2. Outline
Objectives
Introduction
Dawn stream effectors of DAG
PKC induced cellular processes
The DGK, RasGRP, PKD, Chimaerins, and Munc 13
Summary
Reference
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3. Objectives
At the end of this session you will be able to:
Define diacylglycerol
Describe the signaling pathways of diacyleglycerol
List the down stream effectors of DAG and
List some of the cellular processes takes place by DAG through PKC
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4. Introduction
Diacylglycerol (DAG) is a lipid signal messenger which is an
essential second messenger in mammalian cells.
Is glycerol derivative that have two hydroxyl groups esterified by
fatty acids.
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R and R' are saturated or unsaturated hydrocarbon chains
5. Cont...
Key enzymes in most of the signaling processes that generate DAG are the
phosphatidylinositol 4,5-bisphosphate- specific phospholipase C isozymes.
Hydrolysis of Phosphatidyl inositol 4,5 bisphosphate (PIP2) by PLC causes the
formation of DAG and inositol 1,4,5-trisphosphate (IP3).
In compared to IP3, DAG is very nonpolar molecule therefore it remain in the
membrane while IP3 is released into cytosol.
In addition, DAGs are produced from phosphatidylcholine (PC) by two
subsequent reactions involving two mammalian PC-specific phospholipase D
isozymes (PCPLDs) and phosphatidic acid phosphohydrolase.
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7. Cont…
The most prominent downstream effector of diacylglycerol and the functionally
analogous phorbol esters belong to the protein kinase C family, but at least five
alternative types are known:
Protein kinase D,
RasGRPs,
Chimaerins, and
Munc13s.
These can functions independently of protein kinase C isozymes, and regulates
processes as diverse as gene transcription, lipid signaling, cytoskeletal dynamics,
intracellular membrane trafficking, or neurotransmitter release.
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8. Signaling pathway through PKC
Protein kinase C is a family of protein kinase enzymes that are involved in
controlling the function of other proteins through phosphorylation.
They are divided into three subfamilies, based on their second messenger
requirements:
Conventional (c)PKCs contain the isoforms α, βI, βII, and γ. These require
Ca2+, DAG, and a phospholipid such as phosphatidylserine for activation.
Novel (n)PKCs include the δ, ε, η, and θ isoforms, and require DAG, but do
not require Ca2+ for activation.
Atypical (a)PKCs includes ζ and λ isoforms.
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10. PKC signaling induced cellular processes
PKC is involved in regulation of diverse cellular processes including
proliferation, differentiation, neural development, synaptic transmission, axonal
regeneration, smooth muscle contraction, gene expression, etc.
Example: Cell proliferation by PKC signaling
Activation of PKC induces phosphorylation of transcription factor serum
response factor (SRF).
SRF regulates immediate early genes (IEGs) such as Egr-1 (early growth
response gene 1) that control cell proliferation.
Immediate early genes with proteins such as c-jun, c-fos, myc which are
transcription factors and activate target gene resulting in cell proliferation.
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12. cont…
PKCθ plays important roll in T cell activation by activating Nuclear factor kappa-
light-chain-enhancer of activated B cells (NF-kβ), activator protein 1(AP1),
Nuclear factor activating T cell (NFAT), T cell proliferation by IL2( inter leukin
2), Cell apoptosis by FasL.
PKC is anchored to membrane by specific proteins often localized to specific
subcellular sites. This PKC anchoring proteins are named receptors for activated
C kinases (RACKs).
Therefore cells often regulates PKC effect according to their need by RACKs.
Cell type.docx
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13. Other down stream effectors of DAG
Protein kinase Ds The PKD family in mammals consists of PKD1,
PKD2, and PKD3.
All PKDs contain two C1 domains, a negatively charged central
domain, a pleckstrin homology (PH) domain, and a serine/threonine
protein kinase catalytic domain. PKD1 contains an additional
amino-terminal apolar domain.
The major PKD activation mechanism involves phosphorylation by
PKCα, PKCε, and PKCη at sites in the regulatory domain
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15. Cont…
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PKD regulate processes like signal transduction, membrane trafficking, and cell
survival, migration, differentiation, and proliferation.
16. Cont…
Through their RasGEF domains, most RasGRPs(Ras guanyl
nucleotide-releasing protein) promote GDP/GTP exchange and
activation of Ras and related small GTPases, which leads to the
stimulation of the Raf-MEK-ERK cascade and to the regulation of
other signaling pathways.
Through activation of the Raf-MEK-ERK pathway, RasGRPs have
been implicated in cell differentiation, cell proliferation, cell
transformation, T-cell receptor signaling, T cell differentiation, B-
cell receptor signaling, and neuronal differentiation.
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17. Cont…
Chimaerins the first high affinity non-PKC DAG/PE receptors
discovered, By inactivating Rac, necessarily interfere with all
downstream effects of Rac (e.g. formation of lamellipodia and
membrane ruffles, and loss of stress fibers).
Generally, chimaerins are implicated in diverse cellular processes
such as cell adhesion, cytoskeletal dynamics, lamellipodia/
filopodia formation, phagocytosis, and cell proliferation.
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18. Cont…
Munc13s act by unfolding and activating the soluble N-ethyl
maleimide sensitive factor attachment protein receptor (SNARE)
protein Syntaxin and thereby promoting SNARE complex
formation and secretory vesicle priming.
Munc13s rather than PKCs are the only functionally relevant DAG
sensitive presynaptic regulators of transmitter release. Similarly,
insulin secretion from pancreatic b cells is increased by Munc13
action in experiment.
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19. Regulation of DAG
DAG kinases phosphorylate DAG to form phosphatidic acid.
DAG kinases have a major role in intracellular signaling in which
they terminate the DAG signal generated by PLCs and form
phosphatidic acid, which itself is a signaling molecule.
DAG kinases are thought to be active only in spatially restricted
compartments following physiological DAG generation.
One such subcellular compartment is the cytoskeleton where DAG
kinases might regulate cytoskeletal dynamics by producing
phosphatidic acid, which would affect actin capping proteins and
actin polymerization. 6/21/2016by Yonas Teshome
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20. In addition, DAGkinase dependent regulation of DAG and
phosphatidic acid levels affect GTPase activating proteins for Rho
family members and certain DAG kinases interact directly with
Rho GTPase.
DAG kinases(α,β,γ) are also constitutively localized in or recruited
to nuclear compartments where they are thought to regulate a
strictly compartmentalized DAG signaling pathway that controls
cell proliferation.
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21. Summary
DAG is an essential second messenger in mammalian cells.
DAG is formed by hydrolysis of PIP2 by an enzyme called PLC.
Intracellular targets of DAG are: Protein kinase C, Protein kinase D, RasGRPs,
Chimaerins, and Munc13s.
As a second messenger DAG signals diverse cellular processes including
proliferation, differentiation, neural development, synaptic transmission, axonal
regeneration, smooth muscle contraction, gene expression, etc.
DAG kinases have a major role in intracellular signaling in which they terminate
the DAG signal generated by PLCs with receptors for activated C kinases
(RACKs).
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22. References
N. Brose, A. Betz and H. Wegmeyer, Divergent and convergent signaling by the
diacylglycerol second messenger pathway in mammals
Yukiko K. Kaneko* and T. Ishikawa, Diacylglycerol Signaling Pathway in
Pancreatic β-Cells: An Essential Role of Diacylglycerol Kinase in the Regulation
of Insulin Secretion, Shizuoka 422–8526, Japan. Received January 20, 2015
https://en.wikipedia.org/wiki/Protein_kinase_C
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