Báo cáo khoa học: Death-associated protein kinase (DAPK) and signal transduction: additional roles beyond cell death ppt

MINIREVIEW
Death-associated protein kinase (DAPK) and signal
transduction: additional roles beyond cell death
Yao Lin, Ted R. Hupp and Craig Stevens
CRUK p53 Signal Transduction Laboratories, Institute of Genetics and Molecular Medicine, University of Edinburgh, UK
Introduction
Death-associated protein kinase-1 (DAPK-1) is the
prototypic member of a family of death-related kinases
that includes DAPK-1-related protein 1 (also named
DAPK-2), Zipper interacting kinase (ZIPK, also
named DAPK-3), DAP kinase related apoptosis indu-
cing protein kinase 1 (DRAK1) and DRAK2 [1].
These kinases share a high degree of homology in their
catalytic domains. However, the extracatalytic domains
and biological function of these five proteins differ
markedly [1]. DAPK, a calcium ⁄ calmodulin (CaM)-
regulated Ser ⁄ Thr protein kinase, was originally identi-
fied as a factor that regulates apoptosis in response to
the death-inducing cytokine signal interferon-c (INF-c)
[2]. In addition to its role in apoptosis, recent advances
have established an important role for DAPK in a
diverse range of signal transduction pathways, includ-
ing growth factor signalling and autophagy. In this
review we will integrate these new findings with our
existing knowledge of DAPK function and attempt to
highlight the areas that remain unresolved and require
further investigation.
The DAPK interactome
A major goal in biological research is to define the
system within which a signalling protein operates and
Keywords

autophagy; DAPK; growth factor;
immune response; interactome; kinase;
mTOR; peptide
Correspondence
C. Stevens, CRUK p53 Signal Transduction
Laboratories, Institute of Genetics and
Molecular Medicine, University of
Edinburgh, Edinburgh EH4 2XR, UK
E-mail:
(Received 11 March 2009, revised
12 August 2009, accepted 8 September
2009)
doi:10.1111/j.1742-4658.2009.07411.x
Death-associated protein kinase (DAPK) is a stress-regulated protein
kinase that mediates a range of processes, including signal-induced cell
death and autophagy. Although the kinase domain of DAPK has a range
of substrates that mediate its signalling, the additional protein interaction
domains of DAPK are relatively ill defined. This review will summarize
our current knowledge of the DAPK interactome, the use of peptide apta-
mers to define novel protein–protein interaction motifs, and how these new
protein–protein interactions give insight into DAPK functions in diverse
cellular processes, including growth factor signalling, the regulation of
autophagy, and its emerging role in the regulation of immune responses.
Abbreviations
ATM, ataxia telangiectasia mutated; BH3, Bcl-2-homology-3; CaM, calcium ⁄ calmodulin; DAPK, death-associated protein kinase; DIP1, DAPK
interacting protein-1; EGF, epidermal growth factor; ER, endoplasmic reticulum; ERK, extracellular signal-regulated kinase; HSP, heat shock
protein; INF-c, interferon-c; LAR, leukocyte common antigen related; MAP1B, microtubule-associated protein 1B; MCM3, mini-chromosome
maintenance complex component 3; mTOR, mammalian target of rapamycin; NF-jB, nuclear factor kappa-b; PMA, phorbol-12-myristate-13-
acetate; RSK, ribosomal S6 kinase; S6K1, ribosomal protein S6 kinase-1; TGF-b, transforming growth factor-b; TNF, tumour necrosis factor;
TSC, tuberous sclerosis; ZIPK, Zipper interacting kinase.

48 FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS
to use this information to understand developmental
or disease processes. Classically, genetic screens in trac-
table organisms, such as yeast, worms and flies, have
been used for defining the landscape of a protein ⁄ path-
way. However, many cancer- and immunity-related
genes are confined to vertebrates and a full under-
standing of how these proteins operate without the use
of classic genetics has been relatively difficult. Instead,
technologies that define protein–protein interactions
have been used to build a protein interaction map (i.e.
like a genetic interaction pathway) for a target protein.
Such technologies include the yeast two hybrid, mono-
clonal antibody co-immunoprecipitation methods
coupled to protein sequencing, and tap-tagging molecular
biology approaches for trapping a multiprotein com-
plex. The yeast two hybrid, for example, has been used
to discover a novel interaction between extracellular
signal-regulated kinase (ERK) and DAPK, with impli-
cations for pro-apoptotic pathways [3]. Furthermore,
recent ideas in systems biology hold that many pro-
teins have unstructured motifs or linear domains and
that dynamic regulation of protein–protein interactions
is mediated by the diversity in such small signalling
motifs. This property has been exploited using peptide
combinatorial libraries to discover novel complexes
between DAPK and microtubule-associated protein 1B
(MAP1B) [4] and DAPK and tuberous sclerosis 2
(TSC2) [5], with implications for autophagy and mam-
malian target of rapamycin (mTOR) signalling.

Together, using such distinct approaches, the DAPK
interactome is being built up in a range of back-
grounds.
DAPK is a large 160 kDa protein composed of
several functional domains, including a kinase domain,
a CaM regulatory domain, eight consecutive ankyrin
repeats, two putative nucleotide binding domains (P-
loops), a cytoskeletal binding domain and a death
domain (Fig. 1). Proteins that interact with DAPK,
the domain on DAPK that mediates the interaction
and the methods used to discover the interactions are
summarized in Table 1. Given that many regions of
DAPK can form protein–protein interfaces it is unsur-
prising that only a few of the DAPK binding proteins
highlighted in Table 1 are substrates of DAPK, sug-
gesting that in some circumstances protein interaction
alone is sufficient for DAPK to exert its biological
effects. Because of the paucity of DAPK substrates, a
screen aimed at identifying a consensus DAPK phos-
phorylation motif was carried out based on positional
scanning peptide substrate library synthesis and activ-
ity [6]. The preferred consensus motif for DAPK
phosphorylation and substrates for which phospho-
acceptor site(s) have been identified are described in
Table 2. Of note, mini-chromosome maintenance com-
plex component 3 (MCM3), which is a DNA replica-
tion licensing factor, was identified using biochemical
fractionation and MS analysis to purify and identify
potential substrates from Hela cell lysate [7]. This kind
of proteomic approach should expedite the identifica-

tion of novel, physiologically relevant in vivo substrates
of DAPK. Moreover, it could be tailored to reflect
DAPK substrate specificity in response to specific
signalling events, such as growth factor or cytokine
signalling.
It is apparent from Table 2 that not all of the
DAPK substrates identified are a good match to the
identified consensus motif. Chemical genetics, a bio-
chemical approach to develop small peptide-mimetic
ligands to alter how an enzyme functions, was utilized
Ca
2+
/CaM
Ankyrin
P-loopsrepeats
1
1431
Kinase
Death
Cytoskeletal
Fig. 1. Schematic representation of DAPK. DAPK is a large
160 kDa Ser ⁄ Thr Ca2
+
⁄ CaM-regulated kinase that consists of
several functional domains, including a kinase domain, a CaM
regulatory domain, eight consecutive ankyrin repeats, two P-loops,
a cytoskeletal binding domain and a death domain, which enable it
to participate in a wide range of signalling pathways.
Table 1. DAPK binding proteins, the region of DAPK important for
mediating the protein-protein interaction, and the method used to

define the interaction.
Binding protein Binding region on DAPK Binding assay used
14-3-3 [65] Not defined Immunoprecipitation
Actin [66] Cytoskeletal domain Immunostaining
Beclin-1 [36] Not defined Immunoprecipitation
CaM [67] Ca
2
+ ⁄ CaM
regulatory domain
Overlay binding assay
Cathepsin B [61] C-terminal domain Immunoprecipitation
DIP1 [13] Ankyrin repeats Yeast two hybrid
a
ERK [3] Death domain Yeast two hybrid
a
FADD [65] Not defined Immunoprecipitation
Hsp90 [62] Kinase domain Immunoprecipitation
LAR [23] Ankyrin repeats Yeast two hybrid
a
MAP1B [4] Kinase domain Peptide libraries
a
PKD [68] Not defined Immunoprecipitation
RSK [25] Not defined Immunoprecipitation
Src [23] Not defined Yeast two hybrid
a
TNFR-1 [65] Not defined Immunoprecipitation
TSC2 [5] Death domain Peptide libraries
a
UNC5H2 [69] Death domain Yeast two hybrid
a

ZIPK [70] Kinase domain Immunoprecipitation
a
Protein interactions have been confirmed by more physiological
methods.
Y. Lin et al. DAPK and signal transduction
FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS 49
recently to develop selective peptide ligands that mod-
ulate DAPK activity. For example, DAPK binding to
a peptide derived from the amino acid sequence of the
cyclin-dependent kinase inhibitor p21 induces a confor-
mational change in DAPK that enhances its kinase
activity, suggesting that DAPK may require docking in
order to phosphorylate a subset of its substrates [8]. It
is also possible that the interaction of DAPK with
many of its substrates is of too low affinity to detect in
cells. In support of this notion, the ataxia telangiecta-
sia mutated (ATM) protein kinase, a large > 300 kDa
enzyme, does not have an abundance of stable
protein–protein partners that would be expected of a
protein of its large size. However, a recent MS-linked
proteomics screen identifying phospho-Ser-Gln pep-
tides that are phosphorylated by ATM identified over
700 substrates [9]. Therefore, it seems that the previ-
ously available protein interaction methodologies were
not able to faithfully reflect the ATM kinase inter-
actome.
A future challenge will be the identification of lower
affinity or transient DAPK interactions that might
otherwise be overlooked in the more traditional assays
to further elucidate the functional role of DAPK in

diverse signalling pathways.
Signalling to DAPK
DAPK plays an important role in a wide range of sig-
nal transduction pathways with diverse outcomes, such
as apoptosis, autophagy and immune responses. The
functional outcome of DAPK activity depends largely
on the input signal (Fig. 2). For example, DAPK gene
expression and apoptotic activity is increased in
response to transforming growth factor-b (TGF-b) [10]
and to stimuli that activate p53 [11], such as DNA-
damaging agents. Other death signals, such as the
transforming oncogenes E2F1 and Myc [12], also
induce DAPK expression. In addition to its well-docu-
mented role in the regulation of apoptosis, DAPK
may also play a role in survival pathways, reflected in
its activation by growth factor signalling pathways [5],
and its ability to counter tumour necrosis factor
(TNF)-mediated apoptosis [13].
Table 2. DAPK substrates and the amino acid sequence surround-
ing the phosphorylation site. The substrate phosphorylation pattern
preferred by DAPK is highlighted in bold; the basic residues also
preferred by DAPK are underlined.
Substrate Phosphorylation site
Beclin-1[36] RLKVT
119
GDL
CaMKK [71] GSRREERSLS
511
APG
DAPK [72] A R KKW KQS

308
VRLI
MCM3 [7] TKKTIERRYS
160
DLTTL
MLC [73] TTKKRPQRATS
19
NVF
p21 [8] RKRRQT
145
SMTDFYHSK
p53 [8] PPLSQET
18
FS
20
DLWKLL
S6 [27] QIAKRRRLS
235
SLRAS
Syntaxin-1A [38] IIMDSSIS
188
KQALSEIE
Tropomyosin-1 [74] HALNDMTS
283
I
ZIPK [70] KT
299
TRLKEYTIKS
30
9HS

311
S
312
LPPNNS
318
YADFERFS
326
Consensus KRxxxxxKRRxxS ⁄ T
Mitogens
EGF
Short treatment Long treatment
TNF-α TNF-α
IFN-γ
TGF-β
DNA damage
oncogenes
Growth
mTORC1
Gene expression
Kinase activity Kinase activity
?
Degradation
DAPK
over-expression
AutophagyApoptosisApoptosis
Autophagy
Inflammation
Immune response
Blebbing
Autophagy

AB C
DE
Beclin-1
phosphorylation
MAP1B binding
ApoptosisApoptosis
Inflammation ?mTORC1?
Fig. 2. Signalling to DAPK. DAPK plays an important role in a diverse range of signal transduction pathways. The biological outcome of
DAPK activity depends on the input signal and includes cell growth, immune responses, apoptosis and autophagy. (A) Growth factor signal-
ling to DAPK is probably the best defined with respect to the proteins that are involved and includes the activities of Src, LAR, ERK and
RSK (see text and Fig. 3). (B) The functional outcome of increased DAPK activity in response to short-term treatment with TNF-a is currently
unclear, but may contribute to mTORC1 activation and inhibition of inflammatory responses. Longer-term treatment with TNF-a leads to
DAPK degradation coincident with apoptosis, suggesting that DAPK may be a resistance factor to TNF-a-induced cell death in some circum-
stances. (C) DAPK mediates many cellular responses in response to INF-c, but the molecular mechanisms have not yet been defined. (D)
DAPK gene expression and apoptotic activity are increased in response to TGF-b and to stimuli that activate p53, such as DNA-damaging
agents. Other death signals, such as the transforming oncogenes E2F1 and Myc, also induce DAPK expression. (E) Overexpression of DAPK
can promote autophagy and membrane blebbing via binding to MAP1B, or autophagy via the direct phosphorylation of Beclin-1. The signals
that regulate DAPK autophagic activity have yet to be defined.
DAPK and signal transduction Y. Lin et al.
50 FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS
Growth factor signalling/mTOR
Serum-induced activation of DAPK catalytic activity
has been demonstrated recently [3,5,14] and it is
becoming increasingly clear that DAPK is intimately
linked to growth factor signalling pathways (Fig. 3).
For example, serum-induced phosphorylation of
DAPK by ERK enhances its kinase activity and
death-promoting effects [3], whereas serum activation
of DAPK has also been linked to cell death by
suppressing integrin functions and integrin-mediated

survival signals [14]. However, in addition to apoptotic
signalling, we have recently demonstrated a stimula-
tory role for serum-activated DAPK in mTOR signal-
ling [5]. mTOR is a member of the phosphoinositide-
3-kinase-related kinase family, which is centrally
involved in growth regulation, proliferation control
and cell metabolism [15]. In mammalian cells, two
structurally and functionally distinct mTOR-containing
complexes have been identified, mTORC1 and
mTORC2 [15]. mTORC1 directly regulates cell growth
by controlling the phosphorylation of a number of
components of the translational machinery. In particu-
lar, phosphorylation and activation of eukaryotic initi-
ation factor 4E binding protein-1 and ribosomal
protein S6 kinase-1 (S6K1) are stimulated by serum,
insulin and growth factors in an mTORC1-dependent
manner [16].
The TSC complex, formed by two proteins, TSC1
and TSC2, is a major regulator of the mTORC1
signalling pathway [17]. TSC2 contains a GTPase-acti-
vating protein domain that converts the small GTPase
Ras homolog enriched in brain to its inactive
GDP-bound form [18]. mTORC1 activity is stimulated
by the active GTP-bound form of Ras homolog
enriched in brain, thus the TSC complex acts to inhibit
mTORC1 function [18]. Growth factor-induced, inacti-
vating TSC2 phosphorylation results in mTORC1 acti-
vation and is thought to occur primarily through
activation of the RAS–extracellular signal-regulated
kinase kinase (MEK)–ERK and phosphoinositide-3-

kinase–Akt pathways [19,20]. In a protein interaction
screen in our laboratory, we identified TSC2 as a novel
DAPK death domain interacting protein, and in analy-
sing the biological consequences of the DAPK–TSC2
interaction, we were led to the discovery that DAPK
can phosphorylate and inactivate TSC2 and functions
as a positive cofactor in mTORC1 signalling in
response to serum and epidermal growth factor (EGF)
stimulation [5].
ERK can directly interact with and phosphorylate
DAPK at Ser735, which leads to enhanced kinase
activity and pro-apoptotic activity of DAPK [3]. This
Ser735 phosphorylation can be stimulated by serum or
phorbol-12-myristate-13-acetate (PMA) [3], which acti-
vates the RAS–MEK–ERK pathway [21,22]. Interest-
Ras
Raf
MEK
ERK
RSK
DAPK
TSC2
TSC1
Apoptosis ?ApoptosisApoptosis
DAPK
Rheb
S6K
S6
-T389
-S235/236

P
P
Cell growth
Protein synthesis
mTORC1
DAPK
P
-S289
P
-S735
Src
LAR
P
Y491/Y492 -
EGF
Fig. 3. Growth factor regulation of DAPK.
Growth factor signalling to DAPK is complex
and regulates a diverse range of biological
outcomes. For example, phosphorylation by
ERK enhances the apoptotic activity of
DAPK, but Src-mediated phosphorylation of
DAPK suppresses its apoptotic, antimigra-
tion and antiadhesion functions. Under nor-
mal growth conditions, DAPK apoptotic
activity may also be suppressed until such
times as required due to phosphorylation by
RSK. DAPK may also act in concert with
ERK and RSK to inhibit the TSC complex,
resulting in mTORC1 activation. In addition,
DAPK and RSK may co-operate to promote

protein translation via direct phosphorylation
of ribosomal protein S6.
Y. Lin et al. DAPK and signal transduction
FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS 51
ingly, the inactivation of DAPK activity by EGF has
been recently described. Wang et al. [23] demonstrated
that DAPK is a substrate for leukocyte common
antigen related (LAR) tyrosine phosphatase and that
dephosphorylation of Y491 ⁄ Y492, located in the
ankyrin repeat domain, resulted in activation of the
pro-apoptotic activities of DAPK. Reciprocally, Src
kinase phosphorylation of Y491⁄ Y492 inhibited
DAPK activity [23]. Src kinase was activated in
response to EGF stimulation and LAR was downregu-
lated, resulting in DAPK inactivation. The ability of
EGF signalling to inactivate DAPK is inconsistent
with previous findings that DAPK activity can be up-
regulated by serum stimulation and ERK, a down-
stream effector of the EGF pathway [3], and this is
further inconsistent with data showing that in response
to PMA, the DAPK–ERK complex induces apoptosis
[3]. It is important to note, however, that the apopto-
sis-promoting effect of DAPK induced by the ERK
activator PMA was only observed in suspension cells
[3], whereas in adherent cells the co-expression of a
constitutively active mutant of MEK is required for
DAPK to induce apoptosis [24]. Therefore, the apop-
tosis function of the ERK–DAPK complex may only
exist under aberrant conditions, such as when cells are
detached, or when the signal to grow is excessive.

Other signalling pathways can in turn modify these
core activities of DAPK. For example, RAS activation
of the ERK–ribosomal S6 kinase (RSK) pathway can
attenuate the pro-apoptotic function of DAPK. RSK
interacts with DAPK in vitro and in vivo and catalyses
the phosphorylation of DAPK on Ser289 in response
to PMA [25]. The effect of this phosphorylation on the
kinase activity of DAPK was not tested. However,
mutation of Ser289 to a nonphosphorylatable Ala
results in a DAPK mutant with enhanced apoptotic
activity, whereas the phosphomimetic mutation
(Ser289Glu) attenuates its apoptotic activity [25]. The
observation that the Ser289Ala mutant of DAPK is
more apoptotic suggests that phosphorylation inhibits
the catalytic activity of DAPK [25]. Thus, kinase assays
using the Ser289 mutants are required to clearly deter-
mine the function of DAPK Ser289 phosphorylation.
Interestingly, RSK has also been shown to interact with
TSC2, and phosphorylation by RSK inactivates TSC2,
resulting in mTORC1 activation [26].
DAPK has also been directly linked to the control
of protein translation by phosphorylating ribosomal
protein S6 on Ser235 ⁄ 236 [27]. In agreement with this
study, we have shown that DAPK can robustly stimu-
late the phosphorylation of S6 in cells, even in the
presence of the lipophilic macrolide antibiotic rapa-
mycin, a potent inhibitor of mTORC1 activity, indicat-
ing that DAPK can mediate phosphorylation of S6 in
an mTORC1–S6K-dependent and -independent man-
ner. Schumacher et al. [27] demonstrated that DAPK

phosphorylates S6 directly on Ser235 ⁄ 236 and con-
cluded that this is an inhibitory phosphorylation
reducing S6 activity and protein translation in vitro.In
contrast, Roux et al. [28] demonstrated that RSK
kinase phosphorylates the same sites on S6, but they
concluded that this was an activating phosphorylation
that stimulates S6 activity and promotes assembly of
the translation preinitiation complex in cells. Our
results are in agreement with the latter study and point
towards a role for DAPK in activating S6 and protein
translation. Further studies are required to clarify the
role of DAPK in the regulation of S6 activity and pro-
tein translation in vivo, in particular the interplay
between DAPK and RSK signalling to S6 needs to be
addressed, and the ability of DAPK to promote cell
growth needs to be clearly demonstrated.
Taken together, these studies reveal a complex regu-
lation of DAPK activity by growth factor signalling
pathways mediated by Src, LAR, ERK and RSK. A
better understanding of the interplay between signalling
to DAPK and TSC2 may explain how the specific activ-
ity of DAPK can be modulated to control the balance
between pro-apoptotic and pro-survival pathways.
DAPK and autophagy
DAPK was originally identified as a factor that regu-
lates apoptosis in response to various death-inducing
signals, including INF-c [2]. DAPK also has auto-
phagy signalling activity, which can be either pro-sur-
vival or lead to or participate in cell death.
Autophagy is a membrane system involved in pro-

tein and organelle degradation that probably repre-
sents an innate adaptation to starvation. In times of
nutrient deficiency, the cell can self-digest and recycle
some nonessential components to sustain its minimal
growth requirements until a food source becomes
available. Over recent years, autophagy has been impli-
cated in an increasing number of clinical scenarios,
notably infectious diseases, cancer, neurodegenerative
diseases and autoimmunity. In some cell types, the
overexpression of DAPK can lead to the appearance
of autophagic vesicles [29]. However, there is still little
known about how DAPK exerts its effects on auto-
phagy, and as DAPK is not present in yeast, there
have been no classic genetic screens to analyse how
DAPK interacts with the core autophagy pathway.
Recently, peptide combinatorial libraries identified
MAP1B as a DAPK interacting protein that functions
as a positive cofactor in DAPK-mediated autophagic
DAPK and signal transduction Y. Lin et al.
52 FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS
vesicle formation and membrane blebbing [4]. MAP1B
has been most widely studied as a major component of
the neuronal cytoskeleton [30] and relatively little is
known about its role outside of these neuronal
systems. The cotransfection of both genes stimulated
the disruption of microtubules during the induction of
membrane blebbing, suggesting that MAP1B–DAPK-
induced blebbing involves changes in the dynamics of
mictrotubules, as well as changes in the dynamics of
contractile cortical actin [4]. This is even more intrigu-

ing in light of the recently identified interaction
between the essential autophagy protein Atg8 (LC3)
and MAP1B [31], and the observation that micro-
tubules play an important role in autophagy by support-
ing the production and transport of autophagosomes
[32]. Future studies will determine whether MAP1B is
a key factor that switches DAPK activity towards
autophagy induced by certain stresses such as INF-c.
Beclin-1, the first identified mammalian autophagy
gene [33], interacts with several cofactors to activate
the lipid kinase Vps34, thereby inducing autophagy
[34]. Beclin-1 is a Bcl-2-homology-3 (BH3) domain-only
protein that binds to the BH3 domain of the
antiapoptotic proteins Bcl-2 ⁄ Bcl-X
L
[35]. Under normal
conditions, beclin-1 is bound to and inhibited by Bcl-2
or the Bcl-2 homolog Bcl-X
L
and the dissociation of
beclin-1 from Bcl-2 is essential for its autophagic activ-
ity [34]. Nutrient deprivation stimulates the dissociation
either by activating BH3-only proteins (such as Bad),
which can competitively disrupt the interaction, or by
post-translational modification [34]. A recent report
demonstrated that a constitutively activated form of
DAPK triggers autophagy in a beclin-1-dependent
manner [36]. DAPK phosphorylates beclin-1 on Thr119
located at a crucial position within its BH3 domain,
and thus promotes the dissociation of beclin-1 from

Bcl-X
L
and the induction of autophagy [36]. This study
revealed a new substrate for DAPK that acts as one of
the core proteins of the autophagic machinery, and
provides a new phosphorylation-based mechanism for
how DAPK activates autophagy by reducing the inter-
action of beclin-1 with its inhibitor Bcl-X
L
.
DAPK has also been directly linked to the regu-
lation of endocytosis [37], and can phosphorylate
syntaxin-1A, a key component of the soluble N-ethyl-
maleimide-sensitive factor (NSF) attachment protein
receptors complex essential for synaptic vesicle docking
and fusion [38]. Therefore, DAPK may also regulate
autophagy via syntaxin-1A.
Although most evidence suggests that autophagy
acts as a survival response to provide an energy source
maintaining cell survival, it has been proposed that
autophagy can contribute to cell death in a process
termed autophagic (type II) cell death. Disturbance to
endoplasmic reticulum (ER) homeostasis that leads to
irreparable damage activates ER-specific cell death
mechanisms [39]. DAPK was recently identified as an
important component in ER stress-induced cell death
[40]. DAPK ) ⁄ ) mice are protected from kidney dam-
age caused by injection of the ER stress inducer
tunicamycin and the cell death response to tunicamy-
cin is reduced in DAPK ) ⁄ ) mouse embryonic fibro-

blasts [40]. Interestingly, both caspase activation and
autophagy induction are attenuated in DAPK) ⁄ )
mouse embryonic fibroblasts, and depletion of ATG5
or beclin-1, essential autophagic proteins, are protected
from ER-induced death when combined with caspase-3
depletion [40]. These results suggest that under certain
conditions, DAPK-induced autophagy contributes to
cell death, possibly through the induction of apoptosis.
In the model organism Caenorhabditis elegans,it
was recently demonstrated that starvation-induced
autophagy is regulated in part through a DAPK sig-
nalling pathway and that autophagy levels are critical
to drive such cell fate decisions, leading to survival or
death of the organism [41] (see the accompanying
review by Kang and Avery [42]). In C. elegans, mus-
caranic acetylcholine receptor signalling is important
in modulating the level of autophagy during starvation
[43]. In a simplified model, starvation activates MAPK
(MPK-1), the C. elegans ortholog of mammalian
ERK, and activated MPK-1 positively regulates auto-
phagy, at least in part through DAPK and RGS-2
[43]. It will be interesting to determine whether ERK
and DAPK can co-operate to regulate autophagy in
higher organisms.
The pathway that regulates autophagy also acts
through mTORC1 [44]. Rapamycin binds to and inac-
tivates mTORC1, leading to an upregulation of auto-
phagy [45]. The finding that DAPK is a positive
regulator of mTORC1 signalling and a positive regula-
tor of autophagy at first seems counterintuitive. There-

fore, we would predict that DAPK activity should be
activated by starvation, and that its activity would be
inversely correlated with that of mTORC1. However,
in mammalian cells, although DAPK is reported to be
necessary for INF-c-induced autophagy, it seems not
to be a crucial element in starvation or rapamycin-
induced autophagy [46]. The accompanying review by
Kang and Avery [42] proposes an interesting explana-
tion for the seemingly contradictory functions of
DAPK to promote mTORC1 activity and autophagy.
They propose that DAPK may promote mTORC1
activity specifically to mediate S6K activity during
starvation, as S6K activity has been shown to promote
rather than suppress autophagy in Drosophila [47].
Y. Lin et al. DAPK and signal transduction
FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS 53
Clearly, further characterization of the interacting
proteins and direct substrates of DAPK, as well as
differences between simple organisms and complex
mammalian systems, are required to clarify how the
kinase is linked to the autophagic pathways.
DAPK immune responses
DAPK has been shown to participate in cell death in
response to various cytokine signals, including IFN-c-
induced cell death [2], TNF-a and FAS-induced cell
death [48], and TGF-b-induced cell death [10]. There
are two distinct outcomes of TNF-a signalling, an
inflammatory immune response mediated by the
nuclear factor kappa-b (NF-jB) signalling pathway,
and apoptosis [49]. By comparing the response to

TNF-a treatment in DAPK-deficient and wild-type
cells, several groups have demonstrated that DAPK is
neutral against TNF-a-induced apoptosis [2,10]. More
recent studies have indicated that DAPK is in fact a
negative regulator of TNF-a-induced apoptosis. For
example, antisense depletion of DAPK in Hela cells
protects cells from IFN-c-induced apoptosis, but pro-
motes TNF-a-induced apoptosis [50], and the expres-
sion of DAPK interacting protein-1 (DIP1), a
ubiquitin E3 ligase that degrades DAPK, promotes
TNF-a-induced apoptosis [13]. Therefore, although it
functions as a death-promoting kinase, DAPK can
also act as a survival factor and block apoptosis in
response to certain cytokine signals. Interestingly,
DAPK has recently been shown to function as a neg-
ative regulator of T cell activation via NF-jB. How-
ever, DAPK had no effect on NF-jB activation by
TNF-a, only by T cell receptor activation [51]. In
addition, DAPK can act as a negative regulator of
inflammatory gene expression in monocytes [52]. In
C. elegans, wounding of epidermal layers triggers mul-
tiple co-ordinated responses to damage. It was
recently shown that the C. elegans ortholog of DAPK
acts as a negative regulator of barrier repair and
innate immune responses to wounding [53]. Taken
together, these studies suggest an intriguing role for
DAPK, not only as a modulator of cytokine-induced
apoptosis, but as a regulator of various immune
responses.
Future work

It is becoming increasingly clear that DAPK family
members have additional roles beyond their functions
in cell death. The recent findings that DAPK nega-
tively regulates inflammatory gene expression [51,52],
responds to mitogenic signals to regulate mTORC1
activity [5] and negatively regulates epidermal damage
responses in C. elegans in an apoptosis- and auto-
phagy-independent manner [53], highlight the pleo-
trophic role of this kinase.
What are the crucial questions for the future? Of
considerable importance will be to gain a clear under-
standing of the role of DAPK in the RAS–MEK–
ERK growth factor signalling pathway, in particular
the interplay between ERK, RSK and DAPK and
the balance between apoptosis and growth needs to
be addressed. Gaining a better understanding of
DAPK’s role in cancer is particularly important.
DAPK hypermethylation is strongly associated with
various cancers (see the accompanying review by
Michie et al. [54]), but it is not yet clear how reduced
levels of DAPK contribute to carcinogenesis. Possible
mechanisms include DAPK’s ability to suppress extra-
cellular matrix survival signals to regulate anoikis [14]
and its ability to inhibit cell polarization and motility
[55]. DAPK can suppress transformation by oncoge-
nes by activating a pro-apoptotic p53-dependent
checkpoint [12], and it can activate autophagy, which
has been shown to be antitumorigenic [56–58]. Recent
studies indicate that inflammation is an important
contributor to tumorigenesis [59]. Therefore, the anti-

inflammatory function of DAPK may also contribute
to its tumour suppressive activity [52]. Of interest in
this regard are recent studies showing that the TSC–
mTORC1 pathway regulates inflammatory responses
in monocytes, macrophages and primary dendritic
cells [60]. The finding that DAPK regulates mTORC1
activity [5], together with the observation that both
mTORC1 and DAPK can block NF-jB activation
[51,60], raise the intriguing possibility that DAPK
may regulate inflammatory immune responses via an
mTORC1-dependent mechanism. Further studies are
required to determine whether these pathways are
related in this context.
Mechanisms regulating protein stabilization and
turnover are also critical for modulating DAPK activi-
ties. Several studies have shown DAPK degradation to
be dependent on the ubiquitin–proteasome pathway
[13,61–64]. To date, two E3 ubiquitin ligases have been
identified that can promote the ubiquitination of
DAPK; DIP-1, a ring finger containing E3 that inter-
acts directly with the ankyrin repeat region of DAPK
[13], and carboxyl terminus of HSC70-interacting pro-
tein, a U-box containing E3 ubiquitin ligase that can
bind to the heat shock protein (HSP) chaperone pro-
teins HSP70 and HSP90, interacts with DAPK indi-
rectly via Hsp90 [62]. The identification of additional
ubiquitin ligases, and deciphering the degradation
pathways that modulate DAPK stability, will shed
DAPK and signal transduction Y. Lin et al.
54 FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS

further light on the role played by DAPK in the regu-
lation of cell growth control.
There is no doubt that future research into the role
of DAPK will yield new and important insights into
the mechanisms that integrate the apoptotic, auto-
phagic and cell growth regulatory pathways.
References
1 Bialik S & Kimchi A (2006) The death-associated
protein kinases: structure, function, and beyond. Annu
Rev Biochem 75, 189–210.
2 Deiss LP, Feinstein E, Berissi H, Cohen O & Kimchi A
(1995) Identification of a novel serine ⁄ threonine kinase
and a novel 15-kD protein as potential mediators of the
gamma interferon-induced cell death. Genes Dev 9,
15–30.
3 Chen CH, Wang WJ, Kuo JC, Tsai HC, Lin JR, Chang
ZF & Chen RH (2005) Bidirectional signals transduced
by DAPK-ERK interaction promote the apoptotic
effect of DAPK. EMBO J 24, 294–304.
4 Harrison B, Kraus M, Burch L, Stevens C, Craig A,
Gordon-Weeks P & Hupp TR (2008) DAPK-1 binding
to a linear peptide motif in MAP1B stimulates auto-
phagy and membrane blebbing. J Biol Chem 283,
9999–10014.
5 Stevens C, Lin Y, Harrison B, Burch L, Ridgway RA,
Sansom O & Hupp T (2009) Peptide combinatorial
libraries identify TSC2 as a death-associated protein
kinase (DAPK) death domain-binding protein and
reveal a stimulatory role for DAPK in mTORC1 signal-
ing. J Biol Chem 284, 334–344.

6 Velentza AV, Schumacher AM, Weiss C, Egli M &
Watterson DM (2001) A protein kinase associated with
apoptosis and tumor suppression: structure, activity,
and discovery of peptide substrates. J Biol Chem 276,
38956–38965.
7 Bialik S, Berissi H & Kimchi A (2008) A high through-
put proteomics screen identifies novel substrates of
death-associated protein kinase. Mol Cell Proteomics 7,
1089–1098.
8 Fraser JA & Hupp TR (2007) Chemical genetics
approach to identify peptide ligands that selectively
stimulate DAPK-1 kinase activity. Biochemistry 46,
2655–2673.
9 Matsuoka S, Ballif BA, Smogorzewska A, McDonald
ER 3rd, Hurov KE, Luo J, Bakalarski CE, Zhao Z,
Solimini N, Lerenthal Y et al. (2007) ATM and ATR
substrate analysis reveals extensive protein networks
responsive to DNA damage. Science 316, 1160–1166.
10 Jang CW, Chen CH, Chen CC, Chen JY, Su YH &
Chen RH (2002) TGF-beta induces apoptosis through
Smad-mediated expression of DAP-kinase. Nat Cell Biol
4, 51–58.
11 Martoriati A, Doumont G, Alcalay M, Bellefroid E,
Pelicci PG & Marine JC (2005) dapk1, encoding an
activator of a p19ARF-p53-mediated apoptotic check-
point, is a transcription target of p53. Oncogene 24,
1461–1466.
12 Raveh T, Droguett G, Horwitz MS, DePinho RA &
Kimchi A (2001) DAP kinase activates a p19ARF ⁄ p53-
mediated apoptotic checkpoint to suppress oncogenic

transformation. Nat Cell Biol 3, 1–7.
13 Jin Y, Blue EK, Dixon S, Shao Z & Gallagher PJ
(2002) A death-associated protein kinase (DAPK)-inter-
acting protein, DIP-1, is an E3 ubiquitin ligase that
promotes tumor necrosis factor-induced apoptosis and
regulates the cellular levels of DAPK. J Biol Chem 277,
46980–46986.
14 Wang WJ, Kuo JC, Yao CC & Chen RH (2002) DAP-
kinase induces apoptosis by suppressing integrin activity
and disrupting matrix survival signals. J Cell Biol 159,
169–179.
15 Rosner M, Hanneder M, Siegel N, Valli A, Fuchs C &
Hengstschlager M (2008) The mTOR pathway and its
role in human genetic diseases. Mutat Res 659, 284–292.
16 Proud CG (2007) Signalling to translation: how signal
transduction pathways control the protein synthetic
machinery. Biochem J
403, 217–234.
17 Tee AR, Fingar DC, Manning BD, Kwiatkowski DJ,
Cantley LC & Blenis J (2002) Tuberous sclerosis com-
plex-1 and -2 gene products function together to inhibit
mammalian target of rapamycin (mTOR)-mediated
downstream signaling. Proc Natl Acad Sci USA 99,
13571–13576.
18 Tee AR, Manning BD, Roux PP, Cantley LC & Blenis
J (2003) Tuberous sclerosis complex gene products,
Tuberin and Hamartin, control mTOR signaling by act-
ing as a GTPase-activating protein complex toward
Rheb. Curr Biol 13, 1259–1268.
19 Inoki K, Li Y, Zhu T, Wu J & Guan KL (2002) TSC2

is phosphorylated and inhibited by Akt and suppresses
mTOR signalling. Nat Cell Biol 4, 648–657.
20 Ma L, Chen Z, Erdjument-Bromage H, Tempst P &
Pandolfi PP (2005) Phosphorylation and functional
inactivation of TSC2 by Erk implications for tuberous
sclerosis and cancer pathogenesis. Cell 121, 179–193.
21 Thomas SM, DeMarco M, D’Arcangelo G, Halegoua S
& Brugge JS (1992) Ras is essential for nerve growth
factor- and phorbol ester-induced tyrosine phosphoryla-
tion of MAP kinases. Cell 68, 1031–1040.
22 Wood KW, Sarnecki C, Roberts TM & Blenis J (1992)
ras mediates nerve growth factor receptor modulation
of three signal-transducing protein kinases: MAP
kinase, Raf-1, and RSK. Cell 68, 1041–1050.
23 Wang WJ, Kuo JC, Ku W, Lee YR, Lin FC, Chang
YL, Lin YM, Chen CH, Huang YP, Chiang MJ et al.
(2007) The tumor suppressor DAPK is reciprocally
Y. Lin et al. DAPK and signal transduction
FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS 55
regulated by tyrosine kinase Src and phosphatase LAR.
Mol Cell 27, 701–716.
24 Stevens C, Lin Y, Sanchez M, Amin E, Copson E,
White H, Durston V, Eccles DM & Hupp T (2007) A
germ line mutation in the death domain of DAPK-1
inactivates ERK-induced apoptosis. J Biol Chem 282,
13791–13803.
25 Anjum R, Roux PP, Ballif BA, Gygi SP & Blenis J
(2005) The tumor suppressor DAP kinase is a target of
RSK-mediated survival signaling. Curr Biol 15, 1762–
1767.

26 Roux PP, Ballif BA, Anjum R, Gygi SP & Blenis J
(2004) Tumor-promoting phorbol esters and activated
Ras inactivate the tuberous sclerosis tumor suppressor
complex via p90 ribosomal S6 kinase. Proc Natl Acad
Sci USA 101, 13489–13494.
27 Schumacher AM, Velentza AV, Watterson DM & Dres-
ios J (2006) Death-associated protein kinase phosphory-
lates mammalian ribosomal protein S6 and reduces
protein synthesis. Biochemistry 45 , 13614–13621.
28 Roux PP, Shahbazian D, Vu H, Holz MK, Cohen MS,
Taunton J, Sonenberg N & Blenis J (2007) RAS ⁄ ERK
signaling promotes site-specific ribosomal protein S6
phosphorylation via RSK and stimulates cap-dependent
translation. J Biol Chem 282, 14056–14064.
29 Inbal B, Bialik S, Sabanay I, Shani G & Kimchi A
(2002) DAP kinase and DRP-1 mediate membrane
blebbing and the formation of autophagic vesicles dur-
ing programmed cell death. J Cell Biol 157, 455–468.
30 Gordon-Weeks PR (1993) Organization of microtubules
in axonal growth cones: a role for microtubule-associ-
ated protein MAP 1B. J Neurocytol 22, 717–725.
31 Wang QJ, Ding Y, Kohtz DS, Mizushima N, Cristea
IM, Rout MP, Chait BT, Zhong Y, Heintz N & Yue Z
(2006) Induction of autophagy in axonal dystrophy and
degeneration. J Neurosci 26, 8057–8068.
32 Kochl R, Hu XW, Chan EY & Tooze SA (2006)
Microtubules facilitate autophagosome formation and
fusion of autophagosomes with endosomes. Traffic 7,
129–145.
33 Liang XH, Kleeman LK, Jiang HH, Gordon G, Gold-

man JE, Berry G, Herman B & Levine B (1998) Protec-
tion against fatal Sindbis virus encephalitis by beclin, a
novel Bcl-2-interacting protein. J Virol 72, 8586–8596.
34 Levine B, Sinha S & Kroemer G (2008) Bcl-2 family
members: dual regulators of apoptosis and autophagy.
Autophagy 4, 600–606.
35 Pattingre S, Tassa A, Qu X, Garuti R, Liang XH,
Mizushima N, Packer M, Schneider MD & Levine B
(2005) Bcl-2 antiapoptotic proteins inhibit Beclin
1-dependent autophagy. Cell 122, 927–939.
36 Zalckvar E, Berissi H, Mizrachy L, Idelchuk Y, Koren
I, Eisenstein M, Sabanay H, Pinkas-Kramarski R &
Kimchi A (2009) DAP-kinase-mediated phosphorylation
on the BH3 domain of beclin 1 promotes dissociation
of beclin 1 from Bcl-X(L) and induction of autophagy.
EMBO Rep 10, 285–292.
37 Pelkmans L, Fava E, Grabner H, Hannus M, Haber-
mann B, Krausz E & Zerial M (2005) Genome-wide
analysis of human kinases in clathrin- and caveo-
lae ⁄ raft-mediated endocytosis. Nature 436, 78–86.
38 Tian JH, Das S & Sheng ZH (2003) Ca2+-dependent
phosphorylation of syntaxin-1A by the death-associated
protein (DAP) kinase regulates its interaction with
Munc18. J Biol Chem 278, 26265–26274.
39 Wu J & Kaufman RJ (2006) From acute ER stress to
physiological roles of the unfolded protein response.
Cell Death Differ 13 , 374–384.
40 Gozuacik D, Bialik S, Raveh T, Mitou G, Shohat G,
Sabanay H, Mizushima N, Yoshimori T & Kimchi A
(2008) DAP-kinase is a mediator of endoplasmic reticu-

lum stress-induced caspase activation and autophagic
cell death. Cell Death Differ
15, 1875–1886.
41 Kang C, You YJ & Avery L (2007) Dual roles of auto-
phagy in the survival of Caenorhabditis elegans during
starvation. Genes Dev 21 , 2161–2171.
42 Kang C & Avery L (2009) Death-associated protein
kinase (DAPK) and signal transduction: fine-tuning of
autophagy in Caenorhabditis elegans homeostasis.
FEBS J.
43 Kang C & Avery L (2008) To be or not to be, the level
of autophagy is the question: dual roles of autophagy in
the survival response to starvation. Autophagy 4, 82–84.
44 Klionsky DJ (2007) Autophagy: from phenomenology
to molecular understanding in less than a decade. Nat
Rev Mol Cell Biol 8, 931–937.
45 Rubinsztein DC, Gestwicki JE, Murphy LO & Klionsky
DJ (2007) Potential therapeutic applications of auto-
phagy. Nat Rev Drug Discov 6, 304–312.
46 Gozuacik D & Kimchi A (2006) DAPk protein family
and cancer. Autophagy 2, 74–79.
47 Scott RC, Schuldiner O & Neufeld TP (2004) Role and
regulation of starvation-induced autophagy in the
Drosophila fat body. Dev Cell 7, 167–178.
48 Cohen O, Inbal B, Kissil JL, Raveh T, Berissi H,
Spivak-Kroizaman T, Feinstein E & Kimchi A (1999)
DAP-kinase participates in TNF-alpha- and Fas-
induced apoptosis and its function requires the death
domain. J Cell Biol 146, 141–148.
49 Bradley JR (2008) TNF-mediated inflammatory disease.

J Pathol 214, 149–160.
50 Jin Y & Gallagher PJ (2003) Antisense depletion of
death-associated protein kinase promotes apoptosis.
J Biol Chem 278 , 51587–51593.
51 Chuang YT, Fang LW, Lin-Feng MH, Chen RH &
Lai MZ (2008) The tumor suppressor death-associated
protein kinase targets to TCR-stimulated NF-kappa B
activation. J Immunol 180, 3238–3249.
52 Mukhopadhyay R, Ray PS, Arif A, Brady AK, Kinter
M & Fox PL (2008) DAPK-ZIPK-L13a axis constitutes
DAPK and signal transduction Y. Lin et al.
56 FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS
a negative-feedback module regulating inflammatory
gene expression. Mol Cell 32, 371–382.
53 Tong A, Lynn G, Ngo V, Wong D, Moseley SL,
Ewbank JJ, Goncharov A, Wu YC, Pujol N &
Chisholm AD (2009) Negative regulation of
Caenorhabditis elegans epidermal damage responses by
death-associated protein kinase. Proc Natl Acad Sci
USA 106, 1457–1461.
54 Michie AM, McCaig AM, Nakagawa R & Vukovic M
(2009) Death-associated protein kinase (DAPK) and
signal transduction: regulation in cancer. FEBS J.
55 Kuo JC, Wang WJ, Yao CC, Wu PR & Chen RH
(2006) The tumor suppressor DAPK inhibits cell motil-
ity by blocking the integrin-mediated polarity pathway.
J Cell Biol 172, 619–631.
56 Mathew R, Karp CM, Beaudoin B, Vuong N, Chen G,
Chen HY, Bray K, Reddy A, Bhanot G, Gelinas C
et al. (2009) Autophagy suppresses tumorigenesis

through elimination of p62. Cell 137, 1062–1075.
57 Karantza-Wadsworth V, Patel S, Kravchuk O, Chen G,
Mathew R, Jin S & White E (2007) Autophagy miti-
gates metabolic stress and genome damage in mammary
tumorigenesis. Genes Dev 21, 1621–1635.
58 Mathew R, Kongara S, Beaudoin B, Karp CM, Bray
K, Degenhardt K, Chen G, Jin S & White E (2007)
Autophagy suppresses tumor progression by limiting
chromosomal instability. Genes Dev 21, 1367–1381.
59 Coussens LM & Werb Z (2002) Inflammation and
cancer. Nature 420, 860–867.
60 Weichhart T, Costantino G, Poglitsch M, Rosner M,
Zeyda M, Stuhlmeier KM, Kolbe T, Stulnig TM, Horl
WH, Hengstschlager M et al. (2008) The TSC-mTOR
signaling pathway regulates the innate inflammatory
response. Immunity 29, 565–577.
61 Lin Y, Stevens C & Hupp T (2007) Identification of a
dominant negative functional domain on DAPK-1
that degrades DAPK-1 protein and stimulates
TNFR-1-mediated apoptosis. J Biol Chem 282, 16792–
16802.
62 Zhang L, Nephew KP & Gallagher PJ (2007) Regula-
tion of death-associated protein kinase. Stabilization by
HSP90 heterocomplexes. J Biol Chem 282, 11795–
11804.
63 Jin Y, Blue EK & Gallagher PJ (2006) Control of
death-associated protein kinase (DAPK) activity by
phosphorylation and proteasomal degradation. J Biol
Chem 281, 39033–39040.
64 Citri A, Harari D, Shohat G, Ramakrishnan P, Gan J,

Lavi S, Eisenstein M, Kimchi A, Wallach D, Pietrokov-
ski S et al. (2006) Hsp90 recognizes a common surface
on client kinases. J Biol Chem 281, 14361–14369.
65 Henshall DC, Araki T, Schindler CK, Shinoda S, Lan
JQ & Simon RP (2003) Expression of death-associated
protein kinase and recruitment to the tumor necrosis
factor signaling pathway following brief seizures.
J Neurochem 86, 1260–1270.
66 Bialik S, Bresnick AR & Kimchi A (2004) DAP-kinase-
mediated morphological changes are localization depen-
dent and involve myosin-II phosphorylation. Cell Death
Differ 11, 631–644.
67 Cohen O, Feinstein E & Kimchi A (1997) DAP-kinase
is a Ca2+ ⁄ calmodulin-dependent, cytoskeletal-asso-
ciated protein kinase, with cell death-inducing functions
that depend on its catalytic activity. Embo J 16, 998–
1008.
68 Eisenberg-Lerner A & Kimchi A (2007) DAP kinase
regulates JNK signaling by binding and activating
protein kinase D under oxidative stress. Cell Death
Differ 14, 1908–1915.
69 Llambi F, Lourenco FC, Gozuacik D, Guix C, Pays L,
Del Rio G, Kimchi A & Mehlen P (2005) The depen-
dence receptor UNC5H2 mediates apoptosis through
DAP-kinase. Embo J 24, 1192–1201.
70 Shani G, Marash L, Gozuacik D, Bialik S, Teitelbaum
L, Shohat G & Kimchi A (2004) Death-associated pro-
tein kinase phosphorylates ZIP kinase, forming a
unique kinase hierarchy to activate its cell death func-
tions. Mol Cell Biol 24, 8611–8626.

71 Schumacher AM, Schavocky JP, Velentza AV,
Mirzoeva S & Watterson DM (2004) A calmodulin-
regulated protein kinase linked to neuron survival is a
substrate for the calmodulin-regulated death-associated
protein kinase. Biochemistry 43, 8116–8124.
72 Shohat G, Spivak-Kroizman T, Cohen O, Bialik S,
Shani G, Berrisi H, Eisenstein M & Kimchi A (2001)
The pro-apoptotic function of death-associated protein
kinase is controlled by a unique inhibitory autopho-
sphorylation-based mechanism. J Biol Chem 276,
47460–47467.
73 Kuo JC, Lin JR, Staddon JM, Hosoya H & Chen RH
(2003) Uncoordinated regulation of stress fibers and
focal adhesions by DAP kinase. J Cell Sci 116, 4777–
4790.
74 Houle F, Poirier A, Dumaresq J & Huot J (2007) DAP
kinase mediates the phosphorylation of tropomyosin-1
downstream of the ERK pathway, which regulates the
formation of stress fibers in response to oxidative stress.
J Cell Sci 120, 3666–3677.
Y. Lin et al. DAPK and signal transduction
FEBS Journal 277 (2010) 48–57 ª 2009 The Authors Journal compilation ª 2009 FEBS 57