Pharmacological Research 54 (2006) 317–325
Review
The different facets of protein kinases C: old and new
players in neuronal signal transduction pathways
Marialaura Amadio a , Fiorenzo Battaini b , Alessia Pascale a,∗
a
Department of Experimental and Applied Pharmacology, University of Pavia, Pavia, Italy
b Department of Neuroscience, University of Roma “Tor Vergata”, Roma, Italy
Accepted 8 August 2006
Abstract
Signal transduction pathways are crucial for cell-to-cell communication. Various molecular cascades allow the translation of distinct stimuli,
targeting the cell, into a language that the cell itself is able to understand, thus elaborating specific responses. Within this context, a strategic role is
played by protein kinases which catalyze the phosphorylation of specific substrates. The serine/threonine protein kinase C (PKC) enzymes family
(at least 10 isoforms) is implicated in the transduction of signals coupled to receptor-mediated hydrolysis of membrane phospholipids. Within this
molecular pathway, protein–protein interactions play a critical role in directing the distinct activated PKCs towards selective subcellular compartments, in order to guarantee spatio-temporal and localized cellular responses. A space-specific modulation of biochemical events is particularly
important during learning. Among the various mechanisms, the modulation of mRNA decay appears to be an efficient post-transcriptional way
of controlling gene expression during learning, allowing changes to take place in selected neuronal regions, in particular at synaptic level. To this
regard, recent studies have pointed out that PKC activation is also involved in a novel signalling cascade leading to the stabilization of specific
mRNAs. This review will especially focus the attention on the implication of PKC in memory trace formation and how alterations within this
molecular cascade may have consequences on physiological and pathological neuronal aging (i.e. Alzheimer’s disease).
© 2006 Elsevier Ltd. All rights reserved.
Keywords: PKC; RACK1; ELAV proteins; Memory; Aging and Alzheimer’s disease
Contents
1.
2.
3.
4.
Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
PKC family and the importance of the interaction with scaffolding proteins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Role of PKCs within some cellular functions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.1. Nuclear activities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.2. Neurotransmission . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.3. Hypotheses on the involvement of ELAVs in memory and on their relationship with PKC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.4. Physiological and pathological aging target PKC pathway . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1. Introduction
The flow of information from the outside to the inside of the
cell and vice versa allows each cell to adapt to continual changes
∗
Corresponding author. Tel.: +39 0382 987 963; fax: +39 0382 987 405.
E-mail addresses: [email protected] (F. Battaini),
[email protected] (A. Pascale).
1043-6618/$ – see front matter © 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.phrs.2006.08.002
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and requests coming from the external environment, and to
maintain a general equilibrium. Among the multiple molecular
systems belonging to the so-called signal transduction process,
the proteins phosphorylation and dephosphorylation represent a
versatile mechanism to mediate cellular responses. The two main
classes of enzymes controlling these processes are on one side
the protein kinases, which catalyse the phosphotransfer reaction,
and on the other side the phosphatases, which are implicated in
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M. Amadio et al. / Pharmacological Research 54 (2006) 317–325
the subsequent dephosphorylating hydrolysis reaction. Several
signalling networks, operating on both kinases and phosphatases, orchestrate these phosphorylation/dephosphorylation
events. Anchoring proteins also participate in this complex
machinery that has as a final goal the regulation of specific
cellular functions, through adding/removing phosphate group(s)
on selected substrates present in distinct subcellular sites [1].
Localized responses are in fact crucial to guarantee that specific
neuronal regions become theatre of key changes, some of them
leading to synaptic remodelling occurring during learning.
2. PKC family and the importance of the interaction
with scaffolding proteins
The term protein kinase C (PKC) defines a serine/threonine
family of phosphorylating enzymes ubiquitously expressed and
implicated in multiple cellular functions. These kinases transduce signals involved in short-term processes (ion fluxes, neurotransmitter release), mid-term process (receptor modulation), as
well as long-term processes (cell proliferation, synaptic remodelling and gene expression). According to sequence homology
and sensitivity to activators, at least 10 isoforms (encoded by
nine different genes) have been described. The various PKCs
are grouped in three distinct classes: (1) calcium-dependent or
classical cPKCs (␣, ␤I, ␤II and ␥); (2) calcium-independent or
novel nPKCs (␦, ␧, ␩ and ␪); and finally (3) atypical aPKCs (␨
and ␭/␫) (for a review see [2,3]).
The general structure of the different PKCs includes conserved domains (C1–C4) (see Fig. 1) separated by variable
sequences (V1–V5). C1–C2 represent the regulatory portion of
each enzyme, where the specific activators interact with, while
C3–C4 form the catalytic region responsible for both the substrate binding and the kinase activity. The catalytic domain is
characterized by a high degree of homology among the various kinases. The C1 domain bears one (in atypical) or two
(in conventional and novel) cysteine-rich domains, located near
the amino-terminal region, that represent the docking-sites for
phosphatidylserine (PS) and the physiological activator diacylglycerol (DAG) as well as for the analogous phorbol esters (i.e.
PMA), which activate all the different PKCs, with the exception
of the atypical isoforms (that are sensitive/activated by PS and
also ceramide and phosphatidylinositol-3,4,5-trisphosphate).
The C2 region contains the Ca2+ binding-site and it is present
in both the conventional and the novel isoenzymes (as a “C2like” sequence). However, the nPKCs are not responsive to Ca2+
because of the absence in the C2-like domain of amino acids
residues essential for the Ca2+ binding: recruitment to membrane
is dependent on a C1 region with higher affinity for phospholipids (in comparison with cPKCs) [4]. Finally, C3 bears the ATP
binding lobe, while C4 contains the substrate docking sequence
[5]. All known PKCs, at the amino-terminal side, are also characterized by a pseudosubstrate or autoinhibitory region, adjacent
to C1, which keeps each isoform in an inactive conformation, as
indicated in Fig. 1 as a “folded conformation”. The interaction
with the specific activators allows the activation of the enzyme
by opening its folded conformation: these cofactors are in fact
able to decrease the affinity of the pseudosubstrate domain for
the catalytic site that can exert its phosphorylating activity. This
activation mechanism is related to the translocation of PKCs
to different intracellular sites [6], and the subsequent phosphorylation of specific substrates by the translocated enzyme. The
translocation/activation mechanism of each PKC from one to
another subcellular compartment was firstly thought to reflect
only the interaction between PKC and lipids. However, several
proteins can interact with inactive and active PKCs dictating
enzyme location in both basal and stimulated conditions, underlining the additional relevance of protein–protein interactions in
PKCs homeostasis [7,8]. A relevant example is the importance
of receptors for activated C kinase (RACKs) in the localization
of activated PKCs close to pertinent substrates [9–11] (see also
Fig. 2).
RACKs are intracellular scaffold proteins belonging to the
tryptophan-aspartate 40 (WD40) repeat family. They are homologous to the ␤-subunit of heterotrimeric G-proteins. These
anchoring proteins appear to be isoform-specific; so far, four
RACK-like proteins have been described: RACK1 preferentially
binds to PKC␤II [12]; RACK2 (␤ -COP) interacts with PKC␧
[13], PRKCBP1 is a putative anchoring protein for PKC␤I [14]
while p32 (gC1qBP) can bind activated PKC␦ [15]. RACK1,
cloned from a rat brain cDNA library [12], is the best described
member of this family of scaffolding proteins. RACK1 comprises seven WD40 repeats and several studies indicate that bears
various independent binding-sites [16]. The individual repeats
can simultaneously interact with diverse signalling molecules,
suggesting that RACK1 can participate to distinct intracellular
cascades [10,17].
A direct correlation between PKC and RACK1 has been
demonstrated in cardiac myocytes where phorbol ester-activated
PKC␤ colocalizes with RACK1 [18], confirming the key implication of RACKs in PKC-mediated functions. In addition, PKC␤II and RACK1 redistribute synchronously after activation with
both phorbol esters and dopamine in CHO cells overexpressing the long isoform of the dopamine receptor type 2 [19]. In
astrocytes, translocation of PKC from the cytosol to the plasma
membrane has been reported to precede and to be related to
the transformation of polygonal-shaped astrocytes into processbearing cells [20,21]. It is interesting to note that such changes
in astrocyte morphology are documented not only during brain
development, but also in pathological conditions such as reactive
gliosis [22]. We found that in rat astrocytes, following phorbol
esters exposure, PKC-␤II was efficiently translocated from the
cytosol to the membrane compartment [23]. However, RACK1
did not redistribute synchronously with PKC-␤II to the plasma
membrane, as instead reported in studies on activated CHO
(expressing dopamine D2 receptors) and in neuroblastomaglioma hybrid NG108-15 cells [19]. These data may indicate that
the requirement for an asynchronous relocation of PKC-␤II and
its anchoring protein is specific in astrocytes. On the other hand,
we observed that the integrity of the actin-cytoskeleton (but
not of tubulin-cytoskeleton) was required in these cells, since
only the disruption of the actin-based cytoskeleton deranged
the translocation process of PKC-␤II. Additional functional
data on chromaffin cells indicate that the interaction of F-actin
with activated PKC␣ and ␤ through the binding to RACK1 is
M. Amadio et al. / Pharmacological Research 54 (2006) 317–325
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Fig. 1. Signal transduction PKC cascades: the interaction of calcium-dependent PKCs with selective activators [diacylglycerol (DAG), phosphatidylserine (PS) and
calcium (Ca2+ )] reduces the affinity of the pseudosubstrate domain for the catalytic site thus opening the folded conformation and allowing the activation of the enzyme.
The activated PKC can be directed to specific subcellular compartments via anchoring proteins (i.e. RACKs). PKC is also involved in the phosphorylation/activation
of the mRNA-stabilizing ELAV proteins, especially important for gene expression regulation. The correct activation of these molecular pathways triggers specific
responses in the cell; in contrast, changes in any of these steps may affect cellular responses and be implicated in aging-associated physiological or pathological
alterations (i.e. Alzheimer’s disease). [ARE: Adenine-uridine rich element; ELAV: embryonic lethal abnormal vision; PKC: protein kinase C; RACKs: receptors for
activated C kinase.]
essential for the potentiation by phorbol ester of catecholamine
release (Ohara-Imaizumi, Battaini and Kumakura, in preparation). According to all these results, we can hypothesize that,
within some transduction pathways, PKC may translocate, via
actin filaments, to different pools of RACKs located in distinct
compartments. RACKs may then act not only as shuttling proteins for PKCs, but also as protein acceptors. We also propose
that, in some cells, rapidly occurring changes of actin cytoskeleton organization may be implicated in the fast reprogramming
of PKCs targeting to specific subcellular compartments where
these enzymes can phosphorylate appropriate substrates.
A fascinating role proposed for RACK1 is that it may also
assemble signalling complexes on the ribosomes (for a review
see [24]). Within this context, it has been reported that the
PKC-␤II-mediated phosphorylation of the initiation factor eIF6
allows the functional assembling of the ribosome. In fact, the
unphosphorylated form of eIF6 binds to the large subunit of
the ribosome, thus preventing its association with the smaller
ribosomal subunit, while the phosphorylated eIF6 dissociates
from the large ribosomal subunit allowing its association with
the smaller subunit [25].
One of the most exciting aspects of PKC/RACKs interactions,
having relevant pharmacological implications, is the observation
that peptides that specifically inhibit or stimulate PKC binding
to its RACK, inhibit or stimulate the activity of that specific
isozyme in vitro [9]. These peptides with appropriate carriers in
vivo can be delivered to different organs, including brain, where
that PKC isozyme is specifically affected [26].
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M. Amadio et al. / Pharmacological Research 54 (2006) 317–325
Fig. 2. PKC translocation-dependent pathways. The activation of Gq-coupled metabotropic or tyrosine kinase receptors leads to the hydrolysis of membrane
phospholipids (i.e. PIP2, phospatidyl-inositol-4,5-bisphosphate) via phospholipase C activation and the generation of the two second messengers: diacylglycerol
(DAG) and inositol trisphophate (IP3). IP3 induces the following release of calcium (Ca2+ ) from the intracellular store—the endoplasmic reticulum. Ca2+ and DAG
activate PKC. Activated PKC can either bind to soluble RACK and being shuttled to subcellular sites or bind RACK directly close to relevant substrates where it can
exert its specific function (i.e. on synaptic remodelling processes associated with learning; on proliferation and differentiation mechanisms; on translation control,
etc.; see text for more details).
Besides the already mentioned RACKs, proteins that interact with C kinase (PICKs) and substrates that interact with
C kinase (STICKs) have been reported to be also implicated
in PKCs compartmentalization. Although through slightly different mechanisms, the common function of these anchoring
proteins is to recruit the various PKCs within specific signalling
networks where they can affect selected cellular functions
[27].
3. Role of PKCs within some cellular functions
3.1. Nuclear activities
Although most of the biological functions of PKC have
been related to events occurring within the membrane/cytoplasm
compartments, a growing body of evidence point out on a role
of PKC in nuclear functions as DNA replication, RNA synthesis and processing, gene expression [28]. Different PKCs have
been mapped in the nucleus in various tissues and especially
at brain level, where PKC is activated in response to a variety
of mitogenic stimuli. Within this context, several studies have
tried to link the presence of nuclear PKCs with cell proliferation.
The most direct finding has come from investigations in HL60
cells on the nuclear envelope component laminin B, a major
PKC substrate at this level [29]. Laminin B is phosphorylated
by PKC-␤II following its translocation to the nucleus after the
treatment with a DAG-mimicking compound (i.e. Bryostatin).
Functionally, this phosphorylation triggers laminin B solubilization. These results indicate that PKC-␤II stimulation is required
for nuclear laminin phosphorylation and disassembly leading to
the entry into mitosis. In parallel, during the G2 –M transition,
increased levels of nuclear DAG have been also documented
[30].
Increasing evidences indicate that the translocation to the
nucleus of the various PKCs may be implicated in cell differentiation, mainly of hematolymphopoietic lineage (reviewed
in [31]). However, functional studies addressed to how PKCs
may affect cellular differentiation are lacking. One possibility is
that nuclear PKCs may mediate transcriptional changes of genes
involved in this process.
In contrast to necrosis, which is a form of cell death that
results from acute cellular injury, apoptosis, or programmed cell
death, is a normal component of the development and health
of multicellular organisms and it is carried out in an ordered
and highly regulated fashion. Within this context, a role in such
an important mechanism has also been reported for different
PKCs, being either proapoptotic or antiapoptotic depending on
PKC isozyme and cell type (for a review see [28]). A direct
link between PKC and apoptosis was documented in HL60
cells. In these cells, induced to apoptosis, the PKC␣-mediated
M. Amadio et al. / Pharmacological Research 54 (2006) 317–325
phosphorylation of laminin B preceded both laminin proteolytic
degradation and DNA fragmentation [32].
However, although some information have been added within
the field of nuclear PKCs, more details need to be still collected
in order to better understand the physiological significance of
the phosphorylation events taking place in the nucleus.
As far as RACK1 is concerned, although this protein is not
constitutively expressed in nuclei, at least at brain level [33],
it can move to nuclei upon acute ethanol exposure in cultured
neurons and in vivo mouse brain areas involved in memory (hippocampus CA2/3 and cerebellum) [34] and then regulates gene
expression, at least in glioma cells [35].
3.2. Neurotransmission
Release of neurotransmitters from presynaptic terminals
requires a highly specialized mechanism leading to the fusion
of synaptic vescicles with the plasma membrane. This multiplesteps process is regulated by various protein kinases; actually,
all the elementary steps implicated in the exocytosis mechanism
can be modulated by phosphorylation. Within this context, in
PC12 cells the SNARE protein family plays a key role in promoting membrane fusion; indeed, SNAP-25 has been reported to
be a PKC substrate [36]. Growth associated protein-43 (GAP43), a presynaptic PKC target, also seems to interact directly
with the components of the synaptic release machinery, including the SNARE complex, and some of these interactions seem to
depend on PKC-mediated GAP-43 phophorylation [37]. Additionally, an intriguing partner for RACK1 is the SNARE protein Syntaxin1A which binds the dopamine transporter (DAT).
DAT is implicated in the regulation of the extent and duration of the dopamine receptor activation. This process depends
on DAT phosphorylation by PKC, stressing the involvement
of PKC in neuronal exocytosis [38]. Moreover, RACK1 also
interacts with the NR2B subunit of the NMDA receptor, the
major excitatory system in the brain. Modulation of the neuronal NMDA is critical within synaptic transmission; in fact,
the regulation of channel gating and trafficking via PKCs is intimately related to neuronal synaptic plasticity [39]. RACK1 has
also been reported to participate in the functioning of GABA
receptors [40], the primary inhibitory system in the mammalian
brain. Furthermore, it has been documented that, in adrenal
chromaffin cells, PKC-dependent phosphorylation of myristoylated Alanine-rich C-kinase substrate (MARCKS) [41] and
of GAP-43 [42] are involved in the regulation of the rapid
phase of catecholamine release. Finally, PKC inhibitors suppress phorbol-ester mediated potentiation of neurotransmitter
release [43].
3.3. Hypotheses on the involvement of ELAVs in memory
and on their relationship with PKC
Memory processes allow organisms to keep and recall
experiences. Persistent changes in the efficacy of cell-to-cell
communication are assumed to reflect long-lasting biochemical and morphological modifications implicated in encoding
mechanisms.
321
Investigations on animal models and patients suggest that
memory is not a single entity; rather, different kinds of memory
can be dissected and involved in distinct neuronal systems in
the brain (reviewed in [44]). Within this context, several families of protein kinases have been reported to play a key role in
learning and memory processes. In particular, growing evidence
documents the importance of PKC (see also [45]).
The implication of long term potentiation (LTP) in memory
storage has been widely explored and is a well established model
for learning and memory. In this synaptic model of memory,
brief trains of high-frequency stimuli to monosynaptic excitatory pathways trigger a sustained increase in the efficiency of
synaptic transmission (for a review see [46,47]). Several studies
have indicated a role of PKC isoforms both in the induction and
in the maintenance phases of LTP [45] as well as of the constitutive active PKM␨ (deriving from endogenous proteolysis of
PKC␨) specifically in the maintenance step [48].
The prominent involvement of PKC in neuronal changes
related to associative learning was initially pointed out by investigations on the marine snail Hermissenda crassicornis [49].
Pavlovian conditioning was also shown to trigger PKC redistribution in rabbit hippocampus [50], and following behavioural
studies reported that the kinase C pathway is also implicated in
spatial memory [51,52]. In fact, poor spatial learners present
decreased PKC activity in the hippocampus [52], as well as
a reduced amount of the neuron-specific PKC␥ [53]. Moreover, intraventricular injections of phorbol esters improve spatial learning abilities in rodents [54], while kinase inhibitors
administration impairs the same performances [55]. Furthermore, pharmacological investigations proved that nootropic
drugs, known to facilitate cognitive mechanisms, target PKC
[56–58].
Generally speaking, it should be taken into account that learning is a complex mechanism associated with specific biochemical changes taking place in selected neuronal regions. These
changes reflect both transcriptional and post-transcriptional controls of gene expression, although only post-transcriptional
mechanisms permit a localized modulation of biochemical
events occurring, at synaptic level, during learning. Within this
context, literature data suggest the importance of messenger
RNA stabilization and the consequent increase of the amount of
a given gene product. The best characterized RNA binding proteins reported to act post-transcriptionally as positive controllers
of gene expression are the ELAV proteins. ELAVs are highly
conserved proteins that preferentially interact with adenine and
uridine-rich elements (AREs). These ARE motifs are present
in the 3 -untranslated region (3 -UTR) of a subset of mRNAs,
including those of many early responsive genes, which through
their 3 -UTR are targeted for rapid degradation [59]. Following
the binding of ARE-containing mRNAs to ELAV proteins, there
is an increase in their cytoplasmic stability and rate of translation
[60].
Of relevance to this aspect, we showed that in vivo PKC
activation is related to an upregulation of the neuron-specific
ELAV proteins (nELAVs) in the hippocampal regions of rat
brain [61]. Studies performed by our group previously demonstrated that nELAVs play a critical physiological role in control-
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M. Amadio et al. / Pharmacological Research 54 (2006) 317–325
ling gene expression during memory trace formation. Indeed,
only animals that had learned a spatial task showed an upregulation of nELAVs in hippocampal pyramidal cells [62]. This
change was accompanied by enhanced expression of GAP-43
[63], a gene known to be implicated in synaptic plasticity and
whose mRNA is a nELAVs target [64]. The strategic implication of nELAVs in memory was confirmed by experiments
showing a significant impairment in the radial maze performance of mice treated with an antisense specifically raised
against with one of the nELAVs [62]. Finally, we showed a
specific increase, both in vitro and in vivo, of the expression
of the nELAV HuD subsequent to treatment with PKC activators (e.g. DAG-mimicking compounds such as bryostatins)
[61], which were also reported to improve cognitive abilities
in Alzheimer’s disease transgenic mice [65]. All these findings
indicate that PKC signalling and nELAV proteins recruitment
and activation belong to the same molecular pathway that ultimately can affect the expression of genes essential for synaptic
remodelling. It remains to be explored whether RACK1 protein
also participates within this cellular cascade anchoring PKCs
to the specific ELAV substrates. This new scenario opens novel
pharmacological approaches focused to modulate learning and
memory processes, and to eventually counteract age-related
alterations.
3.4. Physiological and pathological aging target PKC
pathway
A growing body of evidence clearly indicates that memory
deteriorates with aging. Some of the prominent changes of the
brain reflect alterations involving a number of neurotransmitter systems leading to modified interneuronal communications.
Taking into account that protein kinases play a key role in numerous signal transduction cascades, alterations in the function of
these enzymes may be directly related to neuronal dysfunctions
taking place in aging.
Considering that PKC stimulation strictly depends on an
increase of intracellular free calcium levels and on cell membrane lipid composition, age-related changes in these parameters
may affect the cellular functions mediated by this kinase family.
In order to avoid the influence of these variables, we explored
the functional response of PKC during senescence in terms of
enzyme translocation. We found that, following phorbol esters
exposure, calcium-dependent brain PKC translocation was preserved up to middle-age (8 months old), but was impaired in aged
rodents (more than 24 months) in both cortex and hippocampus,
this effect being strain-independent as opposed to kinase activity
[66–68]. Additionally, PKC translocation process was blunted
also in terms of calcium-independent activity in cortical tissues
of senescent Wistar rats [69]. These findings indicate that the
age-related alteration of this mechanism is independent of eventual changes in neurotransmitter release, receptor(s) availability
and second messenger(s) production, since phorbol esters challenge bypass receptor activation and the subsequent IP3 /DAG
production. Further results indicate that the amount of the various isoenzymes (both calcium-dependent and -independent) are
preserved during senescence, stressing the concept that only the
enzyme activation undergoes age-related changes irrespective
of the strain investigated [67,70].
Taking into account the presumable implication of RACK
proteins in directing the activated PKCs to relevant targets,
we explored whether the age-mediated impairment observed
in PKC relocation could be associated with additional changes
in protein–protein interactions. Following researches were thus
aimed to assess RACK1 content during senescence. We found
that, in comparison with adult and middle-aged animals, old
Wistar rats presented a reduced amount of RACK1 in the
brain cortex (roughly 50%) [69]. Further results obtained in
aged Sprague–Dawley rats confirmed a decrease in cortical
RACK1 protein content, coupled with a parallel reduction in
mRNA levels, also in this rodent strain [71]. Overall these
data stress the involvement of RACKs in PKC translocation
and indicate that changes occurring in senescence may affect
those cellular components essential for PKCs full activation and
function.
Alzheimer’s disease (AD), the most common form of senile
dementia, is characterized by gradual deterioration of cognitive
functions and memory, associated with widespread neuronal
death [72–74]. Several in vitro studies demonstrated a direct
implication of the PKC transduction cascade also in brain neurodegeneration occurring in AD. For the sporadic form of the
disease, the highest risk factor is represented by aging. The
growing attention devoted to this dramatic pathology is related
to the increase of span-life during these last decades. Neurofibrillary tangles and neuritic plaques are distinct markers of
AD. Intracellular neurofibrillary tangles are characterized by
paired helical filaments that are forms of abnormally phosphorylated tau protein, while neuritic plaques, which are largely
extracellular lesions, are mainly composed of a 40–42 amino
acids protein referred to as ␤-amyloid peptide (A␤). A␤ is produced via an aberrant proteolytic cleavage of a larger precursor
named APP (amyloid precursor protein). The amyloidogenic
cleavage of APP occurs through ␤-secretase that firstly originates a C-terminal fragment (CTF␤), which is subsequently cut
by ␥-secretase to generate A␤. The non-amyloidogenic pathway of APP involves, instead, the ␣-secretase enzyme that
cuts within the A␤ sequence leading to the secretion of a soluble APP fragment (sAPP) [75]. Several in vitro researches
point out a direct role of PKC in the regulation of the nonamyloidogenic metabolism of APP [76]. Relevantly to our
previous findings, PKC activation is deficient in AD brain in
terms of marked loss of translocation of cortical PKC from
the cytosolic compartment to the membrane in response to
phorbol esters and K+ depolarization [77]. Moreover, according to our data obtained from rat brains during senescence,
also in Alzheimer’s disease the expression of RACK1 was
reduced in AD human frontal cortex when compared to agematched controls [78]. In analogy with our previous results
from old rodents (see [68]), no pathology-related changes were
documented in PKC-␤II basal protein contents in AD tissues
[78]. These data strongly indicate that AD is not associated
with a reduced expression of PKCs, rather, the mechanism
of kinase activation-anchoring undergoes pathology-dependent
alterations.
M. Amadio et al. / Pharmacological Research 54 (2006) 317–325
4. Conclusion
Literature data clearly indicate that PKC certainly plays a
critical role in several signal transduction pathways. Within the
multiple functions in which the PKC signalling cascade is implicated, recent investigations indicate also its involvement in a new
neuronal pathway leading to mRNA stabilization (see Fig. 1).
However, how the cell is able to selectively elaborate a precise
response in a distinct neuronal district remains a still partially
unsolved problem. During the last years a more complex scenario is emerging, where different cellular components seem to
be involved in order to assess specific and localized responses.
A primary role within this complex machinery is played by the
so-called anchoring proteins, able to direct the various PKCs
in distinct compartments of the cell (see Fig. 2). Memory and
learning processes represent the best examples where localized
responses play a key importance. In fact, molecular changes
taking place at synaptic level are essential for neuronal plasticity and remodelling mechanisms to occur. Changes in any
of these cellular components or a disruption of the PKC system may then have consequences in cell alterations, such as
those documented in physiological or pathological aging (i.e.
Alzheimer’s disease) or in other nervous system-related abnormal states [79]. A better understanding of the whole picture may
lead to new pharmacological interventions aimed to improve
or modulate those signal transduction pathways affected during
senescence or age-associated pathologies that are responsible for
memory disturbances. A first step in this direction is the observation that activation of PKC with the non tumour promoter
bryostatin decreases brain beta amyloid accumulation, premature death and improves behavioural outcomes in mice models
of Alzheimer’s disease [65].
Acknowledgments
The authors would like to thank Annamaria Pascale-Proksch
for carefully reviewing the manuscript, and Dr. Miriam Duchen
for her precious and constant support. This work was partially
sponsored by a grant from Italian Ministero Sanità/Regione
Lazio (Progetto Alzheimer) to F.B.
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