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SIGNALLING VIA LIPID MESSENGERS – PI3K
Different cell signalling pathways include:
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Phosphorylation of receptors and other molecules downstream of the receptor
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Recruitment of proteins that recognises phosphorylated tyrosine, serine/threonine
residues
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GTPase cycle, i
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Ras
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Phospholipids are amphiphatic – that is, they possess a hydrophilic head, which likes
to be immersed in water, and a hydrophilic tail, which prefers nonaqueous
environments
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This polarity explains the structure of lipid bilayers such as the plasma membrane, in
which the hydrophilic groups face and protrude into the extracellular and cytosolic
aqueous environments while the hydrophobic tails are buried in the middle of the
membrane, from which water is excluded
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Some phospholipids contain, at their hydrophilic heads, an inositol group
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Inositol is a water-soluble carbohydrate molecule, and this inositol moiety can be
modified by the addition of phosphate groups
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The resulting phosphoinositol (PI) can be:
• Cleaved by phosphilapses to remove the phospho-inositol ring, which can
then be a signal in the cytoplasm and at the membrane
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PI3K:
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These two groups differ in
the catalytic subunits – p110-alpha, -beta, -delta or –gamma
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- In turn, they regulate a wide variety of activities
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- They do not seem to acutely stimulated by extracellular stimuli
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This demonstrates that they form a fundamental process in cellular
signalling
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Class I PI3Ks preferred substrate is PI(4,5)P2, whilst that of class II and III preferred
substrate is PI(3)P
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Class I PI3Ks are activated downstream of growth factor receptors
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The p85 regulatory domain of class IA PI3Ks contain an SH2 domain, which is
recruited by the phosphorylated residue on the GFR (which can be receptor tyrosine
kinases or GPCRs)
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This causes the PI3K to undergo a conformational change thus activating the
catalytic domain
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Alternatively, activated Ras (which is activated downstream of the RTK can also bind
to and activate the catalytic domain
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Importantly, Ras is incapable of activating the PI3K catalytic domain isoform p110beta
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By forming the triply phosphorylated inositol “head group” of PIP3, PI3K creates a
docking site for cytoplasmic proteins that carry PH domains
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The most important of these is the plasma membrane via its PH domain, Akt/PKB
becomes doubly phosphorylated by two other kinases PDK1 and PDK2, and thereby
activated
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Akt then proceeds to phosphorylate a variety of substrates regulating cell
proliferation, survival and size
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However, if PTEN is also present and active, it dephosphorylates PIP3 to PIP2,
thereby depriving Akt/PKB of a docking site at the plasma membrane
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SHIP1/2 is a phosphatase that removes the 5’ position phosphate to produce
PI(3,4)P2
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Independent of Akt downstream activation, PIP3 also plays a role in the polarity and
motility of cells
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When a reporter for PIP3 is generated via GFP binding (GFP has a PH domain), you
can see the localisation of PIP3 molecules in response to mitogen signalling
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o It’s said to act a “compass”, in that, cells orientate their direction in
response to migratory signalling
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Generation of PIP3 by PI3K, then PI3K recruits Akt to the membrane via a PH
domain
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Akt is activated by PDK1, which is responsible for phosphorylating Akt and thus
activating it
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Activated Akt then phosphorylates TSC2 in the TSC2/TSC1/Rheb complex, causing a
release of Rheb from the complex
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Free Rheb is now GTP loaded and thus active, allowing GTP-Rheb to bind to and
activate mTOR
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Mammalian Target Of Rapamycin is a serine/threonine kinase discovered via the
Rapamycin drug mechanism of action
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It functions as a biological sensor for: nutrients, stress etc
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It operates at two critical nodes in the control circuitry of mammalian cells, thus
mTor integrates a variety of afferent (that is, incoming signals), including nutrient
availability and mitogens, and then acts to control glucose import and protein
synthesis and a variety of other cell-biological processes
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M-TOR is associated with two alternative partners, called Raptor and Rictor
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The m-TOR-Rictor complex and a third protein, GbL, together form the m-TORC2
complex, which is responsible for activating Akt/PKB
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The m-TOR-Raptor complex with GbL forms the m-TORC1 complex, and is
responsible for activating protein synthesis by:
• Phosphorylating S6K1, 4E-BP1, and 4E-BP2
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Inputs of m-TORC1 pathway:
• Growth factors activate m-TORC1: TSC2 is phosphorylated and functionally
INACTIVATED by Akt in response to signalling via PI3K, thus activating mTORC1, which can then induce protein synthesis
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• Low nutrient/ATP levels inactivate m-TORC1: modulated through the AMPK
pathway, thus allowing a reduction in protein synthesis until the cells are in a
more stable state
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Phosphorylation of S6K1 causes 4E-BP1 to phosphorylate the S6 protein of the small
(40S) ribosomal subunit, enabling this subunit to participate in ribosome formation,
that is, by associating with the large ribosomal subunit and thus in protein synthesis
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Additionally, by phosphorylating 4E-BP1 (and 4E-BP2), m-TOR causes 4E-BP1/2 to
release its grip on the key translational initiation factor Eif4E (eukaryotic initiation
factor 4E)
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Once liberated, Eif4E forms complexes with several other initiation factors, and the
resulting complexes enable ribosomes to initiate translation of certain mRNAs,
specifically increasing cap-dependent translation and translation of 5’-TOP mRNAs
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Conversely, m-TORC2 complex governs the activity of Akt/PKB by adding a critical
second phosphate and thereby gaining control of Akt/PKB’s multiple downstream
effectors
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Akt can thus continue to activate m-TORC1, which phosphorylates S6K that inhibits
m-TORC2, thus functioning as a negative feedback loop
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Several solid and liquid cancers have somatic p100-alpha gene mutations, such as
colon, brain, and breast cancer
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The most frequently observed PI3K mutations include:
• E542K/E545K that blocks regulation
• H1047R that activates the enzyme
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These mutations make p100-alpha protein more active, and presence of these
mutations appear to make cancer cells more sensitive to PI3K inhibitor treatment –
“oncogene addiction”
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PI3K and Cancer Treatment:
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