Directory UMM :Data Elmu:jurnal:P:PlantScience:Plant Science_BioMedNet:201-220:

463

Protein transport within the plant cell endomembrane system: an
update
Ken Matsuoka* and Sebastian Y Bednarek†
The secretory pathway plays a central role in plant
development and morphogenesis. Storage protein deposition,
plant cell division and the expansion of the plasma membrane
and extracellular matrix all require the synthesis and trafficking
of membranes, proteins and polysaccharides through this
network of organelles. Increasing evidence demonstrates that
the plant secretory pathway is more complex than previously
appreciated and that its formation and maintenance are
guided/regulated by many different mechanisms.

In this review, we will highlight advances from the past
year in our understanding of the complex nature of the
plant secretory pathway and of how membrane and proteins necessary for the formation and maintenance of the
endomembrane system are localized. For additional discussions of the various topics touched upon here the reader
is referred to several other recent reviews [1–4].

Addresses
*Laboratory of Biochemistry, Graduate School of Bio-agricultural
Sciences, Nagoya University, Chikusa-ku, Nagoya 464-8601, Japan;
e-mail: matsuoka@nuagr1.agr.nagoya-u.ac.jp
†Department of Biochemistry, University of Wisconsin-Madison, 420
Henry Mall, Madison, WI 53706, USA;
e-mail: bednarek@biochem.wisc.edu

In cereals, two major classes of storage proteins accumulate
in the cells of maturing seeds; prolamins are retained within protein bodies (PBs) that are delimited by the ER
membrane, whereas globulins are transported through the
Golgi apparatus and deposited in vacuoles (see Figure 1).
The different fates of these storage proteins may be linked
to the differential distribution of mRNAs encoding these
proteins and their translocation into distinct subdomains of
the ER [2]. Interestingly, prolamin mRNA charged
polysomes are not directly anchored to the PB membrane,
rather they are found to be associated with the cytoskeleton [5•]. Similarly, elongation factor-1, which is required for
synthesis of the protein, binds to the cytoskeleton surrounding PBs in maize seeds [6]. These observations
suggest that the cytoskeleton plays a direct role in the segregation and localization of PB-ER-specific mRNAs from

messages that are translated on other ER domains.

Current Opinion in Plant Biology 1998, 1:463–469
http://biomednet.com/elecref/1369526600100463
© Current Biology Ltd ISSN 1369-5266
Abbreviations
BiP
endoplasmic reticulum binding protein
CCV
clathrin coated vesicle
CTPP
carboxy-terminal propeptide
ER
endoplasmic reticulum
PACV
precursor-accumulating vesicle
PB
protein body
PSV
protein-storage vacuole

ssVTS sequence-specific vacuolar targeting signal

Introduction
The basic secretory pathway is comprised of several discrete organelles, including the endoplasmic reticulum
(ER) and the Golgi apparatus, that are involved in the
assembly, post-translational modification, trafficking and
correct localization of newly synthesized proteins to the
plasma membrane and vacuole. Trafficking between the
various subcompartments of these organelles is thought to
be primarily mediated by small carrier vesicles. In the past
10 years or so many of the basic molecular components
involved in the formation and consumption of these vesicles have been identified in yeast and mammalian cells
and related components have been identified in plants.
This conservation is not surprising given that the basic
organization of the plant secretory system is morphologically similar to that of other eukaryotes. There are a
number of features, however, that distinguish the plant
pathway. For example, the de novo assembly of the cellplate during plant cell division has no counterpart in other
systems. One cannot simply infer how a plant process
functions, therefore, from studies in other systems.

Sorting of storage protein mRNA to different
ER subdomains in cereal seeds

Storage protein deposition in the ER
The four structurally distinct prolamin-type storage proteins of maize, the zeins (α, β, δ, γ), are deposited and
arranged in an orderly fashion within ER-derived PBs.
Inter- and intramolecular interactions between these subunits are necessary for proper PB formation. Indeed,
coexpression studies have recently shown that γ-zein and
β-zein are necessary for the efficient deposition of α-zein
and δ-zein in ER-PBs, respectively [7,8•]. These interactions, however, must be properly organized within the ER
as demonstrated when a gene encoding a mutant α-zein
that is defective in signal sequence cleavage was
expressed, causing the aberrant PB morphology associated
with the floury2 phenotype [9,10].
Deposition of wheat storage protein aggregates is also
dependent on a retention/aggregation mechanism between
different storage protein subunits in the ER. Expression in
tobacco of only a single gliadin subtype, γ-gliadin, resulted
in its degradation in a post-ER compartment [11]. In contrast to the maize and rice prolamins, however, the dense
gliadin/glutenin deposits are transported via a novel Golgiindependent mechanism from the ER to the proteinstorage vacuole (PSV) [12]. Similarly, the globulin storage

proteins in developing pumpkin cotyledons initially

464

Cell biology

Figure 1

ER

Golgi
PACV
PVC

DV

Vacuole

LV
(a) Vegetative tissues

PSV

(b) Pea cotyledon

PB-ER

(c) Pumpkin cotyledon

PB-ER

ER
Gluterin
Golgi

Prolamin

Vacuole

(d) Rice endosperm

Protein aggregrates

(e) Maize endosperm
Soluble protein

(f) Wheat endosperm

Ribosomes
Current Opinion in Plant Biology

Schematic representation of the different routes involved in the
intracellular trafficking of vacuolar and storage proteins and some of
their unique features that have been observed in various plant cells. (a)
Golgi-mediated transport of soluble proteins to the central vacuole in
vegetative tissues and suspension-cultured cells. It is unknown if this
step is mediated by one or more than one distinct vesicle class. The
recently characterized prevacuolar compartment(s) (PVC) may exist in
other plant cell types as well. (b) Transport of storage proteins in
developing pea cotyledon. Storage protein aggregates form at the
trans-side of the Golgi and are transported via dense vesicles (DV) to

protein-storage vacuole (PSV). Hydrolases are transported from the
Golgi apparatus to the lytic vacuole (LV), possibly by clathrin-coated
vesicles. (c) Transport of proteins to the vacuoles in developing
pumpkin cotyledons. The storage proteins that aggregate in the ER are
packaged and exported from the ER to the PSV in precursoraccumulating vesicles (PACV). Precursor processing proteases are

transported through the Golgi apparatus and maybe directed to the
PACV, which is consistent with the appearance of complex glycans in
these vesicles [13•]. (d) Storage protein sorting in developing rice
endosperm. Prolamins accumulate in the ER and remain in ERmembrane delimited protein bodies (PB-ER), whereas gluterin is
transported to the vacuole via the Golgi apparatus. Gluterin mRNA is
localized to the rER, whereas prolamin mRNA is segregated to the
protein body forming region of the ER (PB-ER). (e) Deposition of
storage proteins in developing maize endosperm. Zeins are deposited
in ER-PB. (f) Storage protein transport in developing wheat
endosperm. The majority of wheat endosperm storage proteins are
initially deposited into the ER. These deposits are then directly
transported to the vacuole, and possibly taken up by an autophagiclike mechanism. A minor portion of the soluble storage proteins in
these cells is transported to the vacuole via the Golgi apparatus.

aggregate in the ER and are subsequently transported via
electron-dense precursor-accumulating vesicles (PACV) to
PSV [13•]. Little is known about how these protein aggregates are distinguished from other secretory proteins and
packaged into ER-to-PSV vesicles. The formation of ERto-Golgi transport vesicles in yeast and mammalian cells is

driven by the COPII (coat protein II) vesicle coat protein
complex [14]. Arabidopsis homologs of COPII components
have been identified and localized to the ER, indicating
that COPII also mediates ER-to-Golgi transport in plant
cells [15]. The mechanisms involved in PACV and COPIIvesicle formation are likely to differ because PACVs are

Protein transport within the plant cell edomembrane system Matsuoka and Bednarek

significantly larger (200–400 nm in diameter) than ER-toGolgi vesicles (60–80 nm) and contain significant amounts
of the ER resident molecular chaperone, binding protein
(BiP) [13•].

ER chaperones and protein folding
BiP, a member of the Hsp70 family, has been strongly
implicated in the assembly of storage protein deposits in

the ER [8•,13•,16,17]. Analysis of the assembly of phaseolin, the homotrimeric vacuolar storage glycoprotein from
common bean, has also demonstrated that BiP associates
with the monomeric protein before it trimerizes [18,19•]
and with misfolded protein subunits in the ER [20•]. In
addition to BiP, the folding and oligomerization of newly
synthesized membrane and soluble secretory protein
requires the coordinate action of the ER-resident molecular chaperones, protein disulfide isomerase (PDI),
calreticulin, and the type-I membrane protein, calnexin.
Mutations that disrupt proper disulfide bond formation
have been shown to affect the synthesis of 11S globulins, a
class of seed storage proteins [21•] and the deposition of
gliadin monomers into PB [22,23]. This process is sensitive
to the redox state of the ER [21•] and is likely to be facilitated by PDI. In tobacco, a large fraction of BiP is
associated with calreticulin [24•]. This interaction maybe
important for the regulation of the level of free BiP in the
ER and/or for the ER-retention of BiP. Calnexin and BiP
have also recently been shown to interact with the vacuolar type-proton pump (V-ATPase) from oat seedlings [25•].
Binding of these molecular chaperones to the V-ATPase
may facilitate the assembly of this large multimeric complex. Another possible function for these two chaperones
could be to inhibit the activity of the V-ATPase in the ER

by gating the opening of the proton-conducting channel.
Indeed BiP has been shown to bind in vitro to the lumenal
side of the Sec61p protein-translocation channel (translocon), that transports nascent polypeptides across the ER
membrane, thereby preventing the free movement of
small molecules out of the ER [26]. Thus, it will be interesting to determine if the proton pump–chaperone
complexes are active.
What is the fate of misfolded secretory proteins in the ER?
Recent studies in yeast and mammalian cells have indicated that the ER has a protein-degradation pathway that can
recognize and selectively remove polypeptides from the
secretory pathway. Misfolded proteins are retrotranslocated through the Sec61p-translocon from the ER to the
cytosol and degraded by the ubiquitin/proteasome system
[27]. The existence of this pathway in plant cells can be
inferred from a recent experiment showing that ERtargeted ricin A-chain, which hydrolyzes a phosphodiester
band of rRNA and destroys ribosome activity, can inactivate protein synthesis in the cytosol [28]. It remains to be
determined, however, whether this pathway is part of the
general ‘quality control’ mechanism in plants for removing
unfolded or misfolded secretory proteins such as assemblydefective phaseolin which was found to be slowly

465

degraded in a brefeldin A (BFA)-insensitive (i.e. Golgiindependent) manner [20•]. Alternatively, some misfolded
proteins may be exported from the ER and directly targeted to the vacuole for degradation through the
BFA-insensitive direct pathway(s) used to deliver pumpkin globulins and wheat gliadins to the PSV.

Golgi-mediated sorting to the different types
of plant vacuoles
Soluble vacuolar proteins that are not directly targeted
from the ER to the vacuole are sorted away from secreted
proteins within the trans-Golgi apparatus. Several independent vacuole sorting mechanisms have been identified
in plants including sequence-specific vacuolar targeting
signal (ssVTS) mediated targeting, carboxy-terminal
propeptide signal (CTPP)-mediated transport, and aggregation-mediated targeting [29]. Interestingly, the ER
retention signal HDEL has recently also been shown to
function as a vacuolar targeting signal [30•] and some peptide sorting sequences may behave as both ssVTS and
CTPP [31]. Cargo proteins containing these different signals are not sorted to the vacuole by the same machinery.
First, wortmannin blocks the sorting of CTPP-containing
proteins but does not affect ssVTS-mediated sorting [32].
Second, some plant cells, such as the parenchyma cells of
developing seed cotyledons, root tip cells and barley aleurone cells, contain at least two distinct vacuolar
compartments [33,34,35•]. In barely root tip cells, a CTPPcontaining protein, barley lectin, and a ssVTS-containing
protease, aleurain, are localized in separate functionally
distinct vacuoles [33]. In addition, storage proteins that
accumulate in PSV of developing pea cotyledons are not
packaged into trans-Golgi derived clathrin coated vesicles
(CCVs), rather they appear to be transported in smooth
dense vesicles (DVs) that are distinct from the ER-derived
PACVs of pumpkin cotyledons [36].
Recognition and packaging of proteins containing vacuolar
sorting signals into Golgi-derived transport vesicles is likely to be mediated by their interaction with specific
transmembrane cargo receptors. The saturable nature of
phaseolin targeting to the vacuole in transgenic tobacco
cells supports this idea [37•]. A putative plant vacuolar
sorting receptor, BP-80, that binds specifically to ssVTS
in vitro, was isolated from CCVs [38,39] and cloned [40•].
Related proteins have also been detected in Arabidopsis
(AtELP, [41]) and pumpkin (PV72/82, [42••]). These
~80 kDa type I transmembrane proteins share common
structural features with the mammalian epidermal growth
factor receptor and the yeast vacuolar sorting receptor,
Vps10p. These proteins have a large lumenal domain containing cysteine-rich repeats thought to be involved in
ligand binding, a single transmembrane spanning domain
and a short cytosolic tail. Within the cytosolic tail are a Tyrcontaining binding motif [43] and a di-acidic motif (DXE)
that functions as an export signal from the ER [44].
Consistent with their putative roles as cargo receptors, BP80 and AtELP have been localized to the trans-Golgi,

Cell biology

466

Figure 2
(a)

CCV
NSF/SNAP
complex

AtVPS45p
Uso1p

Cytosol

AtVTI1p

AtPEP12p

Rab

Prevacuolar compartment
(b)

Cell-plate membrane

Knolle

Cytosol
Knolle

AtCDC48p
Cell-plate membrane

Current Opinion in Plant Biology

Schematic diagram of the critical components involved in NSFdependent heterotypic or Cdc48p-dependent homotypic membrane
fusion. (a) Targeting and fusion of Golgi-derived clathrin-coated
vesicles (CCV) with the prevacuolar compartment. Heterotypic vesicle
fusion at each step of the secretory pathway is mediated by the pairing
of cognate compartment specific v- and t-SNAREs (e.g. AtVtilp and
AtPep12p, respectively) and is modulated by a number of proteins
including the general trafficking factors, the N-ethylmaleimide (NEM)
sensitive factor (NSF), and soluble NSF-attachment proteins (SNAPs)
[62] and specific members of the Rab/Ypt family of small GTP-binding
proteins [63], the Sec1 protein family (including AtVps45p) [64] and
putative ‘velcro’ factors such as p115/Uso1p [65,66]. Following
fusion, NSF and SNAPs catalytically disassemble the energetically
stable SNARE complex. (b) Cell-plate membrane fusion. SNAREs are
not only involved in the heterotypic fusion of secretory vesicles with
their appropriate acceptor compartment but also function in the
homotypic fusion of like-like membranes such as vacuoles [50] and
ER-membranes [67]. The recent identification and localization of
KNOLLE, a putative cell-plate t-SNARE [59••] and AtCDC48 [68], a
fusion factor closely related to NSF, may lend some insight into the
mechanisms of membrane fusion involved in cell-plate formation.
CDC48p has been shown to be required for the homotypic fusion of
ER membranes in yeast [67] and is localized at the cell-plate in
dividing cells [68]. In contrast to NSF-dependent homotypic and
heterotypic membrane fusion, however, which operate through the
pairing of v- and t-SNAREs, Cdc48p-dependent homotypic fusion is
mediated by direct t-/t-SNARE interaction.

CCVs and post-Golgi membranes (prevacuolar compartment; PVC) [38,39,41,45 ••]. Binding studies have
suggested that the Tyr-containing motif of the cytosolic tail
of AtELP interacts with the Golgi-localized clathrin AP-1

adaptor complex [45••]. AtELP and BP-80 are, therefore,
likely to mediate the transport of cargo from the transGolgi to the prevacuolar compartment via CCVs, although
the natural cargo for these receptors remains to be determined. Interestingly, PV72/82 was isolated from
ER-derived PACVs [13•,42••]. This protein has been
shown to recognize the NLPS amino acid sequence of 2S
albumin in PACVs suggesting that PV72/82 is indeed a
bona fide sorting receptor for 2S albumin [42••]. Many
questions about the role of PV72/82 and the formation of
PACVs remain to be resolved, however, including why a
protein that is packaged into ER-to-PSV vesicles has both
a putative AP-1 binding motif and a di-acidic ER-export
motif in its cytosolic tail.
Protein aggregation within the Golgi apparatus has been
suggested to be another way in which proteins destined for
the vacuole are segregated away from secreted proteins
[36,46]. In developing legume seeds, electron-dense storage protein aggregates initially accumulate in the
trans-Golgi and are present in DVs of about 130 nm in
diameter that are thought bud from the Golgi [36]. As
these DVs are similar in function and morphology to
pumpkin PACVs (although their origin is different), it is
possible that receptor-like molecules also function in the
concentration and sorting of storage proteins into DVs.
Following formation of the DVs these putative receptors
may be released and removed from the vesicles by CCVs
that have been observed to bud from the surface of immature DVs [36,47].

Targeting and fusion of Golgi-derived CCVs to
the prevacuolar compartment
Transport of proteins from the trans-Golgi to the vacuole is
likely to occur through the recently identified PVC which
is analogous to the endosomal compartment in yeast and
mammalian cells. Like other vesicular transport steps in
the secretory pathway, the targeting and fusion of the
Golgi-derived CCVs with the PVC is thought to be mediated through the pairing of compartment specific integral
membrane proteins, known as SNAREs, that reside on the
two fusing membrane species (i.e. vesicle v-SNAREs pair
with target membrane t-SNAREs) [48]. Their interaction
in vitro is modulated by a large host of regulatory factors
(see Figure 2). In yeast, docking and fusion of Golgiderived vesicles with the PVC is directed by the
v-SNARE, Vti1p, and the t-SNARE, Pep12p, as well as
the Sec1p-related protein, Vps45p. Another t-SNARE,
Vam3p, is required for the transport step from the PVC to
the vacuole [49] and for vacuole fusion [50]. Plant equivalents of these factors — AtPEP12p, AtVPS45p, AtVAM3p
and AtVTI1p — have been characterized [51•,52,53•]
(Zheng H et al, Abstract 123, 9th International Conference
on Arabidopsis Research, Madison, 24–29 June 1998).
Their subcellular distribution closely approximates the
location expected for the function of each of these proteins
in vacuolar protein sorting. In particular, AtPEP12p and a
fraction of AtELP were found to colocalize on PVCs

Protein transport within the plant cell edomembrane system Matsuoka and Bednarek

suggesting that this compartment serves to recycle plant
vacuolar cargo receptors back to the Golgi apparatus.
Similarly, BP-80 resides both in the Golgi apparatus and
PVCs in pea root tips [40•].

Exocytosis and cell-plate formation
Plant cell growth and expansion is accomplished by the
delivery and fusion of Golgi-derived exocytic vesicles carrying membranes and cell wall components with the
pre-existing plasma membrane. Similarly, the vesicles that
fuse to form the cell-plate in dividing plant cells are
thought to bud from the Golgi [54]. Very little is however
known about the formation, trafficking and fusion of these
vesicles. For example, it remains to be determined
whether exocytosis and cell-plate formation are mediated
by a single homogeneous vesicle population or whether
multiple Golgi-derived vesicle populations exist, carrying
distinct sets of cargo, that in the case of cell-plate formation would converge and fuse to form a new plasma
membrane and cell wall de novo. Recent studies in plants,
yeast and mammalian cells have suggested the existence of
multiple parallel vesicular routes that transport distinct
sets of cargo from the Golgi to the plasma membrane
[55–57]. In expanding cells these vesicles are likely to be
targeted to the growing regions of the cell cortex. During
cell-plate formation, one or more of these pathways may
become polarized toward the plane of division, as they
appear to do during cell division in yeast [58]. In this way,
the time at which plasma membrane and secreted proteins
are synthesized may dictate their deposition at the cell surface or the cell-plate. Consistent with this hypothesis is the
observation that the putative t-SNARE, Knolle, which is
expressed during mitosis, is localized specifically to the
cell-plate of dividing Arabidopsis cells [59••].
To identify components involved in exocytosis from the
Golgi apparatus, Nakano and coworkers have screened for
Arabidopsis cDNAs that would complement the yeast late
secretory pathway mutant, sec15 [60]. Sec15p is a component of the exocyst complex required for the
Golgi-to-plasma membrane transport [61]. Interestingly,
one gene that was identified, RMA1, does not encode a
Sec15p homolog, rather it belongs to a new class of genes
containing the RING-finger motif (a protein motif that
binds two Zn2+ ions in a unique ‘cross-brace’ structure)
that is conserved in all multicellular organisms. Whether
RMA1p is actually involved in secretion in plant cells
requires further study.

Conclusions
A notable strength of the work highlighted above has been
the integration of ideas and observations from a number of
systems and different plant cell types over the last several
years. Nevertheless caution needs to be exercised when it
comes to deciding whether a particular secretory process is a
general one or if it is restricted to only a limited set of cells
(i.e. those from a specific tissue or developmental stage). For
instance, is the direct ER-to-vacuole transport pathway

467

observed in developing wheat and pumpkin seeds utilized in
other cell types? We would like to encourage frequent discussions of such ideas through formal and informal venues
such as the recently formed Secretory Pathway Group listserver (SECPATWY@LIST.MSU.EDU). Morphological,
biochemical and genetic studies of the plant secretory pathway are beginning to provide a detailed molecular
description of how the elaborate endomembrane network of
organelles in plant cells is established and how it functions in
the growth and development of the whole organism.

Acknowledgements
We thank all the colleagues who provided us unpublished information.
We apologize to those authors whose articles could not be cited due to
space limitations.

References and recommended reading
Papers of particular interest, published within the annual period of review,
have been highlighted as:

• of special interest
•• of outstanding interest
1.

Bassham DC, Raikhel NV: Transport proteins in the plasma
membrane and the secretory system. Trends Plant Sci 1996,
1:15-20.

2.

Okita T, Rogers J: Compartmentation of proteins in the
endomembrane system of plant cells. Annu Rev Plant Physiol
Plant Mol Biol 1996, 47:327-350.

3.

Okita TW, Choi SB, Ito H, Muench DG, Wu Y, Zhang F: Entry into
the secretory system — the role of mRNA localization. J Exp Bot
1998, 49:1081-1089.

4.

Ceriotti A, Duranti M, Bollini R: Effects of N-glycosylation on the
folding and structure of plant proteins. J Exp Bot 1998.
49:1091-1103.

Muench DG, Wu Y, Coughlan SJ, Okita TW: Evidence for a
cytoskeleton-associated binding site involved in prolamine mRNA
localization to the protein bodies in rice endosperm tissue. Plant
Physiol 1998, 116:559-569.
Rice prolamin and glutelin mRNAs are localized to morphologically distinct
parts of the ER. In this paper, PB associated polysomes were found to be
resistant to detergent extraction and puromycin treatment suggesting that
the majority of prolamine mRNA is not directly anchored to the ER membrane through ribosome binding sites nor through the binding of the nascent
prolamin polypeptide. Rather, the prolamine polysome-binding activity was
shown to be associated with cytoskeletal elements.

5.
•

6.

Clore AM, Dannenhoffer JM, Larkins BA: EF-1-alpha is associated
with a cytoskeletal network surrounding protein bodies in maize
endosperm cells. Plant Cell 1996, 8:2003-2014.

7.

Coleman CE, Herman EM, Takasaki K, Larkins BA: The maize g-zein
sequesters a-zein and stabilizes its accumulation in protein
bodies of transgenic tobacco endosperm. Plant Cell 1996,
8:2335-2345.

8.
•

Bagga S, Adams HP, Rodriguez FD, Kemp JD, Sengupta-Gopalan C:
Coexpression of the maize d-zein and b-zein genes results in
stable accumulation of d-zein in endoplasmic reticulum-derived
protein bodies formed by b-zein. Plant Cell 1997, 9:1683-1696.
Zeins, the major seed storage proteins of maize, are synthesized on the rER
and are deposited in ER-PB. In this paper β-zein and δ-zein were shown to
be stably expressed and deposited in zein-specific ER-PB in transgenic
tobacco leaves. Coexpression of β-zein and δ-zein together resulted in an
increase in δ-zein accumulation relative to plants expressing d-zein alone and
both proteins colocalized in the same PB. These results suggest that δ-zein
interacts with and stabilizes the assembly of δ-zein into PB.
9.

Gillikin JW, Zhang F, Coleman CE, Bass HW, Larkins BA, Boston RS:
A defective signal peptide tethers the floury-2 zein to the
endoplasmic reticulum membrane. Plant Physiol 1997, 114:345-352.

10. Coleman CE, Clore AM, Ranch JP, Higgins R, Lopes MA, Larkins, BA:
Expression of a mutant alpha-zein creates the floury2 phenotype
in transgenic maize. Proc Natl Acad Sci USA 1997, 94:7094-7097.
11. Napier JA, Richard G, Turner MF, Shewry PR: Trafficking of wheat
gluten proteins in transgenic tobacco plants: gamma-gliadin does

468

Cell biology

not contain an endoplasmic reticulum-retention signal. Planta
1997, 203:488-494.
12. Levanony H, Rubin R, Altschuler Y, Galili G: Evidence for a novel
route of wheat storage proteins to vacuoles. J Cell Biol 1992,
119:1117-1128.

conditions that lead in an increase in the number of unfolded proteins in the
ER. On the basis of antibody accessibility studies the binding of BiP to calreticulin and to unfolded proteins appears to be different. The
BiP–calreticulin complex can be disrupted by low pH but not by divalent
cation chelators and complex formation is independent of the ER retention
signal on BiP.

13. Hara-Nishimura I, Shimada T, Hatano K, Takeuchi Y, Nishimura M:
•
Transport of storage proteins to protein storage vacuoles is
mediated by large precursor-accumulating vesicles. Plant Cell
1998, 10:825-836.
Large 200–400 nm diameter vesicles that accumulate storage protein precursors were purified from pumpkin seeds. These vesicles contain an
electron-dense core of storage proteins surrounded by an electron-translucent
layer. Numerous storage protein aggregates were also found to be present in
the ER suggesting that these vesicles are ER-derived and that they mediate
the transport of the insoluble storage protein aggregates to vacuole.

25. Li X, Su RTC, Hsu HT, Sze H: The molecular chaperone calnexin
•
associates with the vacuolar H+-ATPase from oat seedlings. Plant
Cell 1998, 10:119-130.
The V-ATPase complex consists of a peripheral sector (V1) and a membrane
integral sector (V0). A 64 kDa polypeptide that copurifies with oat V-ATPase
subunits was positively identified as the ER-membrane chaperone, calnexin,
and shown to interact directly with the V-ATPase complex in purified ER
membrane fractions by coimmunoprecipitation. Monoclonal antibodies
against the catalytic subunit of the V1 complex precipitated the entire VATPase complex as well as calnexin and BiP.

14. Bednarek SY, Orci L, Schekman R: Traffic COPs and the formation
of vesicle coats. Trends Cell Biol 1996, 6:468-473.

26. Hamman BD, Hendershot LM, Johnson AE: BiP maintains the
permeability barrier of the ER membrane by sealing the lumenal
end of the translocon pore before and early in translocation. Cell
1998, 92:747-758.

15. Bar-Peled M, Raikhel NV: Characterization of AtSec12 and AtSAR1
proteins likely involved in ER and Golgi traffic. Plant Physiol 1997,
114:315-324.
16. Muench DG, Wu Y, Zhang Y, Li X, Boston RS, Okita TW: Molecular
cloning, expression and subcellular localization of a BiP homolog
from rice endosperm tissue. Plant Cell Physiol 1997, 38:404-412.
17.

Li X, Wu Y, Zhang D-Z, Gillikin JW, Boston RS, Franceschi VR, Okita
TW: Rice prolamine protein body biogenesis: a BiP mediated
process. Science 1993, 262:1054-1056.

27.

Suzuki T, Yan Q, Lennarz WJ: Complex, two-way traffic of molecules
across the membrane of the endoplasmic reticulum. J Biol Chem
1998, 273:10083-10086.

28. Frigerio L, Vitale A, Lord JM, Ceriotti A, Roberts LM: Free ricin A
chain, proricin, and native toxin have different cellular fates
when expressed in tobacco protoplasts. J Biol Chem 1998,
273:14194-14199.

18. Vitale A, Bielli A, Ceriotti A, The binding protein associates with
monomeric phaseolin. Plant Physiol 1995, 107:1411-1418.

29. Matsuoka K, Neuhaus JM: Cis-elements of protein transport to the
vacuole. J Exp Bot 1999, in press.

19. Lupattelli F, Pedrazzini E, Bollini R, Vitale A, Ceriotti A: The rate of
•
phaseolin assembly is controlled by the glucosylation state of its
N-linked oligosaccharide chains. Plant Cell 1997, 9:597-609.
In the ER nascent secretory proteins containing N-glycans are modified by
the addition of an Glc3Man9GlcNAc2 oligosaccharide unit. Subsequently,
the three terminal glucose residues are removed by ER-resident glucosidases. In this paper, the role of glucose trimming on the folding and assembly of
bean phaseolin was analyzed using an in vitro system. In the presence of
specific inhibitors of the glucose trimming reaction, the assembly of phaseolin was found to be accelerated. In contrast, polypeptides bearing partially
trimmed glycans were unable to properly trimerize. Glycan chains and their
accessibility to glucosidases are, therefore, likely to modulate the assembly
of phaseolin.

30. Gomord V, Denmat LA, Fitchette-Laine AC, Satiat-Jeunemaitre B,
•
Hawes C, Faye L: The C-terminal HDEL sequence is sufficient for
retention of secretory proteins in the endoplasmic reticulum (ER)
but promotes vacuolar targeting of proteins that escape the ER.
Plant J 1997, 11:313-325.
The carboxy-terminal tetrapeptides, HDEL and KDEL, serve as retention signals for soluble ER resident proteins. In this paper the authors characterized
a fusion protein containing a secreted form of sporamin and an HDEL C-terminal extension. Addition of the HDEL extension resulted in the
accumulation of sporamin in the ER. In addition, a significant amount of the
fusion protein that escaped from the ER was transported to the vacuole.

20. Pedrazzini E, Giovinazzo G, Bielli A, de Virgilio M, Frigerio L, Pesca M,
•
Faoro F, Bollini R, Ceriotti A, Vitale A: Protein quality control along
the route to the plant vacuole. Plant Cell 1997, 9:1869-1880.
In this paper, the fate of an assembly-defective form of the trimeric storage
protein, phaseolin, was analyzed in the leaves of transgenic tobacco.
Assembly defective phaseolin was bound to BiP in the ER and slowly
degraded by a Brefeldin-A insensitive process, suggesting that its turnover
does not require Golgi-mediated secretory protein transport. These results
provide strong evidence for the presence of a ‘quality control’ mechanism in
the ER of plant cells that eliminates malfolded proteins.
21. Jung R, Nam YW, Saalbach I, Muntz K, Nielsen NC: Role of the
•
sulfhydryl redox state and disulfide bonds in processing and
assembly of 11S seed globulins. Plant Cell 1997, 9:2037-2050.
Seed legumins contain two conserved disulfide bonds that are important for
the initial formation of 9S trimers in the ER and their assembly into 11S hexamers in the PSV. In this paper mutant subunits were constructed in which the
critical cysteine residues for the interchain bond (IE) connecting the acidic
and basic chains and the intrachain bond (IA) were disrupted. Whereas oxidized glutathione stimulated the trimerization of the wild-type protein no
stimulation was detected for the assembly of IE mutant subunits and it was
diminished for the IA mutant. IE mutant trimers were not capable of assembling into hexamers unless they were in the presence of wild-type subunits.
22. Shimoni Y, Galili G: Intramolecular disulfide bonds between
conserved cysteines in wheat gliadins control their deposition
into protein bodies. J Biol Chem 1996, 271:18869-18874.
23. Shimoni Y, Blechl AE, Anderson OD, Galili G: A recombinant
protein of two high molecular weight glutenins alters gluten
polymer formation in transgenic wheat. J Biol Chem 1997,
272:15488-15495.
24. Crofts AJ, Leborgne-Castel N, Pesca M, Vitale A, Denecke J: BiP and
•
calreticulin form an abundant complex that is independent of
endoplasmic reticulum stress. Plant Cell 1998, 10:813-823.
In this paper a large fraction of BiP was found to be in a stable complex with
another soluble ER-resident molecular chaperone, calreticulin. Formation of
this complex was not affected by either the presence or absence of stress

31. Koide Y, Hirano H, Matsuoka K, Nakamura K: The N-terminal
propeptide of the precursor to sporamin acts as a vacuoletargeting signal even at the C terminus of the mature part in
tobacco cells. Plant Physiol 1997, 114:863-870
32. Matsuoka K, Bassham DC, Raikhel NV, Nakamura K: Different
sensitivity to Wortmannin of two vacuolar sorting signals indicates
the presence of distinct sorting machineries in tobacco cells.
J Cell Biol 1995, 130:1307-1318.
33. Paris N, Stanley CM, Jones RL, Rogers JC: Plant cells contain
two functionally distinct vacuolar compartments. Cell 1996,
85:563-572.
34. Hoh B, Hinz G, Jeong BK, Robinson DG: Protein storage vacuoles
form de novo during pea cotyledon development. J Cell Sci 1995,
108:299-310.
35. Swanson SJ, Bethke PC, Jones RL, Barley aleurone cells contain
•
two types of vacuoles. Characterization of lytic organelles by use
of fluorescent probes. Plant Cell 1998, 10:685-698
Light microscopy was used to study the structure and function of vacuoles
in living protoplasts of barley (Hordeum vulgare cv Himalaya) aleurone.
Aleurone protoplasts contain two distinct types of vacuole: the protein storage vacuole and a lysosome-like organelle. In contrast to the two vacuole
compartments in barley root tip cells, both of the vacuole types of aleurone
cells contain the tonoplast marker protein α-TIP. The effects of the phytohormones, ABA and GA, on the formation and morphological change of
these two vacuoles are reported.
36. Hohl I, Robinson DG, Chrispeels MJ, Hinz G: Transport of storage
proteins to the vacuole is mediated by vesicles without a clathrin
coat. J Cell Sci 1996, 109:2539-2550.
Frigerio L, de Virgilio M, Prada A, Faoro F, Vitale A: Sorting of
phaseolin to the vacuole is saturable and requires a short Cterminal peptide. Plant Cell 1998, 10:1031-1042.
Phaseolin is a trimeric glycoprotein that accumulates in the vacuoles of common bean seeds and in transgenic tobacco leaf cells. Golgi-mediated
transport of wild-type phaseolin to the vacuole was found to be saturable
and to lead to the secretion of the overexpressed protein. Deletion of the four
carboxy-terminal residues of phaseolin did not inhibit the trimerization but
37.
•

Protein transport within the plant cell edomembrane system Matsuoka and Bednarek

469

38. Kirsch T, Paris N, Butler JM, Beevers L, Rogers JC: Purification and
initial characterization of a potential plant vacuolar targeting
receptor. Proc Natl Acad Sci USA 1994, 91:3403-3407.

51. da Silva Conceicão A, Marty-Mazars D, Bassham D, Sanderfoot, AA,
•
Marty F, Raikhel NV: The syntaxin homolog AtPEP12p resides on a
late post-Golgi compartment in plants. Plant Cell 1997, 9:571-582.
The intracellular localization of an Arabidopsis thaliana homolog (AtPEP12p)
of the yeast Golgi-to-vacuole t-SNARE, Pep12p, was analyzed. Subcellular
fractionation and immunolocalization studies indicated that AtPEP12 is an integral membrane protein and that it is localized to a post-Golgi compartment.

39. Kirsch T, Saalbach G, Raikhel NV, Beevers L: Interaction of a
potential vacuolar targeting receptor with amino- and carboxylterminal targeting determinants. Plant Physiol 1996, 111:469-474.

52. Bassham DC, Raikhel NV, An Arabidopsis VPS45p homolog
implicated in protein transport to the vacuole. Plant Physiol 1998,
117:407-415.

40. Paris N, Rogers SW, Jiang L, Kirsch T, Beevers L, Phillips TE, Rogers
•
JC: Molecular cloning and further characterization of a probable
plant vacuolar sorting receptor. Plant Physiol 1997, 115:29-39.
A cDNA encoding BP-80, an abundant type I integral membrane protein in
pea (Pisum sativum) from CCVs that binds with high affinity to peptides containing vacuole-targeting determinant of aleurain precursor was isolated.
Sequence analysis indicated that BP-80 defines a novel family of plant-specific proteins that are highly conserved in both monocotyledons and
dicotyledons. Immunolocalization showed that the BP-80 protein is present
in dilated ends of Golgi cisternae and in ‘prevacuoles’.

53. Sato MH, Nakamura N, Ohsumi Y, Kouchi H, Kondo M, Hara•
Nishimura I, Nishimura M, Wada Y, The AtVAM3 encodes a syntaxinrelated molecule implicated in the vacuolar assembly in
Arabidopsis thaliana. J Biol Chem 1997, 272:24530-24535.
In this paper an Arabidopsis thaliana cDNA, AtVAM3, that complements the
yeast vam3 mutation was isolated. AtVAM3p is expressed in various tissues
and was found to co-fractionate with a vacuolar membrane protein H+translocating inorganic pyrophosphatase. Immunoelectron microscopy
showed that AtVam3p was localized to restricted regions on the tonoplast in
the cells at the shoot apical meristem.

41. Ahmed SU, Bar-Peled M, Raikhel NV: Cloning and subcellular
location of an Arabidopsis receptor-like protein that shares
common features with protein-sorting receptors of eukaryotic
cells. Plant Physiol 1997, 114:325-336.

54. Staehelin LA, Hepler PK: Cytokinesis in higher plants. Cell 1996,
84:821-824.

resulted in the secretion of the mutant protein. These results provide strong
evidence that phaseolin to the vacuole is signal-mediated and receptor
dependent process.

42. Shimada T, Kuroyanagi M, Nishimura M, Hara-Nishimura I: A pumpkin
•• 72-kDa membrane protein of precursor-accumulating vesicles
has characteristics of a vacuolar sorting receptor. Plant Cell
Physiol 1997, 38:1414-1420.
Precursor-accumulating vesicles mediate the transport of pro2S albumin to
protein-storage vacuoles in developing pumpkin cotyledons. Two homologous membrane proteins of 72 kDa and 82 kDa were isolated from these
vesicles and shown to bind to peptides derived from pro2S albumin. These
proteins are predicted to be type I membrane protein containing lumenal
EGF receptor-like motifs similar to those found in BP-80 and AtELP. This is
the first report in plants showing that a receptor-like protein and its defined
ligand are packaged into the same transport vesicle.
43. Hoflak B: Mechanisms of protein sorting and coat assembly:
clathrin coated vesicle pathways. Curr Opin Cell Biol 1998,
10:499-503.
44. Nishimura N, Balch WE: A di-acidic signal required for selective
export from the endoplasmic reticulum. Science 1997,
277:556-558.
45. Sanderfoot AA, Ahmed SU, Marty-Mazars D, Rapoport I, Kirchhausen T,
•• Marty F, Raikhel NV: A putative vacuolar cargo receptor partially
colocalizes with AtPEP12p on a prevacuolar compartment in
Arabidopsis roots. Proc Natl Acad Sci USA 1998, 95: in press.
The putative vacuolar cargo receptor, AtELP, from Arabidopsis thaliana
shows significant sequence similarity to BP-80. AtELP was previously
shown to be present in two membrane fractions corresponding to a novel,
undefined compartment and a clathrin coated vesicle fraction. In this paper,
the undefined compartment was shown to correspond to a post-Golgi prevacuolar compartment (PVC) containing the t-SNARE, AtPEP12p, which is
required for Golgi-to-vacuole transport. In addition, AtELP is also present in
the trans-Golgi. The cytosolic tail of AtELP binds to the mammalian AP-1
adaptor complex which connects the clathrin-coat to Golgi-localized receptors. Thus AtELP and its homolog BP-80 are likely to mediate protein
sorting into CCVs and to shuttle back and forth between the trans-Golgi
and the PVC.
46. Robinson DG, Hinz G: Multiple mechanisms of protein body
formation in pea cotyledons. Plant Physiol Biochem 1996,
34:155-163.
47.

Robinson DG, Baeumer M, Hinz G, Hohl I, Clark SE, Williams RW,
Meyerowitz EM, Ultrastructure of the pea cotyledon Golgi
apparatus: origin of dense vesicles and the action of brefeldin A.
Protoplasma 1997, 200:198-209.

48. Weber T, Zemelman BV, McNew JA, Westermann B, Gmachl M,
Parlati F, Söllner TH, Rothman JE: SNAREpins: minimal machinery
for membrane fusion. Cell 1998, 92:759-772.
49. Wada Y, Nakamura N, Ohsumi Y, Hirata A: Vam3p, a new member of
syntaxin related protein, is required for vacuolar assembly in the
yeast Saccharomyces cerevisiae. J Cell Sci 1997, 110:1299-1306.
50. Nichols BJ, Ungermann C, Pelham HRB, Wickner WT, Haas A:
Homotypic vacuolar fusion mediated by t- and v-SNAREs. Nature
1997, 387:199-202.

55. Moore PJ, Swords KM, Lynch MA, Staehelin LA: Spatial organization
of the assembly pathways of glycoproteins and complex
polysaccharides in the Golgi apparatus of plants. J Cell Biol 1991,
112:589-602.
56. Harsay E, Bretscher A: Parallel secretory pathways to the cell
surface in yeast. J Cell Biol 1995, 131:297-310.
57.

Yoshimori T, Keller P, Roth MG, Simons K: Different biosynthetic
transport routes to the plasma membrane in BHK and CHO cells.
J Cell Biol 1996, 133:247-256.

58. Lew DJ, Reed SI: Morphogenesis in the yeast cell-cycle: Regulation
by Cdc28 and cyclins. J Cell Biol 1993, 120:1305-1320.
59. Lauber MH, Waizenegger I, Steinmann T, Schwarz H, Mayer U,
•• Hwang I, Lukowitz W, Jürgens G: The Arabidopsis KNOLLE protein
is a cytokinesis-specific syntaxin. J Cell Biol 1997, 139:1485-1493.
The Arabidopsis gene, KNOLLE, which is required for cytokinesis, encodes
a putative t-SNARE related to the mammalian plasma membrane fusion protein, syntaxin. KNOLLE is membrane associated and was shown by
immunofluorescence microscopy to be specifically expressed during mitosis
and to be localized to the plane of division during cytokinesis. Electron
microscopic analysis indicates that cell-plate vesicle fusion is impaired in
knolle mutant embryos. This is the first report of a syntaxin-like protein that
appears to be involved specifically in the formation of the cell-plate.
60. Matsuda N, Nakano A: RMA1, an Arabidopsis thaliana gene whose
cDNA suppresses the yeast sec15 mutation, encodes a novel
protein with a RING finger motif and a membrane anchor. Plant
Cell Physiol 1998, 39:545-554.
61. Terbush DR, Maurice T, Roth D, Novick P: The exocyst is a
multiprotein complex required for exocytosis in Saccharomyces
cerevisiae. EMBO J 1996, 15:6483-6494.
62. Rothman JE: Mechanisms of intracellular protein transport. Nature
1994, 372:55-62.
63. Novick P, Brennwald P: Friends and family: the role of the Rab
GTPases in vesicular traffic. Cell 1993, 75:597-601.
64. Pevsner J: The role of Sec1p-related proteins in vesicle trafficking
in the nerve terminal. J Neurosci Res 1996, 45:89-95.
65. Cao X, Ballew N, Barlowe C: Initial docking of ER-derived vesicles
requires Uso1p and Ypt1p but is independent of SNARE proteins.
EMBO J 1998, 17:2156-2165.
66. Nakamura N, Lowe M, Levine TP, Rabouille C, Warren G: The vesicle
docking protein p115 binds to GM130, a cis-Golgi matrix protein,
in a mitotically regulated manner. Cell 1998, 89:445-455.
67.

Patel SK, Indig FE, Olivieri N, Levine ND, Latterich M: Organelle
membrane fusion: a novel function for the syntaxin homolog
Ufe1p in ER membrane fusion. Cell 1998, 92:611-620.

68. Feiler HS, Desprez T, Santoni V, Kronenberger J, Caboche M, Traas J:
The higher plant Arabidopsis thaliana encodes a functional
CDC48 homologue which is highly expressed in dividing and
expanding cells. EMBO J 1995, 14:5626-5637.