Role of protein kinase C in stimulus–response coupling

The role of protein kinase C in stimulus–response
coupling was fi rst demonstrated for the release of
serotonin from platelets. The enzyme has since
been shown to be involved in the release of certain
hormones and neurotransmitters, the secretion of
certain enzymes, smooth muscle contraction and
relaxation, activation of T and B lymphocytes, inhibition
of gap junctions, modulation of ion conductance,
interaction and down-regulation of receptors, cell
proliferation, expression of certain genes, steroidogenesis,
lipogenesis and glucose metabolism. When
intracellular Ca2+ concentrations become too high,
protein kinase C activates the Ca2+-transport ATPase
(calcium pump) and the Na+/Ca2+ exchange protein,
both of which remove Ca2+ from the cell (Nishizuka,
1986).
Protein kinase C is synthesized as an unphosphorylated
and catalytically inactive protein of 74 kDa
that is bound to the plasma membrane of the cell. The
protein is converted into an active form of 77 kDa and
then into an 80 kDa form by at least two phosphorylation
steps. The fi rst phosphorylation is initiated by an
as yet unidentifi ed kinase (‘protein kinase C kinase’).
This phosphorylation step takes place on the activation
loop of the newly synthesized protein and renders
it catalytically competent. The membrane-bound
enzyme, when stimulated by phosphatidylserine,
then phosphorylates itself at the carboxy terminus.
This autophosphorylation decreases the enzyme’s
affi nity for the membrane so that it partitions into
the cytosol (Dutil et al., 1994). The doubly phosphorylated
enzyme is allosteric, i.e. regulatable. Binding
of the activator (diacylglycerol) to the allosteric site of
the enzyme causes a conformational change, thereby
54 Vitamins: their role in the human body
exposing the catalytic site that is normally blocked by
the so-called pseudosubstrate region.
There are several subtypes of protein kinase C, each
the product of a separate gene; these are identifi ed by a
Greek letter, the α-subtype being the most ubiquitous.
Isoforms of the β-subtype (β1 and β2) are generated
post-translationally by the alternative splicing of the
primary mRNA transcript. The eleven subspecies of
protein kinase C (PKC) so far identifi ed have been
divided into three groups: the conventional (cPKC)
group comprises α, β1, β2 and γ; the new (nPKC)
group comprises δ, ε, η, θ and μ; and the atypical
(aPKC) group comprises ζ and λ(ι) (Nishizuka, 1995).
The ability of the individual subspecies to elicit different
physiological responses is presumably due to their
localization in different tissues and subcellular sites.
In the absence of diacylglycerol, protein kinase C
resides in the cytosol. When diacylglycerol is generated,
it recruits the kinase from the cytosol to the plasma
membrane. Diacylglycerol, aided by cis-unsaturated
fatty acids, also increases the affi nity of protein kinase
C for Ca2+, thereby causing full enzyme activity when
Ca2+ concentrations are basal (Nishizuka, 1995). A
proportion of the activated enzyme is targeted to the
nucleus where the phosphorylation of nuclear regulatory
proteins (transcription factors) could provide a
mechanism for the direct regulation of gene expression,
leading to changes in cell proliferation and differentiation
(Olson et al., 1993).
The diacylglycerol produced by receptor-mediated
hydrolysis of phosphatidylinositol biphosphate is
short-lived, either being re-incorporated into phosphoinositides
or hydrolysed to arachidonic acid, the
precursor of prostaglandins and thromboxane. Longterm
cellular responses, such as cell proliferation and
activation of T lymphocytes, requires sustained activation
of protein kinase C through other pathways.
For example, hydrolysis of membrane phosphatidylcholine
by phospholipase D produces phosphatidic
acid and choline. Phosphatidic acid is converted to
diacylglycerol by the action of a phosphomonoesterase,
thereby indirectly activating protein kinase C.
Phosphatidic acid may also activate protein kinase C
directly (Nishizuka, 1995).
Protein kinase C exerts negative feedback control
over various steps of its activation pathway, including
down-regulation of the cell-surface receptor. In addition,
protein kinase C exerts negative feedback control
at the level of the receptors for various growth factors,
such as epidermal growth factor.
Protein kinase C is a target for phorbol esters, such
as 12-O-tetradecanoylphorbol-13-acetate (TPA), and
probably serves as a receptor for these compounds. A
number of phorbol esters are well known as potent
tumour promoters. Extremely low concentrations of
phorbol esters are able to substitute for diacylglycerol
and activate protein kinase C. Unlike diacylglycerol,
however, phorbol esters are metabolically stable;
moreover, their entry into the cell is not susceptible
to feedback control. Phorbol esters are therefore able
to activate protein kinase C in a sustained rather than
transient manner. Prolonged treatment of cells with
phorbol esters leads to disappearance of protein
kinase C owing to an increased rate of proteolysis
(Young et al., 1987). Depletion of protein kinase C
would remove the negative feedback control that the
kinase exerts over growth factor receptors, leading to
uncontrolled cell proliferation in the presence of a mitogenic
stimulus.