lecture 24 - insulin signalling Flashcards
protein domain PH
PH = pleckstrin homology domain. proteins with PH domain binds to phosphorylates inositol phospholipid in the plasma membrane
point is to localise the signalling protein to the plasma membrane
PTB domain
binds phosphotyrosine (P-Y) residues
SH2 domain
= src homology 2 domain
binds phosphotyrosine residues surrounded by unique protein sequences
SH3 domain
Src homology 3 domain
binds specifically to proline rich regions
insulin receptor is a
tetrameric tyrosine kinase receptor (2 alpha 2 beta subunits)
alpha subunit at the top so binds to insulin
the IR is expressed as two isoforms
isoform A and B
held together by disulphide bonds
beta subunit has tyrosine kinase residues meaning it can phosphorylate tyrosine when activated
insulin binding
activation of tyrosine kinase activity
phosphorylation of IR generates binding site for proteins with PTB and SH2
ser / thr can be phosphorylated and they inhibit the kinase activity
Shc - SH2 containing adapter protein
has PTB, CH1 ans SH2 domain
Shc can bind to insulin receptor
when they bind to IR, tyrosine in CH1 is phosphorylated, this uncovers a binding site for binding of SH3 domain (e.g. Grb2 from SH3 domain binds to Shc)
IRS = insulin receptor substrate (adapter protein)
4 genes: IRS 1,2,3,4
contains a PH domain and a PTB domain enables binding to phosphorylated insulin receptor
PH binds to membrane
PTB domain binds to insulin receptor
after binding the IRS becomes phosphorylated on tyrosines by the insulin receptor tyrosine kinase activity
IRS has 21 potential tyrosine phosphorylation sites
makes is a docking protein for proteins with SH2 domains
PI3 kinase, Grb2 and SHP2
can bind to IRS as they have SH2 domains
IR –> IRS/Shc –> Grb2 –> SOS –> MAP-Kinase pathway
Grb2 = growth factor receptor bound protein 2
Grb2 has one SH2 domain which binds to phoshporylated IRS or Shc
Grb2 has an SH3 domain which interacts with SOS when activated by binding to IRS/Shc
SOS acts as a GDP/GTP exchange factor
Ras is inactive when bound to GDP
Ras is activated when this is exchanged for GTP (by SOS)
Ras-GTP activates Raf, which activates MAPKK
MAPKK activates MAP- kinase
MAP-kinase drives growth, differentiation and proliferation
PI-3-kinase pathway
PI3 can bind to IRS via SH2 domain, this leads to phosphate being added to an inositol phospholipid in the plasma membrane leading to generation of PIP3
PIP3 is a binding partner for proteins with PH domains (eg protein kinase B (AKT/PKB))
dual phosphorylation of PKB/AKT
PI3K
consists of 2 subunit enzymes
p85 = regulatory unit, contains two SH2 domains and one SH3
p110 = catalytic subunit
action of PI3K
function = adds a phosphate group to the 3 position of the inositol ring
results in PIP3 which remains membrane bound
PIP3 binding site for PH domain proteins
PDK1 binds to PIP 3 and it enables the phosphorylation of a residue on AKT (thr308)
mTORC2 phosphorylates AKT on Ser473
dual phosphorylation activates AKT (PKB)
AKT
3 domains
N-terminal pleckstrin homology (binds to PIP3 in the plasma membrane
central kinase domain (Thr308)
C-terminal hydrophobic regulatory domain (Ser473)
when AKT is phosphorylated it then has the capacity to phosphorylate other proteins
AS160 blocks movement of vesicles containing GLUT4, does this by regulating Rab
for Rab to facilitate movement to the membrane it needs to be bound to GTP, AS160 prevents them binding
phosphorylation of AKT leads to phosphorylation of AS160 leading to its inactivation causing GLUT4 translocation
AKT substrate 2
protein synthesis
AKT phosphorylates TSC2 which regulates mTORC1
AKT phosphorylates and inhibits TSC2
TSC2 in complex with TSC1 acts as an inhibitor of Rheb by converting it from GTP to GDP bound state
Rheb-GTP activates mTORC1
mTORC1 activates eIF4E-BP1 and S6K1 which leads to translation initiation and ribosome biogenesis
AKT substrate: FOXO1
FOXO1 induces G6PC and PEPCK (enzymes of gluconeogenesis)
FOXO1 represses GCK (glucokinase)
AKT phosphorylates FOXO1, FOXO1 then translocates to the cytoplasm and is ubiquitinated and degraded allowing repression of G6PC and PEPCK and induction of GCK
AKT substrate
glycogen synthase kinase 3 (GSK3)
GSK3 is inactivates when phosphorylated by AKT
GSK3 when is active is able to phosphorylate many proteins (glycogen synthase and e1F2B
glycogen synthase is active when is dephosphorylated
GSK3 phosphorylates glycogen synthase meaning its inactive and not much glycogen is formed
GSK phosphorylates e1F2B which is also inactive when phosphorylated so not much protein synthesis
AKT phosphorylates GSK3 making it inactive promoting glycogen and protein synthesis
insulin also activates glycogen synthase phosphatase (GSP) and phosphorylase phosphatase (PP) dont understand how yet
