The Plant Cell, Vol. 17, 3213–3216, December 2005, www.plantcell.org ª 2005 American Society of Plant Biologists
IN THIS ISSUE
Insights into Plant Cellular Mechanisms: Of Phosphate
Transporters and Arbuscular Mycorrhizal Infection
The high demand for phosphorus as an
essential macronutrient coupled with limitations in phosphate availability or accessibility
in soils has led to the evolution in plants of
various mechanisms for facilitating and enhancing the uptake of inorganic phosphate.
Uptake of phosphate into plant cells occurs
against a steep concentration gradient, as
the concentration within plant cells (1 to
10 mM) is ;10,000-fold higher than that of
the soil solution (Rausch and Bucher, 2002).
Two important mechanisms for phosphate
uptake in higher plant roots are the activity of
high affinity phosphate transporters and the
symbiotic association with arbuscular mycorrhizal (AM) fungi. Both of these phenomena involve orchestration of a complex series
of events within the cell. On the one hand,
traffic through the secretory pathway is required to direct localization of phosphate
transporters to the plasma membrane, and
on the other, the plant cell allows for penetration and elaboration of a complex network of fungal arbuscular hyphae that
facilitates exchange of minerals and nutrients between the plant cell and fungal
symbiont. Two articles in this issue of The
Plant Cell present significant new information
on cellular mechanisms associated with
these processes of fundamental importance
to higher plants. In one article, González
et al. (pages 3500–3512) identify an accessory protein associated with targeting of
phosphate transporters to the plasma membrane in Arabidopsis. In a separate article,
Genre et al. (pages 3489–3499) present
a detailed intracellular view of the early
stages of root colonization by AM fungi in
Medicago truncatula and show that the host
plant plays a key role in orchestrating the AM
infection process.
and Bucher, 2002). The PHT1 family includes high affinity phosphate transporters
that have 12 membrane spanning domains
and are presumed to be targeted to the
plasma membrane. PHT1 is a relatively
large family in Arabidopsis (nine members)
and rice (at least 13), and patterns of gene
expression suggest functional specialization within the family (Mudge et al., 2002).
Thus, some PHT1 proteins may function
primarily in phosphate uptake at the soil–
root interface, whereas others may participate predominantly in translocation to
other parts of the plant and/or transport
within certain tissues or cell types. Plant
organelles appear to contain separate families of phosphate transporters; for example, PHT2 proteins and another type called
phosphate translocators may be found in
chloroplasts, whereas PHT3 proteins are
targeted to mitochondria.
González et al. isolated a mutant of
Arabidopsis that displays constitutive expression of a phosphate starvation responsive reporter gene, suggesting mutation in
a gene associated with phosphate perception or transport (i.e., perception and trans-
ACCESSORY PROTEIN GUIDES
ENDOPLASMIC RETICULUM EXIT OF
A PHOSPHATE TRANSPORTER
Figure 1. Subcellular Localization of PHT1 in Wild-Type and phf1 Mutant Plants.
Plants contain a large number of phosphate transporters (reviewed in Rausch
port of phosphate normally inhibits the
expression of phosphate starvation responsive genes, which will only be induced
when the inorganic phosphate concentration falls below a certain threshold). Analysis of the mutant plants suggested that
phosphate accumulation was reduced due
to a specific effect on phosphate uptake
(uptake of other nutrients was unaffected).
The corresponding gene, named PHOSPHATE TRANSPORTER TRAFFIC FACILITATOR (PHF1), was found to encode a
Sec12-related plant-specific protein required for localization of PHT1 to the plasma
membrane. PHT1 was shown to be localized to the plasma membrane in wild-type
cells but was retained in the endoplasmic
reticulum (ER) in the phf mutant (Figure 1).
Localization to the plasma membrane
requires trafficking through the secretory
pathway. From information obtained mainly
in the yeast system, it is known that
proteins bound for secretion are synthesized in the ER and are transported via
COPII vesicles to the Golgi apparatus, then
to the trans-Golgi network where they
are sorted and packed into vesicles for
Micrographs of leaf epidermal cells from transgenic wild type (left) and phf1 mutant (right) harboring
a PHT1:GFP construct driven by the cauliflower mosaic virus 35S promoter. PHT1:GFP is localized to
the plasma membrane in wild-type cells, but is retained in the ER in the phf1 mutant. (Figure courtesy of
Esperanza González and Javier Paz-Ares.)
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further transport to various destinations
(e.g., plasma membrane, vacuoles, and
cell wall). Traffic through the secretory
pathway occurs in a highly complex series
of steps involving a large number of transport and accessory proteins. In the early
secretory pathway, COPII catalyzes formation of transport vesicles (COPII-coated
vesicles) from the ER and segregates cargo
proteins from ER-resident proteins (reviewed in Antonny and Schekman, 2001;
Lee et al., 2004). Sec12 is a guanine
nucleotide exchange factor, originally identified in yeast, which plays a critical role in
the formation of COPII-coated vesicles.
One of the main components of COPII
coats is the Sar1 GTPase, which initiates
coat assembly when activated by Sec12.
GTP binding by Sar1 (catalyzed by Sec12)
causes association with the ER membrane
and subsequent interaction with Sec23/24
(involved in cargo recognition and coat
assembly) and Sec13/31 (involved in polymerization and budding of the COPII
vesicle). Antonny and Schekman (2001)
suggested that Sec12 may be continuously
active to support the constitutive functioning of COPII vesicles.
The ER in yeast and animals also contains a number of substrate- or cargospecific accessory proteins that assist in
various ways with the recruitment, packaging, and transport of cargo in COPII
vesicles (Herrmann et al., 1999). For example, mammalian RanB2 and Drosophila
NinaA, related to cyclophilins known to be
involved in protein folding, are involved in
the ER–Golgi transport of specific seventransmembrane photoreceptors. Various
other plasma membrane–bound proteins
require the presence of accessory proteins
for transport through the secretory pathway. Indeed, the yeast PHT1-related phosphate transporter PHO84 requires the
accessory protein PHO86 for exit from the
ER (Lau et al., 2000).
Although PHF1 shares homology with
Sec12 and no structural or sequence
similarity to PHO86, González et al. show
that it is functionally similar to PHO86,
acting as a substrate-specific accessory
protein for the transport of PHT1 out of the
ER, rather than a general activator of Sar1
GTPase similar to yeast Sec12. First, al-
though PHF1 shows a high degree of
structural similarity to Sec12 and is localized to the ER, it lacks a number of the most
highly conserved residues present in Sec12
proteins from a number of organisms.
Arabidopsis contains a close homolog of
Sec12, STL2, and the authors showed that
heterologously expressed STL2 was able
to complement the sec12 yeast mutant,
whereas PHF1 was not. Secondly, the phf1
mutant did not exhibit generalized impairment of protein trafficking out of the ER, as
would be expected by disruption of Sec12
activation of Sar1 GTPase. The authors
confirmed this observation by visualization
of a green fluorescent reporter fused to the
plasma membrane marker protein PIP2A,
which was correctly targeted to the plasma
membrane in phf1 mutant plants.
Phylogenetic analysis conducted by
González et al., together with the functional
characterization, suggests that PHF1 proteins originated from a plant-specific Sec12
ancestral protein and evolved functional
specialization as accessory proteins guiding ER exit of phosphate transporters. This
work provides new information about the
secretory pathway in plants, as it reveals
the likely existence of plant-specific accessory proteins that participate in targeting of
proteins to the plasma membrane.
A DETAILED VIEW OF THE AM
INFECTION PROCESS
Most terrestrial plants participate in a symbiotic relationship with AM fungi that
facilitates uptake of nutrients, particularly
phosphorus, by plant roots and provides
the fungus with essential carbohydrates
(reviewed in Hause and Fester, 2005). AM
fungi are obligate biotrophs that depend
entirely on the host plant as a carbon
source. All AM fungi belong to the phylum
Glomeromycota, which is divided into four
orders and ;150 known species (Harrison,
2005). Glomus and Gigaspora species are
the best characterized and most studied.
Figure 2. Intracellular Dynamics in the M. truncatula Root Epidermis Preceding AM Infection.
The illustration shows the transient PPA, comprising cytoskeletal/ER components, which is assembled
in response to surface hyphal contact and appressorium formation. The PPA is assembled within the
cytoplasmic column created during transcellular nuclear migration and likely plays a central role in the
synthesis of the apoplastic compartment through which the AM infection hypha subsequently crosses
the epidermal barrier. Color coding: cell nucleus, dark brown; plasma membrane, light brown;
microtubules, green; actin bundles, red; ER, white. (Figure courtesy of Andrea Genre.)
December 2005
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IN THIS ISSUE
Colonization of roots by AM fungi and development of the arbuscular hyphae within
root cortical cells is a highly complex process. Plants must recognize AM fungi as
beneficial nonpathogenic intruders and
facilitate both the initial entry of the fungus
and the subsequent development of the
arbuscular network (reviewed in Harrison,
2005). In legumes such as Medicago and
Lotus, a subset of genes is required for
establishing both the AM association and
root nodulation associated with nitrogenfixing bacteria, indicating that common
pathways are likely to be involved in initiating both types of symbioses (Parniske,
2004; Kistner et al., 2005).
Enhanced uptake of inorganic phosphate is one of the major benefits that
plants derive from AM fungi, and indeed
plants appear to contain phosphate transporters specialized for this purpose. In
M. truncatula, transcript and protein levels
of the high affinity plasma membrane phosphate transporters PT1 and PT2 are induced in roots in response to phosphate
starvation but are not expressed in roots
colonized by AM fungi (Chiou et al., 2001).
Instead, the phosphate transporter MtPT4
is expressed specifically in mycorrhizal
roots, where it is likely localized to the
periarbuscular membrane and functions
specifically in uptake of phosphate released by the AM fungus (Harrison et al.,
2002). Karandashov et al. (2004) found that
AM-specific phosphate transporter genes
related to MtPT4 are conserved across
phylogenetically distant plant species.
Despite knowledge of plant genes that
are required for or whose expression is
altered by the association with AM fungi,
relatively little is known about the mechanisms that control AM development. Genre
et al. studied in vivo cellular dynamics
within M. truncatula root epidermal cells
during the early stages of colonization by
Gigaspora AM fungi, making use of green
fluorescent protein (GFP) reporters to visualize the plant cytoskeleton and ER. Three
different GFP constructs were used to
visualize the microtubular cytoskeleton,
actin filaments, and the ER. The authors
modified a targeted AM inoculation technique developed by Chabaud et al. (2002)
to allow continuous microscopic observa-
tion during AM colonization of roots. The
resulting images provide a detailed picture
of intracellular events associated with AM
infection.
AM root infection initiates with the
formation of surface appressoria at points
of contact between fungal hyphae and the
root epidermal cell. Notably, the results of
Genre et al. have shown that the epidermal
cell nucleus undergoes two distinct phases
of movement, both preceding fungal penetration. After appressoria formation, the
epidermal cell nucleus rapidly moves toward and positions itself directly below the
appressorium contact site. During a second
trans-cellular phase of nuclear movement,
a hollow structure comprising ER and cytoskeletal components is progressively assembled between the appressorium contact
site and the nucleus. This process takes ;4
to 5 h following appressorium formation,
and fungal penetration occurs only after
completion of this structure, termed the
prepenetration apparatus (PPA; Figure 2).
Furthermore, since hyphal growth occurs
strictly along and within the trans-cellular
path created by the PPA, the authors propose that the role of this apparatus is to
synthesize the apoplastic compartment,
which always surrounds and isolates the
infection hypha (Bonfante et al., 2000). After
fungal penetration, the epidermal cell nucleus repositions at the cell periphery. The
authors provide a supplemental animated
video depicting the entire AM infection
process and the roles of the plant cell
nucleus, cytoskeleton, and ER in formation
of the PPA.
In addition, observations using mutants
of M. truncatula impaired in the AM infection process suggest that formation of
the PPA is essential for fungal penetration.
Mutants of M. truncatula called doesn’t
make infections (dmi) have defined three
distinct DMI genes required for both AM
development and nodulation associated
with Sinorhizobium meliloti (Catoira et al.,
2000). Despite the fact that surface appressoria form following contact of dmi
mutant roots with AM fungi, the fungus fails
to penetrate the epidermal cells. Using the
GFP reporter system described above,
Genre et al. show that dmi2 and dmi3 roots
fail to form the PPA following appressoria
formation. The authors hypothesize that
appressorium formation generates a signal,
termed a Myc (mycorrhizal) signal, by way
of analogy with Nod (nodulation) factor signaling in the legume-Rhizobium symbiosis
that is transduced by a DMI-dependent
signaling pathway to activate trans-nuclear
migration and assembly of the PPA.
The work of Genre et al. provides significant new insights into the early stages of
AM infection and in particular the active
role of the host plant in regulating this process. These results also provide direct
evidence for the pivotal role of epidermal
cells in recognition and accommodation of
the AM symbiont and raise the likelihood of
direct analogies with the cellular mechanism involved in the entry of rhizobia via the
root hair apoplastic invagination known as
the infection thread.
Nancy A. Eckardt
News and Reviews Editor
[email protected]
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Insights into Plant Cellular Mechanisms: Of Phosphate Transporters and Arbuscular Mycorrhizal
Infection
Nancy A. Eckardt
Plant Cell 2005;17;3213-3216
DOI 10.1105/tpc.105.039297
This information is current as of June 16, 2017
References
This article cites 18 articles, 7 of which can be accessed free at:
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