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Critical Reviews in Biotechnology
ISSN: 0738-8551 (Print) 1549-7801 (Online) Journal homepage: http://www.tandfonline.com/loi/ibty20
Plant protein phosphatases 2C: from genomic
diversity to functional multiplicity and importance
in stress management
Amarjeet Singh, Amita Pandey, Ashish K. Srivastava, Lam-Son Phan Tran &
Girdhar K. Pandey
To cite this article: Amarjeet Singh, Amita Pandey, Ashish K. Srivastava, Lam-Son Phan Tran &
Girdhar K. Pandey (2015): Plant protein phosphatases 2C: from genomic diversity to functional
multiplicity and importance in stress management, Critical Reviews in Biotechnology, DOI:
10.3109/07388551.2015.1083941
To link to this article: http://dx.doi.org/10.3109/07388551.2015.1083941
Published online: 18 Sep 2015.
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ISSN: 0738-8551 (print), 1549-7801 (electronic)
Crit Rev Biotechnol, Early Online: 1–13
!2015 Taylor & Francis. DOI: 10.3109/07388551.2015.1083941
REVIEW ARTICLE
Plant protein phosphatases 2C: from genomic diversity to functional
multiplicity and importance in stress management
Amarjeet Singh
1
, Amita Pandey
1
, Ashish K. Srivastava
2
, Lam-Son Phan Tran
3
, and Girdhar K. Pandey
1
1
Department of Plant Molecular Biology, University of Delhi South Campus, Benito Juarez Road, Dhaula Kuan, New Delhi, India, ,
2
Nuclear
Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Trombay, Mumbai, Maharashtra, India, and ,
3
Signaling Pathway
Research Unit, RIKEN Center for Sustainable Resource Science, Tsurumi, Yokohama, Kanagawa, Japan
Abstract
Protein phosphatases (PPs) counteract kinases in reversible phosphorylation events during
numerous signal transduction pathways in eukaryotes. Type 2C PPs (PP2Cs) represent the major
group of PPs in plants, and recent discovery of novel abscisic acid (ABA) receptors (ABARs) has
placed the PP2Cs at the center stage of the major signaling pathway regulating plant responses
to stresses and plant development. Several studies have provided deep insight into vital roles
of the PP2Cs in various plant processes. Global analyses of the PP2C gene family in model
plants have contributed to our understanding of their genomic diversity and conservation,
across plant species. In this review, we discuss the genomic and structural accounts of PP2Cs in
plants. Recent advancements in their interaction paradigm with ABARs and sucrose
nonfermenting related kinases 2 (SnRK2s) in ABA signaling are also highlighted. In addition,
expression analyses and important roles of PP2Cs in the regulation of biotic and abiotic stress
responses, potassium (K
+
) deficiency signaling, plant immunity and development are
elaborated. Knowledge of functional roles of specific PP2Cs could be exploited for the genetic
manipulation of crop plants. Genetic engineering using PP2C genes could provide great
impetus in the agricultural biotechnology sector in terms of imparting desired traits, including a
higher degree of stress tolerance and productivity without a yield penalty.
Keywords
Abscisic acid, ABA receptors, abiotic stress,
development, expression, protein
phosphatase 2C, potassium deficiency,
signaling
History
Received 14 March 2015
Revised 27 June 2015
Accepted 4 July 2015
Published online 16 September 2015
Introduction
Reversible protein phosphorylation is recognized as one of the
key events in various signal transduction networks, which
regulates numerous biological processes in eukaryotes (Uhrig
et al., 2013). Protein kinases (PKs) and phosphatases (PPs)
are the key players that mediate these events and maintain the
balance of phosphorylation status of a cell during normal and
adverse conditions (Singh & Pandey, 2012). For a long period
of time, the major attention has been given to PKs in terms of
their structure, function and evolutionary relationship in
eukaryotes, whereas PPs are scarcely studied (Brautigan,
2013). Interestingly, among eukaryotes, plants contain a
higher number of kinase-encoding genes than any other
organisms. For example, rice (Oryza sativa) and Arabidopsis
thaliana genomes are reported to have 1500 (Lehti-Shiu &
Han-Shiu, 2012 ) and 1000 kinases (Zulawski et al., 2014),
respectively. In contrast, the yeast (Saccharomyces cerevisiae)
genome has only 130 kinases. Similarly, mouse (Mus
musculus) and human (Homo sapiens) genomes also encodes
about 500 PKs (Daigle et al., 2014; Sadowski et al., 2013).
Plant PPs are broadly categorized into serine/threonine
(Ser/Thr) phosphatases and tyrosine (Tyr) phosphatases based
on the amino acid residue they dephosphorylate. Molecular,
biochemical and genomic analyses enabled us to further
classify Ser/Thr phosphatases into two major families: (i)
phosphoprotein phosphatases (PPs) comprising PP1, PP2A,
PP2B and other distantly related phosphatases (PP4, PP5, PP6
and PP7) and (ii) PPs requiring metal ion (PPM) for catalysis,
including PP2C and other Mg
2+
-dependent phosphatases (Lee
et al., 2010; Singh et al., 2010). However, PP2B, a Ca
2+
-
dependent phosphatase, also known as calcineurin A (CNA),
is not identified so far in plants (Uhrig et al., 2013).
Similarly, plant protein Tyr phosphatases (PTPs) are also
divided into Tyr-specific PTPs and dual specificity PTPs
(DSPs), the later can dephosphorylate phosphotyrosine, as
well as phosphoserine/phosphothreonine (Bentem & Hirt,
2009).
In the post-genomic era, due to the availability of complete
sequences of several animals and plant genomes, PPs are
explored at the global level. The genome-wide analyses
revealed that the human genome possesses a higher number of
PTPs than any plant species (Kerk et al., 2008). On the other
Address for correspondence: Girdhar K. Pandey, Department of Plant
Molecular Biology, University of Delhi South Campus, Benito Juarez
Road, Dhaula Kuan, New Delhi, 110021 India. E-mail: gkpandey@
south.du.ac.in
Lam-Son Phan Tran, Signaling Pathway Research Unit, RIKEN Center
for Sustainable Resource Science, Tsurumi, Yokohama, Kanagawa 230-
0045, Japan. E-mail: son.tran@riken.jp
Downloaded by [University of Delhi] at 23:39 20 September 2015
hand, plant genomes, such as those of model plants, including
Arabidopsis and rice, encode PP2Cs as a major phosphatase
group, representing 60–65% of the total phosphatase reper-
toire (Kerk et al., 2008; Singh et al., 2010). The high
proportion of PP2C genes is indicative of their evolutionary
significance, requirement and involvement in diverse cellular
plant functions. In line with this hypothesis, most of the
molecular, biochemical, genetic and functional analyses are
carried out for the PP2C group of plant phosphatases, and
many of the PP2Cs are shown to regulate diverse aspects of
plant physiology (Bhaskara et al., 2012; Fuchs et al., 2013).
In this review, we are providing an overview of the
genomic organization of the PP2C gene family in plants.
Their phylogenetic and evolutionary relationship and expres-
sion patterns, as well as their involvement in different plant
processes, such as ABA signaling, biotic and abiotic stress
responses, plant immunity, K
+
nutrient signaling and plant
development, will be discussed in order to comprehend the
diverse function of PP2C in plants.
PP2C family in plants
The whole complement of PP encoding genes has been
explored in different plant species. The Arabidopsis genome
is reported to encode 126 PPs (Kerk et al., 2008), while 132,
113 and 102 PP-encoding genes are identified in rice, tomato
(Solanum lycopersicum) and hot pepper (Capsicum annuum)
genomes, respectively (Kim et al., 2014; Singh et al., 2010).
This set of PPs comprises different groups, such as PPPs
(including PP1, PP2A), PP2Cs, Tyr-specific PTPs, DSPs and
low-molecular-weight PTPs (LMWP). The PP2Cs represent a
major group of the PP-encoding gene family, with a total of
80, 90, 91 and 88 PP2C genes identified in the Arabidopsis,
rice, tomato and hot pepper genome, respectively (Kim et al.,
2014; Singh et al., 2010). In contrast, the human genome
possesses only 16 PP2C genes that code for at least 22
different isoforms (Shi, 2009). Interestingly, lower plants,
including green alga (Chlamydomonas reinhardtii), moss
(Physcomitrella patens) and lycophyte (Selaginella moellen-
dorffii), have a genome size comparable to that of higher
plants, such as Arabidopsis and rice, but the number of PP2C
genes varies from 10 in green alga, 50 in moss and lycophyte
to 80–130 in higher plants such as Arabidopsis, rice and
maize (Zea mays) (Fuchs et al., 2013). Therefore, the
expansion and diversification of the PP2C gene family
could be correlated with evolution of plants from unicellular
to multicellular organisms.
The PP2C family is further divided into 11 subclades
(A–K) in rice and Arabidopsis (Singh et al., 2010), and 13
subgroups in tomato and hot pepper (Kim et al., 2014). In
phylogenetic analyses, different PP groups from Arabidopsis
and rice were shown to be aligned together to form a common
clade, which indicates that the PPs have sequence conserva-
tion, common ancestors and similar evolutionary lineage. The
large extent of division within a group is possibly due to
variations in some specific amino acids or sequences that
might be responsible for specific interaction with target
proteins and functional variability among different PP2C
subgroups. Moreover, gene (intron–exon) structure analysis of
the PP2C families in different plant species, such as
Arabidopsis, rice and maize, supported their structural
conservation. The number of introns in Arabidopsis (Xue
et al., 2008) and maize (Wei & Pan, 2014) PP2Cs ranges
from 0 to 12, whereas rice PP2Cs genes contain 0–18 introns
(Singh et al., 2010; Xue et al., 2008). Members of the same
subgroup share a similar intron–exon structure and gene
length, which indicates their close evolutionary relationship
and gene structure conservation across different plant species.
In addition, a tremendous amount of chromosomal duplica-
tion was detected, both in Arabidopsis and in rice PP2C gene
families, suggesting that duplication contributed to the
expansion and evolution of PP2Cs in plants (Singh et al.,
2010; Xue et al., 2008). Moreover, in rice, besides genomic
diversity, it was observed that chromosomal duplication,
has also led to functional diversification of the PP2C gene
family, as some of the duplicated gene pairs showed
expression retention, while other duplicated pairs exhibited
neo-functionalization and pseudofunctionalization (Singh
et al., 2010).
Sequence and structure of plant PP2Cs
Domain structure analyses of eukaryotic PP2Cs revealed that
the catalytic domain location is variable, and it could be
present either at the N- or C-terminus (Schweighofer et al.,
2004). In Arabidopsis, a high proportion of PP2Cs (58%)
contain catalytic domain at the C-terminus, with different N-
terminal extensions. Some Arabidopsis PP2Cs, especially
those belonging to group F, start with the N-terminal catalytic
domain (Schweighofer et al., 2004). However, the presence
of 11 characteristic subdomains in the catalytic part of all the
eukaryotic PP2Cs is a common feature (Su & Forchhammer,
2013). The domain structures of typical plant PP2C members
are depicted in Figure 1. The structural variability possibly
contributes to their regulatory and functional diversities.
Detail sequence analysis revealed the presence of some
unique domains and motifs in plant PP2C proteins.
For instance, a mitogen-activated protein kinase (MAPK)
interaction motif known as kinase interaction motif (KIM)
[(K/R)
3–4
X
1–6
(L/I)X(L/I)] has been identified in several plant
PP2Cs, which is similar to one found in animal MAPK
kinases (MAPKKs) or MAPK phosphatases (Schweighofer
et al., 2007). This domain is present in most Arabidopsis
group-B PP2C members, including AP2C1 (ARABIDOPSIS
PP2C-TYPE PHOSPHATASE1), AP2C2, AP2C4 and
AP2C3/AtPP2C5 (Umbrasaite et al., 2010). Remarkably,
KIM has been found in PTPs of most organisms, but in
plants, PP2Cs have been evolved with KIM, representing a
point of evolutionary convergence of PP2Cs and PTPs.
Increasing evidence suggests that KIM is essential for
PP2Cs functional activity in regulating MAPK signaling in
plants (Schweighofer & Meskiene, 2008). A specific
Arabidopsis PP2C, kinase-associated protein phosphatase
(KAPP), contains three unique functional domains: (i) type
I membrane anchor, (ii) receptor-like kinase (RLK) associ-
ation domain and (iii) a catalytic domain. At the N-terminus
of the kinase interaction region, a forkhead-associated
homology region is also present which is vital for attachment
with phosphorylated target proteins and therefore helps in
signal transduction (Schweighofer et al., 2004).
2A. Singh et al., Crit Rev Biotechnol, Early Online: 1–13
Downloaded by [University of Delhi] at 23:39 20 September 2015
PP2Cs in ABA signaling module
PP2Cs have been recognized as important regulators of ABA
signaling. However, their exact position and regulatory
mechanism in ABA-signaling pathway were largely unknown
until the recent discovery of novel ABA receptors (ABARs)
in plants. In the breakthrough discovery, 14 members of
pyrabactin resistance (PYR)/pyrabactin-like (PYL)/regulatory
components of the ABA receptor (RCAR) family of
steroidogenic acute regulatory protein (StAR)-related lipid
transfer (START) proteins were identified in Arabidopsis
genome as ABARs (Fujii et al., 2009; Ma et al., 2009; Park
et al., 2009).
After their discovery, a number of reports have provided
details about the structure and interaction mechanisms of
novel ABARs to inhibit PP2Cs in ABA-signaling pathway.
Structural analyses have provided insight into dynamics of
ABA binding to the receptors. The conformational change in
bloops of ABA-free and ABA-bound RCAR proteins was
found as the major event in ABA perception and signal
transduction (Nishimura et al., 2009; Santiago et al., 2009;
Shibata et al., 2010; Sun et al., 2012). Melcher et al. (2009)
provided more refined and in-depth mechanistic details and
revealed a unique gate-latch-lock mechanism in ABA/RCAR
signaling. It was shown that ABA-free (apo) receptors are
featured with an open ligand-binding pocket safeguarded by a
gate that is closed upon ABA binding due to the conform-
ational change of highly conserved bloops. Closure of gate in
the presence of ABA provides a surface on receptor to interact
with a conserved tryptophan (Trp) residue in PP2C and
block PP2C-active site. The gain-of-function mutation in
the critical Trp residue (W385A) of the PP2C HAB1
[HYPERSENSITIVE TO ABA 1] abolishes ABA/PYR-
dependent inhibition of PP2C. Consequently, it leads to
constitutive suppression of OST1 (OPEN STOMATA 1) PK
even in ABA availability, thereby resulting in dominant ABA
insensitivity in vivo (Dupeux et al., 2011). Interestingly,
PP2Cs were found to have direct physical interaction with
ABA in a complex with PYR/RCAR (Miyazono et al., 2009;
Yin et al., 2009), and the conserved Trp residue of group A
PP2Cs was responsible for this interaction (Santiago et al.,
2012). Therefore, the key Trp residue could be targeted for
genetic engineering to generate the dominant group-A PP2Cs
that can bypass the ABA/PYR interactions and lead to ABA
insensitive responses. These findings provide deep insight
into the molecular mechanism of PP2C interaction with
PYR/PYL/RCAR in ABA perception and signaling.
In vitro reconstitution analysis of Arabidopsis ABA-
signaling pathway revealed that except PYL13, all the other
members of PYR/PYL/RCAR family are functional ABARs
(Fujii et al., 2009). Structural analyses showed a highly
conserved lysine residue crucial for ABA binding in all the
ABARs, except PYL13. This possibly accounts for the
nonfunctional behavior of PYL13 (Miyazono et al., 2009;
Nishimura et al., 2009; Yin et al., 2009). However, a recent
study suggested that through mutation in the key residues Q38
in conserved loop 1 (CL1), F71 in CL2 and T135 in CL4 (Yin
et al., 2009) of PYL13, it could be converted into a partially
functional ABAR (Zhao et al., 2013). Furthermore, PYL13
could interact with some group A PP2Cs and inhibit them in
ABA-independent manner (Li et al., 2013; Zhao et al.,
2013). Interestingly, PYL13 also interacts with other PYLs
and perturbs their ABAR activity (Zhao et al., 2013). In
addition, recent report by Fuchs et al. (2014) showed that
RCAR7/PYL13 could regulate the phosphatase activity of
ABI1 (ABA insensitive 1), ABI2 and PP2CA/AHG3 (ABA
hypersensitive germination 3) in vitro. Ectopic expression of
RCAR7/PYL13 led to ABA hypersensitivity and changes in
gene expression, seed germination and stomatal closure.
Thus, RCAR7/PYL13 was proved to function as an ABAR
during early plant development.
Among Arabidopsis ABARs and PP2Cs, different
functional interactions have been reported. PYR1, PYL1,
PYL2 and PYL3 inhibit group-A PP2Cs, including ABI1,
HAB1, HAB2 and PP2CA, in the presence of ABA.
Interestingly, PYL5, PYL6, PYL8, PYL9 and PYL10
inhibit different PP2Cs, even in ABA-independent manner
NC
HsPP2C
ABI1
PP2Cc
PP2Cc
AP2C1
PP2Cc
KIM
POL
KAPP
PP2Cc
PP2Cc
FHA
PP2Cc OsPP2C76
Ser/Thr kinase
Figure 1. Schematic representation of PP2C protein structure. Domain organization of typical PP2C candidates from human and plant species is
shown. HsPP2C is a human PP2C; ABI1 (ABA insensitive 1), AP2C1 (Arabidopsis PP2C-type phosphatase 1), POL (POLTERGEIST) and KAPP
(kinase-associated protein phosphatase) are the well-studied Arabidopsis PP2Cs, whereas OsPP2C76 (Oryza sativa PP2C76) is a unique rice PP2C
containing both catalytic PP2C and Ser/Thr kinase domains. Pink color bar represents a noncatalytic domain; orange bar in KAPP represents putative
MAPK (mitogen-activated protein kinase) docking site. PP2Cc, catalytic PP2C domain; KIM, kinase interaction motif; FHA, forkhead-associated
domain.
DOI: 10.3109/07388551.2015.1083941 Plant protein phosphatases 2C 3
Downloaded by [University of Delhi] at 23:39 20 September 2015
(Antoni et al., 2013; Hao et al., 2011; Miyakawa et al.,
2013; Park et al., 2009). Strikingly, PYL4 inhibits the activity
of HAB2 exclusively in the absence of ABA. Due to
difficulties in recombinant protein expression and purifica-
tion, some other ABARs, such as PYL7, PYL11 and PYL12,
could not be analyzed for their interaction with PP2Cs (Hao
et al., 2011; Miyakawa et al., 2013; Zhang et al., 2012).
Besides Arabidopsis, ABARs and PP2Cs interactions have
been reported in other important plant species, such as rice
(Kim et al., 2012), tomato (Sun et al., 2011), soybean
(Glycine max) (Bai et al., 2013), barley (Hordeum vulgare)
(Seiler et al., 2014) and maize (Wang et al., 2014).
In ABA-signaling pathways, SnRK2 Ser/Thr kinases are
other critical components, functioning downstream of PP2Cs.
Under normal conditions (the absence of ABA), PP2C binds
and blocks the SnRK2 kinase domain, rendering Ser/Thr
kinase inactive. A complete overview of ABA signaling in
plants is depicted in Figure 2. The perception of stress or
developmental stimulus results in ABA production, and ABA-
bound receptors interact with PP2Cs to remove inhibitors of
SnRK2s (Fujii et al., 2009; Umezawa et al., 2009). Auto- or
trans-activated kinase, through phosphorylation, regulates
various downstream components, such as transcription factors,
including ABF, ABI5 and WRKY in the nucleus, to induce
gene expression (Antoni et al., 2011; Cutler et al., 2010;
Miyakawa et al., 2013). On the other hand, in the guard cells,
SnRK2s activate slow anion channel-associated 1 (SLAC1)
and inhibits potassium channel in Arabidopsis thaliana
(KAT1), causing stomatal closure (Miyakawa et al., 2013).
Recent structural studies provided the critical mechanistic
details of SnRK2s regulation by PP2Cs. Two group-A
PP2Cs, ABI1 and HAB1, were found to target and depho-
sphorylate pSer176 in the activation loop of OST1 (SnRK2.6)
kinase. The pSer176 is a critical residue, essential for OST1
autoactivation. After release of PP2C inhibition, conform-
ation changes occur in the activation loop of SnRK2 and lead
to its autoactivation (Ng et al., 2011; Xie et al., 2012; Yunta
et al., 2011). The SnRK2 activation loop docks into the
PP2C-active site, while conserved Trp residue of PP2C is
inserted into the catalytic cleft of SnRK2. Therefore, a
molecular mimicry exists between ABARs and SnRK2s in
binding with PP2Cs, as both utilize a gate-latch-lock mech-
anism (Soon et al., 2012). This mechanism directly couples
ABA binding to SnRK2 activation and thus presents a new
regulatory paradigm in ABA signaling through a kinase–
phosphatase pair. The available lines of evidence indicate that
the ABA-ABAR-PP2C-SnRK2 module forms the core func-
tional unit of ABA-signaling pathway, which is evolutionarily
conserved across different plant species. These components of
ABA signaling interact in different combinations and regulate
their target proteins with varying affinities. Thereby, ABA
and these proteins control, modulate and fine-tune various
stresses and developmental responses, according to the need
of plants in diverse environmental conditions.
Diverse functional role of PP2Cs in plants
PP2Cs in abiotic stress signaling and responses
PP2Cs are important signal transduction components in
different organisms, including human and plants. Over the
last 10–15 years, accumulating evidence has suggested a
significant role for PP2Cs in controlling cellular stress
Figure 2. ABA-ABAR-PP2C module in
plants. Upon perception of stress or devel-
opmental stimulus, ABA (abscisic acid) is
produced in the cell and then it binds and
activates ABAR (ABA receptor). Activated
ABARs interact with PP2Cs either in cyto-
plasm or in nucleus. This interaction leads to
removal of PP2C inhibition from SnRK2
(sucrose nonfermenting related kinase 2),
resulting in SnRK2 activation. Activated
kinase in turn activates downstream compo-
nents, such as ABF/AREB/ABI5 transcrip-
tion factors, which bind to ABRE (ABA-
responsive element) in ABA-responsive
genes and induces gene expression. This
ultimately leads to adaptive physiological
responses, including regulation of growth and
development and stress tolerance.
Simultaneously, SnRK2 regulates some other
components, such as ion channels (SLAC1;
slow anion channel associated 1) and trans-
porters (KAT1; potassium channel in
Arabidopsis thaliana 1), which participate in
ion transport, membrane potential mainten-
ance and stomatal movement.
Cytoplasm
Ser/Thr kinase
(SnRK2) Ser/Thr kinase
(SnRK2)
Nucleus
ABF/AREB/
ABI5
Anion
ABRE Gene
epression
K+
Responses
Growth regulaon and stress tolerance
KAT1
ABARABAR
ABA
PP2C
ABAR
PP2C
P
SLAC1
P
P
4A. Singh et al., Crit Rev Biotechnol, Early Online: 1–13
Downloaded by [University of Delhi] at 23:39 20 September 2015
signaling (Bhaskara et al., 2012; Geiger et al., 2010; Lowe
et al., 2012). Understanding the transcript regulation under
different abiotic stress conditions is critical, as it would
provide a significant clue about their functional role.
Expression analyses in model plants, such as Arabidopsis,
rice, maize and tomato, have revealed that group-A PP2C
genes are highly inducible in response to ABA and different
abiotic stresses, including drought, osmotic, salinity and cold
(Singh et al., 2010; Sun et al., 2011; Wei & Pan, 2014; Xue
et al., 2008). Notably, PP2C genes, both from Arabidopsis
and rice, exhibit specific as well as overlapping expression
during different stress treatments, especially drought and high
salinity (Singh et al., 2010; Xue et al., 2008). This type of
expression pattern strengthens the fact that drought, osmotic
and salinity stresses are manifested through an overlapping
molecular mechanism, and PP2Cs could be point of ‘‘cross-
talk’’ among different signaling pathways.
In Arabidopsis, various members of group-A PP2Cs
regulate ABA signaling triggered under abiotic stresses.
ABI1 and ABI2 are the two most extensively studied PP2Cs
in Arabidopsis and have been characterized as the main
components of ABA signaling under abiotic stresses and
during development (Fuchs et al., 2013; Singh & Pandey,
2012). The dominant mutants abi1 and abi2 harbor glycine
(Gly) to aspartate (Asp) substitution near Mg
2+
-binding site,
which results in reduction in the phosphatase activity and
consequently causes ABA insensitivity, impaired seed dor-
mancy, defects in stomatal movement and eventually poor
drought tolerance (Schweighofer & Meskiene, 2008). Most of
the group-A PP2Cs are induced by ABA and stress treatments
(Fujita et al., 2011). Higher PP2C level is considered a part
of the negative feedback loop mechanism to desensitize the
plants to high ABA levels (Fuchs et al., 2013; Szostkiewicz
et al., 2010).
A significant role of group-A PP2Cs has been appreciated
as regulators of SnRK2 kinase activities. SnRK2s, including
SnRK2.2, SnRK2.3 and SnRK2.6, are known as positive
regulators of ABA and abiotic stress signaling (Fuji et al.,
2009; Fujita et al., 2009; Soon et al., 2012; Umezawa et al.,
2009). Interestingly, the latest finding has shown that group-A
PP2Cs (ABI1 and PP2CA) also control the function of related
SnRK1 kinases under stress conditions through ABA signal-
ing (Rodrigues et al., 2013). This observation hints toward
the reinforcement of stress responses through coordinated
function of ABA in two complementary pathways. In an
interesting study, Komatsu et al. (2013) showed that group-A
PP2Cs are important regulators of desiccation tolerance in
land plants, as elimination of group-A PP2Cs leads to
survival of moss even in complete desiccation conditions.
Moreover, the authors showed that disruption of group-A
PP2Cs insignificantly perturbs ABA-activated kinase activity,
suggesting that unlike other plant species, group-A PP2Cs act
downstream of the kinases in moss.
Additionally, plant PP2Cs are known to regulate abiotic
stress-triggered MAPK-signaling pathways (Danquah et al.,
2014). PP2C enzymes deactivate MAPKs through depho-
sphorylation, and thus block the downstream regulation of the
signaling cascade (Sme
´kalova
´et al., 2014). For instance,
MP2C (medicago sativa phosphatase 2C), a PP2C from
M. sativa, controls the activity of stress-induced MAPK
(SIMK) through dephosphorylation of pThr in pTEpY motif
in the activation loop. Interestingly, human PP2Cs are also
implicated in cellular stress signaling regulated by MAPK
cascades. At least four human PP2Cs [PP2Ca, PP2Cb, PP2Ce
and Wip1 (wild-type p53-induced phosphatase 1)] were found
to negatively regulate the function of SAPKs (stress-
associated protein kinases belonging to a subfamily of
MAPK superfamily) through dephosphorylation (Tamura
et al., 2006). Two distinct classes of SAPKs (JNK and p38
kinases) are present in mammalian cells, and these kinases are
involved in a variety of environmental stress responses.
Stress-triggered SAPK activation is vital for promoting
apoptosis in different cell types, such as primary tumor
cells and transformed cell lines (Tamura et al., 2006). The
Wip1 is a stress-responsive PP2C that dampens the function
of various stress signaling proteins, including p53 (tumor
protein 53), ATM (ataxia telangiectasia mutated), p19 (cyclin-
dependent kinase inhibitor 2D; CDKN2D) and checkpoint
kinases Chk1 and Chk2, through direct binding and depho-
sphorylation (Arino et al., 2011; Zhu & Bulavin, 2012).
Negative regulation of these different proteins by Wip1 leads
to enhanced tumorigenesis, hampered DNA repair process,
cell cycle arrest and apoptosis. In addition, Wip1 has been
found to suppress inflammatory signaling and promote
senescence escape (Lowe et al., 2012). These observations
suggest functional conservation of PP2Cs across plant and
animal kingdoms.
Some other PP2Cs have been investigated for their
expression and in planta functional roles under abiotic
stresses that are summarized in Table 1. Overall, PP2Cs
have emerged as key players of ABA signaling, regulating
plant responses to different abiotic stresses through inter-
action and modulation of different target proteins. Latest
findings have revealed some novel and unique functional roles
of PP2Cs in plant stress signaling and opened a new field for
future investigation.
PP2Cs mediate biotic stress responses and plant
immunity
Plants have devised several mechanisms to combat the
pathogen attack. One of the mechanisms is modulation of
stomatal movement and prevention of pathogen invasion
during early disease stages, as stomata is a natural site of entry
for destructive bacteria and fungi (Montillet et al., 2013).
ABA, like abiotic stresses, has also been involved in plant
responses to biotic stresses and plant pathogen interactions
(Ton et al., 2009; Kim et al., 2010). ABA has been
recognized as a major regulator of stomatal defense against
pathogens (Zeng et al., 2011). However, the role of ABA
varies, and it can act either as a positive or negative regulator,
based on pathogen type, mode of invasion in host cell and
defence response time (Ton et al., 2009; Kim et al., 2010).
Interestingly, stomata-related regulatory functions of ABA in
response to pathogen attack are mediated by PP2Cs.
Alteration of PP2C (ABI1 and ABI2) function to ABA
insensitivity (abi1, abi2) led to 20- to 80-fold lesser
multiplication of bacterial pathogen Pseudomonas syringae
(DC3000). On the other hand, intragenic revertants of abi1
(abi1-1sup5 and abi1-1sup7; recessive alleles of ABI1) were
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Table 1. Summary of diverse functions of PP2Cs in plants.
Gene Plant species Type of study Physiological function References
Abiotic stress
Group-A PP2CsArabidopsis Transcriptome/microarray/RT-PCR ABA, drought, salt, cold stress signaling Xue et al., 2008
Group-A PP2Cs Rice Transcriptome /microarray/MPSS/RT-qPCR ABA, drought, salt, cold stress signaling Singh et al., 2010; Xue et al.,
2008
Group-A PP2Cs Maize Transcriptome /RNAseq ABA, drought, salt signaling Wei & Pan, 2014
Group-A PP2Cs Tomato RT-qPCR-based expression Drought signaling Sun et al., 2011
CsPP2C2 Cucumber RT-qPCR-based expression Drought signaling Wang et al., 2012
ABI1/ABI2 Arabidopsis Expression, molecular and genetic ABA and drought signaling Fuchs et al., 2013
AtPP2CA/AHG3 Arabidopsis Expression, molecular and genetic ABA signaling Kuhn et al., 2006; Yoshida et al.,
2006
HAI1/2/3 Arabidopsis Expression, molecular and physiological Wounding, drought, osmotic stress
signaling
Bhaskara et al., 2012; Zhang
et al., 2013
SAG113 Arabidopsis Molecular and genetic ABA signaling, drought and leaf
senescence
Zhang et al., 2012
Group-A PP2CsPhyscomitrella Molecular, biochemical and genetic ABA signaling and desiccation tolerance Komatsu et al., 2013
ABI1 and PP2CA Arabidopsis Molecular, biochemical and genetic ABA and drought signaling via SnRK1 Rodrigues et al., 2013
MP2C Alfalfa Biochemical Drought signaling via MAPK regulation Meskiene et al., 2003
OsPP108 Rice Molecular and genetic in Arabidopsis ABA, drought, osmotic and salt signaling Singh et al., 2015
ZmPP2C Maize Molecular and genetic in Arabidopsis Negatively regulates osmotic stress
responses
Liu et al., 2009
ZmPP2C2 Maize Molecular and genetic in Tobacco Positively regulates cold stress response Hu et al., 2010
OsPP18 Rice Molecular, genetic and physiological Positively regulates drought/oxidative
stress response
You et al., 2014
Biotic stress/immunity
ABI1/2 Arabidopsis Molecular and physiological Pseudomonas syringae response
regulation
de Torres-Zabala et al., 2007
ERA1 Arabidopsis Molecular and physiological Pseudomonas syringae response
regulation
Goritschnig et al., 2008
PP2CA Arabidopsis Expression, molecular and genetic Response to bacterial pathogen, regulates
plant immunity
Lim et al., 2014
AP2C1 Arabidopsis Molecular, cellular, biochemical and genetic Wounding, pathogen response, plant
immunity
Schweighofer et al., 2007
PIA1 Arabidopsis Expression, molecular and genetic Response to bacterial pathogen through
hormonal regulation
Widjaja et al., 2010
XB15 Rice Molecular, biochemical and genetic Negative regulator of innate immune
response
Park et al., 2008
Potassium deficiency
AIP1 Arabidopsis Molecular and biochemical Negatively regulator of K + uptake
activity
Lee et al., 2007
AtPP2CA Arabidopsis Molecular and biochemical Negatively regulator of K + channel
activity
Wang & wu, 2013
Development
PP2CsArabidopsis Transcriptome/microarray/RT-PCR Various tissues and developmental stages Xue et al., 2008
PP2Cs Rice Transcriptome/microarray/MPSS/RT-qPCR Vegetative and reproductive development
(seeds and panicles)
Singh et al., 2010; Xue et al.,
2008
PP2Cs Maize Transcriptome/RNAseq Various tissues and developmental stages Wei and Pan, 2014
Group- A PP2Cs Tomato RT-qPCR-based expression Fruit development Sun et al., 2011
CsPP2C2 Cucumber RT-qPCR-based expression Fruit development Wang et al., 2012
KAPP Arabidopsis Molecular, biochemical and genetic Luan, 2003
6A. Singh et al., Crit Rev Biotechnol, Early Online: 1–13
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ABA hypersensitive and supported 30-fold higher bacterial
growth compared to wild type (de Torres-Zabala et al.,
2007). Consistently, era1 [enhanced response to ABA1 (ABA-
deficient 1)], an ABA-hypersensitive mutant showed higher
susceptibility to bacterial pathogen (Goritschnig et al., 2008).
These observations propose ABA as a negative regulator of
pathogen attack responses and plant innate immunity. The
negative regulation of plant immunity by ABA could be
accounted by its antagonistic relationship with salicylic acid
(SA), a phytohormone required to develop defense responses.
Bacterial effectors activate ABA biosynthesis, and higher
ABA level inhibits SA synthesis, thereby blocking SA-
mediated defence responses (de Torres Zabala et al., 2009).
Recently, Lim et al. (2014) reported the involvement of
RCAR3- and PP2CA-mediated ABA signaling in response to
P. syringae pv. tomato DC3000 (Pst DC3000) infection in
Arabidopsis. After Pst DC3000 inoculation, the expression
levels of RCAR3 declined, while that of PP2CA and other
PP2C genes, such as HAB1, AHG1 and AIP1 [AKT1
(Arabidopis K
+
transporter 1)-interacting PP2C 1], increased
gradually from 12 to 24 hours postinoculation (hpi). PP2CA
interacted with RCAR3, and the level of ABA sensitivity in
PP2CA loss- and gain-of-function mutants could be directly
correlated with stomatal opening and closure in response to
Pst DC3000 infection. After inoculation of pp2ca mutant
lines, the bacterial population was about 4-fold lower, while
PP2CA overexpression (PP2CA-OX) leaves had only margin-
ally higher bacterial population than wild-type leaves. The
low level of the bacterial population could be correlated with
the smaller stomatal pore size in pp2ca-mutant leaves, and
most of their stomata were closed 3 hpi of Pst DC3000, while
PP2CA-OX exhibited lower stomatal closure. Moreover,
suppression of stomatal reopening in pp2ca mutants was
also maintained after treatment with flg22 (flagellin 22, a
microbial-associated molecular pattern) and coronatine
(COR, virulence factors secreted by Pst DC3000), or ABA
and COR. Thus, along with RCAR3, PP2CA modulates ABA
signaling in response to bacterial pathogen, and critically
participates in plant immunity through stomatal movement.
AP2C1, one of the well-characterized group-B PP2Cs, has
been known to regulate wound and pathogen-related
responses in Arabidopsis (Schweighofer et al., 2007).
AP2C1 dephosphorylates and inactivates MAPK4 and
MAPK6 that are known positive regulators of wounding and
pathogen-triggered signaling and responses. Expression of
AP2C1 is induced in response to the necrotrophic fungus
Botrytis cinerea and wounding. Overexpression of AP2C1
resulted in low wound-responsive MAPK activity, reduced
ethylene level and hampered innate immunity against B.
cinerea. Whereas, higher level of wound-induced jasmonate
(JA) was detected in ap2c1 mutant plants, and as a
consequence, ap2c1 mutant plants exhibited higher resistance
to herbivore, such as phytophagus mites (Tetranychus
urticae). Moreover, ectopic expression of AP2C1 resulted in
the suppression of flg22- and elicitor oligogalacturonide-
induced MAPK function (Galletti et al., 2011; Fuchs et al.,
2013). Overall, these observations suggested that AP2C1
functions as a negative regulator of MAPK signaling and
controls the defense hormone level to regulate plant innate
immunity.
Regulate flower development through
CLAVATA pathway
POL Arabidopsis Molecular, biochemical and genetic Regulate f lower development through
CLAVATA pathway
Yu et al., 2003
POL/PLL Arabidopsis Molecular, biochemical and genetic Early embryo and root meristem
development
Song and Clark, 2005; Wang
et al., 2007;Gagne and Clark,
2010
PLL4/PLL5 Arabidopsis Molecular, biochemical and genetic Leaf development Song et al., 2005
AtPP2C5/AP2C3 Arabidopsis Molecular, biochemical and genetic ABA-mediated seed germination, stomata
development
Brock et al., 2010; Umbrasaite
et al., 2010
Group-D PP2CsArabidopsis Molecular, biochemical and genetic Auxin-mediated regulation of plant cell
expansion
Spartz et al., 2014
AtPP2CF1 Arabidopsis Expression, molecular and genetic Cell proliferation/expansion, accelerated
inflorescence stem growth
Sugimoto et al., 2014
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Similarly, other Arabidopsis PP2Cs are implicated in plant
response to P. syringae. For instance, the PIA1 (PP2C
induced by AvrRpm1), which belongs to group-F PP2Csin
Arabidopsis, is upregulated in response to AvrRpm1 (bacter-
ial type-III effector) (Widjaja et al., 2010). The pia1
knockout mutant shows higher resistance against P. syringae
Pto DC3000 (AvrRpm1), but not to other strains such as Pto
DC3000 (avrB) or Pto DC3000. Moreover, low level of stress
hormones, such as ethylene after wounding and SA after
AvrRpm1 induction, were detected in pia1-mutant plants,
suggesting that PIA1 also regulates stress hormone production
in response to pathogen. Rice PP2C, XB15 (XA21 binding
protein 15; XA, Xanthomonas oryzae pv. oryzae) was found
to regulate the function of the RLK XA21, an important
positive regulator of plant immunity to bacterial pathogen
X. oryzae pv. oryzae (Park et al., 2008). XB15 interacts with
XA21 through a Ser residue in XA21 juxtamembrane domain,
and dephosphorylates autoactivated kinase. Functional ana-
lysis concluded that XB15 acts as a negative regulator of
XA21-mediated innate immune response, as xb15-mutant
plants exhibited strong cell death, induction of PR (patho-
genesis related) genes and higher XA21-mediated resistance
to X. oryzae. Therefore, PP2Cs majorly mediate MAPK, ABA
and RLK signaling to control stomatal movement and the
production of defense hormones, including ethylene, JA and
SA, in response to pathogen attacks to confer immunity in
different plant species.
PP2Cs in K
+
nutrition deficiency
K
+
is the most abundant cation and one of the major
macronutrients required for the normal plant growth and
development. Deficiency of K
+
results in growth arrest,
inhibition of protein synthesis and impaired nitrogen balance,
as well as reduced sugar levels due to inhibition of
photosynthesis and long distance transportation (Che
´rel
et al., 2014). To combat K
+
deficiency, uptake activity of
different K
+
channels and transporters, which are located at
the root cells, is of prime importance. The activity of K
+
transporters and channels under K
+
-deficient conditions is
regulated by reversible phosphorylation mechanism, and thus,
PKs and PPs are the key participants in this regulatory
mechanism (Wang & Wu, 2013). In Arabidopsis, AIP1, by
dephosphorylating AKT1 channel, reverses the action of
CIPK23 [CBL (calcineurin B-like)-interacting protein kinase
23) and therefore negatively regulates K
+
uptake activity
under K
+
-deprived conditions (Lee et al., 2007). However,
Lan et al. (2011) revealed that a PP2C could directly interact
with a CIPK to block its kinase activity, resulting in AKT1
channel inactivation. Phosphorylation is the key mechanism
to regulate AKT2 (a shaker-type K
+
channel) function in K
+
circulation in phloem, acting as a K
+
battery. This K
+
battery
helps H
+
-ATPase in facilitating transmembrane transport
(Gajdanowicz et al., 2011). Interestingly, the kinase respon-
sible for AKT2 phosphorylation has not been identified
currently. However, AtPP2CA was shown to interact and
dephosphorylate AKT2 to inhibit AKT2 channel activity
(Wang & Wu, 2013). Various events triggering plant
responses during K
+
deficiency are depicted in Figure 3.
CIPK9, a close homolog of CIPK23 in Arabidopsis, has been
identified as a critical regulator of low K
+
response, and its
involvement is proposed to be in the K
+
sensing and
utilization processes (Pandey et al., 2007). Through yeast
two-hybrid cDNA library screening, a PP2C was identified as
a potential interactor of CIPK9 in Arabidopsis. Biochemical
and genetic studies concluded that this PP2C could regulate
CIPK9 through dephosphorylation in K
+
-deficient signaling
(Singh et al. unpublished data). Therefore, PP2Cs participate
Figure 3. A model depicting role of reversible
phosphorylation in K
+
-deficient signaling in
plants. K
+
(potassium) deficiency in the soil
results in elevated cytosolic Ca
2+
(calcium)
level. The high Ca
2+
levels are sensed by
CBLs (calcineurin B-likes), which, after
activation in calcium-dependent manner,
interact with CIPKs (CBL-interacting protein
kinases) and perpetuate the calcium signal-
ing. Specific CBLs, such as CBL1/9, interact
with CIPK23, which subsequently interacts
and phosphorylates the plasma membrane-
located AKT1 (Arabidopsis K
+
transporter 1)
channel, resulting in high K
+
uptake from
soil. AIP1 (AKT1-interacting PP2C 1)
reverses this process through dephosphoryla-
tion of AKT1. Similarly, CBL4/CIPK6
module interacts with another channel AKT2
and leads to its translocation from endoplas-
mic reticulum (ER) to plasma membrane,
thereby activating the channel independently
of phosphorylation.
P
P
P
P
PP2CA
K+ deficiency
AKT1
AKT1
CIPK23
CBL1/9
AIP1
CIPK6
CBL4
Ca2+
channel
Ca2+
Ca2+ K+
K+
P
AKT2
CIPK6
Cytoplasm
AKT2
Nucleus
ER
8A. Singh et al., Crit Rev Biotechnol, Early Online: 1–13
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in reversible phosphorylation events to control the activity of
kinases and K
+
channels under low K
+
conditions.
PP2Cs as regulators of plant development
Understanding the complex events of plant development has
been a real challenge to researchers worldwide. Recent
advancements in genomics techniques and high-throughput
expression-based studies have revealed various components
that are potential regulators of plant development at different
stages (Kamenetsky et al., 2015; Zhang et al., 2015). Among
these, PP2Cs have emerged as vital regulatory components of
signaling during critical stages of plant development (Singh &
Pandey, 2012). Various expression detection techniques,
including microarray, massively parallel signature sequence
(MPSS) and expressed sequence tag (EST), revealed that a
large set of 49 PP2C genes in Arabidopsis is highly expressed
in various tissues at various developmental stages, including
leaves, roots, inflorescences and siliques (Xue et al., 2008).
Similarly, genome-wide expression analysis in rice revealed
the differential transcript changes of several PP2C genes
(63%) during the critical stages of reproductive develop-
ment, including panicle (P1–P6; from floral transition stage to
mature pollen) and seed development (S1–S5; early globular
embryo to dormancy desiccation tolerance) (Singh et al.,
2010). Interestingly, significant overlap of expression was also
observed during reproductive development (panicles and
seeds) and abiotic stresses. The overlapping expression can
be attributed to cis-acting regulatory elements, such as ABA-
responsive element (ABRE), which could regulate both plant
stress response and development simultaneously (Singh
et al., 2014). Moreover, a developmentally programmed
dehydration event leading to seed dormancy is triggered
during the later stages of seed maturation (Singh et al., 2013).
Developmental stimuli could also lead to transcript changes
of PP2C genes in other crop plants, including tomato and
cucumber, during vital fruit development stages (Sun et al.,
2011; Wang et al., 2012).
Several distinct members of Arabidopsis PP2C family
have been implicated in the regulation of plant development,
and the most significant ones include KAPP and POL
(POLTERGEIST). Both the PP2Cs participate in
CLAVATA1 (CLV1, a RLK) signaling pathway to regulate
flower development in Arabidopsis (Luan, 2003). KAPP,
through its forkhead-associated domain (FHA), interacts
and dephosphorylates autophosphorylated CLV1 in vitro.
Moreover, overexpression of KAPP in Arabidopsis leads to
clv1 mutant-like phenotype, while reduced KAPP expression
results in reverse phenotype. It was demonstrated that POL
regulated the CLV1 pathway by modulating the activity of
WUSCHEL (WUS) transcription factor. However, detailed
genetic analyses of clv/wus double and pol/clv/wus triple
mutants suggested that POL functions in both WUS-depend-
ent and -independent pathways (Yu et al., 2003), which was
further supported by a broad expression pattern of POL and
POL-like (PLL) genes (Song & Clark, 2005). Moreover, POL
and PLL are implicated in root and shoot meristem as well as
embryo formation, as pol/pll1 double mutants were found to
be seedling lethal (Song & Clark, 2005; Wang et al., 2007).
These observations indicate the broader role for these PP2Cs
in plant growth and development. Later studies further
strengthened this assumption by showing that downstream
of CLV1, POL and PLL1 regulate the fate of stem cells by
controlling the expression of WUS. POL and PLL1 also
mediate the CLE40 [CLV3/ESR (endosperm surrounding
region)-related 40]/WOX5 (WUSCHEL-related homeobox 5)
pathway to modulate early embryo and root meristem
development (Gagne & Clark, 2010; Song et al., 2008).
Therefore, by controlling these important events, POL and
PLL1 participate in asymmetric stem cell division and
regulate induction and maintenance of stem cell polarity
(Fuchs et al., 2013). Moreover, genetic analyses of other PLL
mutants, including PLL4 and PPL5, proposed their role in
leaf development (Song et al., 2005).
Interestingly, PP2Cs have been found to be key players that
integrate the MAPK signaling in response to ABA during
plant development. For instance, AtPP2C5/AP2C3, a MAPK
phosphatase, expresses distinctively in stomata/stomata lin-
eage cells, and positively regulates seed germination, stomatal
development, stomatal closure and ABA-mediated gene
expression through interaction with MAPKs and the regula-
tion of MAPK3, MAPK4 and MAPK6 activities in the
nucleus (Brock et al., 2010; Umbrasaite et al., 2010).
Moreover, enhanced ABA-dependent activation of MAPK3
and MAPK6 was detected in double knockout mutant of
MAPK-interacting phosphatases, the pp2c5/ap2c1, which
rendered plants ABA-insensitive. Thus, it was suggested
that the PP2C-controlled MAPK cascade negatively regulates
ABA signaling in plants (Brock et al., 2010). AtPP2C5/
AP2C3 nuclear localization was found to be crucial for
induction of cell division and conversion of epidermal cells to
stomata guard cells (Umbrasaite et al., 2010).
Recent functional studies have shown that besides classical
PP2Cs, several novel PP2C candidates have emerged as
potential mediators of plant development signaling. A study
by Spartz et al. (2014) revealed that a few Arabidopsis PP2Cs
from group D interact with auxin-induced SAUR19 (SMALL
AUXIN UP RNA 19) and other members of this family.
SAUR19 was found to enhance the activity of a plasma
membrane-located H
+
-ATPase by promoting phosphorylation
at its C-terminal autoinhibitory domain. H
+
-ATPase activa-
tion possibly aids in plant cell expansion by allowing proton
efflux in apoplast, and facilitates the uptake of solutes and
water, consequently leading to elongated hypocotyl in
SAUR19 overexpression plants. Group-D PP2C (PP2C-D)
members act antagonistically to SAURs, as they interact and
deactivate H
+
-ATPase in vivo. The pp2c-d loss-of-function
single and multiple mutants display elongated hypocotyl
phenotype, similar to that of SAUR19-overexpressing plants,
while PP2C-D1 overexpression leads to reduced hypocotyl
length and dwarf phenotype. This suggests that PP2C-D
members exhibit a novel function of modulating auxin-
mediated regulation of plant cell expansion to control plant
growth. Recently, AtPP2CF1, a group-E Arabidopsis PP2C,
was shown to play a novel role in plant growth; specifically, in
activating cell proliferation and expansion and accelerating
inflorescence growth, which was supported by enhanced
biomass production of AtPP2CF1-overexpressing transgenic
plants (Sugimoto et al., 2014). Thus, PP2Cs, such as those
from group D and AtPP2CF1 could be the ideal candidates for
DOI: 10.3109/07388551.2015.1083941 Plant protein phosphatases 2C 9
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the genetic engineering of crop plants for enhanced growth
and sustainable plant biomass production.
Conclusions
PP2Cs form the major group of PPs in plants, and plant
genomes code for more number of PP2C genes than yeast,
mouse and even humans, which advocates the need of PP2Cs
in diverse cellular processes in plants. Recent studies have
revealed that plant genomes are evolved through chromo-
somal duplication events that also resulted in the vast
expansion of PP2C family into several groups.
Discovery of ABARs in plants unveiled the novel function
of PP2Cs as ABA coreceptors, thereby establishing the
ligand-regulated function of PP2Cs. In the newly established
paradigm, ABARs, PP2Cs and SnRK2s interact in different
combinations via specific domains and amino acid residues.
These interactions coordinate ABA signaling in response to
stress and developmental stimuli for specific and overlapping
adaptive responses. Some unique PP2Cs fine-tune the biotic/
abiotic stress responses and developmental events through the
modulation of MAPK-signaling pathways. Distinct PP2Cs
regulating different MAPK-signaling pathways impart the
specificity to intermingled MAPK-signaling cascades in
plants. Roles of plant PP2Cs in different signaling pathways
triggered by biotic and abiotic stresses are summarized in
Figure 4. In addition, involvement of PP2Cs in the regulation
of CLAVATA pathway, cell proliferation and auxin-signaling
pathway advocates their vital functions in controlling plant
growth and development.
Based on the expression profiles, structural features and
functional behavior of well-characterized PP2Cs, several
novel and potential candidates can be identified and
characterized for their functional roles by employing various
molecular, cellular, biochemical and genetic approaches.
During the major signal transduction pathways triggered by
biotic and/or abiotic stresses and/or development, it would be
interesting to decipher whether interactions of PP2Cs with
ABARs are exclusive, or they could also integrate some other
signaling components like calcium and phytohormones,
including cytokinins, auxins, GA and ethylene.
Emerging evidence has suggested a connection between
ABA- with MAPK-signaling cascades. However, most of the
links between these two signaling are poorly characterized.
Since PP2Cs mediate both ABA- and MAPK-signaling
cascades in stress and developmental signaling, they could
be the possible point of connection between ABA and MAPK
cascades. Future research can unravel how and where these
cascades are connected. Some PP2Cs have been shown to be
involved in reversible phosphorylation mechanism to regulate
Gene expression,
ion transport,
adapve response
Gene expression
adapve response
ABA RLK(XA21) ABA
ABARs XB15
RCAR3
PIA
?
MEKK
MKK
ABA-dependent
ABA-independent
PP2C
SnRK1, SnRK2, other kinases
MP2C
Ethylene,
JA
Ethylene,
SA
ABI1/2
era1 PP2CA
MAPKs AP2C1
?? ?
TF, channels, transporters
Resistance to
bacterial pathogen
Defense
response
Phosphorylaon
De-phosphorylaon
Figure 4. A model depicting the involvement of PP2Cs in abiotic and biotic stress signaling and responses in plants. Upon perception of stress, different
signaling cascades, including those of ABA (abscisic acid), MAPK (mitogen-activated protein kinase) and RLK (receptor-like kinase), are activatedin
plant cell, which are shown by green (abiotic) and orange (biotic) compartments. In a major signaling pathway, PP2Cs transduce abiotic stress-triggered
signals either by ABA-dependent (continuous lines) or ABA-independent manner (dotted lines). In ABA-dependent pathway, after binding with ABA,
ABARs (ABA receptors) interact with PP2Cs to remove the inhibition from Ser/Thr kinases (e.g., SnRKs), which results in kinase activation and
subsequent activation of downstream components, such as transcription factors (TFs), channels or transporters, ultimately leading to adaptive response.
On the other hand, in ABA-independent pathway, PP2Cs regulate downstream components without sensing and interacting with ABA/ABARs. ABA
also acts as a signal in biotic stresses imposed by bacterial pathogen, and PP2Cs, such as PP2CA, ABI1/2 (ABA insensitve 1/2) and era1 [enhanced
response to ABA1 (ABA-deficient 1)], control plants responses to bacterial pathogens by regulating some unknown downstream components. PIA
(PP2C induced by AvrRpm1) also transduces the signal from pathogen attack through enhanced production of stress hormones, such as ethylene and SA
(salicylic acid), which participate in defence response. MAPK cascade is another major signaling pathway which is triggered by both biotic and abiotic
stimuli. This MAPK cascade includes many common components, and specific PP2Cs regulating MAPK signaling in biotic stress-dependent [e.g.,
AP2C1 (arabidopsis pp2c-type phosphatase 1)] or abiotic stress-dependent manner [e.g., MP2C (Medicago sativa protein phosphatase 2c], thereby
providing specificity to the overlapping signaling networks. Moreover, pathogen-stimulated RLK [XA21 (Xanthomonas oryzae pv. oryzae 21)] also
integrates signal to MAPK pathway through unknown components. XB15 (XA21 binding protein 15) blocks the activity of XA21, therefore, prevents
downstream signaling and acts as negative regulator of plant immunity.
10 A. Singh et al., Crit Rev Biotechnol, Early Online: 1–13
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the K
+
uptake in plants during low K
+
conditions. However,
these lines of evidence are majorly molecular and biochem-
ical, and no genetic or in planta evidence is available yet for
PP2C functions in these pathways. Therefore, it would be of
great significance to undertake in planta functional analyses
to find out how these PP2Cs control plant responses and
physiology under K
+
-deficient conditions.
In plants, in addition to the PP2Cs from groups A and B,
recently several novel PP2Cs from other groups have been
reported to play significant roles in plant stress tolerance and
development. The homologs of these PP2Cs can be
characterized in related processes and pathways to unearth
the novel PP2C functions. Thereby, in planta functional roles
for specific PP2Cs can be established by employing genetic
and transgenic approaches. Especially, further comprehension
of PP2C-regulated ABA network will foster the efforts of
generating plants with improved stress tolerance without yield
penalty. Pivotal functional information gathered from model
plant like Arabidopsis and rice can be exploited in biotech-
nological applications to genetically engineer important crop
varieties for various desired traits, including stress tolerance,
high yield and improved grain quality. This will ultimately
help address the problems of food security, posed by ever-
increasing world population.
Declaration of interest
The authors declare no conflict and competing interests. The
research work in GKP’s laboratory is supported by grants
from the Council of Scientific and Industrial Research
(CSIR), Department of Biotechnology (DBT), Department
of Science and Technology (DST), India. The research work
in L-SPT’s laboratory is supported by the Scientific Research
C Grant (no. 25440149) from Japan Society for the Promotion
of Science. AS acknowledges CSIR for his research
fellowship.
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