Transcript ATP as a Signaling Molecule - ASAB-NUST

ATP as a Signaling Molecule
CONTENTS
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What is ATP..?
ATP as a Signaling Molecule
ATP Storage and Release
Purinergic Receptors
ATP in Extracellular Signaling
ATP in Intracellular Signaling
Ectonucleotidases
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In times of plenty, need, or stress, cells can release ATP and
other nucleotides into their surroundings.
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This ATP can potentially, via specific purinergic receptors,
influence the ATP-releasing cells or neighboring cells by the
processes of
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neural transmission,
paracrine signaling, and
autocrine signaling
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Physiological and Pathophysiological
associated with purinergic receptors
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synaptic transmission,
pain and touch perception,
vasomotor responses,
platelet aggregation,
endothelial release of vasorelaxants,
immune defense,
cell volume regulation,
cell proliferation and mitogenesis,
apoptosis, and epithelial ion and water transport
processes
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Action of ATP on cells depends on
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storage and release of ATP,
stimulus for the release,
population of the purinergic receptors on cells,
breakdown of ATP by ectonucleotidases,
cellular signaling pathways, and
effector processes within the stimulated cells
Communicating via ATP: the basic steps.
ATP storage and release
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Newly synthesized ATP is first made and transported
out
of the
mitochondrion
via oxidative
phosphorylation.
Steady-state cytosolic ATP conc. ~3–10 mM
Higher concentrations of ATP (up to 100s of mmol/l)
are stored in secretory vesicles of neurons and
released with acetylcholine, noradrenaline, and other
transmitters, including VIP and neuropeptide Y.
Extracellular ATP signaling in epithelial cells
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In addition
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chromaffin granules of the adrenal medulla,
serotonergic granules of platelets, and
insulin-containing granules of pancreatic-cells also store
significant amounts of ATP.
ATP release via exocytosis or
Shear stress, stretch, hypoxia, inflammation, osmotic
swelling, cell death, cholinergic stimulation.
Possible ATP release mechanisms in
epithelia
ATP-permeable release channels
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Anion channels like PM Voltage Gated Anion Channels
(VDAC)
Resides in apical & basolateral membranes
It was proposed that the cystic fibrosis (CF) transmembrane
regulator (CFTR) or other members of the ATP-binding
cassette protein family could transport ATP directly.
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The existence of ion channels, that are permeated by ATP
anions, was underscored by many laboratories investigating
whether CFTR conducted ATP itself in addition to
Clˉ or regulated a separate ion channel that was permeable
to ATPˉ.
One group suggested that CFTR and the multidrug
resistance transporter (mdr or P-glycoprotein) conducted
ATP
Another study proposed
CFTR could control maxi anion channels similar to
mitochondrial porins and/or the outward rectifier Clˉ channel,
either of which could transport ATP.
Connexin hemichannel
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One attractive proposal is the connexin hemichannel, which,
apart from being a building block for the gap junctions, is
implicated in ATP release.
The hemichannels could themselves transport ATP or be
associated with the cytoskeletal organization and movement of
intracellular vesicles.
Adenine nucleotide transporters
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Adenine nucleotide transporters may also exist that may be
carriers or permeases
Nucleotide/Nucleoside exchangers
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Exchangers may exist for ATP in the plasma membrane, as
they do in mitochondrial inner membrane, where ADP may be
the exchanged substrate. Other substrates such as Na+ or Clˉ,
which have favorable entry gradients, may be exchanged for
ATP.
ATP-filled vesicle
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ATP-filled vesicles, which may also contain additional
agonists or co-agonists, may fuse with the plasma membrane
releasing ATP.
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A high content of ATP is found in synaptic vesicles with other
neurotransmitters, mast cell granules with histamine, and
chromaffin granules with epinephrine.
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ATP and its metabolites are known co-transmitters that
modulate the effect of excitatory or inhibitory
neurotransmitters.
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ADP and ATP are also released by platelets on their own via
exocytosis for a ‘‘self-aggregation’’ signal at the clotting zone.
Purinergic receptors
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Purinoceptors, are a family of newly characterized
plasma membrane molecules involved in several
cellular functions such as vascular reactivity,
apoptosis and cytokine secretion.
Purinoceptors are divided into two classes
P1 or adenosine receptors
P2, which recognize primarily extracellular ATP, ADP, UTP
and UDP.
Two subclasses of P2 receptors- P2X
and P2Y
P2X receptors
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P2X receptors are extracellular ATP-gated
calcium-permeable non-selective cation
channels that are modulated by extracellular
Ca2 +, Na+, Mg2 +, H+ and metal ions, such
Zn2 + and/or Cu2 +.
To date, seven separate genes coding for
P2X subunits have been identified, and
named to as P2X1 through P2X7
Two transmembrane domains
connected by a large extracellular
loop and a COOH terminal of
variable length
P2Y receptors
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P2Y receptors (with exception of P2Y12 and P2Y13) couple
to heterotrimeric G proteins and phospholipases (primarily
phospholipase Cβ) to raise intracellular free calcium
concentration.
To date, 12 P2Y receptors have been cloned in humans: P2Y1,
P2Y2, P2Y4, P2Y5, P2Y6, P2Y8, P2Y9 (present in NCBI as
GPR23), P2Y10, P2Y11, P2Y12, P2Y13 and P2Y14.
P2Y receptors are present in almost all human tissues where
they exert various biological functions based on their Gprotein coupling.
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All genuine P2Y receptors (except P2Y12) activate
phospholipase C (PLC), phosphoinositide hydrolysis,
mobilization of intracellular Ca2+, and activation of PKC
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Regarding their selectivity, P2Y1, P2Y11, P2Y12, and P2Y13
are purinoceptors, P2Y6 is a pyrimidinoceptor, and P2Y2 and
P2Y4 receptors have mixed selectivity.
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P2Y11 stimulates PLC and adenylate cyclase, whereas P2Y12
and P2Y13 inhibit adenylate cyclase.
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Other P2Y receptors can also couple to several distinct G
proteins, e.g., P2Y1 and P2Y2 via Gs stimulate PLC, but via
Gi they inhibit adenylate cyclase.
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A multitude of receptors and signaling pathways offers a rich
possibility for a cross-talk mechanism, including various
isoforms of adenylyl cyclases (activated by βγ–subunits
released from G proteins), Ca2+-calmodulin and/or PKC, and
adenosine receptors after ATP is split into adenosine by
ectonucleotidases and 5'-nucleotidase.
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New pathways that may be involved in long-term
regulation for non-epithelial cells
metabolism
cell adhesion
gene expression and
growth
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For example,
In astrocytes UTP/ATP can stimulate a mitogen-activated
protein kinase (MAPK) cascade, such as extracelluar signalregulated kinase (ERK), which involves Ca2+-independent
PKC.
Topology of P2X and P2Y receptors
Mammalian P2Y receptors are 328 to 370 amino acids long, transverse the
membrane 7 times, and activate 1 or more G proteins. Mammalian P2X receptor
monomers are 339 to 595 amino acids long and contain a long COOH terminal and
a large extracellular loop, with disulfide bonds and several N-linked glycosylation
sites.
Adenosine receptors
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All adenosine receptors were shown to activate at least one
subfamily of mitogen-activated protein kinases.
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The actions of adenosine are often antagonistic or synergistic
to the actions of ATP.
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In the CNS, adenosine has multiple functions, such as
modulation of neural development, neuron and glial signalling
and the control of innate and adaptive immune systems
ATP in extracellular signaling
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Once released, ATP is free to bind to P2X and P2Y receptors,
on the same cell or neighboring cells.
Alternatively, ADP, a metabolite, may also bind with high
affinity to a subset of P2 receptors.
UTP and UDP follow similar chemistry and have their specific
P2Y receptor subtypes.
The final metabolite of ATP, adenosine, is also biologically
active and binds to P1 receptors.
Once these receptors are activated, signal transduction begins
and affects cell or tissue function.
ECTONUCLEOTIDASES
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Once ATP leaves the cell, it is also degraded rapidly; thus,
it is thought of as a local mediator that acts in an autocrine
or paracrine manner within tissues and tissue
microenvironments to stimulate its receptors before it is
chemically modified.
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Ecto-enzymes secreted into the extracellular milieu as a
secreted protein or membrane-bound as an ecto-enzyme
target ATP.
Two families of ectonucleotidases
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E-NTPDases are ecto-nucleoside triphosphate
diphosphohydrolases that hydrolyze nucleoside 5'-tri- and
diphosphates.
E-NPP,
are
ectonucleotide
pyrophosphatase/
phosphodiesterases with a broad substrate specificity.
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These can hydrolyze phosphodiester bonds of nucleotides
and nucleic acids and pyrophosphatase bonds of
nucleotides and nucleotide sugars.
e.g., cleavage of ATP to AMP and PPi and conversion of
cAMP to AMP
Resynthesis of ATP
 Secreted or membrane-bound ecto-kinases such
as adenylate kinase or nucleoside monophosphoand
diphosphokinases
can
phosphorylate
nucleosides to remake 5'-AMP, ADP and,
ultimately, the triphosphate nucleotide ATP.
Intracellular ATP signaling
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ATP is critical in signal transduction processes.
It is used by kinases as the source of phosphate groups in their
phosphate transfer reactions.
Kinase activity on substrates such as proteins or membrane
lipids are a common form of signal transduction.
Phosphorylation of a protein by kinase
Mitogen-activated protein kinase cascade
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ATP is also used by adenylate cyclase and is transformed to
the second messenger molecule cyclic AMP, which is
involved in triggering calcium signals by the release of
calcium from intracellular stores.
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This form of signal transduction is particularly important in
brain function, although it is involved in the regulation of a
multitude of other cellular processes.
A signal-transduction pathway for ATP release
from erythrocytes
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ATP is released from erythrocytes in response to a clearly
defined physiological stimulus i.e reduced hemoglobin oxygen
content
Suggests that signaling mechanism that relates this stimulus to
ATP release must be present within that cell.
This pathway includes the heterotrimeric G protein Gi ,
adenylyl cyclase (AC) , protein kinase A and the cystic
fibrosis transmembrane regulator (CFTR)
Proposed pathway for regulated ATP release from erythrocytes in
response to passage of these cells through areas of increased oxygen
demand in skeletal muscle
The increase in oxygen demand
Oxygen release from hemoglobin within the erythrocyte.
Decrease in Hemoglobin oxygen content
Activation of the heterotrimeric G protein, Gi and release of ATP
ATP released from the erythrocyte can bind to purinergic
receptors (P2y) on the vascular endothelium
Release of vasodilators and, an increase in blood flow (oxygen
delivery)