FEATURE ARTICLE

Molecular basis of lithium action: integration of

lithium-responsive signaling and gene expression
networks
RH Lenox and Le Wang
Molecular Neuropsychopharmacology Program, Department of Psychiatry, University of Pennsylvania School of Medicine,
Philadelphia, PA, USA
The clinical efficacy of lithium in the prophylaxis of recurrent affective episodes in bipolar
disorder is characterized by a lag in onset and remains for weeks to months after
discontinuation. Thus, the long-term therapeutic effect of lithium likely requires reprogram-
ming of gene expression. Protein kinase C and glycogen synthase kinase-3 signal
transduction pathways are perturbed by chronic lithium at therapeutically relevant concentra-
tions and have been implicated in modulating synaptic function in nerve terminals. These
signaling pathways offer an opportunity to model critical signals for altering gene expression
programs that underlie adaptive responses of neurons to long-term lithium exposure. While
the precise physiological events critical for the clinical efficacy of lithium remain unknown, we
propose that linking lithium-responsive genes as a regulatory network will provide a strategy
to identify signature gene expression patterns that distinguish between therapeutic and
nontherapeutic actions of lithium.
Molecular Psychiatry (2003) 8, 135144. doi:10.1038/sj.mp.4001306
Keywords: lithium; gene; protein kinase C (PKC); glycogen synthase-3 (GSK-3); myristoylated
alanine-rich C-kinase sustrate (MARCKS)
Introduction
Lithium (Li

) has been the standard pharmacological

treatment for bipolar disorder (BPD) (manic-depres-
sive illness) over the last 50 years (see reviews:
Goodwin and Ghaemi,
1
Lenox and Manji,
2
and Lenox
and Hahn
3
). Dysregulation of the balance of activation
of signaling pathways in critical brain regions such as
limbic and associated cortical/subcortical areas may
underlie the often-wild oscillations in the behavioral
states of mania and depression in patients genetically
predisposed to BPD.
4,5
Furthermore, it is thought that
once the disease has been triggered and clinically
manifest in the absence of treatment, long-term
compensatory changes in the central nervous system
set the stage for more frequent and severe affective
episodes over time.
6
The clinical efficacy of lithium
in preventing recurrent affective episodes requires a
lag period for onset and is reversed upon disconti-
nuation of treatment only after weeks or months.
79
By
virtue of its prophylactic properties, lithium is
thought to target the underlying pathophysiology
of the disease, yet the precise molecular mechanism
for this therapeutic action remains elusive. Evi-
dence from both in vitro and in vivo studies has
demonstrated that lithium exerts multiple effects on
neurotransmitter/receptor-mediated signaling, ion
transport, signal transduction cascades, hormonal
and circadian regulation, and profoundly alters gene
expression patterns (see reviews Lenox and Hahn,
3
Lenox et al,
10
Manji and Lenox,
11
Jope,
12
Williams and
Harwood,
13
and Manji and Lenox
14
). Intense interest
has been focused upon phosphoinositide (PI) meta-
bolism and glycogen synthase kinase-3 (GSK-3)-
mediated signal transduction that are perturbed by
chronic lithium at therapeutically relevant concentra-
tions.
13
Lithium acting through these pathways is
thought to provide initial signals that could elicit
extensive reprogramming of gene expression, a para-
digm necessary for the long-term therapeutic effect of
lithium treatment.
15
Furthermore, since the activity of
a given gene is likely regulated by a chain of proteins
encoded by other genes, a signature gene network
may be used to better represent the reprogrammed
lithium-responsive gene response. Identifying such a
lithium-responsive gene network in brain would
allow us to distinguish between subsets of genes
underlying therapeutic and nontherapeutic actions of
lithium. In this article, we discuss the rationale for
using a well-defined neurobiologically and pharma-
cologically relevant target to identify a lithium-
responsive element and critical transcription factor(s)
that may ultimately define a network of genes
Received 3 July 2002; revised 23 October 2002; accepted 28
October 2002
Correspondence: Dr RH Lenox, Mailstop: BWL-103A, CNS Drug
Discovery Program, Aventis Pharmaceuticals, Route 202-206
North, Bridgewater, NJ 08807, USA.
E-mail: Robert.Lenox@aventis.com
Molecular Psychiatry (2003) 8, 135144
& 2003 Nature Publishing Group All rights reserved 1359-4184/03 $25.00
www.nature.com/mp
implicated in mood stabilization for the treatment of
bipolar disorder.
Lithium and phosphoinositide/protein kinase C
signaling pathway
Lithium has been shown to be an inhibitor of a
number of structurally similar magnesium-dependent
phosphomonoesterases at K
i
values within the
therapeutically relevant range of concentrations
(0.81.2 mM).
16,17
It was recognized over two decades
ago that lithium is a potent inhibitor of the intracel-
lular enzyme, inositol monophosphatase (IMPase)
(K
i
0.8 mM), which converts inositol monopho-
sphate to inositol.
1820
Biochemical and Genetic
studies subsequently identified the upstream inositol
polyphosphatase as an additional target for
lithium.
21,22
Thus, receptor G-protein-coupled PI
hydrolysis has been extensively investigated as a site
for the action of lithium as a mood stabilizer (see
review Manji and Lenox;
23
see Figure 1). Furthermore,
since there is evidence that the mode of enzyme
inhibition of IMPase is uncompetitive, the preferen-
tial site of action for lithium was proposed to be the
most overactive receptor-mediated neuronal path-
ways undergoing the highest rate of phosphatidyl
(4,5) bisphosphate (PIP
2
) hydrolysis.
24,25
While it was
posited that lithium produces its therapeutic effects
via a consequent reduction in relative concentrations
of myo-inositol and PIP
2
concentrations, data-driven
support for this hypothesis has been highly depen-
dent upon the cell and animal models under
investigation.
2631
Much of this inconsistency may
be related to the relatively small size of the signal-
Figure 1 Lithium-responsive signal transduction pathways focused on PI/PKC and GSK-3 signaling. Lithium directly
inhibits the enzyme inositol monophosphate phosphatase (IMPase; K
i
E0.8 mM). Binding of ligands to G-protein-coupled
receptors (GPCRs) activates PLC and causes hydrolysis of PIP
2
to inositol-1,4,5 trisphosphate (IP
3
) and DAG. IP
3
stimulates
Ca
2
release from cellular store, and DAG activates DAG-dependent protein kinase C (PKC). In the presence of receptor
ligands, long-term inhibition of IMPase results in a depletion of myo-inositol (myo-I) and an accumulation of DAG followed
by a downregulation of PKC isozymes a and e. On the other hand, lithium is a potent inhibitor of glycogen synthase kinase-3b
(GSK-3b; K
i
E2 mM) leading to the stabilization of b-catenin, which enters the nucleus to activate LEF/TCF-dependent genes.
Lithium-responsive gene network is hypothetically proposed to be activated through PKC and GSK-3 signaling pathways and
their crosstalk at therapeutically relevant concentrations. Abbreviations: A, receptor agoinst; Akt/PKB, a serine/threonine
kinase; APC, adenomatous polyposis protein; axin, a homolog of Drosophila product of the fused locus; CMP-PA, cytidine
monophosphate-phosphatidate; DAG, diacylglycerol; Dsh, dishevelled; G, G protein; GBP, GSK-3 binding protein; GSK-3,
glycogen synthase kinase-3; IP3, inositol-1,4,5-trisphosphate; LEF/TCF, LEF(lymphoid enhancer element)/TCF(T cell factor-
1) family transcription factors; LRE, lithium-responsive promoter element; MUNC, a synaptic protein that contains DAG
binding domain; PI, phosphatidylinositol; PIP
2
, phosphatidylinositol 4,5 bisphosphate; PP2A, protein phosphatase 2A; PLC,
phospholipase C; Wnt, a secreted glycoprotein ligand for Wnt/b-catenin signaling pathway.
Molecular basis of lithium action
RH Lenox and Le Wang
136
Molecular Psychiatry
dependent pools of myo-inositol and PIP
2
, as well as
the fact that the lithium-induced reduction of myo-
inositol appears to be dependent upon multiple
factors including cell-type and tonic activity of the
receptor-coupled PI signaling pathway.
3133
It is of
interest in this regard that a significant lithium-
induced reduction in myo-inositol levels in the right
frontal lobe has been observed in bipolar patients
prior to clinical response using proton magnetic
resonance spectroscopy.
34
However, these and other
data suggest that while inhibition of IMPase may
represent an initial effect of lithium, reducing myo-
inositol levels may be more critical to the specificity
of the cellular site of action for lithium than its long-
term therapeutic efficacy (see review, Lenox and
Manji).
2
Such data in turn supported the hypothesis that
short-term lithium exposure in selective neuronal
populations in brain undergoing heightened activa-
tion would result in diacylglycerol (DAG)-mediated
activation of protein kinase C (PKC), subsequent
downregulation of PKC isozymes, and long-term
downstream changes in brain.
5,35
PKC isozymes have
been implicated in the regulation of neurotransmitter
release, neuronal excitability, and long-term changes
in PKC-regulated protein function, neuroplasticity,
and gene expression. PKC activity is tightly regulated
by virtue of not only the distinct distribution of its
family of isozymes, but its required translocation to
the membrane associated with phosphorylation and
binding to a receptor for activated C-kinase (RACK)
proteins, and ultimate autocatalysis upon prolonged
activation. Several laboratories including our own
have demonstrated that short-term lithium treatment
activates PKC, and long-term lithium treatment down-
regulates PKC isozymes in a PI-signaling-dependent
manner in brain.
5,35
Studies of chronic lithium (1mM)
exposure in both in vivo and in vitro models from our
laboratory and others have demonstrated a reduction
in PKC isozymes a and e in rat subiculum and in CA1
regions of the hippocampus, which has been repli-
cated in immortalized hippocampal cell model
system.
19,36,37
Furthermore, administration of myo-
inositol to rats has been reported to reverse the
downregulation of PKCe in brain following chronic
lithium. These studies have led to investigations
examining PKC regulation in man (see reviews, Manji
and Lenox,
19,35
Jope and Song,
38
Wang et al,
39
Soares
et al
40
). Recent studies using human post-mortem
brain homogenates revealed an increased association
of the receptor for activated C kinase-1 (RACK1) with
PKC isozymes in frontal cortex of subjects with BPD,
suggesting that interactions between these proteins
may be also altered in bipolar disease.
37
In addition,
there is intriguing evidence that tamoxifen, a drug
known to inhibit PKC in vitro, was effective in an
initial clinical study of acutely manic patients with
BPD.
41
Thus, it appears that chronic lithium-depen-
dent modulation of receptor-coupled PIPKC signal-
ing pathways may contribute to the long-term action
of lithium in the brain.
Lithium and Wnt signaling pathway
Glycogen synthase kinase 3b (GSK-3b) is a serine/
threonine kinase that is constitutively active in cells
and negatively regulates its substrates, one of which is
b-catenin, a downstream effector of the Wnt signaling
pathway controlling dorsalventral axis specification
and cell fate determination in various organisms
including Dictyostelium, sea urchins, zebrafish, and
Xenopus.
17
GSK-3 activity is regulated through multi-
ple proteins in a complex (Figure 1), where GSK-3 is
activated in the presence of axin, adenomatous
polyposis coli (APC), dishelveled (Dsh) and pro-
motes phosphorylation of b-catenin for ubiquitin
proteosome-mediated degradation.
43
On the contrary,
GSK-3 binding protein (GBP), which is required
for axis formation in Xenopus, could join the comp-
lex to inhibit GSK-3 activity, in part, by preventing
axin from binding GSK-3.
44
When b-catenin
accumulates in cytoplasm, it translocates into nu-
cleus and activates T-cell factor (TCF) and lymphoid
enhancer element (LEF)-dependent gene transcrip-
tion, through which the Wnt pathway controls
numerous developmental processes. The full reper-
toire of the genes containing LEF/TCF responsive
elements in their promoters has not been defined.
Patterning genes Ultrabithorax in Drosophila, sia-
mois, and nodal-related gene-3 in Xenopus,
4547
hu-
man genes c-myc and cyclin D1,
4850
and connexin43
and E-cadherin
51,52
represent major LEF/TCF-respon-
sive genes discovered to date. Other possible
targets of GSK-3 may include AP-1, CREB, NF-kB,
heat shock protein 1, and CCAAT/enhancer
binding proteins.
53
LEF1 was recently found to be
critical to the generation of dentate gyrus granule
cells and the development of the hippocampus in
mice.
4547,54
In addition, GSK-3 could phosphorylate
NF-AT, facilitating its export from the nucleus,
and thus antagonizing NF-AT-dependent gene tran-
scription.
55
Lithium mimics Wnt signals by inhibition of GSK-3
both in vitro and in vivo. This suggests that the often-
observed developmental effects of lithium, which
could not be explained by inhibition of IMPase, may
be in fact a result of inhibition of GSK-3b.
56,57
Valproic
acid also inhibits GSK-3,
58
but its control over the
b-catenin stability differs from lithium. While lithium
stabilizes b-catenin via inhibition of GSK-3, VPA-
induced b-catenin accumulation is through increased
expression of b-catenin rather than stabilization of
the protein.
59
GSK-3 has also been implicated in
synaptic function. For example, GSK-3b inhibition
induces the synapsin I clustering in the formation of
synapses and neurotransmitter release.
60
In addition,
GSK-mediated phosphorylation of MAP-B and tau
proteins regulate microtubule assembly and stabiliza-
tion at synapses.
57,6163
Clinical studies have demon-
strated that specific GSK-3 inhibitors mimic the
therapeutic action of mood stabilizers and might
therefore be plausible drugs in treating bipolar
patients.
64
Molecular basis of lithium action
RH Lenox and Le Wang
137
Molecular Psychiatry
Crosstalk between PKC and GSK-3b signaling
pathways
Certain PKC isoforms have been shown to phosphor-
ylate and inactivate GSK-3b in vitro.
65
However, Wnt-
induced b-catenin accumulation is only partially
inhibited in vivo by PKC inhibitors and by chronic
treatment of cells with phorbol ester.
66,67
This sug-
gests that Wnt/b-catenin pathway may comprise two
components, one is mimicked by lithium and the
other involves PKC. Further studies demonstrated
that some PKC isoforms (TPA-sensitive) are involved
in the Wnt-induced GSK-3b inactivation, whereas
others (atypical PKC) contribute to b-catenin degrada-
tion.
43,67
On the other hand, certain Wnt and Frizzled
homologs have been reported to regulate PKC differ-
entially; i.e. some stimulate PKC but have no effect on
expression of b-catenin target genes, while others,
capable of activating b-catenin target genes, do not
activate PKC.
68
New data also establish that Frizzleds
are bonafide G-protein-coupled receptors.
69,70
For
example, Frizzled-1 couples via G-protein G
o
and G
q
to the b-cateninLEF-TCF pathway. Frizzled-2 cou-
ples via G
q
and G
t
to downstream effectors including
Ca
2
mobilization. In addition, the disheveled pro-
teins may also mediate phosphoinositol signal trans-
duction, leading to the activation of PKC.
13
Thus, the
mutual interaction between several components of
PKC and GSK-3 signaling pathways suggests a model
of crosstalk between the two pathways (Figure 1). In
view of the fact that the relative K
i
for lithium
inhibition of IMPase vs that for GSK-3b favors an
interaction of lithium in the PI/PKC signaling path-
way in brain, chronic lithium might interact with
GSK-3b in vivo by virtue of crosstalk at therapeuti-
cally relevant concentrations.
Lithium and synaptic function
While there is evidence for effects of lithium on the
expression of numbers of genes in brain, including
the DNA binding and transactivation of AP-1 tran-
scription factors
71
and both pro- and antiapoptotic
genes through the AKT/PKB signaling pathway,
72,73
for the purpose of this discussion we will focus upon
the long-term action of lithium in the brain that stems
from its interaction with the PI/PKC signaling path-
way. There is significant evidence that alterations in
downstream targets in this pathway have functional
consequences related to synaptic signaling in the
brain, which actually may be amplified through
crosstalk with GSK-3b signaling as noted above.
PKC isozymes have been implicated in neurotrans-
mitter release, and a number of synaptic proteins are
possible substrates for PKC. Phorbol ester (PMA) has
been shown to evoke synaptic potentiation (PESP) at
the nerve terminal by presynaptic PKC acting on the
release machinery.
74
Inhibition of protein kinase C at
the presynaptic terminal attenuates the phorbol ester-
induced synaptic potentiation (PESP), supporting a
role for PKC in synaptic potentiation.
74
However,
chelating presynaptic Ca
2
does not significantly
affect PESP, indicating that only certain PKC isoforms
such as PKCe, whose activation does not require Ca
2
,
are important in mediating the neurotransmitter
release at nerve endings. In fact, chronic lithium-
facilitated neurotransmitter release has been linked to
the isoform-specific decrease in PKC a and e in the
absence of significant alterations in other PKC iso-
forms.
23
Synaptic transmission is regulated by two
mechanisms: an increase in the Ca
2
sensitivity of
exocytosis (release probability) and an increase in the
amount of releasable synaptic vesicles (the pool size).
DAG, which activates PKC, specifically increases the
pool size but not the release probability. Recently, a
new family of proteins, Munc13/unc13 proteins,
initially identified in C. elegans, was found to contain
DAG receptor. This raises an interesting possibility
that chronic lithium action in part may be mediated
through interaction of DAG with Munc proteins. In
support of this hypothesis, presynaptic loading of a
synthetic peptide with the sequence of the N-terminal
domain of Doc2a interacting with Munc13 attenuates
PESP.
75
In addition, Munc18-1 was found to interact
with syntaxin1, a member of SNARE complex that
promotes large dense-core vesicle docking.
76
There-
fore, lithium-induced changes in synaptic transmis-
sion can be mediated by a complex network of
synaptic proteins through both PKC and Munc
signaling pathways. In addition to direct modulation
of Munc through DAG, lithium has also been shown
to enhance the expression of a gene encoding cysteine
string proteins (CSPs) at therapeutically relevant
concentrations.
77
CSPs are novel synaptic vesicle
components conserved in evolution,
78
which also
interact with the SNARE neurotransmitter release
complex.
79
Lithium and MARCKS: a downstream target
for PI/PKC signaling
Accumulating evidence over the past several years
has identified a target in the brain for the action of
chronic lithium that is in the PI/PKC signaling
cascade and possesses pharmacological characteris-
tics consistent with the therapeutic properties of
lithium in man. Activation of PKC results in the post-
translational phosphorylation of various proteins,
most prominent of which is the myristoylated
alanine-rich C-kinase substrate (MARCKS) in brain
(Blackshear, 1993). MARCKS has been implicated in
processes requiring signal-dependent changes in
actin-membrane plasticity and cytoskeletal restruc-
turing.
80,81
It can crosslink actin, bind calcium/
calmodulin, interact with the plasma membrane,
and has been linked to organization of polypho-
sphoinositide (PI (4,5)P
2
) signaling domains,
82,83
regulation of phospholipase D,
84
secretion,
85,86
and
phagocytosis.
87
MARCKS is preferentially expressed
in dendritic branches and axon terminals within
limbic and limbic-associated regions of brain, as well
as neuronal growth cones necessary for normal brain
Molecular basis of lithium action
RH Lenox and Le Wang
138
Molecular Psychiatry
development.
8890
Within brain and neurons in cul-
ture, MARCKS protein is expressed in neurites and
synaptosomes, and is colocalized with synaptic
vesicles.
9193
PKC-MARCKS signaling has been im-
plicated in vesicular trafficking of neurotransmitter in
living neurons,
94
and in the regulation of various
processes including synaptic transmission at nerve
terminals.
80,81
Electrophysiological and behavioral
studies in heterozygote mutant mice have shown that
50% reduction in MARCKS significantly affects
long-term potentiation (LTP) and hippocampally
dependent behavior.
9597
These data indicate that
MARCKS plays an important role in the mediation
of neuroplastic processes in the developing and
mature CNS.
Chronic treatment with lithium at therapeutically
relevant concentrations significantly downregulates
MARCKS protein and mRNA (50%) in both rat brain
and immortalized hippocampal cells.
27,98,99
Direct
activation of PKC by phorbol esters in immortalized
hippocampal cells also downregulates the MARCKS
protein,
100
suggesting a role for PKC in the regulation
of MARCKS gene (Macs) expression in brain. The
lithium-induced downregulation of MARCKS protein
is dependent upon the myo-inositol concentration
and the level of activation of receptor-coupled PI
signaling.
31
Addition of inositol completely reverses
the lithium-induced downregulation of MARCKS,
indicating that the downregulation of MARCKS was
mediated via the phosphoinositol pathway.
31
Further-
more, lithium-induced down regulation of MARCKS
is only apparent after chronic, but not acute, admin-
istration and persists beyond abrupt withdrawal of
the drug for an extended period of time; paralleling
the clinical time course for the therapeutic effects of
lithium during initial treatment as well as its
discontinuation. Of interest, the structurally unre-
lated mood-stabilizer, valproic acid (VPA), shares the
property of lithium in reducing the MARCKS expres-
sion in hippocampus, but carbamazepine (CBZ) and
other psychotropic agents (antianxiety, analgesic,
antipsychotic, antidepressant, calcium-channel
blocker) as well as other monovalent cations such as
rubidium do not alter MARCKS regulation.
101,102
On
the other hand, VPA-induced downregulation of
MARCKS is inositol-independent, and has an addi-
tive effect on MARCKS regulation, consistent with
clinical data supporting greater efficacy of the
combined treatment in refractory bipolar patients.
101
These findings point to a common mechanism, in
which MARCKS may be a shared target for both
lithium and VPA. Thus, we have proposed that the
reduction of MARCKS protein following long-term
lithium administration alters pre/postsynaptic mem-
brane structure and stabilizes aberrant neuronal
signaling in key brain regions of patients with
BPD.
3,5
The regulation of MARCKS expression repre-
sents a highly valued target for the long-term action of
mood stabilizers, not only by virtue of its underlying
neurobiological properties but also in light of its
pharmacological sensitivity and selectivity for struc-
turally unrelated drugs with efficacy in the prophy-
lactic treatment of BPD. While MARCKS remains an
attractive target for illustrative purposes in defining a
lithium-responsive gene network, it is evident that
other targets, such as prolyl oligopeptidase (POase),
which appears to mediate the common effect of
lithium, CBZ and VPA on growth cone stability and
has been implicated in affective disorders, may at
some point represent a similar opportunity.
103
Perspective: linking the lithium-responsive genes
as a network
The mechanisms underlying the prophylactic treat-
ment of BPD by lithium are likely to be hardwired in
the genomic DNA. Despite all the reports of how
individual gene expression can be modulated in
response to lithiums exposure, there is as yet no
strategy available for the identification of the lithium-
responsive genomic regulatory network in brain.
Many studies have focused on determining the effect
of lithium on one or a few genes at a time, an
approach that is not adequate for the analysis of large
regulatory control system organized as networks.
Gene-specific expression is controlled by specific
cis-regulatory target sequence embedded in promoters
and the cognate binding transcription factors encoded
by a set of regulatory genes. The functional linkages of
such a genomic regulatory network include elements
that exhibit multiple interactions between the outputs
of regulatory genes and corresponding cis-regulatory
genomic sequences. Thus, identifying networks of
transcription factors and the genes they regulate in
response to lithium exposure is important to under-
stand the biological responsiveness of an organism to
lithium and the prophylactic properties of its action
in brain.
Studies in our laboratory have demonstrated that
the lithium-induced reduction of MARCKS protein
was accompanied by a downregulation of MARCKS
mRNA with no evidence for a change in the half-life
of the mRNA.
99
Furthermore, synthesis of nascent
RNA for MARCKS and the Macs promoter activity
were also found to be significantly reduced in
chronic, but not acute, lithium-treated immortalized
hippocampal cells. Both reductions were signifi-
cantly enhanced in the presence of activation of
receptor-coupled PI signaling.
99
We have identified
for the first time a lithium-responsive promoter
sequence located in the upstream region (993/
713) of the Macs promoter.
99
The mutant promoter
lacking the 993/713 fragment not only did not
respond to chronic lithium expose but also had a
significantly reduced promoter activity, suggesting
that chronic lithium represses the transcriptional
activator(s) bound to this region. Until now, however,
no lithium-responsive transcription factor directly
bound to this region has been identified. While
specific transcription factors that bind to lithium-
responsive promoter element (LRE) remain to be
identified (Figure 2), current data suggest that the
Molecular basis of lithium action
RH Lenox and Le Wang
139
Molecular Psychiatry
mechanism by which chronic lithium represses the
Macs gene transcription are likely through: (a)
lithium-induced downregulation of DNA binding
activity and/or expression of transcription activator(s)
interacting with LRE; (b) lithium-induced down-
regulation of activator function of transcription
activator(s) acting at LRE; (c) lithium-induced disrup-
tion of the coordinate interaction between the distal
activating sequence-bound activator(s) and the prox-
imal core promoter sequence-bound basal transcrip-
tional machinery. The lithium-responsive 993/713
fragment contains significant enhancer/activator DNA
elements that are necessary to sustain the optimal
Macs gene transcription in cells.
104
While an atypical
Sp1 site, characterized by the presence of a prominent
GA-rich sequence, was identified within the 993/
713, a classical Sp1 site was found in the middle of
the GC-rich box close to a potential Z-DNA-forming
segment near the transcription initiation site.
104
Presence of Z-DNA forming segment signifies both
compositional and conformational changes influen-
cing the genomic landscape, the accessibility of cis-
acting elements, and the activation of gene transcrip-
tion in the core promoter region.
105
As summarized in
Figure 2, Macs promoter lacks typical TATA box,
and in the absence of a TATA box multiple Sp1 sites
in the promoter may control the transcription of Macs
gene.
A recent microarray experiment further showed
that the expression of as many as 37 genes from 4132
rat genes is altered during the chronic lithium
treatment.
106
If these data were extrapolated to the
entire genome, there would be as many as 750 genes
that could potentially be regulated by lithium.
However, the majority of such studies using differ-
ential display and microarray analysis do not distin-
guish between direct activation of target genes by a
transcription factor and indirect effects resulting from
one transcription factor inducing the expression of a
second. Furthermore, the functional value of the vast
majority of these potential targets remains unknown.
Using our knowledge regarding a high-value pharma-
cologically relevant target such as MARCKS for the
action of chronic lithium in brain to identify lithium-
responsive elements that could serve to link lithium-
responsive genes as a regulatory network is both
intriguing and challenging. To establish such a gene
network, some studies have coupled the overexpres-
sion of a given transcription factor with microarray
experiments. However, the genes affected by the
overexpressed transcription factors may not represent
the true targets of those factors under physiological
conditions. Recently, using finite perturbations of
gene expression in conjunction with microarray
experiments that may mimic physiological conditions
has been advocated; thus differing from the drastic
Figure 2 Lithium-responsive promoter of the Macs gene. The Promoter region upstream of Macs is pharmacologically
responsive to chronic, but not acute, lithium exposure. A hypothetical activator complex binds to the upstream region of
Macs promoter and crosstalk with proteins in proximal promoter region including TBP, TAFs, Sp1, hypothetical tethering
proteins, etc. The gray rectangle box represents cis-regulatory enhancer elements (LRE) that may interact with specific
lithium-responsive transcription factors. Long-term treatment with lithium is postulated to repress the Macs transcription
through: (a) lithium-induced reduction in the DNA binding of the transcription activator(s) acting at the LRE; (b) lithium-
induced alteration in the expression and/or the transactivation function of lithium-responsive transcription activator(s)
acting at the LRE; (c) lithium-induced disruption of the interaction between enhancer complex formed at LRE and the
proximal transcription initiator. In the figure, # refers to potential intermediary factors mediating the indirect action of
chronic lithium on the Macs promoter.
Molecular basis of lithium action
RH Lenox and Le Wang
140
Molecular Psychiatry
changes incurred during transgenic overexpression or
knockouts.
107
In such studies, one can modulate
single gene expression using techniques such as
RNA interference (RNAi) one at a time and in a
systematic fashion among all the genes of interest.
Once the perturbed systems are in a new steady state,
the levels of gene expression is measured against that
of reference by microarray or Taqman RCR. The
resulting data are arranged as a regulatory strength
matrix, from which a regulatory network exhibiting
genegene interactions could be inferred using ap-
proaches adapted from metabolic control analysis.
108
The method, however, lacks genome-wide efficiency
and does not provide direct evidence of in vivo
interaction of given transcription factors with their
respective cis-regulatory elements across the genome.
To address these questions, a new technology known
as chIpchip was developed, in which chromatin
immunoprecipitation (chIp) is coupled with micro-
array experiment (chip) to accelerate the genome-
wide detection of DNAprotein interactions in real
time and in real space. Using such a combined
approach as chIpchip, genome-wide mapping of
DNA binding sites for transcription factors (STE12,
GAL4, RAP1, SCB, MCB, MCM1, SFF, and SW15) has
been achieved in yeast.
109,110
This experimental
approach has also been successfully extended to
human genome, where DNA binding sites for E2F
and GATA-1 transcription factors are mapped genome
wide.
111113
A lithium-responsive gene network will offer an
opportunity to define a pathway associated with the
long-term prophylactic properties of lithium, distinct
from its side-effect profile, which will drive the
discovery of novel agents for stabilization of mood
in patients with BPD (Figure 3). Recently, a regulatory
gene network that directs specific developmental
events has been identified in developing sea urchin
embryo.
114,115
This provides a heuristic model for
constructing a lithium-responsive gene network as
a means to identify signature genes that direct
the therapeutic or nontherapeutic action of lithium.
While chromosome immunoprecipitation could start
with antibodies against several known lithium-
responsive transcription factors such as AP-1, CREB,
NF-kB, LEF/TCF, these are general transcription
Figure 3 Model for a gene network for lithium-responsive genes: defining the therapeutic effect of chronic lithium. A gene
network can be reconstructed by perturbation microarray and chlpchip methods (see test for details). A lithium-responsive
gene network will consist of a distinctive subset of genes that share a lithium-responsive element (LRE) as defined within the
Macs gene which is regulated by specific transcription factors referred to as TFs that are yet to be identified. Such a lithium-
responsive gene network (TF1) may subserve critical processes in the brain underlying cytoskeletal remodeling and
regulation of synaptic signaling, while a different subset of genes defined by lithium-responsive elements regulated by
transcription factors (TF2, TF3) may underlie processes such as cell proliferation. Since these different network of genes
underlie different physiological sequela of chronic lithium, modulation of gene expression involved in the cytoskeletal
restructuring and neuroplasticity by lithium may prove to be fundamental to the mood-stabilizing properties of lithium in the
brain, whereas regulation of genes involved in cell proliferation and immune response may be linked to lithiums
leukocytotic and antiviral effects.
Molecular basis of lithium action
RH Lenox and Le Wang
141
Molecular Psychiatry
factors that elicit far more gene expression than
necessary for the therapeutic action of lithium. Thus,
identifying lithium-responsive transcription factors
or promoter elements from highly valued targets such
as the Macs gene becomes extremely desirable. We
demonstrated that Macs promoter is pharmacologi-
cally responsive to chronic, but not acute, lithium
treatment at therapeutically relevant concentrations,
and possesses pharmacological properties consistent
with its expressed protein and clinical properties of
lithium.
99
We further showed that the sites for lithium
responsiveness lie within an enhancer element of the
promoter that has the potential to interact with
a unique lithium-responsive activator complex
(Figure 3).
104
This raises the possibility that unique
transcription factors sensitive to lithium treatment
may be identifiable through this promoter. While
genes like Macs involved in the cytoskeletal restruc-
turing, synaptic transmission, and neuroplasticity
may relate preferentially to the long-term mood-
stabilizing properties of chronic lithium in the brain,
genes underlying cell proliferation and immune
response may serve to identify pathways associated
with lithiums leukocytotic and antiviral effects
(Figure 3). Thus, defining a lithium-responsive gene
network will provide the data for pathway mapping of
novel targets with better-defined mood-stabilizing
properties for the long-term treatment of BPD.
References
1 Goodwin FK, Ghaemi SN. The impact of the discovery of lithium
on psychiatric thought and practice in the USA and Europe. Aust
NZ J Psychiatry 1999; 33(Suppl): S54S64.
2 Lenox RH, Manji HK. Lithium. In: Nemeroff C, Schatzberg A
(eds). APA Textbook of Psychopharmacology. Washington, DC:
APA Press, Inc: 1998, pp. 303314.
3 Lenox RH, Hahn CG. Overview of the mechanism of action of
lithium in the brain: fifty-year update. J Clin Psychiatry 2000; 61:
515.
4 Hudson CJ, Young LT, Li PP, Warsh JJ. CNS signal transduction in
the pathophysiology and pharmacotherapy of affective disorders
and schizophrenia. Synapse 1993; 13: 278293.
5 Lenox RH. Role of receptor coupling to phosphoinositide
metabolism in the therapeutic action of lithium. In: Ehrlich Y
(ed). Molecular Mechanisms of Neuronal Responsiveness. New
York: Plenum. Adv Exp Med 1987; 221: 515530.
6 Post RM. Transduction of psychosocial stress into the neuro-
biology of recurrent affective disorder. Am J Psychiatry 1992; 149:
9991010.
7 Suppes T, Baldessarini RJ, Faedda GL, Tohen M. Risk of
recurrence following discontinuation of lithium treatment in
bipolar disorder. Arch Gen Psychiatry 1991; 48: 10821088.
8 Faedda GL, Tondo L, Baldessarini RJ, Suppes T, Tohen M.
Outcome after rapid vs gradual discontinuation of lithium
treatment in bipolar disorders. Arch Gen Psychiatry 1993; 50:
448455.
9 Tondo L, Baldessarini RJ, Hennen J, Floris G. Lithium main-
tenance treatment of depression and mania in bipolar I and
bipolar II disorders. Am J Psychiatry 1998; 155: 638645.
10 Lenox RH, McNamara RK, Papke RL, Manji HK. Neurobiology of
lithium: an update. J Clin Psychiatry 1998; 59: 3747.
11 Manji HK, Lenox RH. Lithium: a molecular transducer of mood-
stabilization in the treatment of bipolar disorder. Neuropsycho-
pharmacology 1998; 19: 161166.
12 Jope RS. Anti-bipolar therapy: mechanism of action of lithium.
Mol Psychiatry 1999; 4: 117128.
13 Williams RS, Harwood AJ. Lithium therapy and signal transduc-
tion. Trends Pharmacol Sci 2000; 21: 6164.
14 Manji HK, Lenox RH. Signaling: cellular insights into the
pathophysiology of bipolar disorder [In Process Citation]. Biol
Psychiatry 2000; 48: 518530.
15 Hyman SE, Nestler EJ. Initiation and adaptation: a paradigm for
understanding psychotropic drug action. Am J Psychiatry 1996;
153: 151162.
16 York JD, Ponder JW, Majerus PW. Definition of a metal-
dependent/Li(+)-inhibited phosphomonoesterase protein family
based upon a conserved three-dimensional core structure. Proc
Natl Acad Sci USA 1995; 92: 51495153.
17 Phiel C, Klein P. Molecular targets of lithium action. Annu Rev
Pharmacol Toxicol 2001; 41: 789813.
18 Hallcher LM, Sherman WR. The effects of lithium ion and other
agents on the activity of myo-inositol-1-phosphatase from bovine
brain. J Biol Chem 1980; 255: 1089610901.
19 Sherman WR, Gish BG, Honchar MP, Munsell LY. Effects of
lithium on phosphoinositide metabolism in vivo. Fed Proc 1986;
45: 26392646.
20 Irvine RF. Nuclear lipid signaling. Sci STKE 2002; 2002: RE13.
21 Majerus PW. Inositol phosphate biochemistry. Annu Rev Bio-
chem 1992; 61: 225250.
22 Acharya JK, Labarca P, Delgado R, Jalink K, Zuker CS. Synaptic
defects and compensatory regulation of inositol metabolism in
inositol polyphosphate 1-phosphatase mutants. Neuron 1998; 20:
12191229.
23 Manji HK, Lenox RH. Ziskind-Somerfeld Research Award.
Protein kinase C signaling in the brain: molecular transduction
of mood stabilization in the treatment of manic-depressive
illness. Biol Psychiatry 1999; 46: 13281351.
24 Nahorski SR, Ragan CI, Challiss RA. Lithium and the phosphoi-
nositide cycle: an example of uncompetitive inhibition and its
pharmacological consequences. Trends Pharmacol Sci 1991; 12:
297303.
25 Berridge MJ, Downes CP, Hanley MR. Lithium amplifies agonist-
dependent phosphatidylinositol responses in brain and salivary
glands. Biochem J 1982; 206: 587595.
26 Kendall DA, Nahorski SR. Acute and chronic lithium treatments
influence agoinst and depolarization-stimulated inositol phos-
pholipid hydrolysis in rat cerebral cortex. J Pharmacol Exp Ther
1987; 241: 10231027.
27 Kennedy ED, Challiss RA, Nahorski SR. Lithium reduces the
accumulation of inositol polyphosphate second messengers
following cholinergic stimulation of cerebral cortex slices.
J Neurochem 1989; 53: 16521655.
28 Kennedy ED, Challiss RA, Ragan CI, Nahorski SR. Reduced
inositol polyphosphate accumulation and inositol supply in-
duced by lithium in stimulated cerebral cortex slices. Biochem J
1990; 267: 781786.
29 Kofman O, Belmaker RH, Grisaru, N, Alpert C, Fuchs I, Katz V,
Rigler O. Myo-inositol attenuates two specific behavioral effects
of acute lithium in rats. Psychopharmacol Bull 1991; 27: 185
190.
30 Tricklebank MD, Singh L, Jackson A, Oles RJ. Evidence that a
proconvulsant action of lithium is mediated by inhibition of
myo-inositol phosphatase in mouse brain. Brain Res 1991; 558:
145148.
31 Watson DG, Lenox RH. Chronic lithium-induced down-regula-
tion of MARCKS in immortalized hippocampal cells: potentia-
tion by muscarinic receptor activation. J Neurochem 1996; 67:
767777.
32 Fisher SK, Novak JE, Agranoff BW. Inositol and higher inositol
phosphates in neural tissues: homeostasis, metabolism and
functional significance. J Neurochem 2002; 82: 736754.
33 Gani D, Downes CP, Batty I, Bramham J. Lithium and myo-
inositol homeostasis. Biochim Biophys Acta 1993; 1177:
253269.
34 Moore GJ, Bedchuk JM, Parrish JK, Faulk MW, Arjken CL, Strahl-
Bevacqua J, Marji HK. Temporal dissociation between lithium-
induced changes in frontal lobe myo-inositol and clinical
response in manic-depressive illness. Am J Psychiatry 1999;
156: 19021908.
Molecular basis of lithium action
RH Lenox and Le Wang
142
Molecular Psychiatry
35 Manji HK, Lenox RH. Long-term action of lithium: a role for
transcriptional and posttranscriptional factors regulated by
protein kinase C. Synapse 1994; 16: 1128.
36 Manji HK, Etcheberrigaray R, Chen G, Olds JL. Lithium decreases
membrane-associated protein kinase C in hippocampus: selectiv-
ity for the alpha isozyme. J Neurochem 1993; 61: 23032310.
37 Manji HK, Bersudsky Y, Chen G, Belmaker RH, Potter WZ.
Modulation of protein kinase C isozymes and substrates by
lithium: the role of myo-inositol. Neuropsychopharmacology
1996; 15: 370381.
38 Jope RS, Song L. AP-1 and NF-kappaB stimulated by carbachol in
human neuroblastoma SH-SY5Y cells are differentially sensitive
to inhibition by lithium. Brain Res Mol Brain Res 1997; 50: 171
180.
39 Wang HY, Markowitz P, Levinson D, Undie AS, Freidman E.
Increased membrane-associated protein kinase C activity and
translocation in blood platelets from bipolar affective disorder
patients. J Psychiatr Res 1999; 33: 171179.
40 Soares JC, Chen G, Dippold CS et al. Concurrent measures of
protein kinase C and phophoinositides in lithium-treated bipolar
patients and healthy individuals: a preliminiary study. Psychia-
try Res 2000; 95: 109118.
41 Wang H, Freidman E. Increased association of brain protein
kinase C with the receptor for activated C kinase-1 (RACK1) in
bipolar affective disorder. Biol Psychiatry 2001; 50: 364370.
42 Bebchuk JM, Arfken CL, Dolan-Manji S, Murphy J, Hasanat K,
Manji HK. A preliminary investigation of a protein kinase C
inhibitor in the treatment of acute mania. Arch Gen Psychiatry
2000; 57: 9597.
43 Orford K, Crockett C, Jensen JP, Weissman AM, Byers SW. Serine
phosphorylation-regulated ubiquitination and degradation of
beta-catenin. J Biol Chem 1997; 272: 2473524738.
44 Farr III GH, Ferkey DM, Yost C, Pierce SB, Weaver C, Kimelman
D. Interaction among GSK-3, GBP, axin, and APC in Xenopus axis
specification. J Cell Biol 2000; 148: 691702.
45 McKendry R, Hsu SC, Harland RM, Grosschedl R. LEF-1/TCF
proteins mediate wnt-inducible transcription from the Xenopus
nodal-related 3 promoter. Dev Biol 1997; 192: 420431.
46 Brannon M, Gomperts M, Sumoy L, Moon RT, Kimelman D. A
beta-catenin/XTcf-3 complex binds to the siamois promoter to
regulate drosal axis specification in Xenopus. Genes Dev 1997;
11: 23592370.
47 Riese J, Yu X, Munnerlyn A et al. LEF-1, a nuclear factor
coordinating signaling inputs from wingless and decapentaple-
gic. Cell 1997; 88: 777787.
48 Shtutman M, Zhurinsky J, Simcha I et al. The cyclin D1 gene is a
target of the beta-catenin/LEF-1 pathway. Proc Natl Acad Sci
USA 1999; 96: 55225527.
49 Tetsu O, McCormick F. Beta-catenin regulates expression of
cyclin D1 in colon carcinoma cells. Nature 1999; 398: 422426.
50 He TC, Sparks AB, Rago C et al. Identification of c-MYC as a
target of the APC pathway. Science 1998; 281: 15091512.
51 van der Heyden MA, Rook MB, Hermans MM et al. Identification
of connexin43 as a functional target for Wnt signalling. J Cell Sci
1998; 111: 17411749.
52 Bradley RS, Cowin P, Brown AM. Expression of Wnt-1 in PC12
cells results in modulation of plakoglobin and E-cadherin and
increased cellular adhesion. J Cell Biol 1993; 123: 18571865.
53 Mai L, Jope RS, Li X. BDNF-mediated signal transduction is
modulated by GSK3beta and mood stabilizing agents. J Neuro-
chem 2002; 82: 7583.
54 Galceran J, Miyashita-Lin EM, Devaney E, Rubenstein JL,
Grosschedl R. Hippocampus development and generation of
dentate gyrus granule cells is regulated by LEF1. Development
2000; 127: 469482.
55 Graef IA, Mermelstein PG, Stankunas K et al. L-type calcium
channels and GSK-3 regulate the activity of NF-ATc4 in
hippocampal neurons. Nature 1999; 401: 703708.
56 Hedgepeth CM, Conrad LJ, Zhang J, Huang HC, Lee VM, Klein PS.
Activation of the Wnt signaling pathway: a molecular mechanism
for lithium action. Dev Biol 1997; 185: 8291.
57 Klein PS, Melton DA. A molecular mechanism for the effect of
lithium on development. Proc Natl Acad Sci USA 1996; 93:
84558459.
58 Yuan P-X, Huang L-D, Jiang Y-M, Gutkind JS, Manji HK, Chen G.
The mood stabilizer valproic acid activates mitogen-activated
protein kinases and promotes neurite growth. J Biol Chem 2001;
276: 3167431683.
59 Phiel CJ, Zhang F, Huang EY, Guenther MG, Lazar MA, Klein PS.
Histone deacetylase is a direct target of valproic acid, a potent
anticonvulsant, mood stabilizer, and teratogen. J Biol Chem 2001;
276: 3673436741.
60 Hall AC, Brennan A, Goold RG et al. Valproate regulates
GSK-3-mediated axonal remodeling and synapsin I cluster-
ing in developing neurons. Mol Cell Neurosci 2002; 20:
257270.
61 Lovestone S, Davis DR, Webster MT et al. Lithium reduces tau
phosphorylation: effects in living cells and in neurons at
therapeutic concentrations. Biol Psychiatry 1999; 45: 9951003.
62 Hong M, Chen DC, Klein PS, Lee VM. Lithium reduces tau
phosphorylation by inhibition of glycogen synthase kinase-3.
J Biol Chem 1997; 272: 2532625332.
63 Munoz-Montano JR, Moreno FJ, Avila J, Diaz-Nido J. Lithium
inhibits Alzheimers disease-like tau protein phosphorylation in
neurons. FEBS Lett 1997; 411: 183188.
64 Eldar-Finkelman H. Glycogen synthase kinase 3: an emerging
therapeutic target. Trends Mol Med 2002; 8: 126132.
65 Goode N, Hughes K, Woodgett JR, Parker PJ. Differential
regulation of glycogen synthase kinase-3 beta by protein kinase
C isotypes. J Biol Chem 1992; 267: 1687816882.
66 Chen RH, Ding WV, McCormick F. Wnt signaling to beta-catenin
involves two interactive components. Glycogen synthase kinase-
3beta inhibition and activation of protein kinase. C. J Biol Chem
2000; 275: 1789417899.
67 Cook D, Fry M, Hughes K, Sumathipala R, Woodgett J, Dale T.
Wingless inactivates glycogen synthase kinase-3 via an intracel-
lular signalling pathway which involves a protein kinase C.
EMBO J 1996; 15: 45264536.
68 Sheldahl LC, Park M, Malbon CC, Moon RT. Protein kinase C is
differentially stimulated by Wnt and Frizzled homologs in a G-
protein-dependent manner. Curr Biol 1999; 9: 695698.
69 Malbon CC, Wang H, Moon RT. Wnt signaling and heterotrimeric
G-proteins: strange bedfellows or a classic romance? Biochem
Biophys Res Commun 2001; 287: 589593.
70 Liu T, DeCostanzo AJ, Liu X et al. G protein signaling from
activated rat frizzled-1 to the beta-catenin-Lef-Tcf pathway.
Science 2001; 292: 17181722.
71 Chen G, Hasanat KA, Bebchuk JM, Moore GJ, Glitz D, Manji HK.
Regulation of signal transduction pathways and gene expression
by mood stabilizers and antidepressants. Psychosom Med 1999;
61: 599617.
72 Chen RW, Chuang DM. Long term lithium treatment suppresses
p53 and Bax expression but increases Bcl-2 expression. A
prominent role in neuroprotection against excitotoxicity. J Biol
Chem 1999; 274: 60396042.
73 Chalecka-Franaszek E, Chuang DM. Lithium activates the serine/
threonine kinase Akt-1 and suppresses glutamate-induced in-
hibition of Akt-1 activity in neutrons. Proc Natl Acad Sci USA
1999; 96: 87458750.
74 Berglund K, Midorikawa M, Tachibana M. Increase in the pool
size of releasable synaptic vesicles by the activation of protein
kinase C in goldfish retinal bipolar cells. J Neurosci 2002; 22:
47764785.
75 Hori T, Takai Y, Takahashi T. Presynaptic mechanism for phorbol
ester-induced synaptic potentiation. J Neurosci 1999; 19: 7262
7267.
76 Voets T, Toonen RF, Brain EC et al. Munc18-1 promotes large
dense-core vesicle docking. Neuron 2001; 31: 581591.
77 Cordeiro ML, Umbach JA, Gundersen CB. Lithium ions enhance
cysteine string protein gene expression in vivo and in vitro. J
Neurochem 2000; 74: 23652372.
78 Buchner E, Gundersen CB. The DnaJ-like cysteine string protein
and exocytotic neurotransmitter release. Trends Neurosci 1997;
20: 223227.
79 Heckmann M, Adelsberger H, Dudel J. Evoked transmitter release
at neuromuscular junctions in wild type and cysteine string
protein null mutant larvae of Drosophila. Neurosci Lett 1997; 228:
167170.
Molecular basis of lithium action
RH Lenox and Le Wang
143
Molecular Psychiatry
80 Aderem A. The MARCKS brothers: a family of protein kinase C
substrates. Cell 1992; 71: 713716.
81 Blackshear PJ. The MARCKS family of cellular protein kinase C
substrates. J Biol Chem 1993; 268: 15011504.
82 Laux T, Fukami K, Thelen M, Golub T, Frey D, Caroni P. GAP43,
MARCKS, and CAP23 modulate PI(4,5)P(2) at plasmalemmal
rafts, and regulate cell cortex actin dynamics through a common
mechanism. J Cell Biol 2000; 149: 14551472.
83 Glaser M, Wanaski S, Buser CA et al. Myristoylated alanine-rich
C kinase substrate (MARCKS) produces reversible inhibition of
phospholipase C by sequestering phosphatidylinositol 4,5-bi-
sphosphate in lateral domains. J Biol Chem 1996; 271: 26187
26193.
84 Morash SC, Rose SD, Byers DM, Ridgway ND, Cook HW.
Overexpression of myristoylated alanine-rich C-kinase substrate
enhances activation of phospholipase D by protein kinase C in
SK-N-MC human neuroblastoma cells. Biochem J 1998; 332:
321327.
85 Goodall AR, Turner NA, Walker JH, Ball SG, Vaughan PF.
Activation of protein kinase C-alpha and translocation of the
myristoylated alanine-rich C-kinase substrate correlate with
phorbol ester-enhanced noradrenaline release from SH-SY5Y
human neuroblastoma cells. J Neurochem 1997; 68: 392401.
86 Li Y, Martin LD, Spizz G, Adler KB. MARCKS protein is a key
molecule regulating MUCIN secretion by human airway epithe-
lial cells in vitro. J Biol Chem 2001; 276: 4098240990.
87 Allen LH, Aderem A. A role for MARCKS, the alpha isozyme of
protein kinase C and myosin I in zymosan phagocytosis by
macrophages. J Exp Med 1995; 182: 829840.
88 McNamara RK, Lenox RH. Distribution of the protein kinase C
substrates MARCKS and MRP in the postnatal developing rat
brain. J Comp Neurol 1998; 397: 337356.
89 Stumpo DJ, Bock CB, Tuttle JS, Blackshear PJ. MARCKS
deficiency in mice leads to abnormal brain development and
perinatal death. Proc Natl Acad Sci USA 1995; 92: 944948.
90 Swierczynski SL, Siddhanti SR, Tuttle JS, Blackshear PJ.
Nonmyristoylated MARCKS complements some but not all of
the developmental defects associated with MARCKS deficiency
in mice. Dev Biol 1996; 179: 135147.
91 Wang JK, Walaas SI, Sihra TS, Aderem A, Greengard P.
Phosphorylation and associated translocation of the 87-kDa
protein, a major protein kinase C substrate, in isolated
nerve terminals. Proc Natl Acad Sci USA 1989; 86:
22532256.
92 Lu D, Yang H, Lenox RH, Raizada MK. Regulation of angiotensin
II-induced neuromodulation by MARCKS in brain neurons. J Cell
Biol 1998; 142: 217227.
93 Ouimet CC, Wang JK, Walaas SI, Albert KA, Greengard P.
Localization of the MARCKS (87kDa) protein, a major specific
substrate for protein kinase C, in rat brain. J Neurosci 1990; 10:
16831698.
94 Yang H, Wang X, Sumners C, Raizada MK. Obligatory role of
protein kinase Cbeta and MARCKS in vesicular trafficking in
living neurons. Hypertension 2002; 39: 567572.
95 McNamara RK, Stumpo DJ, Morel LM et al. Effect of reduced
myristoylated alanine-rich C kinase substrate expression
on hippocampal mossy fiber development and spatial
learning in mutant mice: transgenic rescue and interactions
with gene background. Proc Natl Acad Sci USA 1998; 95: 14517
14522.
96 Ramakers GM, McNamara RK, Lenox RH, De Graan PN.
Differential changes in the phosphorylation of the protein kinase
C substrates myristoylated alanine-rich C kinase substrate and
growth-associated protein-43/B-50 following Schaffer collateral
long-term potentiation and long-term depression. J Neurochem
1999; 73: 21752183.
97 Hussain R, McNamara RK, Stumpo DJ et al. Effect of MARCKS
overexpression on hippocampal long-term potentiation and
hippocampal-dependent learning in transgenic mice. Soc Neu-
rosci Abstracts 2000; 26: 1511.
98 Lenox RH, Watson DG, Patel J, Ellis J. Chronic lithium
administration alters a prominent PKC substrate in rat hippo-
campus. Brain Res 1992; 570: 333340.
99 Wang L, Liu X, Lenox RH. Transcriptional down-regulation of
MARCKS gene expression in immortalized hippocampal cells by
lithium. J Neurochem 2001; 79: 816825.
100 Watson DG, Wainer BH, Lenox RH. Phorbol ester- and retinoic
acid-induced regulation of the protein kinase C substrate
MARCKS in immortalized hippocampal cells. J Neurochem
1994; 63: 16661674.
101 Watson DG, Watterson JM, Lenox RH. Sodium valproate down-
regulates the myristoylated alanine-rich C kinase substrate
(MARCKS) in immortalized hippocampal cells: a property of
protein kinase C-mediated mood statilizers. J Pharmacol Exp
Ther 1998; 285: 307316.
102 Lenox RH, McNamara RK, Watterson JM, Watson DG. Myristoy-
lated alanine-rich C kinase substrate (MARKCS): a molecular
target for the therapeutic action of mood stabilizers in the brain?
J Clin Psychiatry 1996; 57: 2331; discussion 3233.
103 Williams RS, Cheng L, Mudge AW, Harwood AJ. A common
mechanism of action for three mood-stabilizing drugs. Nature
2002; 417: 292295.
104 Wang L, Liu X, Lenox RH. Transcriptional regulation of mouse
MARCKS promoter in immortalized hippocampal cells. Biochem
Biophys Res Commun 2002; 292: 969979.
105 Herbert A, Rich A. The biology of left-handed Z-DNA. J Biol
Chem 1996; 271: 1159511598.
106 Bosetti F, Seemann R, Bell JM et al. Analysis of gene expression
with cDNA microarrays in rat brain after 7 and 42 days of oral
lithium administration. Brain Res Bull 2002; 57: 205209.
107 de la Fuente A, Brazhnik P, Mendes P. Linking the genes:
inferring quantitative gene networks from microarray data.
Trends Genet 2002; 18: 395398.
108 Bowden AC. Metabolic control analysis in biotechnology and
medicine. Nat Biotechnol 1999; 17: 641643.
109 Iyer VR, Horak CE, Scafe CS, Botstein D, Snyder M, Brown PO.
Genomic binding sites of the yeast cell-cycle transcription factors
SBF and MBF. Nature 2001; 409: 533538.
110 Ren B, Robert F, Wyrick JJ et al. Genome-wide location and
function of DNA binding proteins. Science 2000; 290: 23062309.
111 Weinmann AS, Yan PS, Oberley MJ, Huang TH, Famham PJ.
Isolating human transcription factor targets by coupling chroma-
tin immunoprecipitation and CpG island microarray analysis.
Genes Dev 2002; 16: 235244.
112 Horak CE, Mahajan MC, Luscombe NM, Gerstein M, Weissman
SM, Snyder M. GATA-1 binding sites mapped in the beta-globin
locus by using mammalian chlpchip analysis. Proc Natl Acad
Sci USA 2002; 99: 29242929.
113 Weinmann AS, Farnham PJ. Identification of unknown target
genes of human transcription factors using chromatin immuno-
precipitation. Methods 2002; 26: 3747.
114 Davidson EH, Rast JP, Oliveri P et al. A provisional regulatory
gene network for specification of endomesoderm in the sea
urchin embryo. Dev Biol 2002; 246: 162190.
115 Oliveri P, Carrick DM, Davidson EH. A regulatory gene network
that directs micromere specification in the sea urchin embryo.
Dev Biol 2002; 246: 209228.
Molecular basis of lithium action
RH Lenox and Le Wang
144
Molecular Psychiatry