*Corresponding author. Tel: 86-21-5080-2786; Fax: 86-21-5080-2783;
Received 25 July 2008, Accepted 29 September 2008
Keywords: AP-1, Lck, NFAT, PKD2, TCR
PKD2 interacts with Lck and regulates NFAT activity in T cells
Qing Li1, Xiaoqing Sun2, Jun Wu1,2,3, Zhixin Lin1 & Ying Luo1,2,3,*
1Department of Life Science and Biotechnology, Shanghai Jiaotong University, 1954 Huashan Road, Shanghai, 200030, China, 2Shanghai
Genomics, Inc. and Functional Genomics Ⅱ of the Chinese National Human Genome Center, Zhangjiang Hi-Tech Park, Shanghai, 201203,
China, 3GNI, 4-2-12 Toranomon, Tokyo, 1050001, Japan
Protein kinase D2 (PKD2) is a member of the PKD serine/
threonine protein kinase family that has been implicated in the
regulation of a variety of cellular processes including pro-
liferation, survival, protein trafficking and immune response. In
the present study, we report a novel interaction between PKD2
and Lck, a member of the Src tyrosine protein kinase family
that is predominantly expressed in T cells. This interaction in-
volved the C-terminal kinase domains of both PKD2 and Lck.
Moreover, co-expression of Lck enhanced the tyrosine phos-
phorylation of PKD2 and increased its kinase activity. Finally,
we report that PKD2 enhanced T cell receptor (TCR)-induced
nuclear factor of T cell (NFAT) activity in Jurkat T cells. These
results suggested that Lck regulated the activity of PKD2 by ty-
rosine phosphorylation, which in turn may have modulated
the physiological functions of PKD2 during TCR-induced T cell
activation. [BMB reports 2009; 42(1): 35-40]
Induction of cellular growth and the functional repertoire of
resting T cells requires engagement of the T cell receptor
(TCR)/CD3. One of the earliest signaling events in T cell activa-
tion via the TCR/CD3 complex is the activation of the Src and
Syk families of protein tyrosine kinases (PTKs), which leads to
the phosphorylation of many cellular proteins (1). Various
members of the Src family of protein tyrosine kinases have been
implicated in the regulation of cell growth and differentiation.
Lck is a member of the Src family of kinases expressed predom-
inantly in T cells, and plays a pivotal role in T cell activation and
differentiation (2). Through interaction with CD4 and CD8
co-receptor glycoproteins, Lck initiates signal transduction fol-
lowing engagement of the TCR complex (1, 3). Mice deficient in
Lck are markedly depleted of T cells, and those T cells that do
develop show a blunted response to TCR stimulation (4). Lck
has also been shown to bind to the IL-2 receptor complex in T
cells and participate in IL-2-mediated signaling (5).
Protein kinase D (PKD) is a family of serine/threonine kinases
that belong to a subfamily of the calcium-calmodulin kinase-like
superfamily. Three members of the PKD family have been identi-
fied so far: PKD1 (PKCμ) (6, 7), PKD2 (8) and PKD3 (PKCν) (9).
The PKDs are implicated in the regulation of fundamental bio-
logical processes including signal transduction, membrane traf-
ficking, cell survival, migration, differentiation and proliferation
(10). The salient features of PKD structure include a tandem re-
peat of zinc-finger-like cysteine-rich motifs (referred to as cys1 or
C1a, and cys2 or C1b, respectively) that are involved in recruit-
ing PKD to the membrane, a pleckstrin-homology (PH) domain
that plays an inhibitory role in the regulation of catalytic activity,
and a C-terminal catalytic domain (11-14). Although PKD1, 2
and 3 share a similar modular structure, they have overlapping
and yet distinct functional roles in various tissues and cell types
(15-17). PKDs are activated by numerous stimuli including re-
active oxygen species (ROS), growth factors (i.e. PDGF), activa-
tors of G-protein-coupled receptors and triggering of immune-
cell receptors such as the B-cell receptor (BCR) or TCR com-
plexes (15, 18-24). PKD1 is the most extensively characterized
member among the three PKD isoforms, with much less being
known about the regulation and biological roles of the other
members of the PKD family. Similar to PKD1, PKD2 is activated
by nPKCs (8, 25) and undergoes reversible translocation from the
cytosol to the plasma membrane in response to GPCR stim-
ulation (17). In contrast to PKD1, PKD2 is largely detectable in
the cytoplasm after dissociation from the plasma membrane.
A recent study reports that PKD2 is predominantly ex-
pressed in T cells and contributes to IL-2 promoter regulation
upon TCR stimulation (26). To further elucidate the regulation
and biological functions of PKD2, we performed a yeast
two-hybrid screen to identify novel PKD2 interacting partners.
In this study, we demonstrated that Lck interacts with PKD2
and increases its activity. We also showed that PKD2 regulates
NFAT activity upon TCR stimulation in Jurkat T cells.
PKD2 is associated with Lck
To identify proteins that interact with PKD2, a yeast two-hy-
brid screen of a human lymphoid cDNA library was performed
using full-length PKD2 as the bait. From a total of 7.5 × 105
PKD2 interacts with Lck and regulates NFAT activity in T cells
Qing Li, et al.
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