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What Can Plant
Autophagy Do for an Innate
Immune Response?
Andrew P. Hayward1 and S.P. Dinesh-Kumar2∗
1
Department of Molecular, Cellular, and Developmental Biology, Yale University,
New Haven, Connecticut 06520-8103
2
Department of Plant Biology and The Genome Center, College of Biological Sciences,
University of California, Davis, California 95616; email: [email protected]
Annu. Rev. Phytopathol. 2011. 49:557–76
Keywords
The Annual Review of Phytopathology is online at
phyto.annualreviews.org
hypersensitive response, senescence, innate immunity, programmed
cell death, chlorophagy
This article’s doi:
10.1146/annurev-phyto-072910-095333
c 2011 by Annual Reviews.
Copyright All rights reserved
0066-4286/11/0908/0557$20.00
Abstract
Autophagy plays an established role in the execution of senescence, starvation, and stress responses in plants. More recently, an emerging role
for autophagy has been discovered during the plant innate immune response. Recent papers have shown autophagy to restrict, and conversely,
to also promote programmed cell death (PCD) at the site of pathogen infection. These initial studies have piqued our excitement, but they have
also revealed gaps in our understanding of plant autophagy regulation,
in our ability to monitor autophagy in plant cells, and in our ability
to manipulate autophagic activity. In this review, we present the most
pressing questions now facing the field of plant autophagy in general,
with specific focus on autophagy as it occurs during a plant-pathogen interaction. To begin to answer these questions, we place recent findings
in the context of studies of autophagy and immunity in other systems,
and in the context of the mammalian immune response in particular.
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INTRODUCTION
UPR: unfolded
protein response
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Cytoplasm-tovacuole (CVT)
pathway: selectively
targets
proaminopeptidase I
to the vacuole in yeast
Phagophore
assembly site/
preautophagosomal
structure (PAS): the
site of autophagosome
formation
PAMP: pathogenassociated molecular
pattern
TLR: Toll-like
receptor
MHCII: major
histocompatibility
complex class 2
PI3K:
phosphoinositide-3kinase
558
Autophagy might be loosely defined as the bulk
degradation of cellular components during
periods of stress and starvation. However, this
definition is at best unrefined. Christian de
Duve, who coined the term autophagy in the
early 1960s, himself described the possibility
of a more selective autophagy for targeting of
dysfunctional proteins and organelles (100).
Such musings foretold a developing role for
autophagy in the removal and degradation
of aberrant proteins during the unfolded
protein response (UPR), where it is now
pursued in regard to a range of UPR disorders
such as Alzheimer’s and Parkinson’s disease
(60). The molecular era has ushered in the
further split of bulk macroautophagy (herein
autophagy) from the selective autophagies,
including the cytoplasm-to-vacuole (CVT)
pathway, chaperone-mediated autophagy, and
the organellar mito-, pexo-, and piecemeal
nuclear autophagies (104).
The core marker for autophagy is the
double membrane–bound autophagosome.
The autophagosome is formed de novo within
the cell, beginning from a platform called the
phagophore assembly site/preautophagosomal
structure (PAS) (95). The autophagosome
expands from the PAS, encircling nearby
components of the cell. The autophagosome
then fuses to the vacuole directly (yeast and
plants), or first to the lysosome (forming the
acidic autophagolysosome) and then to the
vacuole (animals) for further processing and
recycling of its cargo (95).
In the mammalian field, recent studies have
revealed a major role for autophagy in both the
innate and adaptive immune responses. Autophagosomes are able to engulf intracellular
bacteria and viruses (xenophagy), and in some
cases these pathogens have adapted to sequester
autophagic membranes for their own replication (50, 51, 75). Other pathogens, including
HIV, appear to specifically inhibit autophagy
in order to promote infection (4). Autophagy
also promotes the presentation of pathogenassociated molecular patterns (PAMPs) to
Hayward
·
Dinesh-Kumar
Toll-like receptors (TLRs) during an innate
immune response (51). During the adaptive
immune response, autophagy can deliver cytosolic antigens and phagocytosed extracellular
antigens to major histocompatibility complex
class 2 (MHCII) loading compartments, promoting antigen presentation on the cell surface
(15, 47).
In plants, the study of autophagy is reaching
the end of its first decade. Arabidopsis homologs
of most of the core AuTophaGy (ATG) genes
have been identified, and many have been well
described (2, 28, 86). As with other systems, we
now know that plant autophagy has essential
roles not only during starvation, but also during
development, senescence, stress, and pathogen
response. Despite a delayed start, the study of
plant autophagy is catching up with study of
autophagy in yeast and mammals. As it does so,
we find ourselves facing many of the same questions as our colleagues: How is autophagy regulated? Is autophagy pro-survival or pro-death?
Can it be both? Do we have the tools we need
to measure autophagy, to manipulate it, and to
draw strong conclusions from our results?
Here we present what we foresee to be the
most pressing questions facing the field of plant
autophagy. Although we focus on autophagy
during plant-pathogen interactions in particular, many of these challenges are shared along
the breadth of plant autophagy studies.
HAVE THE CORE REGULATORS
OF PLANT AUTOPHAGY
BEEN IDENTIFIED?
The Expansion of Core Autophagy
Machinery in Plants
The core machinery of autophagy is conserved
among yeast, mammals, and plants. This
machinery can be broken down into three
functional units: the ATG1 activation complex
coupled with the ATG2, ATG9, and ATG18
shuttling and membrane delivery system; the
ATG6/Beclin1 and phosphoinositide-3-kinase
(PI3K)/VPS34 autophagosome nucleation
complex; and the ATG8-PE and ATG5-12-16
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conjugation systems (Figure 1). Many excellent
reviews discuss the identification and function
of these core components in yeast and mammals (95), and in plants (2, 28), and we discuss
each functional unit in greater detail below.
Although plants retain the majority of the
autophagy machinery first identified in yeast,
expansion has occurred in several core protein families. In the activation and membrane
delivery complexes, ATG1(4), ATG13(2) and
ATG18(8) have expanded, whereas the two
conjugation systems have increased representation of ATG4(2), ATG8(9), and ATG12(2)
(2, 26, 28). Although several of these families
have been only marginally explored (ATG13,
ATG18), in at least three cases (ATG1, ATG4,
and ATG12) the additional family members
appear to provide functional redundancy (9,
97, 101).
It is tempting to suggest that the expansion of these core families in plants might allow for an increased breadth and specificity of
autophagy function, although little molecular
evidence is available to support this hypothesis.
We have previously proposed that the expanded
ATG8 and ATG18 protein families in plants
may provide for specificity of targeting for autophagosomes, and there is clear evidence of
differential expression of these proteins under
different environmental conditions (28, 76). In
fact, evidence for such substrate targeting exists
in the mammalian field. The LC3(ATG8) interacting protein, Nbr-1, appears to specifically
target autophagy to ubiquitinated substrates in
order to promote their degradation (39).
Others have also noted that although the
Arabidopsis (At) ATG homologs retain high
amino acid conservation of core sequences,
they have low sequence homology relative to
their yeast counterparts, and only ATG4 and
ATG6 have been shown to complement yeast
knockouts (26, 56). We will not begin to learn
the relevance of these nonhomologous regions
(if any) until larger scale interaction screens
or detailed protein-protein interaction studies have been conducted using the AtATG
proteins.
Is the AtTOR/ATG1 Interaction
the Molecular Switch for
Plant Autophagy?
In yeast and mammals, the primary molecular switch for starvation-induced autophagy is
the interaction between target of rapamycin
(TOR) and ATG1 (for review, see 29, 42).
The TOR complex integrates nutrient signaling and, under basal conditions, associates with
the ATG1 complex to inhibit its activity. This
inhibition appears to be mediated by TORinduced phosphorylation events on both the
ATG1 and ATG13 proteins, effectively placing
autophagy in an “off ” state (Figure 1a). Upon
starvation, the TOR complex dissociates from
the ATG1 complex, causing a chain of dephosphorylation events within the ATG1/ATG13
complex and (in yeast) a consequent increase
in ATG13 binding affinity (29). The now activated ATG1 complex will freely localize to
the nascent phagophore, where its activity is
required to recruit ATG9 and possibly its lipid
complement. These activities ultimately flip autophagy to “on” and enable autophagosome
formation.
Homologs of the primary TOR and ATG1
complex constituents have been identified
in Arabidopsis (73). Although ATG1 is less
well studied (with four putative homologs
of the gene), AtTOR knockouts have shown
embryonic lethality, whereas knockdown and
overexpression studies primarily suggest a
function of the protein in regulation of organ
and cell size (14, 59). Despite the existence of
both complexes, however, a clear display of
AtTOR-based autophagy regulation has been
difficult to produce. Initial studies showed an
absence of AtTOR expression in postembryonic cells (59). AtTOR RNAi plants also share
the early senescence phenotype shown in many
plant ATG knockouts (discussed below), suggesting positive rather than negative regulation
of autophagy (14). However, a recent study by
Liu & Bassham (55) strongly affirms that TOR
functions as a negative regulator of autophagy
in plant systems. Using AtTOR RNAi, they
show constitutive autophagy activation and
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TOR: target of
rapamycin
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autophagosome
accumulation
compared
to control plants (55). Further supporting
ATG1-dependent activation of autophagy is
the finding that both AtATG9 and AtATG18a
are transcriptionally upregulated in TOR
RNAi plants (55).
In other systems, the activation of TOR is
primarily mediated by the KOG1/RAPTOR
gene, which binds TOR directly and recruits TOR substrates for phosporylation
(73). There are two raptor-like genes in
Arabidopsis, one of which (AtRAPTOR1)
a
Regulatory complexes
Activation
?
ATG5-12-16
ATG8-PE
LST8
PE
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ATG8
PI
PI(3)P
RAPTOR
TOR
?
TOR
ATG1
Membrane
ATG13
ATG13
ATG1 inhibition
ATG1
?
ATG18
ATG2
?
ATG9
Membrane recruitment
and vesicle expansion
c
ATG12
ATG5
ATG16
PE
ATG12
ATG8
ATG5
ATG10
ATG3
b
ATG5
ATG8*
Activation?
ATG7
?
ATG5
PI3K
ATG6/Bec1
ATG8*
?
ATG8
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Dinesh-Kumar
VPS15
ATG4
ATG6/Bec1
Activation?
CVT pathway?
?
UVRAG
Vesicle nucleation
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is an embryonic lethal knockout, and thus likely
a functional TOR regulator (1, 13). Other
homologs of TOR complex members (namely,
AtLST8) have also been detected in lower
plants, but many remain to be identified (16).
Further study of TOR-based regulation of plant
autophagy will undoubtedly be informative,
particularly regarding the potential for ectopic
induction of autophagy through TOR inactivation (discussed below). It is also important to
note that, although ATG1/TOR is a primary
switch for nutrient stress–induced autophagy,
initial evidence from other organisms suggests
that autophagy regulation during an immune
response may in fact be TOR-independent (93).
What Role Do ATG6 Complexes Play
in Modulating Plant Autophagy?
Rather than a molecular switch, ATG6/Beclin1
appears to act as the primary determinant of the
level of cellular autophagy that occurs in yeast
and mammalian cells (30). Mammalian Beclin1
was first identified in a two-hybrid screen for
Bcl-2-interacting proteins (54). Prior to this
screen, Bcl-2 was known to function endogenously in mammals as an antiapoptotic protein
by inhibiting cytochrome c release from the mitochondria. It was later shown to bind Beclin1
so as to disrupt its autophagy function, suggesting that differential Beclin1 activity might
regulate cell fate decisions between apoptosis
and autophagy (69). ATG6/Beclin1 is required
for nucleation of autophagosomal vesicles,
probably by acting as a scaffold for PI3K
activity, which supplies phosphoinositol-3phosphate (PI3P) to the nascent phagophore
and promotes recruitment of other ATG
proteins required for autophagosome formation (Figure 1b). In the absence of the
ATG6/Beclin1 complex, vesicle nucleation,
and in turn autophagy itself, cannot occur.
Following the identification of the Bcl-2/
Beclin1 interaction, several other Bcl-2 family proteins were also discovered to interact
through the Bcl-2-homology-3 (BH3) domain
of Beclin1, and these interactions appear to be
collectively antiautophagic in nature. (30, 69).
The Bcl-2 family was the first of many other
Beclin1 interactions to be discovered and suggest that Beclin1 is a hub of autophagy regulation. The importance of Beclin1-mediated
autophagy regulation is also illustrated by the
discovery of interactions between Beclin1 and
the MyD88/TIR-domain-containing adapterinducing interferon-β (TRIF) proteins, which
appear to lift Bcl-2-based inhibition (78).
MyD88 and TRIF are essential for integrating signaling downstream of mammalian TLRs,
and this interaction suggests that, although the
TOR pathway may not be pathogen responsive,
Beclin1 is. These particular interactions may
←−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−
Figure 1
The core complexes and regulatory nodes of plant autophagy. The core functional units of autophagy are
conserved in plants, and we use what is known from the yeast and mammalian systems to hypothesize the
functions of these complexes. The interaction of the target of rapamycin (TOR) and ATG1 complexes
inhibits autophagy under basal conditions (a). Components of the TOR complex identified in plants include
RAPTOR and LST8. During stress, TOR dissociates from the ATG1 complex, releasing autophagy
inhibition. The ATG1 complex, including ATG13, localizes to the phagophore and promotes membrane
recruitment and autophagosome expansion. Membrane recruitment is also dependent on ATG9, ATG18,
and ATG2. The initial nucleation of the autophagosome requires the ATG6/Beclin1 nucleation complex (b).
This complex may be shared between the autophagy and CVT pathways, with the relative levels of each
determining the intensity of the autophagy response. ATG6/Beclin1 complex members identified in plants
include PI3K, VPS15, and UVRAG. This complex promotes conversion of phosphoinositol (PI) to
phosphoinositol-3-phosphate (PI3P), necessary for the recruitment of other core components that include
the ATG1 complex. Finally, the two ubiquitin-like conjugation systems (c) promote vesicle maturation. After
modification by ATG4, ATG8 is conjugated to phosphatidyl ethanolamine (PE) by ATG7 and ATG3.
ATG5 is conjugated to ATG12 by ATG7 and ATG10. The ATG5–12 conjugate then forms a complex with
ATG16, and its membrane localization promotes ATG8-PE integration.
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VPS: vacuolar protein
sorting
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PE: phosphatidyl
ethanolamine
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explain the revelation that the viral pathogen
Herpes simplex virus 1 (HSV-1) specifically inhibits autophagy in mammalian neuronal and
immune cells by direct interaction with Beclin1
(49, 67).
The core ATG6/Beclin1 complex has two
other constituents, PI3K/VPS34 and VPS15
(30). In yeast, ATG6 activity is shared between
the autophagy and vacuolar protein sorting
(VPS) pathways, with its activity determined by
the alternative binding of either ATG14 (autophagy) or VPS38 (VPS). Mammalian Beclin1
can restore autophagy, but not protein sorting, in atg6 mutant yeast, and its role in vesicle trafficking pathways outside of autophagy
remains controversial (53). However, a recent
study found that Beclin1 interacting protein
UVRAG mediates endocytic vesicle trafficking,
and the possibility remains that Beclin1 has
roles outside of autophagy in mammals (35). At
least three biochemically distinct Beclin1 complexes have been identified in mammals, some
of which are proposed to be mutually exclusive
(30). Among the variable Beclin1 complex components are Ambra1 (potentially a core component), Atg14 (promotes autophagy), UVRAG
(controversial function in mammals, appears to
be VPS specific in yeast; see 19), and Rubicon
(19, 30).
Plant ATG6 has been shown to restore the
autophagy pathway in atg6 mutant yeast (20,
56). One group has shown that the CVT pathway can also be restored (20), although a more
recent paper could not replicate this result using
an alternative complementation assay (27). The
potential role of ATG6 in protein sorting, independent of autophagy, has been proposed to
explain the embryonic lethality of atg6 T-DNA
insertion knockouts (68, 71). Although representatives of the ATG6 complex core components ATG6, PI3K, VPS15, and UVRAG are
present in plants (Figure 1b), demonstration
of their interactions in the complex is lacking.
These studies will be necessary to determine if
ATG6 activity is involved in both autophagy
and protein sorting in plants. Given the proliferation of autophagy proteins in plants and
the diversity of functions already established, it
Hayward
·
Dinesh-Kumar
is probable that more ATG6-interacting proteins remain to be identified. Any characterization of such complexes would be valuable to all
eukaryotic studies, as it is still unclear if these
variable complexes provide a distinct function
to autophagy beyond its level of induction.
DOES AN AUTOPHAGY-NULL
MUTANT EXIST IN PLANTS?
Genetic Knockouts in the ATG8-PE
and ATG 5–12-16 Conjugation
Systems
Two ubiquitin-like protein conjugation systems are required for autophagosome formation (95) (Figure 1c). In the first, ATG8 is
conjugated to phosphatidyl ethanolamine (PE)
on both the inner and outer membranes of
the phagophore, proceeding through intermediates with the E1-like ATG7 and the E2-like
ATG3. This ATG8-PE conjugate is thought
to provide for membrane tethering and hemifusion, facilitating fusion of autophagosomes
to lysosomal and vacuolar membranes (65).
ATG8 fusion may also mediate targeting of
the autophagosomes to specific substrates, as
discussed above, and more generally dictate
the site of autophagosome formation (96). In
the second conjugation system, ATG5 is conjugated to ATG12, again relying on ATG7
E1-like activity, followed by ATG10 E2like processing. Further interaction of the
ATG5–12 complex with ATG16 enables localization to the phagophore, where together
these proteins promote ATG8-PE conjugation
and provide membrane curvature, enabling expansion and completion of the autophagosome
(22, 25, 61).
In plants, as discussed above, there has been
an expansion in many of the core components of
the two conjugations systems. The Arabidopsis
homologs from both systems retain their functions in vitro, suggesting that the fundamentals
of these conjugations systems are conserved in
plants as in yeast and mammals (21). Several
groups have isolated knockouts of individual
members of the conjugation systems, and there
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is evidence that in the absence of any of its individual components the conjugation system no
longer functions (28).
Knockouts of ATG4a/b, ATG5, ATG7,
ATG10, and ATG12 all have similar phenotypes, displaying stunted growth and early onset of senescence, and all have been described
as autophagy null (9, 18, 70, 85, 101). The evidence for elimination of autophagy in plants
has relied principally on detection of ATG8,
either in the form of the PE-conjugated adduct
or by the accumulation of ATG8 in the plant
vacuole. Adapted Western blotting assays initially showed an absence of the putative ATG8PE adduct in atg4a/b-1 and atg7–1 seedlings,
whereas the adduct was still present in atg5–
1 and atg10–1 seedlings (70, 101). Recently,
Chung et al. reported a less ambiguous assay
that shows the putative ATG8-PE adduct in
wild-type plants, but not in atg4a/b, atg5, atg7,
atg10, or atg12a/b seedlings (9). ATG8 vacuolar
accumulation assays (discussed below) similarly
show a failure of ATG8 to be delivered to the
plant vacuole in the presence of inhibitors of
vacuolar degradation in atg4a/b, atg5, atg7, and
atg10, suggesting that the ATG8 conjugation
system is no longer intact (70, 85, 101).
Although this cumulative evidence suggests
that knockouts in the two conjugation systems prevent conjugation of ATG8-PE and inhibit autophagy, can we assume that autophagy
no longer occurs at all in these plants? Recent results in other systems suggest that this
assumption may not be wise. In the mammalian system, Atg5 and Atg7 knockout mice
survive until the perinatal period before succumbing to lethality (41, 43). Although these
mice were thought to be autophagy knockouts,
Nishida et al. (66) recently reported that this
was not the case. By isolating embryonic fibroblasts from these mice, they show that autophagosomes can still form in Atg5−/− and
Atg7−/− cells, and that some autophagosomemediated protein degradation pathways are still
intact (66). Importantly, this held true despite the fact that LC3(ATG8)-PE conjugation
did not occur during Atg5/Atg7-independent
macroautophagy.
Some evidence is available among published
material suggesting that autophagy does indeed still occur, although to a lesser extent,
in conjugation knockout plants. For example,
autophagosomes appear to form in atg4a/b-1
plants, as detected by microscopic analysis
of green fluorescent protein (GFP)-tagged
ATG8e, ATG8a, and ATG8i (101). These vesicles appear despite the absence of the putative
ATG8-PE conjugate as detected by Western
blotting (101). However, clear distinction between autophagosomes and unconjugated aggregates in this and similar assays is lacking.
Autophagy-associated vacuolar aggregates are
also still present in atg5–3 mutants upon starvation, although again to a lesser degree than
in wild-type plants (33). Although these findings may be accounted for by nonautophagy
pathways, we note that the majority of the experiments conducted on the conjugation systems have been performed using roots to monitor autophagy in plants. Our own experiments
with GFP-tagged ATG8a in atg5–1 knockout
Arabidopsis suggest that autophagy is only
knocked down, not knocked out, in mature leaf
tissue (A. Hayward and S.P. Dinesh-Kumar,
unpublished data).
Autophagy knockouts in the conjugation
system are embryonic lethal in mammalian systems, and autophagy has clear roles in development in both lower and higher eukaryotes
(62). The lack of more striking phenotypes
for ATG gene knockouts in plants is a primary and puzzling exception. One possibility
is that alternative pathways exist in plants that
enable macroautophagy to occur at some (albeit repressed) level while bypassing the conjugation systems altogether. If this is indeed
the case, it might explain why knockouts of
core components in the ATG6 complex are
embryonic lethal, whereas knockouts in the
conjugation systems are not. Alternatively, it
may be that macroautophagy, by strict definition, is indeed null in these plants, whereas
other autophagy variants necessary for development, such as mitophagy or chlorophagy, remain unhindered. In support of this scenario,
we note that although the formation of ATG8+
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rubisco-containing bodies (RCBs) appears to be
autophagy-dependent, rubisco degradation itself is uncompromised in atg4a/b-1 plants (92).
In this case, a double membrane is already available in the form of the chloroplast envelope, and
only a subpopulation of autophagy proteins may
be required to transport and degrade rubisco.
RCB: rubiscocontaining body
Genetic Knockouts in the ATG9
Shuttling System
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Autophagy knockouts have also been described
within the ATG9 shuttling and membrane
delivery system (Figure 1a). For example,
atg9–1 knockout displays the same early senescence phenotype observed for knockouts in
the conjugation systems (26). However, atg9–1
plants have been reported not to be null for
autophagy (101). In fact, the atg9–1 line shows
an increase in both unconjugated ATG8 and
conjugated ATG8-PE in Western blot analyses
(101). Yoshimoto et al. (102) describe a slower
accumulation of autophagic bodies in atg9–1
roots, finally concluding that the autophagic
defect in these plants is leaky (101). Separately,
Inoue et al. (33) showed little compromise of
autophagy in atg9–2 mutants as measured by
accumulation of cytoplasmic material in the
plant vacuoles. atg2–1 mutants and ATG18atargeted RNAi plants, which should also be
impaired for membrane delivery, appear more
profoundly affected than the atg9 mutants,
although the latter still retain a small number of
putative autophagosomes as detected by monodansylcadaverine (MDC) staining (33, 97).
Are There Other Potential Sources
of Autophagy-Null Mutants?
Although it remains to be determined whether
plants are able to compensate for knockouts
in the conjugation and membrane delivery systems, there are two other core complexes that
may also provide avenues to an autophagy-null
system. Obtaining a knockout in the ATG1
core complex is hindered in Arabidopsis by the
presence of four putative genes. However,
initial results suggest that a double atg1
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Dinesh-Kumar
knockout is at least partially compromised for
autophagy (S. Patel and S.P. Dinesh-Kumar,
unpublished data). Alternatively, the ATG6
complex may be the best source of a knockout
for both macroautophagy and its many potential permutations. Targeting of this system is
complicated by the fact that both ATG6 and
PI3K knockouts are embryonic lethal (20, 27,
48, 68, 71). This embryo lethality, generally
attributed to nonautophagy functions for these
core proteins, might alternatively suggest
that these are the only true sources of an
autophagy-null system in plants, whereas the
other components can be drawn from redundant sources. Inducible on/off or silencing
systems targeting ATG6 and PI3K may enable
us to eventually differentiate these scenarios.
DO WE HAVE THE TOOLS
WE NEED TO STUDY
AUTOPHAGY IN PLANTS?
Do We Have the Markers We Need
to Monitor Autophagy?
As in other systems, the primary marker used
to monitor plant autophagy has been the
ATG8 protein. ATG8 is integrated into the
phagophore during the early stages of autophagosome formation, and it marks the autophagosome in untreated tissue until vacuolar
fusion (Figure 1c) (95). Although initial studies
used LysoTracker or MDC staining to identify
acidic autophagosomes, ATG8 has proven to
be a more reliable marker, with no potential for
cross identification of other acidic bodies in the
tissue (40).
Autophagosomes are short lived in the cell,
making quantification difficult (40, 103). Most
studies in plants have relied on tagging of
ATG8 in combination with inhibitors of vacuolar degradation (E64D or concanamycin-A) to
determine if autophagy is occurring. This combination has been useful in characterizing ATG
knockout lines, providing either an “on” or
“off ” phenotype for macroautophagy, but it is
only marginally informative for the study of intermediate up- or downregulation phenotypes.
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The plant autophagy field also has yet to fully
come to terms with the proliferation of ATG8
species in Arabidopsis (77, 81, 85). The nine homologs identified in Arabidopsis display differential expression, particularly during defense
responses, and as discussed above, LC3/ATG8
species are able to act as cargo specification
modules in other systems (28, 38, 77, 85). The
assumption that any given fluorescently tagged
ATG8 protein will suffice must be revisited.
More recently, several groups have taken a
more biochemical approach to autophagy quantification in plants, using modified Western
blotting assays to identify the ATG8-PE conjugate (9). This approach appears to complement
results obtained by microscopy but still relies
on either a single fluorescently tagged species
of ATG8 or, alternatively, an aggregation of
putative ATG8 species identified using ATG8
antibodies. Even if we can successfully differentiate ATG8 from the ATG8-PE adduct, the
ATG8 blotting assay cannot easily differentiate changes in synthesis from changes in degradation, and true monitoring of the ephemeral
autophagic flux remains elusive (74). ATG8 itself may be prone to aggregation, further complicating fluorescence-based assays (44). Some
groups have made inroads in plants by tagging
aggregation-prone proteins whose removal is
autophagy dependent (90), and in mammals
there is evidence that even more sophisticated autophagy monitoring systems can be
designed by tagging cargo specification modules like p62/SQSTM1 (46). The quantification
of autophagy-dependent aggregates in combination with ATG8 vacuolar aggregation and
ATG8-PE conjugation may provide a more accurate picture of the state of autophagy at any
given moment within the plant cell.
Can We Induce Autophagy in Plants?
In other systems, autophagy can be induced
by direct manipulation of the TOR pathway
(discussed above, Figure 1a). Treatment with
the antibiotic rapamycin inhibits TOR through
a tertiary complex involving a second protein,
FKBP12. Binding of rapamycin to FKBP12 and
TOR disrupts the TOR/RAPTOR complex,
mimicking conditions of nutrient starvation
and leading to induction of autophagy (17).
However, evolutionary changes to the FKBP12
homologs in Arabidopsis and other higher plants
have rendered them rapamycin insensitive. One
solution to this problem has been demonstrated
by Sormani et al. (83), who restored rapamycin
sensitivity in Arabidopsis by constitutive expression of the exogenous yeast ScFKBP12
protein. More recently, Liu & Bassham (55)
were able to avoid rapamycin altogether using
AtTOR-targeted RNAi lines. These lines
show constitutive induction of autophagy and
could serve as a model for inducible autophagy
induction lines in the future.
In the absence of chemical induction, most
papers in the plant field have relied on nutrient
starvation to induce autophagy. Starvation
is achieved by plating seeds on a nitrogendepleted medium, growing plants in darkness,
or by excising whole leaves (reviewed in 103).
Although these assays are appropriate for studying true phenomena of nutrient stress, which
are likely to be controlled through the TOR
pathway, they are clumsy as a general-purpose
means of induction. Effects are seen over an
extended timescale (many days), and the treatments are very likely to induce systemic changes
unrelated to autophagy itself. Furthermore,
although TOR appears to be a rather general
inducer of autophagy, there are many other
potential pathways to autophagy induction that
are modulated for specific phenomena (29).
Examples in the mammalian field include viraland bacterial-induced autophagy, which appear
to signal through protein kinase R or TLRs
and to directly activate proteins downstream
of TOR, including Beclin1 and ATG4 (29).
In the place of TOR inactivation, other
potential chemical inducers of autophagy may
be more appropriately used for nonstarvation
studies. For example, Xiong et al. (98) successfully induced autophagy using both hydrogen
peroxide (H2 O2 ) and methyl viologen to mimic
oxidative stress (98). These and other small
molecules, such as nitric oxide or salicylic
acid (SA), might also be excellent mimics of
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PCD: programmed
cell death
HR: hypersensitive
response
4 July 2011
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pathogen defense–related signaling. Pathogens
themselves, both viral and bacterial, have been
shown to induce autophagy (discussed below)
and could be used in a standardized infection
assay to monitor autophagy and its targets (32,
56, 68, 102).
IS AUTOPHAGY PRO-DEATH
OR PRO-SURVIVAL?
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Bcl-2, Type II Cell Death, and
Programmed Cell Death in Plants
The most outstanding example of the interplay
between death and survival pathways and autophagy was the discovery of an interaction between proapoptotic protein Bcl-2 and Beclin1
in mammals (69). This interaction was resolved
through a model in which the binding status of
Bcl-2 to Beclin1 served as a central determinant
of cell fate—be it either apoptotic (Type I) or
autophagic (Type II) cell death—with the two
fates being mutually exclusive (58). Initially, it
was not clear whether autophagic cell death
was a byproduct occurring as a result of inhibition of apoptosis or a true phenomenon. However, examples from developmental remodeling
Drosophila melanogaster and neuronal necrosis in
Caenorhabditis elegans provide strong evidence
for true autophagic cell death phenomena (3,
12, 89). Autophagy, although broadly a pathway
of survival, can in select circumstances double
as a pathway that promotes cell death (52).
In plants, the line between survival and death
remains blurred. Even differentiating would-be
autophagic cell death from apoptotic cell death
is not a simple task, as cells undergoing programmed cell death (PCD) display hallmarks of
both processes (reviewed in 45, 91). Hallmarks
of apoptotic cell death in plants include chromatin condensation, nuclear fragmentation,
and DNA laddering. However, caspase activity
(particularly caspase 3) is critical to apoptotic
death in animals, and a true caspase has yet to
be identified in the plant kingdom (64, 72). This
does not strictly preclude caspase-like cell death
pathways, as caspase inhibitors can inhibit hy-
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persensitive response (HR) PCD (discussed below) in plants (8). Very recently, Coll et al. (10)
showed that type I metacaspase AtMC1 has a
primary role in the execution of HR-PCD (10).
Despite these apoptotic characteristics,
PCD in plants also displays characteristics of
autophagic or Type II cell death, including
vacuolization and the appearance of doublemembrane vesicles, putatively autophagosomes
(64). A plant cell undergoing PCD will ultimately succumb to vacuolar and plasma
membrane collapse, separation of the plasma
membrane from the cell wall, and finally
leakage of the remaining cellular contents
into the apoplast (45, 91). This fate is more
characteristic of animal necrosis than apoptotic
or autophagic cell death (57).
Plant Autophagy is Pro-Survival
During Nutrient Stress
and Senescence
Autophagy is clearly upregulated during periods of starvation and darkness, as shown in
both recent publications and public microarray data (reviewed in 28). These data, when
coupled with a wealth of data from autophagy
gene knockouts that show a premature senescence phenotype (reviewed in 28, 103), logically dictate that autophagy promotes nutrient remobilization and tissue survival during
periods of stress. Autophagy gene knockouts
also show premature expression of senescence
markers, such as SEN1, SAG12, and PED1,
again directly suggesting autophagy staves off
the cell death (70, 85). Most recently, Phillips
et al. (70) showed definitive evidence of premature cell death by lactophenol blue staining and
DNA fragmentation analysis in atg10–1, atg5–
1, and atg7–1 plants compared to wild-type Arabidopsis during conditions of nutrient starvation.
Although less well studied, autophagy during
petal senescence seems to share the prosurvival
profile as do plant leaves (79, 99). Cumulatively,
these data show that autophagy is primarily
a pro-survival phenomenon during periods of
both nutrient stress and tissue senescence.
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Does Autophagy Promote Death or
Survival During HR-PCD?
In order to establish infection, plant pathogens
must first circumvent their host’s basal immune
response. A typical bacterial pathogen will
deliver 15–30 effector proteins into a host cell
via the type III secretion system, and these
effectors will act to inhibit host-pathogen
recognition (36). In response, plants have
evolved a specific set of resistance (R) genes
that encode intracellular immune receptor
proteins able to recognize effector activity.
The two predominant classes of R proteins
are the Toll-interleukin-1 receptor homologynucleotide
binding-leucine-rich
repeat
(TIR-NB-LRR) proteins and the coiled-coil
domain containing CC-NB-LRRs (reviewed
in 5). Upon effector recognition, R proteins
initiate a signaling cascade that culminates in
HR-PCD at the site of pathogen infection (36).
Effector-triggered immunity (ETI) signaling
includes a prolonged influx of Ca2+ , mitogenactivated protein kinase (MAPK)-dependent
production of reactive oxygen species (ROS),
and local production of SA (11, 82). The
ultimate induction of HR-PCD, which is
specific to ETI, is believed to restrict further
pathogen ingress.
Although autophagy appears to promote cell
survival during nutrient stress and senescence,
the role of autophagy during HR-PCD is still
an issue of some conjecture. In Liu et al. (56),
we provided the first evidence that autophagy
plays an important role in the restriction of
HR-PCD to sites of pathogen infection (56).
The execution of HR-PCD was uncompromised in Nicotiana benthamiana plants silenced
for NbATG6/Beclin1 when challenged with
the tobacco mosaic virus (TMV) pathogen.
However, after the HR-PCD lesion was established, the death associated with the lesion
spread beyond the lesion itself and into adjacent tissue. This runaway cell death eventually spread throughout the infected leaf and
even into upper uninfected leaves of the plant.
This result was not limited to ATG6/Beclin1
silencing, being recapitulated in VPS34/
PI3K-, ATG3-, and ATG7-silenced plants
(56).
This initial result was later confirmed in
Arabidopsis ATG6 RNAi lines (68). As in
N. benthamiana, HR-PCD induced by the avirulent pathogen Pseudomonas syringae pv. tomato
(Pst) DC3000 (avrRPM1) became unrestricted
in these plants, spreading beyond the site of
bacterial inoculation. In 2009, Yoshimoto et al.
confirmed these results in Arabidopsis atg5–1 genetic knockouts (102). Again using Pst DC3000
(avrRPM1), they showed that HR-PCD escapes
the site of bacterial infection and spreads into
adjacent tissue. Notably, this runaway cell death
is suppressed in double mutants for atg5 sid2 and
atg5 npr1 (discussed below) (102).
These results together seem to again suggest
a pro-survival role for autophagy, this time in
the inhibition of HR-PCD. However, Hofius
et al. (32) reported that cell death is suppressed
during the first 25 hours of Pst DC3000 (avrRPS4) infection of atg7–1 and atg9–1 knockout Arabidopsis (32). This result appeared to be
specific to TIR-NB-LRR immune receptor signaling, as there was no difference seen in the
CC-NB-LRR signaling triggered by AvrRpt2,
and the differential response to CC-NB-LRR
triggered by AvrRpm1 was muted compared to
the AvrRPS4 response (32).
We offer two hypotheses to explain these
results in their totality. First, there are alternative explanations as to why there is a delay (not
loss) in the induction of cell death in atg7–1 and
atg9–1 Arabidopsis lines that do not invoke autophagy as an executioner of cell death. Primary
among these is the fact that these lines show
gross disruption of cellular homeostasis, manifesting in rapid loss of proteins from the chloroplasts, mitochondria, and cytoplasm compared
to wild-type plants (70, 85). An important result
of this disruption of homeostasis appears to be a
more rapid induction of senescence-associated
death in atg5–1 and atg7–1 lines, as has been
described for other ATG genetic knockouts
(70). Although premature disruption of cellular organelles may prime plant tissue for senescence, it is not surprising that the opposite effect
would be seen in the case of HR-PCD. Here the
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ETI: effectortriggered immunity
ROS: reactive oxygen
species
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disruption of organelles that produce pro-death
signaling, especially the plant chloroplasts and
mitochondria (discussed below), would likely
impede the rapid induction of pro-death signals
necessary for PCD to occur. This slower induction of pro-death signals would yield the delayed cell death phenotype described by Hofius
et al. (32) in autophagy deficient atg5–1 and
atg7–1 plants (32).
We also note that the atg5–1 and atg7–1 lines
showed delays in HR-PCD induction on the
scale of hours, whereas there is no evidence that
ultimate formation of the HR-PCD lesion was
compromised. Drawing from recent studies in
mammalian autophagy (discussed above), these
autophagy-deficient plants may also be compromised for pathogen recognition and basal
immune responses (51). In fact, both TMV and
Pst DC3000 show increased replication in autophagy deficient backgrounds, suggesting failure of the host to recognize or clear initial infection (56, 68). This compromise of basal defense
signaling might be sufficient to delay (but not
inhibit) the induction of HR-PCD.
Alternatively, we also feel that these results are not necessarily contradictory. The
timescales at which the experiments are performed are markedly different, to the point
that data measured by Hofius et al. (32) rarely
overlaps that produced by Liu et al. (56) and
Yoshimoto et al. (102) (32, 56, 102). The
possibility exists that at the site of pathogen
infection, a marked induction of autophagy
indeed contributes to autophagic or type II
cell death. As the signaling associated with this
local induction spreads to neighboring tissue, it
likely becomes depleted. This depleted signaling would diminish the induction of autophagy
to a point that it is no longer sufficient to cause
cell death but instead assumes a cytoprotective
role in the elimination of pro-death signals that
would otherwise ultimately lead to runaway cell
death. These results would fit well with cumulative data obtained from recent studies of aging
and autophagy in animals, where both overactivation and inhibition of autophagy are sufficient
to promote neuronal degeneration (84).
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CAN WE DIFFERENTIATE
AUTOPHAGIC PHENOMENA
THROUGH THE SIGNALING
HAZE?
Differentiating Senescence and
HR-PCD Phenomena: The Enigma
of Salicylic Acid Signaling
Elevated SA accumulation is common in plants
displaying spontaneous death/lesion mimic
phenotypes (reviewed in 64). SA biosynthesis
also increases dramatically upon recognition of
a pathogen effector (reviewed in 24). However,
early studies have shown that plants deficient
in SA accumulation retain the ability to execute
HR-PCD, and it is likely that SA promotes cell
death in conjunction with other primary signals including ROS (63). During their experiments, Yoshimoto et al. (102) found that there
was a high accumulation of SA in atg5 mutants
compared to wild type (102). Furthermore, they
were able to eliminate the runaway cell death associated with infection of atg5 plants by crossing them to sid2 and npr1 knockouts, respectively deficient in SA synthesis and signaling
(102).
Hofius et al. (32) also show an association
between the induction of cell death and the accumulation of SA in their study. As discussed
above, they find that the contribution of autophagy to HR-PCD is most apparent during
infection with Pst DC3000 (avrRPS4), which is
detected in the plant by the TIR-NB-LRR immune receptor RPS4 (32). TIR-NB-LRR signaling is integrated through enhanced disease
susceptibility 1 (EDS1), leading the group to
hypothesize that the induction of autophagic
cell death is EDS1 dependent. In fact, eds1
knockout plants exhibit the same inhibited local cell death phenotype as shown for atg7–1
and atg9–1 plants (32). Like salicylic acid induction deficient-2 (SID2) and non-expressor
of pathogenesis-related 1 (NPR1), EDS1 is
also an amplifier of SA (94). Taken together,
these results suggest a dependency of both local
HR-PCD lesions and proximal runaway cell
death on SA signaling. SA also induces
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autophagy (A. Hayward and S.P. DineshKumar, unpublished data), explaining the apparent epistatic relationship between eds1 and
atg7/atg9 mutants described by Hofius et al.
(32). Ultimately, this SA-induced autophagy
would then suppress or eliminate further accumulation of SA, containing the HR-PCD lesion
to the infection site.
Interestingly, crossing atg5 to npr1 and sid2
mutants also suppressed the premature senescence phenotype shared between atg5 and many
other autophagy gene knockouts (102). The authors found that the runaway cell death phenotype of atg5 plants was age dependent, occurring only in older plants, and they proposed that
this age dependency is a result of accumulation
of SA in older leaves (103). Hofius et al. (32)
used relatively young plants for their experiments and were unable to produce a runaway
cell death phenotype. It remains to be determined if the age of the plants used explains the
absence of a runaway cell death phenotype in
atg7–1 and atg9–1 plants (32). Alternatively, it
is notable that atg9–1 plants may not be null for
autophagy, as discussed above (101). Sufficient
autophagic activity may remain in both atg7–1
and atg9–1 plants to prevent runaway cell death,
whereas atg5–1 knockouts and ATG6-targeted
RNAi plants are, in fact, further compromised
for autophagy.
These studies highlight the difficulty of
determining where HR-PCD ends and senescence signaling begins. Biosynthesis of SA
initially occurs at the sites of developing HRPCD lesions. However, following HR-PCD,
SA biosynthesis is also upregulated throughout
the plant and is essential for the establishment
of systemic acquired resistance (SAR) (reviewed
in 6). The establishment of SAR seems to temporally align with the escape of HR-PCD
from local lesions in both atg5 mutants and
ATG6/Beclin1-silenced N. benthamiana (56,
102). If this accumulation of SA during SAR is
sufficient to induce senescence programming,
this may further explain escape of cell death to
distal tissues described by Liu et al. (56).
The ultimate determination as to the nature of the runaway cell death in autophagy-
deficient plants may be impossible. As well
as SA signaling, there exists an abundance of
other shared signaling between senescence in
HR-PCD, including TOR signaling and upregulation of senescence-associated genes like
SAG1 (reviewed in 23). The death associated
with autophagy silencing may in fact be neither
senescence nor HR-PCD, but instead a SARinduced hybrid resulting from abnormal SA signaling in the absence of appropriate costimulation from other pathways. Determining the true
nature of the phenotype, if it is to occur, may
require determined phenotypic and expression
analysis of tissue as it undergoes PCD, and
HR-PCD and senescence themselves may have
to be concurrently revisited.
Systemic acquired
resistance (SAR):
primes defense
responses to prevent
future infection
Reactive Oxygen Species, Salicylic
Acid, and the Hypersensitive
Response Signaling Hub
ROS are highly diffusible secondary messenger
molecules. Although SA is one of the primary
signals during HR-PCD initiation, ROS are
also highly induced and may help transduce
pro-death signaling (reviewed in 6, 64, 88). A
primary ROS burst has been shown to occur immediately after effector recognition, followed
by a sustained period of upregulation, and
ectopic application of antioxidants can delay or
inhibit HR-PCD (37, 64). Importantly, the secondary oxidative bursts that occur at the site of
HR-PCD also occur in tissue distal to the site of
pathogen infection during the establishment of
SAR, which may be relevant to the occurrence
of runaway cell death (31, discussed above).
Autophagy may function to reduce these
pro-PCD signals in order to delimit HR lesions
and prevent runaway cell death. Autophagy
is upregulated in N. benthamiana plants adjacent to sites of pathogen infection, and
ATG6-silenced Arabidopsis show constitutive
upregulation of defense genes in the absence
of pathogens (56, 71). Autophagy is strongly
induced in response to ectopic application of
H2 O2 in Arabidopsis, and ATG18a-targeted
RNAi lines are hypersensitive to methyl
viologen–induced oxidative stress (98). We
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Nicotinamide
adenine dinucleotide
phosphate-oxidase
(NADPH): produces
superoxide during a
defense response
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similarly find that ectopic application of ROS
can induce autophagy in N. benthamiana and
is sufficient to induce runaway cell death in
autophagy-compromised plants (A. Hayward
and S.P. Dinesh-Kumar, unpublished data).
Finally, a dramatic accumulation of H2 O2 can
be detected in unchallenged atg5 and atg2
plants, as shown by diaminobenzidine (DAB)
staining (102). This mirrors the autophagydependent accumulation of SA found in these
same mutant lines. Cumulatively, these results
suggest that autophagy specifically suppresses
or dampens defense-related signaling.
How does autophagy limit these pro-death
signals? Given the highly diffusible nature of
these signals, particularly H2 O2 , it seems unlikely that the signals themselves are the targets
of autophagosomal degradation. Instead, it is
likely that autophagy targets the organellar
sources of SA and ROS signaling. Although
the membrane bound nicotinamide adenine
dinucleotide phosphate-oxidase (NADPH)
oxidase was once thought to be the primary
source of ROS during HR-PCD, recent results
show that an NADPH-derived superoxide may
actually inhibit PCD (28, 87, 88). Therefore,
although the initial burst may be triggered
by NADPH oxidase, it is likely that the
mitochondria, chloroplasts, and peroxisomes
sustain the prolonged secondary phase of ROS
upregulation during HR-PCD signaling in
plant cells. These organelles may even work in
concert, with evidence that mitochondria can
promote chloroplast-localized production of
SA (reviewed in 80).
Mitophagy and pexophagy are known to occur in mammals and yeast (reviewed in 73). In
plants, although these two processes are relatively unexplored, chlorophagy has been particularly well documented (28, 76). Chlorophagy
appears to take place through two distinct
mechanisms. First, a piecemeal autophagy of
the chloroplast occurs, which may specifically
target rubisco to the vacuole during senescence (7). These ATG8-tagged RCBs are disrupted in both atg5–1 and atg4a4b-1 mutants,
although rubisco can still be delivered to the
vacuole in these plants (34, 92). This piecemeal
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chlorophagy causes shrinkage of the chloroplast, ultimately leading the engulfment of the
entire organelle (92). As discussed previously,
the second of these two processes, but not
the first, appears to be disrupted in atg4a4b1 plants. This may suggest either a secondary
pathway to rubisco degradation, or, alternatively, an autophagy-dependent pathway that
does not require membrane delivery through
the two conjugation systems. Rubisco-targeted
autophagy may be a primary example of the potential for expansive and selective targeting of
autophagy in plants, as well as an unexplored
subcategory of specific degradation pathways.
CONCLUSION
Remarkable progress has been made in the
last few years, and the study of plant autophagy has clearly passed its infancy. However, specific needs must be met for the field
to reach adulthood, particularly at the intersection of autophagy and innate immunity. Primary among these needs is the establishment of
an autophagy-null system. Although this system
may already be in hand among the ATG8-PE
and ATG5–12–16 conjugation system knockouts, at the least more evidence is required to
support this conclusion with conviction. Alternatively, we find the ATG1 or ATG6 complexes
to be the most compelling source for future development of an autophagy null.
Similarly, in order to study autophagy in
plants we must continue to strive towards a reliable quantitative assay for measuring autophagy
induction. Analysis of ATG8-PE conjugation
by Western blotting is promising, but the
multiple bands associated with the conjugate
using current antibodies complicate the results
and can lead to subjective interpretation. We
are not alone in lamenting the absence of a true
“autophagometer,” as this problem persists in
every system in which autophagy is studied (74).
However, we have ample examples available
to expand the limited tools now available. The
field of plant autophagy has also focused heavily on roots, seedlings, and protoplasts when
quantifying autophagy phenotypes. Although
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these systems appear robust for the study
of some macroautophagy phenomena, many
primary processes, such as photosynthesis and
pathogen responsiveness, occur in the plant
leaf. We are concerned that some conclusions
drawn from the study of these specific structures may not prove applicable to the plant in
its entirety and are excited to see further studies
of autophagy in the plant leaves specifically.
Therefore, a robust quantitative autophagy
detection assay needs to be established in
mature leaf tissue.
ATG8 localization and conjugation studies
should continue to have a primary place in the
study of plant autophagy, but we hope that with
time the use of this marker can be refined. Further clarification of the individual roles of the
many ATG8 species in plants, which are currently a burden on plant studies, is likely to pay
off in the discovery of carefully regulated and
closely defined degradation pathways. These
pathways, undoubtedly exciting for plant autophagy, are also very likely to have implications in other eukaryotic systems.
SUMMARY POINTS
1. The molecular machinery of autophagy appears to be conserved in plants. Many individual ATG genes previously identified in yeast and mammals have expanded into gene
families in plants, and the implications of this expansion are still unknown.
2. Plant autophagy has important functions during nutrient stress, senescence, and response
to pathogens. Much of our core understanding of plant autophagy comes from T-DNA
insertion knockouts of individual ATG genes. These knockout plants display stunted
growth and premature senescence, although only knockouts of ATG6 complex proteins
are embryonic lethal.
3. As in other systems, autophagy can be monitored in plants by tagging the ATG8 protein, which labels autophagosomes. Although higher plants are insensitive to rapamycin,
autophagy can be induced by nutrient starvation, ectopic application of ROS, and by
bacterial and viral pathogens.
4. Autophagy is important to proper execution of the plant innate immune response.
Although the exact role of autophagy is still unknown, autophagy may limit pathogen
growth as part of a basal immune response, while also limiting cell death during
HR-PCD.
5. Caution must be applied when differentiating process-specific roles for autophagy from
systemic disruption of homeostasis.
6. Shared signaling occurs during the autophagy-regulated process of senescence and HRPCD, as well as during the establishment of SAR. This shared signaling may obfuscate
the true nature of autophagy-dependent phenotypes.
FUTURE ISSUES
1. Is plant autophagy regulated by TOR signaling, and if so, are there also other proteins
that regulate autophagy in response to specific stimuli?
2. Do the expanded plant ATG gene families, particularly the ATG8, ATG18, and ATG1
families, simply provide redundancy? Alternatively, what are the different roles of the
individual family members?
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3. Do genetic knockouts in the two ubiquitin-like conjugation systems provide autophagynull systems? Can whole-organelle or piecemeal autophagies of the mitochondria,
chloroplast, etc. persist in the absence of canonical macroautophagy?
4. How will we measure autophagic flux? Is a single ATG8 family member sufficient? Are
there aggregation-prone proteins that could be used to help quantify autophagy in plants?
5. Is autophagy pro-death or pro-survival during HR-PCD? Can it be both? Are cell death
pathways characterized with sufficient detail in plants to differentiate one type of cell
death from another?
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6. How does autophagy regulate pro-death/pro-senescence signaling? Are specific signals
or specific signal sources targeted, or is bulk degradation sufficient? If autophagy is being
targeted to specific proteins or organelles, how is this targeting achieved?
DISCLOSURE STATEMENT
The authors are not aware of any affiliations, memberships, funding, or financial holdings that
might be perceived as affecting the objectivity of this review.
ACKNOWLEDGMENTS
We thank Meenu Padmanabhan for critical reading of the manuscript. Innate immunity work in
S.P.D.-K. lab is supported by grants from NIH-GM062625 and NSF.
LITERATURE CITED
1. Anderson GH, Hanson MR. 2005. The Arabidopsis Mei2 homologue AML1 binds AtRaptor1B, the plant
homologue of a major regulator of eukaryotic cell growth. BMC Plant Biol. 5:2
2. Bassham DC, Laporte M, Marty F, Moriyasu Y, Ohsumi Y, et al. 2006. Autophagy in development and
stress responses of plants. Autophagy 2:2–11
3. Berry DL, Baehrecke EH. 2007. Growth arrest and autophagy are required for salivary gland cell degradation in Drosophila. Cell 131:1137–48
4. Blanchet FP, Moris A, Nikolic DS, Lehmann M, Cardinaud S, et al. 2010. Human immunodeficiency
virus-1 inhibition of immunoamphisomes in dendritic cells impairs early innate and adaptive immune
responses. Immunity 32:654–69
5. Caplan J, Padmanabhan M, Dinesh-Kumar SP. 2008. Plant NB-LRR immune receptors: from recognition to transcriptional reprogramming. Cell Host Microbe 3:126–35
6. Carr JP, Lewsey MG, Palukaitis P. 2010. Signaling in induced resistance. Adv. Virus Res. 76:57–121
7. Chiba A, Ishida H, Nishizawa NK, Makino A, Mae T. 2003. Exclusion of ribulose-1,5-bisphosphate
carboxylase/oxygenase from chloroplasts by specific bodies in naturally senescing leaves of wheat. Plant
Cell Physiol. 44:914–21
8. Chichkova NV, Kim SH, Titova ES, Kalkum M, Morozov VS, et al. 2004. A plant caspase-like protease
activated during the hypersensitive response. Plant Cell 16:157–71
9. Chung T, Phillips AR, Vierstra RD. 2010. ATG8 lipidation and ATG8-mediated autophagy in Arabidopsis
require ATG12 expressed from the differentially controlled ATG12A and ATG12B loci. Plant J. 62:483–
93
10. Coll NS, Vercammen D, Smidler A, Clover C, van Breusegem F, et al. 2010. Arabidopsis type I metacaspases control cell death. Science 330:1393–97
11. Dangl JL, Jones JD. 2001. Plant pathogens and integrated defence responses to infection. Nature
411:826–33
572
Hayward
·
Dinesh-Kumar
Annu. Rev. Phytopathol. 2011.49:557-576. Downloaded from www.annualreviews.org
by Helsinki University on 03/16/12. For personal use only.
PY49CH26-DineshKumar
ARI
4 July 2011
16:35
12. Denton D, Shravage B, Simin R, Mills K, Berry DL, et al. 2009. Autophagy, not apoptosis, is essential
for midgut cell death in Drosophila. Curr. Biol. 19:1741–46
13. Deprost D, Truong HN, Robaglia C, Meyer C. 2005. An Arabidopsis homolog of RAPTOR/KOG1 is
essential for early embryo development. Biochem. Biophys. Res. Commun. 326:844–50
14. Deprost D, Yao L, Sormani R, Moreau M, Leterreux G, et al. 2007. The Arabidopsis TOR kinase links
plant growth, yield, stress resistance and mRNA translation. EMBO Rep. 8:864–70
15. Deretic V. 2009. Multiple regulatory and effector roles of autophagy in immunity. Curr. Opin. Immunol.
21:53–62
16. Diaz-Troya S, Florencio FJ, Crespo JL. 2008. Target of rapamycin and LST8 proteins associate with
membranes from the endoplasmic reticulum in the unicellular green alga Chlamydomonas reinhardtii.
Eukaryot. Cell 7:212–22
17. Diaz-Troya S, Perez-Perez ME, Florencio FJ, Crespo JL. 2008. The role of TOR in autophagy regulation
from yeast to plants and mammals. Autophagy 4:851–65
18. Doelling JH, Walker JM, Friedman EM, Thompson AR, Vierstra RD. 2002. The APG8/12-activating
enzyme APG7 is required for proper nutrient recycling and senescence in Arabidopsis thaliana. J. Biol.
Chem. 277:33105–14
19. Farre JC, Mathewson RD, Manjithaya R, Subramani S. 2010. Roles of Pichia pastoris Uvrag in vacuolar
protein sorting and the phosphatidylinositol 3-kinase complex in phagophore elongation in autophagy
pathways. Autophagy 6:86–99
20. Fujiki Y, Yoshimoto K, Ohsumi Y. 2007. An Arabidopsis homolog of yeast ATG6/VPS30 is essential for
pollen germination. Plant Physiol. 143:1132–39
21. Fujioka Y, Noda NN, Fujii K, Yoshimoto K, Ohsumi Y, Inagaki F. 2008. In vitro reconstitution of plant
Atg8 and Atg12 conjugation systems essential for autophagy. J. Biol. Chem. 283:1921–28
22. Fujita N, Itoh T, Omori H, Fukuda M, Noda T, Yoshimori T. 2008. The Atg16L complex specifies the
site of LC3 lipidation for membrane biogenesis in autophagy. Mol. Biol. Cell 19:2092–100
23. Guiboileau A, Sormani R, Meyer C, Masclaux-Daubresse C. 2010. Senescence and death of plant organs:
nutrient recycling and developmental regulation. C R Biol. 333:382–91
24. Hammerschmidt R. 2009. Systemic acquired resistance. Adv. Bot. Res. 51:173–222
25. Hanada T, Noda NN, Satomi Y, Ichimura Y, Fujioka Y, et al. 2007. The Atg12-Atg5 conjugate has a
novel E3-like activity for protein lipidation in autophagy. J. Biol. Chem. 282:37298–302
26. Hanaoka H, Noda T, Shirano Y, Kato T, Hayashi H, et al. 2002. Leaf senescence and starvation-induced
chlorosis are accelerated by the disruption of an Arabidopsis autophagy gene. Plant Physiol. 129:1181–93
27. Harrison-Lowe NJ, Olsen LJ. 2008. Autophagy protein 6 (ATG6) is required for pollen germination in
Arabidopsis thaliana. Autophagy 4:338–48
28. Hayward AP, Tsao J, Dinesh-Kumar SP. 2009. Autophagy and plant innate immunity: defense through
degradation. Semin. Cell Dev. Biol. 20:1041–47
29. He C, Klionsky DJ. 2009. Regulation mechanisms and signaling pathways of autophagy. Annu. Rev.
Genet. 43:67–93
30. He C, Levine B. 2010. The Beclin 1 interactome. Curr. Opin. Cell Biol. 22:140–49
31. Heath MC. 2000. Hypersensitive response-related death. Plant Mol. Biol. 44:321–34
32. Hofius D, Schultz-Larsen T, Joensen J, Tsitsigiannis DI, Petersen NH, et al. 2009. Autophagic components contribute to hypersensitive cell death in Arabidopsis. Cell 137:773–83
33. Inoue Y, Suzuki T, Hattori M, Yoshimoto K, Ohsumi Y, Moriyasu Y. 2006. AtATG genes, homologs
of yeast autophagy genes, are involved in constitutive autophagy in Arabidopsis root tip cells. Plant Cell
Physiol. 47:1641–52
34. Ishida H, Yoshimoto K, Izumi M, Reisen D, Yano Y, et al. 2008. Mobilization of rubisco and stromalocalized fluorescent proteins of chloroplasts to the vacuole by an ATG gene-dependent autophagic
process. Plant Physiol. 148:142–55
35. Itakura E, Kishi C, Inoue K, Mizushima N. 2008. Beclin 1 forms two distinct phosphatidylinositol
3-kinase complexes with mammalian Atg14 and UVRAG. Mol. Biol. Cell 19:5360–72
36. Jones JD, Dangl JL. 2006. The plant immune system. Nature 444:323–29
www.annualreviews.org • Autophagy in Innate Immune Response
573
ARI
4 July 2011
16:35
37. Kiraly L, Hafez YM, Fodor J, Kiraly Z. 2008. Suppression of tobacco mosaic virus–induced
hypersensitive-type necrotization in tobacco at high temperature is associated with downregulation of
NADPH oxidase and superoxide and stimulation of dehydroascorbate reductase. J. Gen. Virol. 89:799–
808
38. Kirkin V, Lamark T, Johansen T, Dikic I. 2009. NBR1 cooperates with p62 in selective autophagy of
ubiquitinated targets. Autophagy 5:732–33
39. Kirkin V, Lamark T, Sou YS, Bjorkoy G, Nunn JL, et al. 2009. A role for NBR1 in autophagosomal
degradation of ubiquitinated substrates. Mol. Cell 33:505–16
40. Klionsky DJ, Abeliovich H, Agostinis P, Agrawal DK, Aliev G, et al. 2008. Guidelines for the use and
interpretation of assays for monitoring autophagy in higher eukaryotes. Autophagy 4:151–75
41. Komatsu M, Waguri S, Ueno T, Iwata J, Murata S, et al. 2005. Impairment of starvation-induced and
constitutive autophagy in Atg7-deficient mice. J. Cell Biol. 169:425–34
42. Kroemer G, Marino G, Levine B. 2010. Autophagy and the integrated stress response. Mol. Cell 40:280–
93
43. Kuma A, Hatano M, Matsui M, Yamamoto A, Nakaya H, et al. 2004. The role of autophagy during the
early neonatal starvation period. Nature 432:1032–36
44. Kuma A, Matsui M, Mizushima N. 2007. LC3, an autophagosome marker, can be incorporated into
protein aggregates independent of autophagy: caution in the interpretation of LC3 localization. Autophagy
3:323–28
45. Lam E. 2004. Controlled cell death, plant survival and development. Nat. Rev. Mol. Cell Biol. 5:305–15
46. Larsen KB, Lamark T, Overvatn A, Harneshaug I, Johansen T, Bjorkoy G. 2010. A reporter cell system
to monitor autophagy based on p62/SQSTM1. Autophagy 6:784–93
47. Lee HK, Mattei LM, Steinberg BE, Alberts P, Lee YH, et al. 2010. In vivo requirement for Atg5 in
antigen presentation by dendritic cells. Immunity 32:227–39
48. Lee Y, Bak G, Choi Y, Chuang WI, Cho HT. 2008. Roles of phosphatidylinositol 3-kinase in root hair
growth. Plant Physiol. 147:624–35
49. Leib DA, Alexander DE, Cox D, Yin J, Ferguson TA. 2009. Interaction of ICP34.5 with Beclin 1
modulates herpes simplex virus type 1 pathogenesis through control of CD4+ T-cell responses. J. Virol.
83:12164–71
50. Levine B. 2005. Eating oneself and uninvited guests: autophagy-related pathways in cellular defense. Cell
120:159–62
51. Levine B, Deretic V. 2007. Unveiling the roles of autophagy in innate and adaptive immunity. Nat. Rev.
Immunol. 7:767–77
52. Levine B, Kroemer G. 2009. Autophagy in aging, disease and death: the true identity of a cell death
impostor. Cell Death Differ. 16:1–2
53. Liang XH, Jackson S, Seaman M, Brown K, Kempkes B, et al. 1999. Induction of autophagy and inhibition
of tumorigenesis by beclin 1. Nature 402:672–76
54. Liang XH, Kleeman LK, Jiang HH, Gordon G, Goldman JE, et al. 1998. Protection against fatal Sindbis
virus encephalitis by beclin, a novel Bcl-2-interacting protein. J. Virol. 72:8586–96
55. Liu Y, Bassham DC. 2010. TOR is a negative regulator of autophagy in Arabidopsis thaliana. PLoS ONE
5:e11883
56. Liu Y, Schiff M, Czymmek K, Talloczy Z, Levine B, Dinesh-Kumar SP. 2005. Autophagy regulates
programmed cell death during the plant innate immune response. Cell 121:567–77
57. Lockshin RA, Zakeri Z. 2004. Apoptosis, autophagy, and more. Int. J. Biochem. Cell Biol. 36:2405–19
58. Maiuri MC, Le Toumelin G, Criollo A, Rain JC, Gautier F, et al. 2007. Functional and physical interaction between Bcl-X(L) and a BH3-like domain in Beclin-1. EMBO J. 26:2527–39
59. Menand B, Desnos T, Nussaume L, Berger F, Bouchez D, et al. 2002. Expression and disruption of the
Arabidopsis TOR (target of rapamycin) gene. Proc. Natl. Acad. Sci. USA 99:6422–27
60. Menzies FM, Moreau K, Rubinsztein DC. 2010. Protein misfolding disorders and macroautophagy.
Curr. Opin. Cell Biol. doi:10.1016/j.ceb.2010.10.010
61. Mizushima N, Kuma A, Kobayashi Y, Yamamoto A, Matsubae M, et al. 2003. Mouse Apg16L, a novel
WD-repeat protein, targets to the autophagic isolation membrane with the Apg12-Apg5 conjugate.
J. Cell Sci. 116:1679–88
Annu. Rev. Phytopathol. 2011.49:557-576. Downloaded from www.annualreviews.org
by Helsinki University on 03/16/12. For personal use only.
PY49CH26-DineshKumar
574
Hayward
·
Dinesh-Kumar
Annu. Rev. Phytopathol. 2011.49:557-576. Downloaded from www.annualreviews.org
by Helsinki University on 03/16/12. For personal use only.
PY49CH26-DineshKumar
ARI
4 July 2011
16:35
62. Mizushima N, Levine B. 2010. Autophagy in mammalian development and differentiation. Nat. Cell Biol.
12:823–30
63. Mur LA, Bi YM, Darby RM, Firek S, Draper J. 1997. Compromising early salicylic acid accumulation
delays the hypersensitive response and increases viral dispersal during lesion establishment in TMVinfected tobacco. Plant J. 12:1113–26
64. Mur LA, Kenton P, Lloyd AJ, Ougham H, Prats E. 2008. The hypersensitive response; the centenary is
upon us but how much do we know? J. Exp. Bot. 59:501–20
65. Nakatogawa H, Ichimura Y, Ohsumi Y. 2007. Atg8, a ubiquitin-like protein required for autophagosome
formation, mediates membrane tethering and hemifusion. Cell 130:165–78
66. Nishida Y, Arakawa S, Fujitani K, Yamaguchi H, Mizuta T, et al. 2009. Discovery of Atg5/Atg7independent alternative macroautophagy. Nature 461:654–58
67. Orvedahl A, Alexander D, Talloczy Z, Sun Q, Wei Y, et al. 2007. HSV-1 ICP34.5 confers neurovirulence
by targeting the Beclin 1 autophagy protein. Cell Host Microbe 1:23–35
68. Patel S, Dinesh-Kumar SP. 2008. Arabidopsis ATG6 is required to limit the pathogen-associated cell
death response. Autophagy 4:20–27
69. Pattingre S, Tassa A, Qu X, Garuti R, Liang XH, et al. 2005. Bcl-2 antiapoptotic proteins inhibit Beclin
1-dependent autophagy. Cell 122:927–39
70. Phillips AR, Suttangkakul A, Vierstra RD. 2008. The ATG12-conjugating enzyme ATG10 is essential
for autophagic vesicle formation in Arabidopsis thaliana. Genetics 178:1339–53
71. Qin G, Ma Z, Zhang L, Xing S, Hou X, et al. 2007. Arabidopsis AtBECLIN 1/AtAtg6/AtVps30 is essential
for pollen germination and plant development. Cell Res. 17:249–63
72. Reape TJ, McCabe PF. 2008. Apoptotic-like programmed cell death in plants. New Phytol. 180:13–26
73. Reumann S, Voitsekhovskaja O, Lillo C. 2010. From signal transduction to autophagy of plant cell
organelles: lessons from yeast and mammals and plant-specific features. Protoplasma 247:233–56
74. Rubinsztein DC, Cuervo AM, Ravikumar B, Sarkar S, Korolchuk V, et al. 2009. In search of an “autophagomometer”. Autophagy 5:585–89
75. Schmid D, Munz C. 2007. Innate and adaptive immunity through autophagy. Immunity 27:11–21
76. Seay M, Hayward AP, Tsao J, Dinesh-Kumar SP. 2009. Something old, something new: plant innate
immunity and autophagy. Curr. Top. Microbiol. Immunol. 335:287–306
77. Seay M, Patel S, Dinesh-Kumar SP. 2006. Autophagy and plant innate immunity. Cell. Microbiol. 8:899–
906
78. Shi CS, Kehrl JH. 2008. MyD88 and Trif target Beclin 1 to trigger autophagy in macrophages. J. Biol.
Chem. 283:33175–82
79. Shibuya K, Yamada T, Ichimura K. 2009. Autophagy regulates progression of programmed cell death
during petal senescence in Japanese morning glory. Autophagy 5:546–47
80. Singh DP, Moore CA, Gilliland A, Carr JP. 2004. Activation of multiple antiviral defence mechanisms
by salicylic acid. Mol. Plant Pathol. 5:57–63
81. Slavikova S, Shy G, Yao Y, Glozman R, Levanony H, et al. 2005. The autophagy-associated Atg8 gene
family operates both under favourable growth conditions and under starvation stresses in Arabidopsis
plants. J. Exp. Bot. 56:2839–49
82. Soosaar JL, Burch-Smith TM, Dinesh-Kumar SP. 2005. Mechanisms of plant resistance to viruses. Nat.
Rev. Microbiol. 3:789–98
83. Sormani R, Yao L, Menand B, Ennar N, Lecampion C, et al. 2007. Saccharomyces cerevisiae FKBP12
binds Arabidopsis thaliana TOR and its expression in plants leads to rapamycin susceptibility. BMC Plant
Biol. 7:26
84. Takacs-Vellai K, Bayci A, Vellai T. 2006. Autophagy in neuronal cell loss: a road to death. Bioessays
28:1126–31
85. Thompson AR, Doelling JH, Suttangkakul A, Vierstra RD. 2005. Autophagic nutrient recycling in
Arabidopsis directed by the ATG8 and ATG12 conjugation pathways. Plant Physiol. 138:2097–110
86. Thompson AR, Vierstra RD. 2005. Autophagic recycling: lessons from yeast help define the process in
plants. Curr. Opin. Plant Biol. 8:165–73
87. Torres MA, Jones JD, Dangl JL. 2005. Pathogen-induced, NADPH oxidase-derived reactive oxygen
intermediates suppress spread of cell death in Arabidopsis thaliana. Nat. Genet. 37:1130–34
www.annualreviews.org • Autophagy in Innate Immune Response
575
ARI
4 July 2011
16:35
88. Torres MA, Jones JD, Dangl JL. 2006. Reactive oxygen species signaling in response to pathogens. Plant
Physiol. 141:373–78
89. Toth ML, Simon P, Kovacs AL, Vellai T. 2007. Influence of autophagy genes on ion-channel-dependent
neuronal degeneration in Caenorhabditis elegans. J. Cell Sci. 120:1134–41
90. Toyooka K, Moriyasu Y, Goto Y, Takeuchi M, Fukuda H, Matsuoka K. 2006. Protein aggregates are
transported to vacuoles by a macroautophagic mechanism in nutrient-starved plant cells. Autophagy
2:96–106
91. van Doorn WG, Woltering EJ. 2005. Many ways to exit? Cell death categories in plants. Trends Plant
Sci. 10:117–22
92. Wada S, Ishida H, Izumi M, Yoshimoto K, Ohsumi Y, et al. 2009. Autophagy plays a role in chloroplast
degradation during senescence in individually darkened leaves. Plant Physiol. 149:885–93
93. Wang Y, Weiss LM, Orlofsky A. 2009. Host cell autophagy is induced by Toxoplasma gondii and contributes to parasite growth. J. Biol. Chem. 284:1694–701
94. Wiermer M, Feys BJ, Parker JE. 2005. Plant immunity: the EDS1 regulatory node. Curr. Opin. Plant
Biol. 8:383–89
95. Xie Z, Klionsky DJ. 2007. Autophagosome formation: core machinery and adaptations. Nat. Cell Biol.
9:1102–9
96. Xie Z, Nair U, Klionsky DJ. 2008. Atg8 controls phagophore expansion during autophagosome formation. Mol. Biol. Cell 19:3290–98
97. Xiong Y, Contento AL, Bassham DC. 2005. AtATG18a is required for the formation of autophagosomes
during nutrient stress and senescence in Arabidopsis thaliana. Plant J. 42:535–46
98. Xiong Y, Contento AL, Nguyen PQ, Bassham DC. 2007. Degradation of oxidized proteins by autophagy
during oxidative stress in Arabidopsis. Plant Physiol. 143:291–99
99. Yamada T, Ichimura K, Kanekatsu M, van Doorn WG. 2009. Homologs of genes associated with
programmed cell death in animal cells are differentially expressed during senescence of Ipomoea nil
petals. Plant Cell Physiol. 50:610–25
100. Yang Z, Klionsky DJ. 2010. Eaten alive: a history of macroautophagy. Nat. Cell Biol. 12:814–22
101. Yoshimoto K, Hanaoka H, Sato S, Kato T, Tabata S, et al. 2004. Processing of ATG8s, ubiquitin-like
proteins, and their deconjugation by ATG4s are essential for plant autophagy. Plant Cell 16:2967–83
102. Yoshimoto K, Jikumaru Y, Kamiya Y, Kusano M, Consonni C, et al. 2009. Autophagy negatively regulates
cell death by controlling NPR1-dependent salicylic acid signaling during senescence and the innate
immune response in Arabidopsis. Plant Cell 21:2914–27
103. Yoshimoto K, Takano Y, Sakai Y. 2010. Autophagy in plants and phytopathogens. FEBS Lett. 584:1350–
58
104. Yu L, Strandberg L, Lenardo MJ. 2008. The selectivity of autophagy and its role in cell death and
survival. Autophagy 4:567–73
Annu. Rev. Phytopathol. 2011.49:557-576. Downloaded from www.annualreviews.org
by Helsinki University on 03/16/12. For personal use only.
PY49CH26-DineshKumar
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Hayward
·
Dinesh-Kumar
PY49-FrontMatter
ARI
8 July 2011
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Contents
Volume 49, 2011
Not As They Seem
George Bruening p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1
Norman Borlaug: The Man I Worked With and Knew
Sanjaya Rajaram p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p17
Chris Lamb: A Visionary Leader in Plant Science
Richard A. Dixon p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p31
A Coevolutionary Framework for Managing Disease-Suppressive Soils
Linda L. Kinkel, Matthew G. Bakker, and Daniel C. Schlatter p p p p p p p p p p p p p p p p p p p p p p p p p p p47
A Successful Bacterial Coup d’État: How Rhodococcus fascians Redirects
Plant Development
Elisabeth Stes, Olivier M. Vandeputte, Mondher El Jaziri, Marcelle Holsters,
and Danny Vereecke p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p69
Application of High-Throughput DNA Sequencing in Phytopathology
David J. Studholme, Rachel H. Glover, and Neil Boonham p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p87
Aspergillus flavus
Saori Amaike and Nancy P. Keller p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 107
Cuticle Surface Coat of Plant-Parasitic Nematodes
Keith G. Davies and Rosane H.C. Curtis p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 135
Detection of Diseased Plants by Analysis of Volatile Organic
Compound Emission
R.M.C. Jansen, J. Wildt, I.F. Kappers, H.J. Bouwmeester, J.W. Hofstee,
and E.J. van Henten p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 157
Diverse Targets of Phytoplasma Effectors: From Plant Development
to Defense Against Insects
Akiko Sugio, Allyson M. MacLean, Heather N. Kingdom, Victoria M. Grieve,
R. Manimekalai, and Saskia A. Hogenhout p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175
Diversity of Puccinia striiformis on Cereals and Grasses
Mogens S. Hovmøller, Chris K. Sørensen, Stephanie Walter,
and Annemarie F. Justesen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 197
v
PY49-FrontMatter
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8 July 2011
9:55
Emerging Virus Diseases Transmitted by Whiteflies
Jesús Navas-Castillo, Elvira Fiallo-Olivé, and Sonia Sánchez-Campos p p p p p p p p p p p p p p p p p 219
Evolution and Population Genetics of Exotic and Re-Emerging
Pathogens: Novel Tools and Approaches
Niklaus J. Grünwald and Erica M. Goss p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 249
Evolution of Plant Pathogenesis in Pseudomonas syringae:
A Genomics Perspective
Heath E. O’Brien, Shalabh Thakur, and David S. Guttman p p p p p p p p p p p p p p p p p p p p p p p p p p p 269
Annu. Rev. Phytopathol. 2011.49:557-576. Downloaded from www.annualreviews.org
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Hidden Fungi, Emergent Properties: Endophytes and Microbiomes
Andrea Porras-Alfaro and Paul Bayman p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 291
Hormone Crosstalk in Plant Disease and Defense: More Than Just
JASMONATE-SALICYLATE Antagonism
Alexandre Robert-Seilaniantz, Murray Grant, and Jonathan D.G. Jones p p p p p p p p p p p p p 317
Plant-Parasite Coevolution: Bridging the Gap between Genetics
and Ecology
James K.M. Brown and Aurélien Tellier p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 345
Reactive Oxygen Species in Phytopathogenic Fungi: Signaling,
Development, and Disease
Jens Heller and Paul Tudzynski p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 369
Revision of the Nomenclature of the Differential Host-Pathogen
Interactions of Venturia inaequalis and Malus
Vincent G.M. Bus, Erik H.A. Rikkerink, Valérie Caffier, Charles-Eric Durel,
and Kim M. Plummer p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391
RNA-RNA Recombination in Plant Virus Replication and Evolution
Joanna Sztuba-Solińska, Anna Urbanowicz, Marek Figlerowicz,
and Jozef J. Bujarski p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 415
The Clavibacter michiganensis Subspecies: Molecular Investigation
of Gram-Positive Bacterial Plant Pathogens
Rudolf Eichenlaub and Karl-Heinz Gartemann p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 445
The Emergence of Ug99 Races of the Stem Rust Fungus is a Threat
to World Wheat Production
Ravi P. Singh, David P. Hodson, Julio Huerta-Espino, Yue Jin, Sridhar Bhavani,
Peter Njau, Sybil Herrera-Foessel, Pawan K. Singh, Sukhwinder Singh,
and Velu Govindan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 465
The Pathogen-Actin Connection: A Platform for Defense
Signaling in Plants
Brad Day, Jessica L. Henty, Katie J. Porter, and Christopher J. Staiger p p p p p p p p p p p p p p p 483
vi
Contents
PY49-FrontMatter
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Understanding and Exploiting Late Blight Resistance in the Age
of Effectors
Vivianne G.A.A. Vleeshouwers, Sylvain Raffaele, Jack H. Vossen, Nicolas Champouret,
Ricardo Oliva, Maria E. Segretin, Hendrik Rietman, Liliana M. Cano,
Anoma Lokossou, Geert Kessel, Mathieu A. Pel, and Sophien Kamoun p p p p p p p p p p p p p p p 507
Water Relations in the Interaction of Foliar Bacterial Pathogens
with Plants
Gwyn A. Beattie p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 533
Annu. Rev. Phytopathol. 2011.49:557-576. Downloaded from www.annualreviews.org
by Helsinki University on 03/16/12. For personal use only.
What Can Plant Autophagy Do for an Innate Immune Response?
Andrew P. Hayward and S.P. Dinesh-Kumar p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 557
Errata
An online log of corrections to Annual Review of Phytopathology articles may be found at
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Contents
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