isoforms of

COPII proteins (23,24,32,33), and analysis of their expression

Table 1: Proteins of the ER–Golgi interface and their putative functionsa

Protein Accession

number

Reference Function or putative

function

Coat and related proteins

Sec12 At2g01470 (6) Sar1p GTP exchange factor on ER

membrane

AtSARA1a At1g09180 (18,33) Initiation of COPII coat assembly

(Sar1 isoforms)AtSARA1b At1g56330 (18,33)

Sar1BTNt (leaf) AF210431 (13)

NtSAR1 (BY-2) BAA13463 (14)

Sec23 At3g23660 (9,12) Sar1p GTPase-activating protein on

COPII coat

Sec24 At3g07100 (12) Cargo binding protein on COPII coat

Sec13 At2g30050,

At3g01340

(11,24) COPII coat protein

Sec31 At1g18830,

At3g63460

(11,24)

Sec16 At5g47480,

At5g47490

Not yet

characterized (24)

Definition of ERES?

Regulatory proteins and fusion proteins

RabD2A At1g02130 (31) Regulation of ER–Golgi transport

Sec22 At1g11890 (30) ER SNARE

Memb11 At2g36900 (30) Golgi SNARE

Bet11 At3g58170 (30)

Sed5 (Syp 31) At5g05760 (30)

Matrix and movement

Myosin XIK At5g20490 (82–84) Tail domain severely perturbs Golgi

movementMyosin XIE At1g54560 (83)

Myosin Mya2 At5g43900 (82)

AtCASP At3g18480 (46) Putative tethering factor

Golgin-84 2 isoforms (GC1 and GC2) At2g19950,

At1g18190

(47)

AtP115 (GC6) At3g27530 (47) Tethering between ER and Golgi?

TRAPP1 and COG complexes Various (45) Tethering between ER and Golgi.

Organization of transferases?

ERD2 At1g29330

(L23573)

(2,90) Putative H/KDEL receptor

aThe table does not include cis-located transferases, sugar transporters or COPI complex proteins.

Figure 1: Visualization of Golgi bodies, ERES and Golgi stack movement in plant tissues. Plant Golgi bodies and ERES can be

visualized by expression of fluorescent protein fusions, allowing analysis of their location and tracking of their movement using confocal

laser scanning microscopy. In tobacco leaf epidermal cells, the Golgi marker ST-cyan fluorescent protein (CFP) (A) and the COPII coat

protein and putative ERES marker yellow fluorescent protein (YFP)–Sec24 (B) behave as mobile secretory units. YFP–Sec24 labels the

cytosol and punctate structures that colocate with ST-CFP (C). Tethering factors could be involved in maintaining the close relationship

between the ER and the Golgi bodies as well as in keeping the cisternae together during stack movement. GFP-AtCASP (D) coexpressed

with the Golgi marker ST monomeric red fluorescent protein (E) in tobacco leaf epidermal cells locates to ring-like structures around the

Golgi bodies (F). A slight shift between the signals reflects their different distribution within the Golgi stack, with GFP-AtCASP being

located towards the cis-Golgi. Fluorescent Golgi body markers were used to track movement over time. Their motility is depicted in

Arabidopsis root meristems (G), Arabidopsis root elongate cells (H) and tobacco epidermal cells (I) as a series of sequential images over

a period of 10 seconds. Each image is false coloured green, red, blue or magenta in sequence. White therefore indicates that Golgi bodies

have not moved (because of colocation of several false-coloured images over time) or as a trail of slightly overlapping colours (see arrow in

I). Movement is more apparent in elongated root and leaf epidermal cells. Coexpression of the tail domain of myosin XIK (magenta, J) with

a Golgi marker (green, K, merged) in tobacco epidermal cells perturbs Golgi movement as indicated in the merged sequential images

(L, compare with control I). (A–C) bars ¼ 5 mm and (D–L) bars ¼ 2 mm.

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Plant ER–Golgi Interface

profiles showed tissue specificity for some of them, such

as the Arabidopsis Sar1 isoform At1g09180 that appears to

be expressed exclusively in stamen and pollen (24). COPII

isoforms can have different intracellular locations and

might also differ in their function, as the Arabidopsis Sar1

isoform AtSARA1a was found to be more cytosolic than

AtSARA1b, and AtSARA1b affected ER export less when

both isoforms were expressed in a GTP-locked form (18).

Little is known about the function of other COPII isoforms,

but it has been described recently that four isoforms of

human Sec24 exhibited preferential binding to different

cargo transport motifs, which could increase the complex-

ity of cargo recognition (34).

As Sec12 and most Sar1 isoforms locate to the ER

membrane, it has been suggested that the whole ER

surface might be competent for ER export (24), as originally

postulated by Boevink et al. (2). However, a combination of

photobleaching studies on moving Golgi (19) and the

colocalization of NtSar1Bt (19) and of Sec24 (12,17) with

Golgi markers on the ER supports the motile export site

complex hypothesis that involves the Golgi stacks plus

ERES moving in synchrony on or with the ER surface

(19,22), a situation very different to the more static exit site

reported in mammalian cells (35). Tissue specificity of Sar1

isoformsmight explain the contradictory evidence regarding

the relationship of Golgi bodies to ERES as previous studies

were undertaken with different Sar1 isoforms (24). The

tobacco leaf isoform NtSar1Bt (13) was observed in the

cytosol and in punctate structures colocating with the Golgi

marker ERD2-green fluorescent protein (GFP) (19), whereas

the NtSar1 isoform isolated from BY-2 cells (8) labelled the

ER and punctate structures that only partially colocated with

the Golgi marker ManI–red fluorescent protein (24). Per-

haps, the discrepancy in location of the Sar1 isoforms

reflects differences in the relationship betweenGolgi bodies

and ER in leaf and root tissue, as Golgi bodies in tobacco leaf

epidermal cells moved with the surface of the ER (36),

whereas Golgi stacks in root cells seem to be able to

dissociate from the ER (3).

Differentiation of ER Exit Sites

The exact processes leading to formation and differenti-

ation of ERES are still unknown. The first step of COPII coat

assembly is the recruitment of Sar1 by Sec12 to the ER (5).

In Pichia pastoris, however, COPII coat formation re-

mained restricted to a specific ER domain termed the

transitional ER (tER) even when Sec12 was dispersed over

the ER membrane (37). Therefore, it is likely that additional

proteins are required to establish the identity of ERES,

maybe by forming an ER membrane scaffold structure

(35). It has been speculated that Sec16 could be part of

such a scaffold (38) as upon expression of GTP-locked Sar1

in animal cells, Sec16 accumulated together with Sec24

and Sec31 not only on juxtanuclear membranes previously

described as clustered ERES (39) but also on additional

peripheral structures on the ER membrane (38). The

authors suggested that those juxtanuclear membrane

structures resembled clustered free COPII carriers and

that Sec16 might constitute a more reliable ERES marker.

Clearly, the choice of the marker protein is critical in

studying the relationship between ERES and Golgi bodies

as in animal cells, it has been suggested that the majority

of total COPII proteins expressed labelled free tubules and

vesicles (40). The Arabidopsis genome encodes two

putative Sec16 isoforms (24), but their function in plants

still needs to be established.

A putative ERES scaffold might also incorporate cis-Golgi

matrix proteins that could play a role both in differentiation

of ERES and in the nucleation and regulation of Golgi stack

formation (41,42). Golgins are large coiled-coil proteins

implicated in the tethering of vesicles or other membrane

compartments and could provide a first level of vesicle

recognition and specificity before SNARE-mediated vesicle

fusion (43). Several golgin homologues have recently been

identified in plants (44–47), and two of them, AtCASP and

golgin-84, have been located to the cis-Golgi (47) (Figure

1D–F). A p115/Uso1p homologue was also described (47),

and this is a matrix protein that has been implicated in

a tethering role at the cis-Golgi (43,48). It has been shown

that p115 tethers COPII vesicles to Golgi membranes (49).

Another task of p115 is to form bridging tethers by linking

giantin (present in recycling COPI vesicles) to GM130

(present on cis-Golgi membranes) (50). Recent results have

shown that p115 forms, together with Rab1, a SNARE

complex (51). These matrix proteins are therefore ideal

candidates for tethering factors at the Golgi/export site

complex to the ER membrane and also for organizing the

export site complex during Golgi biogenesis (42).

Birth of a Golgi

A major step during the cell cycle is the partitioning of

different organelles between daughter cells during the

division process. Organelles with endosymbiotic origins

such as mitochondria and chloroplasts cannot form de novo

and are replicated by division. The Golgi apparatus however,

with a close structural and functional relationship to the ER,

displays different mechanisms of inheritance (52).

Different Models of Golgi Biogenesis

Two ways by which Golgi stacks could multiply are

discussed in the literature: either by de novo formation

from the ER or through fission of an existing stack (53).

In animal cells, mitosis leads to a complete breakdown of

Golgi stacks during prophase and remnant mitotic vesicu-

lar Golgi clusters may be formed (54); however, the

relationship of these small clusters with the ER, and the

importance of the ER in the organization of Golgi remnants

and in the reconstruction of the Golgi, is a hotly debated

topic (55,56). It has been reported that such Golgi clusters

formed in telophase are segregated in pairs between

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daughter cells and fuse just before completion of cyto-

kinesis (57). Alternatively, Golgi stacks may form de novo

either from ERES, as in the yeast P. pastoris (58–61), or

from mitotic vesicular clusters (56,62,63). A model for

Golgi disassembly and reassembly during mitosis in mam-

malian cells has been proposed in which sequential

inactivation of Sar1 and Arf1 leads to disruption of ERES

and redistribution of Golgi enzymes to the ER, whereas

sequential activation of those two proteins initiates Golgi

reformation (55). In Toxoplasma gondii, an intracellular

protozoan parasite, the Golgi apparatus is a single copy

organelle that grows by lateral extension and undergoes

medial fission during cell division (64). Studies on other

protozoan parasites like Trypanosoma brucei have shown

de novo synthesis, suggesting that both models of Golgi

biogenesis can exist in protists (65–67).

Golgi Biogenesis in Plants and Algae

It is well documented in plant cells that Golgi bodies can

reform on washout of Brefeldin A (BFA) from treated

material (68), indicating that the ER has the capacity to

generate Golgi de novo. Hanton et al. (17) have shown,

using Sec24 as a marker, that de novo export site

formation can be cargo induced, indicating that perhaps

Golgi bodies can form in response to cargo production if

export sites and Golgi stacks do behave as a single unit.

During mitosis and cytokinesis in plants, Golgi bodies and

membranes do not disaggregate as in mammalian cells.

Whether secretion per se stops is not known, but from

late anaphase onwards, the Golgi apparatus is highly

active in producing new cell wall membrane and poly-

saccharide for the phragmoplast region (69,70). Data on

Golgi inheritance in higher plant cells are contradictory.

Golgi stacks were reported to double during metaphase

in onion root meristems (71), while duplication was

claimed to occur during cytokinesis in synchronized

cultures of Catharanthus roseus (72). More recently,

a tomographic analysis of Arabidopsis shoot meristem

cells demonstrated a doubling of the number of Golgi

stacks in G2 just prior to mitosis (73). Cells with high

secretory activity such as pollen tubes and root hairs

seem to produce large numbers of new Golgi stacks

depending on their task and growth status, and this is not

related to the cell cycle or division (74).

Recent studies in the single-celled alga Chlamydomonas

noctigama, which has non-motile Golgi stacks around

the nucleus (75), and in BY-2 cells with mobile Golgi (21)

have shown that de novo Golgi biogenesis and Golgi

fission can take place within the same system (Figure 2).

Experiments were based on a complete deconstruction

of Golgi stacks with BFA and reformation after BFA

washout. Initially, in both systems, vesicle clustering

was a first indication of Golgi reformation. After the first

fusion events, mini-Golgi stacks were formed, starting at

200 nm diameter with up to five cisternae (Figure 2A–

C,H,I). An increase in ERES number on the tER accom-

panied the early reformation events in C. noctigama.

Mini-Golgi stacks displayed a very early cis-to-trans polarity,

and in BY-2 cells, this could also be observed in Golgi stacks

with a 250 nm diameter. In both studies, there was no clear

indication that COPII-coated vesicles ormembrane took part

in early stages of biogenesis. Although budding sites on the

ER were observed (Figure 2A), they did not label with

antibodies to the Sec13 component of the COPII coat. From

immunogold labelling, it was however shown that COPI

proteins may play a role in the early membrane fusion

events forming initial cisternae.

After stack formation, Golgi cisternae increase in size. The

growth seems to be related to an increased number of

budding sites on the ER in C. noctigama (75), and there

appeared to be an increased formation of budding profiles

on the ER in BY-2 cells with mobile Golgi stacks. In both

Chlamydomonas and BY-2 cells, reforming Golgi stacks

continued to grow to double the size of those in control

cells and then divided vertically in a cis-to-trans direction

(21,75). There is however no indication as to what triggers

the overgrowth of the stacks or induced their division, but

we have to hypothesize on the existence of molecular

regulators of Golgi stack size. Could this be a putative role

for some of the matrix proteins?

Inmammalian cells, Golgi matrix proteins, mainly GM130 and

p115, have been implicated in Golgi biogenesis (76), and as

discussed earlier, homologues of Golgi matrix proteins have

been described for plant cells (45,47), although a GM130

homologue does not exist. However, the p115 homologue is

most likely situated towards the cis-Golgi and is a good

candidate for a tether involved in early Golgi biogenesis. In

Figure 3, we propose a sequence of events that may be

involved in the birth of an individual Golgi stack from the ER.

First, an exit site differentiates on the ER surface through

interplay of Sec16, Sec12 and Sar1 (Figure 3A). Thismay also

involve cis-Golgi matrix or tethering factors. A COPII-coated

bud forms from the ER membrane and is tethered to the ER

through the proto-Golgi matrix (Figure 3B). The bud or buds

grow to form a tubulovesicular complex, which contains

COPI buds, vesicles and SNARES, and is surrounded by

a matrix (Figure 3C). Whether this is fed by direct membrane

connections to the ER or by vesicles is still to be ascertained

but quickly differentiates into a mini proto-Golgi stack with

structural characteristics of both cis- and trans-faces, includ-

ing clathrin-coated buds (Figure 3D). At some stage, mem-

brane-boundGolgi enzymes are transferred into this structure

from the ER and are anchored in the correct cisternae as the

stack continues to mature. This whole complex is most

likely motile with the ER surface (see subsequently).

Is the ER–Golgi Interface Implicated inMotility?

A large number of plant cells display highly dynamic

organelle movement. However, the requirement for

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organelle motility is not completely understood, although

environmental stresses such as light affecting chloroplast

and nuclear positioning (77,78) and fungal infection affecting

peroxisome location (79) have been implicated. Studies in

tobacco epidermal and BY2 cells using GFP technology have

shown that numerous individual Golgi bodies display a range

of motilities from remaining stationary, slow to fast, plus uni-

and bidirectional movements (2,80). These movements

appear to occur over the ER, in what was coined ‘stacks

on tracks’, where Golgi bodies are ‘stacks’ on the ER/actin

‘tracks’. Obviously, one key question to be asked is what is

the additional contribution of moving Golgi stacks to the

secretory process over static stacks considering that in

undifferentiated meristematic cells, there appears to be less

Golgi movement? A question that is as yet unanswered.

Movement of Golgi and ER

Owing to the intricate nature of ER-to-Golgi trafficking

(antero- and retrograde transport) and the membrane

Figure 2: Golgi biogenesis and fission in tobacco BY-2 cells (A–G) and Chlamydomonas noctigama (H–M) in BFA washout

experiments. An increased number of ER-budding sides (A) and a vesicular cluster (B) are the first steps of Golgi recovery in BY-2 cells

15 min after washing out BFA. These vesicle clusters tend to fuse (C) and form mini-Golgi stacks (D, size around 250 nm) within the first

hour of recovery. Some of these mini-stacks show very early cis (c)–trans (t) polarity. Mini-stacks often appear in groups (E). After maturing

and forming double-sized larger Golgi stacks, the majority divide in a cis-to-trans direction about 180 min after BFA washout (F–G)

arrowheads point to intercisternal filaments. Golgi biogenesis in C. noctigama starts with the formation of vesicular tubular clusters (H) and

continues as described for BY2 cells: formation of mini-Golgi stacks (I), lateral growth (J), double stacks (K), division (L) and the appearance

of normal-sized stacks 3 h after BFA washout (M). D, F and G are taken from Langhans et al. (21). Copyright American Society of Plant

Biologists. (A–B) and (H–J) bars ¼ 100 nm and (C–G) and (K–M) bars ¼ 200 nm. H, K, L and M are taken from Hummel et al. (75).

Copyright German Botanical society.

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Hawes et al.

equilibrium required to maintain Golgi homeostasis, both

compartments are functionally and possibly structurally

linked (see previously). Interesting questions raised from

these observations relate to whether the Golgi body is

then simply a subdomain of the ER (3) and not an organelle

in its own right, and whether the movement of the two

organelles is also intimately linked or co-ordinated.

Comparisons between ER and Golgi body movements have

resulted in the following observations: Golgi bodies are

associated with the three-way junctions and move along

the tubular ER network (2); occasionally, Golgi bodies break

free from the ER ‘track’, and in some cases, the ER tubules

remodel and follow the path of the Golgi body (81). Photo-

activation studies of ER membrane protein shows that the

Golgi bodies move in a similar direction to the underlying

activated ER membrane (36). These observations have

resulted in the development of a model where ER–Golgi exit

site and Golgi movement is a co-ordinated process and

resulted in questions relating to potentially shared or distinct

motors, whereby the ER drags the Golgi bodies or vice versa

through the action of a motor protein either on the ER and/or

on the Golgi bodies.

Myosin-Driven Golgi?

Cytoskeletal depolymerization experiments have indicated

that both ER and Golgi body movements are in higher plants

dependent on actin and not on microtubules (2,80,81).

Comparative studies of ER and Golgi movements in cells

expressing a fluorescent marker for the actin network

confirm a close association between the organelles and the

actin (81). These observations indicate that movement is

driven either by myosins or by actin polymerization/depoly-

merization. Only recently, it was shown that expression of

truncated variants of 3 of the 17 postulated Arabidopsis

myosins (XIK, XIE and MYA2) severely perturbs Golgi,

peroxisome and mitochondrial movement (82–84) (Figure

1I–L) in tobacco epidermal cells. XIK also plays a role in ER

movement and remodelling (Sparkes et al. unpublished data),

and a further myosin tail fragment partially labelled Golgi

bodies (85). The effects of XIK on organelle movement were

further corroborated through Arabidopsis T-DNA insertional

mutant and RNA interference knockdown studies, although it

was not discussed how the phenotype from overexpression

of a tail domain having a dominant-negative effect could

be the same as that from knockdown of the same protein

(84). Interestingly, although XIK and XIE affect movement

of several organelles, they do not appear to be completely

colocated with these organelles and based on fluorescent

markers do not appear to affect the global architecture of

the actin cytoskeleton (83). It therefore remains to be seen

how these motors control movement. While it appears

that microtubules are not involved per se in Golgi body

movement, other cytoskeletal interacting proteins such as

kinesin 13A (86) and an actin-binding protein KATAMARI 1/

MURUS3 (87) have been identified, which could be potential

components of a complex required for an actin–microtubule

linkage. Alternatively, kinesin 13A may be required for main-

taining Golgi stack integrity or division as several studies in

mammals have highlighted interplay between myosin and

kinesins in Golgi motility and maintenance (88).

Another interesting question pertaining to Golgi stack

movement is how the ER–Golgi linkage and the cisternae

themselves are held together during rapid movement?

Considerable shear forces must be exerted on the Golgi

stack during the vectorial movement, which besides

simple membrane connections might require tethering

factors to hold the system together. Obvious candidates

are Golgi matrix proteins as discussed above (47).

While the majority of motility and ER–Golgi interface

studies have been carried out in tobacco epidermal and

BY-2 cells, it is important to note that there are tissue and

cell type differences; actively dividing cells in the root

meristem display slower movement (Figure 1G) compared

with elongating cells (Figure 1H), which could be because

of cell volume restrictions, cell volume/surface area ratio

Figure 3: Proposed model for the

early stages of the biogenesis of

a Golgi stack from the ER. A) Differ-

entiation of exit sites on the ER. B)

Formation of a tethered COPII bud at

the exit site. C) Formation of a tubulove-

sicular complex with associated COPI.

D) Differentiation of a small proto-Golgi

stack. It is not known if mysosins are

associated with the ER or Golgi.

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requiring slower movement or altered metabolic demand

in these cells or a different more remote relationship with

the ER (see previously). Quantitative analysis of Golgi body

movement in roots versus leaf epidermal cells in Arabi-

dopsis also indicated reduced motility in roots compared

with leaves (84). To compare potential effectors on move-

ment, we have analysed microarray data to try and

determine whether movement rates correspond to altered

regulation of components involved in the ER–Golgi inter-

face in specific tissue types. Analysis indicates that

expression of the genes detailed in Table 1 (potentially)

implicated in the ER–Golgi interface is fairly uniform in all

the tissues assessed. There are a few exceptions: TMF

and a Sec16 isoform are upregulated in pollen, Sed5

expression across all tissues is higher than other SNAREs

(Table 1) and myosin XIE is upregulated in stamens.

Whether these differences reflect functional attributes in

Golgi motility and the requirement to tether or hold the

cisternae together, for example, or have any bearing on the

level of protein present in these tissues remains to be

answered. It is important to note however that different

cell types have different cell volume/cytoplasm ratios

owing to the volume occupied by the vacuole. Therefore,

a cell with a low ratio may require protein upregulation not

necessarily in order for increased function but to maintain

the protein/cytoplasm ratio for interaction with binding

partners within the cytoplasm.

Conclusions

In 1996, we posed ‘stacks of questions’ on the working of

the plant Golgi apparatus (89). Twelve years later, with the

completion of genome sequences and the application of

fluorescent proteins to live cell imaging, we are starting to

answer some of these questions. For plants, it is becoming

clear that although at the molecular level they express many

of the proteins described at the yeast and mammalian

ERES, the structural organization and dynamics of this

interface may be very different. Plant Golgi stacks appear

to have the ability to form ‘de novo’ at exit sites, which in

many tissues are closely apposed to the stacks, and this

may be in response to demands imposed by cargo and/or

growth conditions, although they can also divide by fission.

The exit site/Golgi complex is highly motile on the ER

surface and may move with the ER membrane on an actin

scaffold somehow driven by myosin motors. We still do not

understand why this movement is necessary, but one could

envisage a moving Golgi stack more readily shedding its

secretory vesicles than a static stack. Perhaps, in a further

12 years, another stack of questions will be answered.

Acknowledgments

We thank Benoit Binctin for movies of Arabidopsis root tissue. Some of the

work described in this study was supported by Biotechnology and Biological

Sciences Research Council (BBSRC) and Leverhulme Trust grants to C. H.

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