Download PLANT PROTEIN PHOSPHATASES

Annu. Rev. Plant Physiol. Plant Mol. Biol. 1996. 47:101–25
Copyright © 1996 by Annual Reviews Inc. All rights reserved
PLANT PROTEIN PHOSPHATASES
Robert D. Smith
AgBiotech Center, Rutgers University, New Brunswick, New Jersey 08903-0231
John C. Walker
Division of Biological Sciences, University of Missouri, Columbia, Missouri 65211
KEY WORDS: phosphorylation, signal transduction, protein kinase, metabolism, okadaic acid
ABSTRACT
Posttranslational modification of proteins by phosphorylation is a universal
mechanism for regulating diverse biological functions. Recognition that many
cellular proteins are reversibly phosphorylated in response to external stimuli or
intracellular signals has generated an ongoing interest in identifying and characterizing plant protein kinases and protein phosphatases that modulate the
phosphorylation status of proteins. This review discusses recent advances in our
understanding of the structure, regulation, and function of plant protein phosphatases. Three major classes of enzymes have been reported in plants that are
homologues of the mammalian type-1, -2A, and -2C protein serine/threonine
phosphatases. Molecular genetic and biochemical studies reveal a role for some
of these enzymes in signal transduction, cell cycle progression, and hormonal
regulation. Studies also point to the presence of additional phosphatases in plants
that are unrelated to these major classes.
CONTENTS
INTRODUCTION.....................................................................................................................
PROTEIN PHOSPHATASE-1 .................................................................................................
Structure and Regulation.......................... .........................................................................
Heterologous Expression of Plant PP1...............................................................................
PROTEIN PHOSPHATASE-2A...............................................................................................
Structure and Regulation.......................... .........................................................................
Role in Cellular Metabolism..................... .........................................................................
PROTEIN PHOSPHATASE-2C...............................................................................................
Structure, Regulation, and Function ...................................................................................
1040-2519/96/0601-0101$08.00
102
104
104
107
108
108
110
111
111
101
102 SMITH & WALKER
OTHER PROTEIN PHOSPHATASES .................................................................................... 114
INHIBITOR STUDIES ............................................................................................................. 116
CONCLUDING REMARKS .................................................................................................... 119
INTRODUCTION
It is well recognized that reversible phosphorylation of proteins controls many
cellular processes in plants and animals. The phosphorylation status of proteins is regulated by the opposing activities of protein kinases and protein
phosphatases. Phosphorylation of eukaryotic proteins occurs predominantly
(97%) on serine and threonine residues and to a lesser extent on tyrosine
residues (107). Phosphohistidine phosphorylation has also been reported in
plants (50), fungi (86), and animals (25), but its relative contribution to the
total phosphoamino acid content of eukaryotic cells is not known. In animals,
protein phosphorylation plays well-known roles in diverse cellular processes
such as glycogen metabolism, cell cycle control, and signal transduction (9,
23, 83, 106, 107). Clearly, there has been a similar interest in examining the
role of protein phosphorylation in plant cellular regulation and in identifying
the protein kinases and protein phosphatases that modulate the phosphorylation status of target substrate molecules. An increasing number of plant protein
kinases have been reported in recent years. Some of these enzymes play
pivotal roles in the control of plant defense mechanisms, signal transduction,
and metabolism (11, 13, 50, 122). In plants, as in animals, recognition that a
number of protein kinases respond directly to second messengers such as Ca2+
led to the view that protein kinases were primarily responsible for regulating
the phosphorylation status of proteins and that protein phosphatases merely
reverse the effects of protein kinases. Molecular genetic and biochemical
studies have greatly advanced our knowledge of protein phosphatases and
provide compelling evidence that these enzymes perform essential regulatory
functions. The objective of this review is to examine recent advances in our
knowledge of the structure and regulation of plant protein phosphatases as
well as initial insights into their physiological roles.
Protein phosphatase activities have been reported in most plant subcellular
compartments, including mitochondria, chloroplast, nuclei, and cytosol, and
are associated with various membrane and particulate fractions (50, 70). Some
protein phosphatases are poorly characterized and may represent novel enzymes that are unique to plants, such as the chloroplast thylakoid protein
phosphatase (124). Others have biochemical properties that are very similar to
well-known mammalian protein phosphatases, such as the mitochondrial pyruvate dehydrogenase phosphatase (80) and cytosolic protein serine/threonine
phosphatases (69). Because some plant and animal protein phosphatases are
recognized to be evolutionarily conserved, biochemical characterization and
molecular cloning of distinct phosphatases in plants that correspond to the
PROTEIN PHOSPHATASES
103
mammalian type-1 (PP1) and type-2 (PP2) protein serine/threonine phosphatases have been achieved over a short period. Understandably, the plant
PP1 and PP2 represent only a subset of the total phosphatases that will subsequently be discovered in plants, but their essential roles in diverse cellular
processes are already becoming evident. Biochemical and genetic studies in
plants implicate PP1 and/or PP2 activity in signal transduction, hormonal
regulation, mitosis, and control of carbon and nitrogen metabolism.
Classification of mammalian PP1 and PP2 is based on their unique substrate specificities and sensitivities to various inhibitors (52). PP1 dephosphorylates the β-subunit of mammalian phosphorylase kinase preferentially and is
inhibited by the endogenous proteins, inhibitor-1 (I-1) and inhibitor-2 (I-2).
PP2 has greater activity toward the α-subunit of phosphorylase kinase and is
resistant to I-1 and I-2. PP2 is further divided into three subgroups, PP2A,
PP2B, and PP2C, depending on their subunit structure, divalent cation requirements, and substrate specificities (23). PP2A is a heterotrimer of a catalytic
C-subunit and two distinct regulatory A- and B-subunits and does not require
divalent cations for activity. PP2B, a Ca2+-activated phosphatase, exists as a
heterodimer consisting of a catalytic A-subunit and a regulatory B-subunit
belonging to the EF-hand family of calcium-binding proteins. PP2C is found
as a monomer and activity requires Mg2+. Mammalian PP1, PP2A, PP2B, and
PP2C catalytic subunits are products of distinct genes, and examination of
their primary structures indicates that PP1, PP2A, and PP2B are related enzymes (106). They share no structural homology with PP2C. The structure,
regulation, and function of the protein serine/threonine phosphatases in animals and fungi have been the subject of many recent reviews (9, 23, 73, 83,
106, 107, 118).
Protein phosphatases are inhibited by a number of natural toxins such as
okadaic acid and cyclosporin A (21, 73). These compounds have proven
extremely useful in analyzing the differing actions of specific classes of phosphatases in vitro, as well as in vivo, because many of them are readily taken up
by animal and plant cells (73). The marine toxin okadaic acid, for instance, is a
potent inhibitor of PP2A (IC50 ≈ 0.1–1.0 nM) and inhibits PP1 at 10- to
100-fold higher concentrations (IC50 ≈ 10–100 nM). Okadaic acid is marginally effective against PP2B at micromolar concentrations and has no effect on
PP2C. The immunosuppressant cyclosporin A complexes with endogenous
immunophilins and specifically targets PP2B for downregulation (103). PP2C
is insensitive to these drugs. Increasing use of these drugs to study the role of
reversible protein phosphorylation in diverse cellular processes has generated
new insights into the regulation and physiological function of protein serine/
threonine phosphatases. Inhibitor studies also confirm the presence in plants
and animals of protein phosphatases that do not belong to the PP1 and PP2
families of protein phosphatases.
104 SMITH & WALKER
PROTEIN PHOSPHATASE-1
Structure and Regulation
PP1 is a ubiquitous and highly conserved enzyme found in all eukaryotes. The
native mammalian and fungal enzyme is a complex of a catalytic subunit and
one or more regulatory subunits. Genetic studies have shown yeast PP1 catalytic subunit genes to be essential for cellular processes as diverse as glycogen
accumulation, mitosis, and translational control (118). Regulatory subunits
define specific functions of PP1 catalytic activity in vivo by controlling the
subcellular location and substrate specificity of the enzyme complex. For
instance, the Saccharomyces cerevisiae GAC1 protein, a homologue of the
mammalian RG1 subunit, targets PP1 to glycogen particles and enhances
dephosphorylation of glycogen-bound phosphoenzymes required for glycogen
synthesis and accumulation (35). A nuclear-localized PP1 regulatory subunit,
sds22+, which is essential for completion of mitosis in S. pombe, increases
phosphohistone H1 phosphatase activity of PP1 and downregulates its activity
against phosphorylase a (120). Other regulatory subunits, such as mammalian
I-2, have been characterized biochemically. I-2 is hypothesized to function in
the cell as a chaperone that binds and activates newly synthesized PP1 catalytic subunits via a process that requires phosphorylation of I-2 by glycogen
synthase kinase-3 (GSK-3) in the presence of ATP-Mg (107). The active
catalytic subunit may then complex with various targeting or regulatory
subunits, thereby displacing I-2. On the basis of these and additional studies in
animals and fungi, a targeting subunit has been hypothesized for regulation of
PP1 in vivo (24, 45). The hypothesis is that distinct regulatory subunits are
present in the cell that bind transiently to the catalytic subunit and dictate its
subcellular location and substrate specificity. The hypothesis is supported by
the findings that in S. pombe PP1 is present in the cell as high molecular
complexes ranging in size from 80 to 200 kD, of which only the 80-kD
complex has phosphorylase a phosphatase activity (59).
Protein phosphatase activities have been reported in a number of plant
species and in different tissue extracts through the use of mammalian phosphoprotein substrates. In Brassica napus seed extracts, over 60% of the phosphatase activity is inhibited by I-1 (IC50 = 0.6 nM), I-2 (IC50 = 2.0 nM), and
okadaic acid (IC50 = 10 nM) and dephosphorylates the β-subunit of rabbit
phosphorylase kinase preferentially, indicating that it belongs to the PP1 class
of protein phosphatases (69). The PP1 activity is associated predominantly
with membranes of the endoplasmic reticulum and other unidentified particulate fractions in B. napus seed extracts, and gel filtration analyses indicate it
exists as a high molecular weight complex (70). PP1 is almost exclusively
cytosolic in pea leaves and carrot cells and is associated with microsomes in
wheat leaves (70). It has also been reported in isolated nuclei (104) and in
PROTEIN PHOSPHATASES
105
plasma membranes (135). Very little is known about the structure or regulation
of native PP1 in plants, nor have any physiological substrates been identified.
Studies reveal a striking similarity between the biochemical properties and
subcellular distribution of animal and plant PP1 and suggest the possibility
that mechanisms for controlling PP1 activity and function may be equally well
conserved. A key goal in attempting to understand PP1 function in plants is to
identify and characterize regulatory subunits.
CATALYTIC SUBUNIT PP1 catalytic subunit cDNA and genomic clones have
been reported in several plant species (Table 1). Eight distinct isoforms have
been identified in Arabidopsis (4, 34, 84, 114, 116), and additional related genes
appear to be present in its genome (114, 116). The presence of PP1 multigenes
is not unique to Arabidopsis, and they appear to be present in most plant species.
The remarkable similarity (>70%) between plant and mammalian PP1 primary
sequences and the sensitivity of a bacterially expressed recombinant maize
ZmPP1 to I-2 (IC50 = 0.1 nM) and okadaic acid (IC50 = 200 nM) (113) confirms
that the plant genes encode PP1 catalytic subunits. Unlike the native mammalian
enzyme, however, the recombinant maize PP1 requires Mn2+ for activation
(113). Mn2+-dependent activity is also observed in recombinant rabbit PP1 or
when native PP1 is converted to a Mn2+-dependent form by incubation with
NaF (1). The modified enzymes can be restored to their Mn2+-independent
forms by incubation with I-2 and GSK-3/ATP-Mg, which supports the hypothesis that I-2 acts as a chaperone to activate PP1 in vivo and raises the possibility
that plant PP1 catalytic subunits may need to be activated by a plant homologue
of I-2.
Multiple PP1 isogenes are present in most eukaryotes, with the exception of
S. cerevisiae which contains a single PP1, GLC7 (118). The physiological
advantage of having multiple PP1 isogenes remains unknown. Disruption of
the two S. pombe PP1 genes is lethal, but deletion of either gene, singly, has no
effect on cell viability (85), which indicates that the yeast PP1 isoforms have
overlapping functions. In contrast, loss of one of four PP1 genes (30) in
Drosophila inhibits chromosome separation (5), which suggests a selective
role for this isoform in mitosis. Unique roles for some isogenes may be
governed by their spatial and temporal expression. For instance, the rat PP1γ1
is predominantly expressed in brain tissues, whereas PP1γ2 is almost exclusively expressed in testes (108, 109). Upregulation of certain PP1 isogenes has
been reported in plants as well. BoPP1 expression is enhanced at different
stages of microspore development in B. oleracea, and mature trinucleate microspores contain a unique BoPP1 transcript not found at other stages of the
plant life cycle (100). Arabidopsis AtPP1bg is constitutively expressed at low
levels in all tissues with upregulation in male and female tissues (4).
106 SMITH & WALKER
Table 1 Plant protein phosphatases
Species
Name
Protein phosphatase-1
Catalytic subunit
Arabidopsis TOPP1c
TOPP2d
TOPP3
TOPP4
TOPP5e
TOPP6
AtPP1bgf
TOPP8
Alfalfa
PP1Ms
B. napus
PP1Bn
B. oleracea BoPP1
Maize
ZmPP1
AcetabuPP1Ac1
laria
PP1Ac2
Protein phosphatase-2A
Catalytic subunit
Arabidopsis PP2A-1
PP2A-2
PP2A-3
PP2A-4
Alfalfa
pp2aMs
B. napus
PP2ABn
Sunflower PP2AHa
Acetabularia PP2AAc
Regulatory A-subunit
Arabidopsis pDF1
pDF2
RCN1g
Pea
PP2A-Ps
Regulatory B-subunit
Arabidopsis AtBα
Protein phosphatase-X
Catalytic subunit
Arabidopsis PPX-At1
PPX-At2
Protein phosphatase-2C
Arabidopsis ABI1
KAPP
PP2C-At
Clone
#AA
%IDa Expressionb
Database Reference(s)
accession
number
cDNA
cDNA
cDNA
cDNA
cDNA
cDNA
cDNA
cDNA
cDNA
cDNA
cDNA
cDNA
genomic
318
312
322
321
312
—
322
—
321
—
316
316
319
72
76
73
72
76
—
71
—
74
72
71
70
71
R,L,S,F,C
R,L,S,F,C
R,L,S,F
R,L,F
R,L,S,F,C
—
F
—
R,L,S,N,FB,MF
—
R,L,Co,A,P
R,S,L,H,T,C
—
M93408
M93409
M93410
M93411
M93412
—
Z46253
—
X80788
X57438
X63558
M60215
Z28627
genomic
319
71
—
Z28632
cDNA
cDNA
cDNA
cDNA
cDNA
cDNA
cDNA
genomic
306
306
308
313
313
–
305
307
82
82
81
81
79
72
—
—
R,L,S,F
R,L,S,F
R,L,S,F
R,L,S,F
R,L,S,FB,N,
—
—
—
M96732
M96733
M96734
M96841
X70399
X57439
Z26041
Z26654
3
3
3
16
90
74
cDNA
cDNA
genomic
cDNA
587
—
588
—
—
—
58
55
—
—
seedlings
E,R,S,L,C
X82001
X82002
U21557
Z25888
112
112
54, 112
32
cDNA
513
46
R,L,S,Co,F,FB
U18129
102
cDNA
cDNA
305
307
83
83
R,L,S,F
R,L,S,F
Z22587
Z22596
89
89
genomic
cDNA
cDNA
434
582
399
35
19
35
R,L,S,Si
R,L
—
X77116
U09505
D38109
64, 79
121
62
34, 84, 114
33, 114
114
114
114
116
4, 116
116
88
74
100
112
PROTEIN PHOSPHATASES
107
Functional differences between PP1 isoforms may also arise from structural
variations in their primary sequence that control activity, substrate specificity,
and/or regulatory subunit interactions. Phosphorylation of S. pombe PP1,
dis2+, at a conserved cdc2 protein kinase consensus phosphorylation site [S/TP-X-Z: X = polar and Z = basic; (82)] downregulates its activity in vitro (141).
In contrast, S. pombe PP1, sds21+, lacks a cdc2 phosphorylation motif and is
neither phosphorylated nor downregulated by cdc2 protein kinases (141).
Therefore, isoforms that harbor the phosphorylation recognition site may be
selectively targeted for inactivation by cdc2-like protein kinases. The phosphorylation site is found at a conserved position in about half of the known
plant PP1 catalytic subunits (116), but whether they are subject to this form of
regulation in vivo remains unknown.
Heterologous Expression of Plant PP1
One approach that has been taken to address the functional importance of
structural variations in the primary sequences of the catalytic subunits is to
express individual PP1 isoforms in heterologous systems and ask whether they
can fully complement the multiple functions of their host PP1. Expression of
distinct plant PP1 clones in fungal systems reveals that isoforms differ in their
ability to complement various PP1 functions (Table 2). A lethal trait caused by
disruption of S. cerevisiae PP1, glc7- (14), is rescued by Arabidopsis TOPP2
but not TOPP1 (116). Failure of TOPP1 to complement the lethal trait cannot
be attributed to improper transcription or translation because TOPP1 protein is
detected in wild-type cells expressing the TOPP1 clone. However, only S.
cerevisiae strains expressing TOPP2 have increased PP1 activity, which suggests that TOPP1 may be downregulated in yeast cells by an unknown posttranslational mechanism. TOPP2 is unable to suppress an S. cerevisiae glc7-1
glycogen deficiency phenotype (14, 116, 123), which indicates that TOPP2
can dephosphorylate substrates that are essential for completion of mitosis but
is unable to dephosphorylate a substrate (glycogen synthase) of GLC7 required for glycogen accumulation in yeast. Glycogen accumulation is restored
a
Percent identity between plant PP1 catalytic subunits and rabbit PP1α, plant PP2A catalytic
subunits and rabbit PP2α, plant PP2A regulatory A-subunits and human PR65α, plant PP2A
B-regulatory subunit and human B-subunit, plant PPX catalytic subunits and rabbit PPX, plant PP2C
catalytic domains and rat PP2C.
b
Expression of plant genes in different tissues: R, root; L, leaf; S, stem; F, flower; FB, flower bud;
Si, silique; N, node; MF, mature flower; Co, coleoptile; C, cell culture.
c
TOPP1 independently isolated and named PP1-At (84), and PP1A-At2 (33).
d
TOPP2 independently isolated and named PP1A-At1 (34).
e
TOPP5 independently isolated and named PP1A-At3 (34).
f
AtPP1bg independently isolated and named TOPP7 (116).
g
RCN1 independently isolated and named regA (112).
108 SMITH & WALKER
Table 2 Complementation of fungal mutations by plant PP1 clones
S. cerevisiae
TOPP1
TOPP2
TOPP3
AtPP1bg
PP1Ms
glc7-1
No (116)
No (116)
No (116)
glc7No (116)
Yes (116)
No (116)
S. pombe
dis2-11
Yes (84)
No (34)
wee1-
Aspergillus
nidulans
bimG11
Yes (34)
Yes (4)
No (88)
in glc7-1 strains, however, by expressing a chimeric PP1 consisting of the
N-terminal 1-93 amino acid residues of GLC7 fused to residues 98-312 of
TOPP2 (116). This suggests that unique structural sequences located at the
N-terminus may be important for controlling substrate specificity and/or regulatory subunit interactions.
Functional differences among plant PP1 isoforms have also been reported
in S. pombe and Aspergillus (Table 2). A cold-sensitive dis2-11 mutation that
blocks exit from mitosis is suppressed by TOPP1 but not TOPP2 (34, 84).
However, TOPP2 is able to restore a temperature-sensitive S. pombe
cdc25ts/wee1- double mutation (34) that inhibits entry into mitosis (10), indicating that failure of TOPP2 to complement the dis2-11 mutation is not due to
the absence of active protein phosphatase activity but probably results from a
failure of TOPP2 to dephosphorylate substrates of dis2+/sds21+ that are required for entering mitosis. Furthermore, expression of Arabidopsis AtPP1bg
in a temperature-sensitive Aspergillus bimG11 PP1 mutant supports vegetative
growth but not conidia development at nonpermissive temperatures (4). These
results demonstrate that plant PP1 isoforms have limited ability to functionally
complement fungal PP1 enzymes despite their remarkable structural and biochemical similarities and suggest that even small structural differences in their
primary sequences may have profound effects on their activity and function.
PROTEIN PHOSPHATASE-2A
Structure and Regulation
Native PP2A in animals is found either as a heterodimer of a 36-kD catalytic
C-subunit and a 65-kD regulatory A-subunit, or as a heterotrimer in which a
variable regulatory B-subunit (50–70 kD) complexes to the core heterodimer
(23, 77, 106). In S. cerevisiae, a PP2A A-subunit encoded by tpd3 is essential
for cytokinesis (134), and a B-subunit encoded by cdc55 is required for cellular morphogenesis (44). Single A-subunit and B-subunit genes are present in
Drosophila and both appear to be essential for pattern formation (132). Ge-
PROTEIN PHOSPHATASES
109
netic and biochemical studies indicate that regulatory A- and B-subunits control substrate specificity of PP2A (77).
PP2A has been reported in a number of plant species and is found in most
tissues (53, 66, 69–71, 91, 104). Like PP1, PP2A is found in many subcellular
locations including the nucleus (104) and cytosol and is associated with various membranes and insoluble fractions (70, 135). Neither PP1 nor PP2A is
found in chloroplasts (70, 125). Native PP2A from B. napus seed extracts
dephosphorylates the α-subunit of phosphorylase kinase preferentially and is
potently inhibited by okadaic acid (IC50 = 0.1 nM) in vitro (53, 69). A partially
purified PP2A catalytic subunit from maize seedlings readily dephosphorylates phosphocasein, phosphohistone H1, and phosphorylase a; displays no
appreciable activity toward pNPP; and fails to bind to heparin-Sepharose, a
distinct binding characteristic of PP1 (53). These results indicate that plant
PP2A is biochemically nearly indistinguishable from mammalian PP2A.
Four genes encoding PP2A catalytic subunits (Csubunit) have been isolated from Arabidopsis, and Southern blot analyses
indicate a fifth isoform may be present in the genome (3, 16). C-subunit clones
have also been identified in alfalfa (90), B. napus (74), sunflower, and the green
alga Acetabularia cleftonii (Table 1). Plant and mammalian PP2A share about
80% identity. Transcripts corresponding to each of the Arabidopsis PP2A
catalytic subunits are found in all tissues examined, although the level of
expression of some isogenes appears to be developmentally regulated (16, 89).
Thus, unique roles for some PP2A genes may result from enhanced expression
in some tissues. Differential regulation of the C-subunits may also occur from
posttranslational modifications (98). Phosphorylation of tyrosine (17, 18) or
threonine (27, 41) residues downregulates in vitro activity of mammalian PP2A.
Tyrosine and threonine residues are found at a similar position in all plant PP2A
isoforms. Methylation of the C-terminal leucine, conserved in all PP2A, may
provide an additional level of regulation (107).
CATALYTIC SUBUNIT
REGULATORY SUBUNITS Three A-subunit clones have been isolated from
Arabidopsis (112), and a partial cDNA clone has also been identified in pea (32)
(Table 1). The plant A-subunits are approximately 58% identical to the human
regulatory A-subunit. In addition, a B-subunit homologue has been reported in
Arabidopsis that shares 46% identity with the human Bα-subunit (102). Expression of the A-subunit genes has not been thoroughly examined (112), and the
B-subunit appears to be uniformly expressed throughout the plant (102). Interactions between plant catalytic and regulatory subunits have been examined, but
the presence of multiple catalytic and regulatory subunits in plants suggests that
plants may contain a number of different PP2A complexes with distinct physiological functions.
110 SMITH & WALKER
Insight into a potential role for PP2A in plants comes from the recent
discovery that an Arabidopsis polar auxin transport mutant rcn1 encodes a
PP2A regulatory A-subunit homologue (54) that shares 58% identity with the
human PR65α (54). RCN1 rescues the S. cerevisiae temperature-sensitive tpd3
mutant, which indicates that it is a functional A-subunit homologue capable of
controlling the activity of the yeast PP2A catalytic subunit. A T-DNA insertion into the coding region of RCN1 results in normal expression of a truncated
transcript that may encode a protein lacking a significant portion of its C-terminus. This may represent a loss-of-function mutation because the C-terminal
end of the mammalian regulatory A-subunit is known to be required for
interaction with the catalytic domain (99). The rcn1 recessive mutation causes
an altered morphological response in seedlings to N-1-naphthylphthalamic
acid (NPA), a polar auxin transport inhibitor (54). Auxin transport in stems of
rcn1 mutants also shows increased sensitivity to NPA. These results point to a
unique role for RCN1 in controlling the activity of an endogenous PP2A
toward a key phosphosubstrate(s) involved in polar auxin transport.
Role in Cellular Metabolism
Control of key metabolic enzymes by reversible phosphorylation has been
studied extensively in plants (13, 93). Activation of sucrose phosphate synthase (SPS), nitrate reductase (NR), and phosphoenolpyruvate carboxylase
(PEPC) in light is associated with a decrease in the phosphorylation status of
SPS and NR and an increase in phosphorylation of PEPC (50). Phosphorylation of quinate dehydrogenase (QDH) and hydroxymethylglutaryl-CoA reductase kinase (HMG-CoA reductase kinase) and dephosphorylation of HMGCoA reductase also activates these enzymes in vivo (70, 75, 95). Several lines
of evidence point to PP2A activity in dephosphorylating these enzymes.
Dephosphorylation-mediated changes in activity of each enzyme are blocked
by PP1/PP2A inhibitors, such as okadaic acid, microcystin-LR, and calyculin
A (15, 39, 47, 55, 56, 60, 78, 110). Okadaic acid also prevents in vivo
activation of SPS (47, 110) and NR (46, 48) in spinach leaves. Furthermore,
addition of mammalian PP2A catalytic subunit to cell extracts enhances SPS
(110) and NR activities (71) and downregulates PEPC (78) and QDH (70). In
contrast, addition of the mammalian PP1 catalytic subunit does not alter their
activities. These studies suggest that PP2A, and not PP1, is responsible for
dephosphorylating these enzymes in vivo.
SPS, PEPC, and NR are key enzymes that control nitrogen and carbon
assimilation in plants. Because these pathways compete for carbon skeletons
and energy sources, mechanisms must exist to regulate their relative activities
in response to changing environmental and metabolic conditions. Coordination
of these pathways could result from tight control of SPS, NR, and PEPC
activities in the cytosol, which raises the possibility that their respective pro-
PROTEIN PHOSPHATASES
111
tein kinases and phosphatases respond differentially to signals in the cell.
Emerging evidence indicates that the SPS PP2A may be distinct from the
PP2A that dephosphorylates NR. Activation of SPS, for instance, is inhibited
by Pi, sulfate, and tungstate, but NR activation is unaffected by these compounds (49). In contrast, NR activation is inhibited by Mg2+ and stimulated by
5′-AMP (55, 56). These results could be explained by the presence of multiple
PP2A enzymes in the cytosol that respond differentially to effectors such as Pi,
shown to be a potent inhibitor of the SPS phosphatase (47, 139). Alternatively,
these effectors may interact directly with the protein substrates to modulate
activation. Light does not appear to control the protein kinase and phosphatase
activities directly because feeding mannose to excised leaves in the dark
activates SPS and NR (49, 119). However, a recent study suggests that the NR
PP2A may be light-activated by a process that requires de novo protein synthesis, because it can be blocked with cycloheximide (46).
PROTEIN PHOSPHATASE-2C
PP2C is the least well-characterized member of the protein serine/threonine
phosphatases (106). PP2C demonstrates high activity toward enzymes of the
cholesterol biosynthetic pathway in mammals (106, 107), and genetic studies
in fission yeast implicate a possible role for PP2C in growth (76). Physiological substrates for PP2C remain to be identified in animals or plants. PP2C
demonstrates relatively high activity toward phosphocasein, a commonly used
substrate for measuring PP2C activity in vitro. Mg2+-dependent and okadaicinsensitive phosphocasein phosphatase activity has been reported in several
plant species (70, 75), but its distribution in plants appears more restrictive
than for PP1 and PP2A. Activity is predominantly cytosolic in carrot cells,
cauliflower inflorescence, and leaves from pea and wheat (70, 75). However,
PP2C activity was reported to be absent in maize seedlings (53) and B. napus
seed extracts (69). Cauliflower PP2C and PP2A readily dephosphorylate
HMG-CoA reductase kinase, a key enzyme in isoprenoid biosynthesis (75).
There is presently no indication whether the reductase kinase is a physiological substrate for either phosphatase.
Structure, Regulation, and Function
Three novel PP2C phosphatases have been cloned from Arabidopsis (Figure
1). Each contains a catalytic domain that is structurally related (20–35% identical) to mammalian PP2C. In addition, the plant genes contain N-terminal
extensions of variable lengths that share no homology with one another, or to
protein sequences in the data banks (Figure 1). These unique structural domains among plant PP2C are not found in any of the known fungal or animal
PP2C. One of the Arabidopsis phosphatases, PP2C-At, was identified via a
112 SMITH & WALKER
Figure 1 Structural organization of rat and Arabidopsis thaliana type-2C protein phosphatases.
Conserved catalytic domains are indicated as PP2C. The putative EF-hand calcium binding found
in ABI1 is shown as a solid box. The box containing horizontal lines in KAPP designates a signal
anchor.
genetic screen for genes that complement the sterile phenotype of the S. pombe
pde1 mutant, which is defective in cAMP phosphodiesterase, a component of
the cAMP-dependent protein kinase cascade (62). Among three distinct cDNA
clones isolated from this screen, a PP2C that shares 35% identity with the rat
PP2C was recovered. Expression of PP2C-At in S. pombe pde1 mutants also
restores expression of a transcription factor (ste11) that is required for sexual
development and whose expression is inhibited by the activity of a cAMP-protein kinase (PKA). This suggests that the plant PP2C-At may be counteracting
the activity of a PKA in S. pombe. Its role in Arabidopsis, however, remains
unknown.
Abscisic acid (ABA) regulates multiple
functions in plants including embryo maturation, seed dormancy, stomatal
closure, and mitosis in root meristems. Several abscisic acid–insensitive (abi)
Arabidopsis mutants that are unresponsive to elevated concentrations of ABA
have been extensively characterized (61). The dominant mutant, abi1, has been
cloned and found to encode a PP2C homologue with two distinct domains: an
N-terminal domain containing a putative Ca2+-binding site and a C-terminal
PP2C catalytic domain that is 35% identical to the rat PP2C (64, 79). This
represents a novel putative Ca2+-regulated protein serine/threonine phosphatase
that is structurally unrelated to the PP2B class of Ca2+-dependent phosphatases.
Bacterially expressed ABI1 is Mg2+-dependent, but Ca2+ regulation of ABI
activity has not been reported (79). The presence of a putative Ca2+-binding site
suggests that ABI1 may be responsive to ABA-mediated changes in cellular
Ca2+ levels. The involvement of Ca2+ in ABA-dependent stomatal closure is
ABSCISIC ACID RESPONSE GENE—ABI1
PROTEIN PHOSPHATASES
113
well established (40), and recent observations indicate that the abi1-1 mutant
protein interferes with ABA-dependent regulation of K+ channels in guard cells
of transgenic tobacco (38). A Ca2+-activated phosphatase activity has previously been reported in Vicia faba guard cells that is inhibited by cyclosporin
A-cyclophilin protein complexes (CyP-CsA complex) (67). Cyclosporin A is
an immunosuppressant that binds to endogenous cyclophilin proteins and inhibits Ca2+/calmodulin-activated PP2B (105). It is interesting to note that the
CyP-CsA complex blocks Ca2+-induced inactivation of inward K+ channel
activity in V. faba guard cells (67), which suggests that a PP2B homologue, or
possibly ABI1, may be involved in the control of K+ channel activity. Future
studies may resolve whether ABI is a target for CyP-CsA.
The lesion in the abi1-1 mutant is a single nucleotide substitution that
converts Gly-180 to Asp within the catalytic domain (64, 79). The mutation
does not alter the ubiquitous expression of ABI1 (64). The dominant nature of
the mutation could possibly be explained by an alteration in the response of
abi1 to ABA-mediated cellular changes in Ca2+ levels. Constitutive activation
of the phosphatase, for instance, could inhibit ABA-stimulated signaling pathways. Alternatively, protein dephosphorylation may be required to turn on the
pathways, and activation of ABI1 by Ca2+ may be altered in the mutant protein
phosphatase.
KINASE-ASSOCIATED PROTEIN PHOSPHATASE—KAPP Additional evidence that
type-2C protein phosphatases are involved in plant signaling pathways comes
from the identification of a third novel PP2C in Arabidopsis, termed “KAPP”
for kinase-associated protein phosphatase (121). KAPP was isolated via an in
vitro protein interaction screen for proteins that interact with RLK5, a membrane-bound receptor-like protein kinase (RLK) (137). The predicted structure
of KAPP indicates that it contains three distinct domains, an N-terminal signal
anchor, a kinase interaction domain (KI), and a C-terminal PP2C domain (121).
A bacterially expressed KAPP fusion protein demonstrates Mg2+-dependent
and okadaic-insensitive activity in vitro, consistent with the classification of this
enzyme as a PP2C (121). The signal anchor predicts that KAPP is membrane
localized, and translational insertion of KAPP into membrane vesicles, in vitro,
supports this hypothesis (JM Stone & JC Walker, personal communication).
These results suggest that PP2C is not exclusively cytosolic in the cell. Furthermore, ubiquitous expression of KAPP in Arabidopsis suggests that PP2C
distribution in plants is more extensive than previously indicated from PP2C
activities measured in cell extracts. A possible explanation for this difference is
that native KAPP may have negligible activity toward nonphysiological substrates commonly used to measure PP2C activity.
The KI domain is both necessary and sufficient for interaction with the
receptor-like protein kinase, RLK5 (121). However, interaction between
114 SMITH & WALKER
RLK5 and KAPP occurs only upon autophosphorylation of the receptor kinase
domain. In vitro dephosphorylation of RLK5 with recombinant maize PP1
blocks binding by the KI domain. This indicates that sequences bearing phosphoamino acids may act as high-affinity binding sites for KAPP. RLK5 binding to KAPP is akin to the interaction between autophosphorylated protein
tyrosine kinases and proteins containing src-homology-2 (SH2) domains in
mammalian systems (133). However, unlike SH2 domains that bind exclusively to sites bearing phosphotyrosyl residues, the KI domain may belong to a
novel class of protein-protein interacting domains that bind to sequences containing either a phosphoserine or phosphothreonine residue. The KI domain
bears no sequence homology with SH2 domains.
By analogy with the regulation of SH2-phosphotyrosine phosphatase (SH2PTP) activity in animals, it is possible that binding of KAPP to autophosphorylated RLK5 modulates KAPP phosphatase activity in vivo. Phosphorylation
of the KI domain by RLK5, which has been observed in vitro, may have
additional effects on its binding affinity to RLK5 or on KAPP phosphatase
activity. It is not known whether RLK5 is a physiological substrate for KAPP.
The structure of KAPP and its interaction with RLK5 suggests that KAPP may
control early steps of the RLK5 signaling pathway. RLK5 is a member of a
family of related protein kinases that participate in diverse biological functions
including self-incompatibility, defense responses, and plant development
(121). KAPP and related proteins, therefore, may represent a novel class of
protein phosphatases in plants that are early components of RLK-mediated
signaling pathways. It will be of interest to learn whether KAPP acts as a
positive or a negative regulator of the pathway and whether it interacts specifically with RLK5 or additional phosphorylated receptor and nonreceptor protein kinases in the cell.
OTHER PROTEIN PHOSPHATASES
Molecular cloning studies have revealed the presence, in animals and fungi, of a number of protein serine/threonine phosphatases
that are structurally related to PP1, PP2A, and PP2B phosphatases but that
cannot be placed in any of these classes because of unique structural and/or
biochemical features (107). Among these is the rabbit PPX (26), a PP2A-related
protein phosphatase that localizes to centrosomes and is thought to play a role
in nucleation of microtubules (12). The substrate specificity of rabbit PPX and
its sensitivity to various phosphatase inhibitors is similar to PP2A; however,
PPX fails to bind PP2A regulatory A-subunits (12). Two cDNA clones showing
83% amino acid identity to the rabbit PPX have been isolated from Arabidopsis
by hybridization (Table 1) (89). The Arabidopsis genes, ppx1 and ppx2, are
PROTEIN PHOSPHATASE-X
PROTEIN PHOSPHATASES
115
expressed at low levels in flowers, leaves, stems, and roots. The function of ppx1
and ppx2 in Arabidopsis is not known.
The plant mitochondrial pyruvate dehydrogenase complex (PDC) is regulated in part by reversible phosphorylation of
one of its subunits, pyruvate dehydrogenase E1α (PDH) (80). Phosphorylation
by an intrinsic protein kinase inactivates PDH in the light. Conversely, dephosphorylation of PDH in the dark by a loosely associated Mg2+-dependent protein
serine/threonine phosphatase activates the complex (81). Calmodulin, monovalent cations, and polyamines do not affect the phosphatase, and inorganic
phosphate (Pi) is the only known metabolite that has a slight inhibitory effect at
physiological concentrations (92). The plant PDH-phosphatase is similar in its
biochemical properties to the mammalian phosphatase with the exception of its
response to Ca2+, which enhances Mg2+-activation of the mammalian PDHphosphatase (28) but antagonizes the plant enzyme (81). Structural characterization of the bovine PDH-phosphatase indicates that it is a novel PP2C that
contains a Ca2+-binding site (Kd ≈ 8 µM) (63). Recombinant bovine PDH-phosphatase expressed in Escherichia coli is Mg2+-dependent and Ca2+-stimulated.
The similarity of the plant and mammalian PDH-phosphatases suggests the plant
enzyme may also be a PP2C homologue, but unlike the mammalian enzyme the
plant phosphatase may lack a Ca2+ regulatory domain.
MITOCHONDRIAL PDC-PHOSPHATASE
Phosphorylation of chloroplast proteins has been studied extensively over the past decade, but knowledge of the requisite protein kinases and phosphatases is limited (2, 7). Proteins
are phosphorylated on serine or threonine almost exclusively, with the exception
of pyruvate, Pi dikinase, which contains a phosphorylated histidine residue (97).
Phosphorylation of thylakoid proteins occurs by light- and redox-dependent
kinase(s), whereas redox-independent dephosphorylation in the dark is catalyzed by a membrane-bound protein phosphatase (or phosphatases), which is
sensitive to sodium fluoride and molybdate ions (6, 20, 111, 124). The chloroplast thylakoid phosphatase is probably unrelated to the cytosolic serine/threonine phosphatases, because it fails to dephosphorylate phosphohistone or
phosphorylase a and is not inhibited by okadaic acid or microcystin-LR (70,
125). A detailed characterization of the thylakoid phosphatase has been hampered by the lack of a suitable substrate (142). An alternate approach has been
the use of synthetic peptides (126), which reveals that at least one thylakoidbound phosphatase is able to dephosphorylate multiple thylakoid phosphoproteins in vitro (20).
CHLOROPLAST THYLAKOID PROTEIN PHOSPHATASE
Phosphotyrosine content of proteins in eukaryotes accounts for only a fraction (<3%) of the total phosphoamino acids in
the cell, yet reversible phosphorylation on tyrosine residues is essential for
PROTEIN TYROSINE PHOSPHATASE
116 SMITH & WALKER
cellular growth and differentiation in animals (51, 127). Phosphotyrosine residues have also been reported in plants (19, 131), but little is known of the
physiological role of reversible tyrosine phosphorylation in plants. Dual specificity protein kinases that phosphorylate on serine, threonine, and tyrosine
residues have been reported in plants (8, 87). Some of these kinases are
homologues of mammalian GSK-3, which requires phosphorylation of a specific tyrosine residue for full activation. A tyrosine residue is found at a
conserved location on the plant GSK-3 homologues, suggesting that phosphorylation on the residues may also control activity in vivo. A homologue of
mammalian mitogen-activated protein kinases (MAPK) has been identified in
tobacco and is transiently activated and tyrosine phosphorylated in response to
a fungal elicitor (128). Activation of MAPK in animals requires phosphorylation
of both a threonine residue and a tyrosine residue. It is not known whether the
tobacco MAPK is also phosphorylated on threonine/serine residues and whether
this is important for activation. Downregulation of the tobacco MAPK is
associated with tyrosine dephosphorylation and indicates that plants may contain endogenous protein tyrosine phosphatases and/or dual specificity phosphatases that can dephosphorylate both phosphoserine/threonine and
phosphotyrosine residues (22). Addition of calyculin A to elicitor-treated tobacco sustains the activation and tyrosine phosphorylation of the protein kinase
(128), which indicates that a PP1/PP2A may also be involved in turning off the
kinase because protein tyrosine phosphatases and dual-specificity phosphatases
are resistant to calyculin A. One possible explanation for these results is that a
PP1/PP2A is activating an intermediate protein tyrosine phosphatase or a
dual-specificity phosphatase. Inactivation of the tobacco kinase may also require
dephosphorylation of phosphoserine/threonine residues directly by a PP1/
PP2A. Identifying homologues of mammalian protein tyrosine phosphatases in
plants has been hampered by the lack of structural information on putative
tyrosine phosphatases and because phosphotyrosine phosphatase activity in
plants is often attributed to acid phosphatases (36, 53). A 90-kD protein was
partially purified from pea nuclei that specifically dephosphorylates phosphotyrosine-containing peptides, is inactive toward phosphoserine and phosphothreonine residues, and has little activity toward pNPP, a common substrate
used to measure acid phosphatase activity (42). The pea nuclear phosphatase is
biochemically more similar to mammalian protein tyrosine phosphatases than
the acid or serine/threonine phosphatases, although its exact classification
awaits structural characterization.
INHIBITOR STUDIES
The discovery of protein phosphatase inhibitors has significantly advanced our
understanding of the biological processes that are controlled by reversible
PROTEIN PHOSPHATASES
117
protein phosphorylation and has aided in the identification of physiological
substrates for specific classes of protein phosphatases. Most notable among
these inhibitors is okadaic acid, a marine toxin that inhibits PP2A (IC50 =
0.1–1.0 nM) at concentrations 10- to 100-fold lower than PP1 (IC50 = 10–100
nM). Okadaic acid is readily taken up by cells, which provides a useful
molecular probe to study the role of PP1 and PP2A in many cellular processes.
In vivo sensitivity of PP1 and PP2A to okadaic acid, however, is decreased by
high protein concentrations in the cell (107). Thus, while okadaic acid sensitivity of a specific cellular event is indicative of PP1/PP2A involvement, the
concentration of okadaic acid used may not distinguish between these two
classes of phosphatases. Insight into the differing actions of PP1 and PP2A in
particular cellular events has been furthered by the additional use of chemically distinct toxins such as calyculin A, microcystin-LR, and tautomycin (21,
73). Calyculin A and microcystin-LR inhibit PP1 and PP2A with equal potency (IC50 ≈ 0.1–0.3 nM), whereas tautomycin inhibits PP1 (IC50 = 0.16 nM)
at concentrations fivefold lower than PP2A (IC50 = 8 nM). Comparison of the
biological effects of these cell-permeable compounds with those of okadaic
acid can provide an initial evaluation of the relative contribution of each
phosphatase activity. For example, the use of calyculin A and okadaic acid to
study light-dependent signaling pathways in maize protoplasts suggests that a
PP1 is involved in activating multiple transcription factors involved in lightinduced gene expression in plants (104). The addition of nodularin, acanthifolicin, cantharidin, and the herbicide endothal to the growing arsenal of
PP1/PP2A inhibitors should provide additional confidence in our abilities to
study the differing actions of PP1 and PP2A in intact cells. However, identification of novel protein phosphatases that are sensitive to these same compounds may ultimately limit identification of specific phosphatases based on
inhibitor studies alone and will require additional verification by genetic and/
or biochemical experiments. Nonetheless, these inhibitors provide a very powerful approach for the initial assessment of the role of protein phosphorylation
in controlling numerous cellular events. More than 40 publications describing
the effects of these toxins on diverse biological processes in plants have
appeared within the last four years. A summary of many of these studies is
outlined in Table 3 and implicates PP1/PP2A or related phosphatase activities
in hormonal- (31, 94), carbohydrate- (129), light- (104), and elicitor-mediated
signaling pathways; cell cycle regulation (43, 140, 143); growth and development (104, 115); ion channel control (65, 67, 130, 144); pollination (57, 101);
and cellular metabolism (15, 39, 46, 47, 60, 70, 71, 75, 78, 110, 138).
A different class of phosphatase inhibitors is found in microbially derived
immunosuppressants cyclosporin A and FK506, which, upon binding to endogenous immunophilin proteins, cyclophilin and FK506-binding protein, form
inhibitory complexes that target Ca2+/calmodulin-activated PP2B (102). Their
118 SMITH & WALKER
Table 3 Use of phosphatase inhibitors in plants
Biological Species
function
Signal transduction
Auxin
N. plumbaginifolia
Sugar
Potato
Tissuea
Ib
Response
C
O
LP
O, C,
M
Blocks feedback inhibition of auxin- 31
stimulated gene expression
Blocks expression of sporamin,
129
β-amylase, ADP-glucose phosphorylase small subunit. Increases
expression of sucrose synthase
68
Increases expression of αmy3
Rice
C
O
Light
Maize
P
O, C
Ethylene
Tobacco
L
O
Pathogens Tomato
C
O
Tomato
C
C
Tobacco
C
C
Potato
T
O
Parsley
P
O
Soybean
C, Ct
O, C,
M
Soybean
C
O
Regulation of membrane channels
GC
K+ chan- Vicia faba
nel
Vicia faba
GC
Vicia faba
Anion
Tobacco
channel
Control of enzyme activity
SPS
Spinach
NR
Spinach
Pea
B. campestris
PEPCK
Cucumber
Refs
Blocks expression of lightinducible genes
Increases expression of ethyleneinduced PR genes
Blocks elicitor-induced changes in
NADH oxidase and ascorbate
perodixase activities
Causes growth medium
alkalinization,
hyperphosphorylation of cellular
proteins and increased ACC-S
activity
Causes sustained activation of dualspecificity kinase
Increases expression of PR-10a and
binding of PBF-1
Blocks elicitor-induced
furanocoumarin accumulation
Increases PAL expression, medium
alkalinization and production of
isoflavonoid phytoalexins
Increases expression of PAL, CHS,
and HRGP
104
67, 130
94
136
33, 117
128
29
96
72
37
MC
O, C
O
Blocks Ca2+-induced inactivation
of inward current
Blocks inward current; no effect on
outward current
Increases outward current
Alters voltage gating and kinetics
L
L
R
L
O, M
O, M
O
C
Blocks light-activation
Blocks light-activation
Blocks light-activation
Blocks light-activation
47, 110
46, 71
39
60
Ct
M
Blocks dephosphorylation
138
F, Cs
O, C
65
66
144
PROTEIN PHOSPHATASES
119
Table 3 (continued)
Tissuea
Ib
Response
Maize
QDH
Carrot
HMGCauliflower
CoA red
Growth and development
Root
Arabidopsis
L
C
I
O
O, M
O
Blocks inactivation
78
Blocks inactivation
70
Blocks activation of HMG-CoA red 75
R, RH
O, C
ChlorMaize
oplast
Cell cycle Tobacco
Tradescantia
Ca2+
C. roseus
uptake
Pollination B. napus
B. napus;
B. oleracea;
B. campestris
L
O
C
SH
O, C
O, M
C
O
Inhibits cell division and cortical
cell elongation. Blocks root hair
development
Blocks light-induced chlorophyll
accumulation
Causes cell cycle arrest
Alters metaphase transit times and
chromosome separation
Blocks Ca2+ uptake and callose syn
Pi
F, FB
O, M
O, M
Biological
function
Species
Refs
115
104
43, 143
140
58
Blocks pollen growth
57
Inhibits pollen tube growth in cross- 101
pollinated flowers. Alters selfincompatability in flower buds
a
Tissues: A, aleurone; C, cell culture; Ct, cotyledon; F, flower; FB, flower bud; GC, guard cell; I,
inflorescence; L, leaf; MC, mesophyll cell; LP, leaf petiole; P, protoplast; Pi, pistil; PM, plasma
membrane; S, seed; SH, stamen hair cell; T, tuber.
b
Phosphatase inhibitor: O, okadaic acid; C, calyculin A; M, microcystin-LR; F, FK506 binding
protein-FK506 complex; Cs, cyclosporin-cyclophilin complex.
limited use in plants has been discussed above and suggests that a Ca2+-activated protein phosphatase, possibly a PP2B homologue or ABI1, is involved
in regulating K+ channels in guard cells of V. faba (67).
CONCLUDING REMARKS
The objective of this review was to provide an overview of recent advances in
the study of protein phosphatases in plants. Progress has been made in cloning
a number of plant homologues belonging to three major classes of the mammalian protein serine/threonine phosphatases, and a handful of physiological
substrates for PP2A have been identified. Still, very little is known about the
structure and regulation of the native enzymes or their physiological roles.
Molecular genetic and inhibitor studies have provided powerful avenues to
begin addressing these areas, but progress on understanding the role of specific
protein phosphatases in plants will depend, in large part, on the identification
of physiological substrates and the purification of native enzyme complexes
using traditional biochemical methods. Future studies will no doubt reveal that
120 SMITH & WALKER
many novel protein phosphatases exist in plants that do not belong to the
archetypal classes of serine/threonine protein phosphatases. Ultimately, identification and characterization of the protein phosphatases and protein kinases
that modulate the phosphorylation status and function of key regulatory proteins in the cell will generate new insights into the role of protein phosphorylation in controlling hundreds of diverse biological events in plants.
ACKNOWLEDGMENTS
We would like to thank Drs. Doug Bush, Dieter Söll, and Sabine Rundle for
communication of results prior to publication, as well as Julie Stone and Qing
Lin for critical comments on the manuscript.
Any Annual Review chapter, as well as any article cited in an Annual Review chapter,
may be purchased from the Annual Reviews Preprints and Reprints service.
1-800-347-8007; 415-259-5017; email: [email protected]
Literature Cited
1. Alessi DR, Street AJ, Cohen P, Cohen
PTW. 1993. Inhibitor-2 functions like a
chaperone to fold three expressed isoforms
of mammalian protein phosphatase-1 into a
conformation with the specificity and regulatory properties of the native enzyme. Eur.
J. Biochem. 213:1055–66
2. Allen JF. 1992. Protein phosphorylation in
regulation of photosynthesis. Biochim. Biophys. Acta 1098:275–35
3. Ariño J, Pérez-Callejón E, Cunillera N,
Camps M, Posas F, Ferrer A. 1993. Protein
phosphatases in higher plants: multiplicity
of type 2A phosphatases in Arabidopsis
thaliana. Plant Mol. Biol. 21:475–85
4. Arundhati A, Feiler H, Traas J, Zhang H,
Lunness PA, Doonan JH. 1995. A novel
Arabidopsis type 1 protein phosphatase is
highly expressed in male and female tissues
and functionally complements a conditional cell cycle mutant of Aspergillus.
Plant J. 7:823–34
5. Axton JM, Dombradi V, Cohen PTW,
Glover DM. 1990. One of the protein phosphatase 1 isoenzymes in Drosophila is essential for mitosis. Cell 63:33–46
6. Bennett J. 1980. Chloroplast phosphoproteins: evidence for a thylakoid-bound phosphoprotein phosphatase. Eur. J. Biochem.
104:85–89
7. Bennett J. 1991. Phosphorylation in green
plant chloroplast. Annu. Rev. Plant Physiol.
Plant Mol. Biol. 42:281–331
8. Bianchi MW, Guivarc’h D, Thomas M,
Woodgett JR, Kreis M. 1994. Arabidopsis
homologs of the shaggy and GSK-3 protein
kinases: molecular cloning and functional
9.
10.
11.
12.
13.
14.
15.
16.
17.
expression in Escherichia coli. Mol. Gen.
Genet. 242:337–45
Bollen M, Stalmans W. 1992. The structure, role, and regulation of type 1 protein
phosphatases. Crit. Rev. Biochem. Mol.
Biol. 27:227–81
Booher R, Beach D. 1989. Involvement of
a type 1 protein phoshatase encoded by
bws1+ in fission yeast mitotic control. Cell
57:1009–16
Bowler C, Chua N-H. 1994. Emerging
themes of plant signal transduction. Cell
6:1529–41
Brewis ND, Street AJ, Prescott AR, Cohen
PTW. 1993. PPX, a novel protein serine/
threonine phosphatase localized to centrosomes. EMBO J. 12:987–96
Budde RJA, Randall DD. 1990. Protein
kinases in higher plants. In Inositol Metabolism in Plants, ed. DJ Moore, WF Boss,
pp. 351–67. New York: Liss
Cannon JF, Klemens K, Morcos P, Nair B,
Pearson J, Khalil M. 1995. Type 1 protein
phosphatase systems in yeast. Adv. Protein
Phosphatases 9:215–36
Carter P, Nimmo H, Fewson C, Wilkins M.
1990. Bryophyllum fedtschenkoi protein
phosphatase type 2A can dephosphorylate
phosphoenolpyruvate carboxylase. FEBS
Lett. 263:233–36
Casamayor A, Pérez-Callejón E, Pujol G,
Ariño J, Ferrer A. 1994. Molecular characterization of a fourth isoform of the catalytic subunit of protein phosphatase 2A
from Arabidopsis thaliana. Plant Mol.
Biol. 26:523–28
Chen J, Martin BL, Brautigan DL. 1992.
PROTEIN PHOSPHATASES
18.
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
29.
30.
31.
32.
Regulation of protein serine/threonine
phosphatase type-2A by tyrosine phosphorylation. Science 257:1261–64
Chen J, Parsons S, Brautigan DL. 1994.
Tyrosine phosphorylation of protein phosphatase 2A in response to growth stimulation and v-src transformation of fibroblasts.
J. Biol. Chem. 269:7957–62
Cheng HF, Tao M. 1989. Purification and
characterization of a phosphotyrosyl-protein phosphatase from wheat seedlings.
Biochem. Biophys. Acta 998:271–76
Cheng L, Stys D, Allen JF. 1995. Effects of
synthetic peptides on thylakoid phosphoproteins. Physiol. Plant. 93:173–78
Cicirelli M. 1992. Inhibitors of protein serine/threonine phosphatases. Focus 14:
16–20
Clarke PR. 1994. Switching off MAP kinases. Curr. Biol. 4:647–50
Cohen P. 1989. Structure and regulation of
protein phosphatases. Annu. Rev. Biochem.
58:453–508
Cohen PTW, Cohen P. 1989. Protein phosphatases come of age. J. Biol. Chem. 264:
21435–38
Crovello CS, Furie BC, Furie B. 1995. Histidine phosphorylation of p-selectin upon
stimulation of human platelets: a novel
pathway for activation-dependent signal
transduction. Cell 82:279–86
da Cruz e Silva OB, da Cruz e Silva EF,
Cohen PTW. 1988. Identification of a novel
phosphatase catalytic subunit by cDNA
cloning. FEBS Lett. 242:10610
Damuni Z, Guo H. 1993. Autophosphorylation activated protein kinase phosphorylates and inactivates protein phosphatase
2A. Proc. Natl. Acad. Sci. USA 90:2500–4
Denton RM, Randle PJ, Martin BR. 1972.
Stimulation by calcium ions of the pyruvate
dehydrogenase phosphatase. Biochem. J.
128:161–63
Després C, Subramaniam R, Matton DP,
Brisson N. 1995. The activation of the potato PR-10a gene requires the phosphorylation of the nuclear factor PBF-1. Plant
Cell 7:589–98
Dombradi V, Mann DJ, Saunders RD, Cohen PT. 1993. Cloning of the fourth functional gene for protein phosphatase 1 in
Drosophila melanogaster from its chromosomal location. Eur. J. Biochem. 212:
177–83
Dominov JA, Stenzler L, Lee S, Schwarz
JJ, Leisner S, Howell SH. 1992. Cytokinins
and auxins control the expression of a gene
in Nicotiana plubaginifolia cells by feedback regulation. Plant Cell 4:451–61
Evans IM, Fawcett T, Boulter D, FordhamSkelton AP. 1994. A homologue of the 65kDa regulatory subunit of protein phosphatase 2A in early pea (Pisum sativum L.)
embryos. Plant Mol. Biol. 24:689–95
121
33. Felix G, Regenass M, Spanu P, Boller T.
1994. The protein phosphatase inhibitor calyculin A mimics elicitor action in plant
cells and induces rapid hyperphosphorylation of specific proteins as revealed by
pulse labeling with [33P]phosphate. Proc.
Natl. Acad. Sci. USA 91:952–56
34. Ferreira PCG, Hemerly AS, Van Montagu
M, Inzé D. 1993. A protein phosphatase 1
from Arabidopsis thaliana restores temperature sensitivity of a Schizosaccharomyces pombe cdc25ts/wee1- double
mutant. Plant J. 4:81–87
35. François JM, Thompson-Jaeger S, Skroch
J, Zellenka U, Spevak W, Tatchell K. 1992.
GAC1 may encode a regulatory subunit for
protein phosphatase type 1 in Saccharomyces cerevisiae. EMBO J. 11:87–96
36. Gellatly KS, Moorhead GBG, Duff SMG,
Lefebvre DD, Plaxton WC. 1994. Purification and characterization of a potato tuber
acid phosphatase having significant phosphotyrosine phosphatase activity. Plant
Physiol. 106:223–32
37. Gianfagna TJ, Lawton MA. 1995. Specific
activation of soybean defense genes by the
phosphoprotein phosphatase inhibitor
okadaic acid. Plant Sci. 109:165–70
38. Giraudat J. 1995. Abscisic acid signaling.
Curr. Opin. Cell Biol. 7:232–38
39. Glaab J, Kaiser WM. 1993. Rapid modulation of nitrate reductase in pea roots. Planta
191:173–79
40. Guilroy S, Fricker MD, Read ND, Trewavas AJ. 1991. Role of calcium in signal
transduction of Commelina guard cells.
Plant Cell 3:333–44
41. Guo H, Reddy SAG, Damuni Z. 1993. Purification and characterization of a distinct
autophosphorylation-activated protein kinase that phosphorylates and inactivates
protein phosphatase 2A. J. Biol. Chem.
268:11193–98
42. Guo Y-L, Roux SJ. 1995. Partial purification and characterization of an enzyme
from pea nuclei with protein tyrosine phosphatase activity. Plant Physiol. 107:167–75
43. Hasezawa S, Nagata T. 1992. Okadaic acid
as a probe to analyse the cell cycle progression in plant cells. Bot. Acta 105:63–69
44. Healy AM, Zolnierowicz S, Stapleton AE,
Goebl M, DePaoli RA, Pringle JR. 1991.
CDC55, a Saccharomyces cerevisiae gene
involved in cellular morphogenesis: identification, characterization, and homology to
the B subunit of mammalian type 2A protein phosphatase. Mol. Cell. Biol. 11:
5767–80
45. Hubbard MJ, Cohen P. 1993. On target with
a new mechanism for the regulation of protein phosphorylation. Trends Biochem. Sci.
18:172–77
46. Huber JL, Huber SC, Campbell WH, Redinbaugh MG. 1992. Reversible light/dark
122 SMITH & WALKER
47.
48.
49.
50.
51.
52.
53.
54.
55.
56.
57.
58.
59.
modulation of spinach leaf nitrate reductase activity involves protein phosphorylation. Arch. Biochem. Biophys. 296:58–65
Huber SC, Huber JL. 1990. Activation of
sucrose-phosphate synthase from darkened
spinach leaves by an endogenous protein
phosphatase. Arch. Biochem. Biophys. 282:
421–26
Huber SC, Huber JL, Campbell WH, Redinbaugh MC. 1992. Comparative studies of
the light modulation of nitrate reductase
and sucrose-phosphate synthase activities
in spinach leaves. Plant Physiol. 100:
706–12
Huber SC, Huber JL, Kaiser WM. 1994.
Differential response of nitrate reductase
and sucrose-phosphatase synthase-activation to inorganic and organic salts, in vitro
and in situ. Physiol. Plant. 92:302–10
Huber SC, Huber JL, McMichael RW.
1994. Control of plant enzyme activity by
reversible protein phosphorylation. Int.
Rev. Cytol. 149:47–98
Hunter T. 1995. Protein kinases and phosphatases: the yin and yang of protein phosphorylation and signaling. Cell 80:225–36
Ingebritsen TS, Cohen P. 1983. Protein
phosphatases: properties and role in cellular regulation. Science 221:331–38
Jagiello I, Donella-Deana A, Szczegielniak
J, Pinna LA, Muszynska G. 1992. Identification of protein phosphatase activities in
maize seedlings. Biochim. Biophys. Acta
1134:129–36
Johnson K, DeLong A, Garbers C, Simmons C, Söll D. 1995. Auxin physiology
mutants rcn1 and rgr1. Abstr. Int. Meet.
Arabidopsis Res., 6th, Madison, Wis., p.
299
Kaiser WM, Huber SC. 1994. Modulation
of nitrate reductase in vivo and in vitro:
effects of phosphoprotein phosphatase inhibitors, free Mg2+ and 5′-AMP. Planta
193:358–64
Kaiser WM, Huber SC. 1994. Posttranslational regulation of nitrate reductase in
higher plants. Plant Physiol. 106:817–21
Kandasamy MK, Thorsness MK, Rundle
SJ, Goldberg MI, Nasrallah JB, Nasrallah
ME. 1993. Ablation of papillar cell function
in Brassica flowers results in the loss of
stigma receptivity to pollination. Plant Cell
5:263–75
Kauss H, Jeblick W. 1991. Induced Ca2+
uptake and callose synthesis in suspensioncultured cells of Catharanthus roseus are
decreased by the protein phosphatase inhibitor okadaic acid. Physiol. Plant. 81:
309–12
Kinoshita N, Ohkura H, Yanagida M. 1990.
Distinct, essential roles of type 1 and type
2A protein phosphatases in the control of
the fission yeast cell cycle. Cell 63:
405–15
60. Kojima M, Wu S-J, Fukui H, Sugimoto T,
Nanmori T, Oji Y. 1995. Phosphorylation/
dephosphorylation of Komatsuna (Brassica campestris) leaf nitrate reductase in
vivo and in vitro in response to environmental light conditions: effects of protein
kinase and protein phosphatase inhibitors.
Physiol. Plant. 93:139–45
61. Koornneef M, Reuling G, Karssen CM.
1984. The isolation and characterization of
abscisic acid–insensitive mutants of Arabidopsis thaliana. Physiol. Plant. 61:377–83
62. Kuromoni T, Yamamoto M. 1994. Cloning
of cDNAs from Arabidopsis thaliana that
encode putative protein phosphatase 2C
and a human Dr1-like protein by transformation of a fission yeast mutant. Nucleic
Acids Res. 22:5296–301
63. Lawson JE, Niu X-D, Browning KS, Trong
HL, Yan J, Reed LJ. 1993. Molecular cloning and expression of the catalytic subunit
of bovine pyruvate dehydrogenase phosphatase and sequence similarity with protein phosphatase 2C. Biochemistry 32:
9887–93
64. Leung J, Bouvier-Durand M, Morris P-C,
Guerrier D, Chefdor F, Giraudat J. 1994.
Arabidopsis ABA response gene ABI1:
features of a calcium-modulated protein
phosphatase. Science 264:1448–52
65. Li W, Luan S, Schreiber SL, Assmann SM.
1994. Evidence for protein phosphatase 1
and 2A regulation of K+ channels in two
types of leaf cells. Plant Physiol. 106:
963–70
66. Lin PP-C, Mori T, Key JL. 1988. Phosphoprotein phosphatase from soybean hypocotyls. Plant Physiol. 66:368–74
67. Luan S, Li W, Rusnak F, Assmann SM,
Schreiber SL. 1993. Immunosuppressants
implicate protein phosphatase regulation of
K+ channels in guard cells. Proc. Natl.
Acad. Sci. USA 90:2202–6
68. Lue M-Y, Lee H. 1994. Protein phosphatase inhibitors enhance the expression
of an α-amylase gene, αAmy3, in cultured
rice cells. Biochem. Biophys. Res. Commun. 205:807–16
69. MacKintosh C, Cohen P. 1989. Identification of high levels of type 1 and type 2A
protein phosphatases in higher plants. Biochem. J. 262:335–39
70. MacKintosh C, Coggins J, Cohen P. 1991.
Plant protein phosphatase: subcellular distribution, detection of protein phosphatase
2C and identification of protein phosphatase 2A as the major quinate dehydrogenase phosphatase. Biochem. J. 273:
733–38
71. MacKintosh C. 1992. Regulation of spinach leaf nitrate reductase by reversible
phosphorylation. Biochim. Biophys. Acta
1137:121–26
72. MacKintosh C, Lyon GD, MacKintosh
PROTEIN PHOSPHATASES
73.
74.
75.
76.
77.
78.
79.
80.
81.
82.
83.
84.
85.
RW. 1994. Protein phosphatase inhibitors
activate anti-fungal defence responses of
soybean cotelydons and cell cultures. Plant
J. 5:137–47
MacKintosh C, MacKintosh RW. 1994. Inhibitors of protein kinases and phosphatases. Trends Biochem. Sci. 19:
444–48
MacKintosh RW, Haycox G, Hardie DG,
Cohen PTW. 1990. Identification by molecular cloning of two cDNA sequences
from the plant Brassica napus which are
very similar to mammalian protein phosphatases-1 and -2A. FEBS Lett. 276:
156–60
MacKintosh RW, Davies SP, Clarke PR,
Weekes J, Gillespie JG, et al. 1992. Evidence for a protein kinase cascade in higher
plants. Eur. J. Biochem. 209:923–31
Maeda T, Tsai AYM, Saito H. 1993. Mutations in protein tyrosine phosphatase gene
(PTP2) and a protein serine/threonine
phosphatase gene (PTC1) cause a synthetic
growth defect in Saccharomyces cerevisiae. Mol. Cell. Biol. 13:5408–17
Mayer-Jaekel RE, Hemmings BA. 1994.
Protein phosphatase 2A—a ‘menage à
trois.’ Trends Cell Biol. 4:287–91
McNaughton GAL, MacKintosh C,
Fewson CA, Wilkins MB, Nimmo HG.
1991. Illumination increases the phosphorylation state of maize leaf phosphoenolpyruvate carboxylase by causing
an increase in the activity of a protein kinase. Biochim. Biophys. Acta 1093:189–95
Meyer K, Leube MP, Grill E. 1994. A protein phosphatase 2C involved in ABA signal transduction in Arabidopsis thaliana.
Science 264:1452–55
Miernyk JA, Camp PJ, Randall DD. 1985.
Regulation of plant pyruvate dehydrogenase complexes. Curr. Top. Plant Biochem. Physiol. 4:175–90
Miernyk JA, Randall DD. 1987. Some
properties of pea mitochondrial phospho-pyruvate
dehydrogenase-phosphatase. Plant Physiol. 83:311–15
Moreno S, Nurse P. 1990. Substrates for
P34cdc2 in vivo veritas? Cell 61:549–51
Mumby M, Walter G. 1993. Protein serine/threonine phosphatases: structure,
regulation and functions in cell growth.
Physiol. Rev. 73:673–99
Nitschke K, Fleig U, Schell J, Palme K.
1992. Complementation of the cs dis2-11
cell cycle mutant of Schizosaccharomyces
pombe by a protein phosphatase from
Arabidopsis
thaliana.
EMBO
J.
11:1327–33
Ohkura H, Kinoshita N, Miyatani S, Toda
T, Yanagida M. 1989. The fission yeast
dis2+ gene required for chromosome disjoining encodes one of two putative type 1
protein phosphatases. Cell 57:997–1007
123
86. Ota IM, Varshavsky A. 1993. A yeast protein similar to bacterial two-component
regulators. Science 262:566–69
87. Páy A, Jonak C, Bögre L, Meskiene I,
Mairinger T, et al. 1993. The msK family
of alfalfa protein kinase genes encodes
homologues of shaggy/glycogen synthase
kinase-3 and shows differential expression
patterns in plant organs and development.
Plant J. 3:847–56
88. Páy A, Pirck M, Bögre L, Hirt H, HeberleBors E. 1994. Isolation and characterization of phosphoprotein phosphatase 1
from alfalfa. Mol. Gen. Genet. 244:176–82
89. Pérez-Callejón E, Casamayor A, Pujol G,
Clua E, Ferrer A, Ariño J. 1993. Identification and molecular cloning of two homologues of protein phosphatase X from
Arabidopsis thaliana. Plant Mol. Biol. 23:
1177–85
90. Pirck M, Páy A, Heberle-Bors E, Hirt H.
1993. Isolation and characterization of a
phosphoprotein phosphatase 2A gene from
alfalfa. Mol. Gen. Genet. 240:126–31
91. Polya GM, Haritou M. 1988. Purification
and characterization of two wheat-embryo
protein phosphatases. Biochem. J. 251:
357–63
92. Randall DD, Miernyk JA, David NR,
Budde RJA, Schuller KA, et al. 1990. Phosphorylation of the leaf mitochondrial pyruvate dehydrogenase complex and inactivation of the complex in the light. Curr. Top.
Plant Biochem. Physiol. 9:313–28
93. Ranjeva R, Boudet AM. 1987. Phosphorylation of protein in plants: regulatory effects and potential involvement in stimuli/response coupling. Annu. Rev. Plant
Physiol. 38:73–93
94. Raz V, Fluhr R. 1993. Ethylene signal is
transduced via protein phosphorylation
events in plants. Plant Cell 5:523–30
95. Refeno R, Ranjeva R, Boudet AM. 1982.
Modulation of quinate:NAD+ oxidoreductase activity through reversible phosphorylation in carrot cell suspensions. Planta
154:193–98
96. Renelt A, Colling C, Hahlbrock K, Nurnberger T, Parker JE, et al. 1993. Studies on
elicitor recognition and signal transduction
in plant defense. J. Exp. Bot. 44:257–68
97. Roeske CA, Kutny RM, Budde RJA,
Chollet R. 1988. Sequence of the phosphothreonyl regulatory site peptide from
inactive maize leaf pyruvate, orthophosphate dikinase. J. Biol. Chem. 263:
6683–87
98. Ruediger R, Hood JEV, Mumby M, Walter
G. 1991. Constant expression and activity
of protein phosphatase type 2A in synchronized cells. Mol. Cell. Biol. 11: 4282–85
99. Ruediger R, Roeckel D, Fait J, Bergqvist A,
Magnusson F, Walter F. 1992. Identification of binding sites on the regulatory A
124 SMITH & WALKER
subunit of protein phosphatase 2A for the
catalytic C subunit and for tumor antigens
of simian virus 40 and polyomavirus. Mol.
Cell. Biol. 12:4872–82
100. Rundle SJ, Nasrallah JB. 1992. Molecular
characterization of a type 1 serine/threonine phosphatase from Brassica oleracea.
Plant Mol. Biol. 20:367–75
101. Rundle SJ, Nasrallah ME, Nasrallah JB.
1993. Effects of inhibitors of protein serine/threonine phosphatases on pollination
in Brassica. Plant Physiol. 103:1165–71
102. Rundle SJ, Hartung AJ, Corum JW III,
O’Neill M. 1995. Characterization of the
55-kDa B regulatory subunit of Arabidopsis protein phosphatase 2A. Plant Mol.
Biol. 28:257–66
103. Schreiber SL. 1992. Immunophilin-sensitive phosphatase action in cell signaling
pathways. Cell 70:365–68
104. Sheen J. 1993. Protein phosphatase activity
is required for light-inducible gene expression in maize. EMBO J. 12:3497–505
105. Shenolikar S. 1992. A window opens on
immunosuppression. Curr. Biol. 2:549–51
106. Shenolikar S, Nairn AC. 1991. Protein
phosphatases: recent progress. Adv. Second
Messenger Phosphoprotein Res. 23:1–121
107. Shenolikar S. 1994. Protein serine/threonine phosphatases: new avenues for cell
regulation. Annu. Rev. Cell Biol. 10:55–86
108. Shima H, Haneji T, Hatano Y, Kasugai I,
Sugimura T, Nagao M. 1993. Protein phosphatase 1γ2 is associated with nuclei of
meiotic cells in rat testis. Biochem. Biophys. Res. Commun. 194:930–37
109. Shima H, Hatano Y, Chun Y-S, Sugimura T,
Zhang Z, et al. 1993. Identification of PP1
catalytic subunit isotypes PP1γ, PP1δ and
PP1α in various rat tissues. Biochem. Biophys. Res. Commun. 192:1289–96
110. Siegl G, MacKintosh C, Stitt M. 1990. Sucrose-phosphatse synthase is dephosphorylated by protein phosphatase 2A in spinach
leaves. FEBS Lett. 270:198–202
111. Silverstein T, Chang L, Allen JF. 1993.
Chloroplast thylakoid protein phosphatase
reactions are redox-independent and kinetically heterogeneous. FEBS Lett. 334:101–5
112. Slabas AR, Fordham-Skelton AP, Fletcher
D, Martinez-Rivas JM, Swinhoe R, et al.
1994. Characterization of cDNA and
genomic clones encoding homologues of
the 65-kDa regulatory subunit of protein
phosphatase 2A in Arabidopsis thaliana.
Plant Mol. Biol. 26:1125–38
113. Smith RD, Walker JC. 1991. Isolation and
expression of a maize type 1 protein phosphatase. Plant Physiol. 97:677–83
114. Smith RD, Walker JC. 1993. Expression of
multiple type 1 protein phosphoprotein
phosphatases in Arabidopsis thaliana.
Plant Mol. Biol. 21:307–16
115. Smith RD, Wilson JE, Walker JC, Baskin
TI. 1994. Protein-phosphatase inhibitors
block root hair growth and alter cortical cell
shape of Arabidopsis roots. Planta 194:
516–24
116. Smith RD, Lin Q, Cannon JF, Walker JC.
1995. Type-1 and Type-2C protein phosphatases of higher plants. Adv. Protein
Phosphatases 9:105–20
117. Spanu P, Grosskopf DG, Felix G, Boller T.
1994. The apparent turnover of 1-aminocyclopropane-1-carboxylate synthase in tomato cells is regulated by protein phosphorylation and dephosphorylation. Plant
Physiol. 106:529–35
118. Stark MJR, Black S, Sneddon AA, Andrews PD. 1994. Genetic analyses of yeast
protein serine/threonine phosphatases.
FEMS Microbiol. Lett. 117:121–30
119. Stitt M, Wilke I, Feil R, Heldt HW. 1988.
Coarse control of sucrose-phosphatae synthase in leaves: alteration of the kinetic
properties in response to the rate of photosynthesis and the accumulation of sucrose.
Planta 174:217–30
120. Stone EM, Yamano H, Kinoshita N,
Yanagida M. 1993. Mitotic regulation of
protein phosphatases by the fission yeast
sds22 protein. Curr. Biol. 3:13–26
121. Stone JM, Collinge MA, Smith RD, Horn
MA, Walker JC. 1994. Interaction of a protein phosphatase with an Arabidopsis serine-threonine receptor kinase. Science 266:
793–95
122. Stone JM, Walker JC. 1995. Protein kinase
families and signal transduction. Plant
Physiol. 108:451–57
123. Stuart JK, Frederick DL, Varner CM,
Tatchell K. 1994. The mutant type 1 protein
phosphatase encoded by glc7-1 from Saccharomyces cerevisiae fails to interact productively with the GAC1-encoded regulatory subunit. Mol. Cell. Biol. 14:
896–905
124. Sun G, Bailey D, Jones MW, Markwell J.
1989. Chloroplast thylakoid protein phosphatase is a membrane surface-associated
activity. Plant Physiol. 89:238–43
125. Sun G, Markwell J. 1992. Lack of type 1
and type 2A protein serine(P)/threonine (P)
phosphatase activities in chloroplasts.
Plant Physiol. 100:620–24
126. Sun G, Sarath G, Markwell J. 1993. Phosphopeptides as substrates for thylakoid protein phosphatase activity. Arch. Biochem.
Biophys. 304:490–95
127. Sun H, Tonks NK. 1994. The coordinated
action of protein tyrosine phosphatases and
kinases in cell signaling. Trends Biochem.
Sci. 19:480–85
128. Suzuki K, Shinshi H. 1995. Transient activation and tyrosine phosphorylation of a
protein kinase in tobacco cells treated with
a fungal elicitor. Plant Cell 7:639–47
129. Takeda S, Mano S, Ohto MA, Nakamura K.
PROTEIN PHOSPHATASES
Inhibitors of protein phosphatase 1 and 2A
block sugar-inducible gene expression in
plants. Plant Physiol. 106:567–74
130. Theil G, Blatt MR. 1994. Phosphatase antagonist okadaic acid inhibits steady-state
K+ currents in guard cells of Vicia faba.
Plant J. 5:523–30
131. Torruella M, Casano LM, Vallejos RH.
1986. Evidence of the activity of tyrosine
kinase(s) and the presence of phosphotyrosine proteins in pea plantlets. J. Biol.
Chem. 261:6651–53
132. Uemura T, Shiomi K, Togashi S, Takeichi
M. 1993. Mutation of twins encoding a
regulator of protein phosphatase 2A leads
to pattern duplication in Drosophila imaginal discs. Genes Dev. 7:429–40
133. van der Geer P, Hunter T, Lindberg RA.
1994. Receptor protein-tyrosine kinases
and their signal transduction pathways.
Annu. Rev. Cell. Biol. 10:251–337
134. van Zyl WH, Huang W, Sneddon AA, Stark
M, Camier S, et al. 1992. Inactivation of the
protein phosphatase 2A regulatory subunit
A results in morphological and transcriptional defects in Saccharomyces cerevisiae.
Mol. Cell. Biol. 12:4946–59
135. Vera-Estrella R, Higgins VJ, Blumwald E.
1994. Plant defense response to fungal
pathogens. Plant Physiol. 106:97–102
136. Viard M-P, Martin F, Pugin A, Ricci P, Blein
J-P. 1994. Protein phosphorylation is induced in tobacco cells by the elicitor cryptogein. Plant Physiol. 104:1245–49
137. Walker JC. 1994. Structure and function of
the receptor-like protein kinase of higher
125
plants. Plant Mol. Biol. 26:1599–609
138. Walker RP, Leegood RC. 1995. Purification, and phosphorylation in vivo and in
vitro, of phosphoenolpyruvate carboxykinase from cucumber cotyledons. FEBS
Lett. 362:70–74
139. Weiner H, Stitt M. 1993. Sucrose-phosphate synthase phosphatase, a type 2A protein phosphatase, changes its sensitivity towards inhibition by inorganic phosphatase
in spinach leaves. FEBS Lett. 333:159–64
140. Wolniak SM, Larsen PM. 1992. Changes in
the metaphase transit times and the pattern
of sister chromatid separation in stamen
hair cells of Tradescantia after treatment
with protein phosphatase inhibitors. J. Cell
Sci. 102:691–715
141. Yamano H, Ishii K, Yanagida M. 1994.
Phosphorylation of dis2 protein phosphatase at the C-terminal cdc2 consensus
and its potential role in cell cycle regulation. EMBO J. 13:5310–18
142. Yang C-M, Danko SJ, Markwell JP. 1987.
Thylakoid acid phosphatase and protein
phosphatase activities in wheat (Triticum
aestivum L.). Plant Sci. 48:17–22
143. Zhang K, Tsukitani Y, John CL. 1992. Mitotic arrest in tobacco caused by the phosphoprotein phosphatase inhibitor okadaic
acid. Plant Cell Physiol. 33:677–88
144. Zimmermann S, Thomine S, Guern J, Barbier-Brygoo H. 1994. An anion current at
the plasma membrane of tobacco protoplasts shows ATP-dependent voltage regulation and is modulated by auxin. Plant J.
6:707–16