Page 54 - 85 cell signalling pathways

Cell Signalling Biology Michael J. Berridge Module 2 Cell Signalling Pathways 2 54

In general, Gβγ is less efficacious than Gα in stimulating lowing B-cell antigen receptor (BCR) activation (Module
PLC. 9: Figure B cell activation). This translocation to the cell
The two G protein subunits interact with PLCβ at dif- surface has two important consequences for the activation
ferent sites. The α subunit interacts with a region within of the enzyme. Firstly, it brings the enzyme close to its
the long C-terminal tail, whereas the βγ subunit interacts substrate in the membrane. Secondly, by interacting with
with a site within the Y region of the catalytic site (Module tyrosine kinases, PLCγ1 is itself phosphorylated on tyr-
2: Figure PLC structure and function). osine residues located at positions 771, 783 and 1254 (black
PLCβ also contains GAP (GTPase-activating protein) triangles in Module 2: Figure PLC structure and function).
activity that stimulates the intrinsic GTPase activity of the Phosphorylation at Tyr-783 is particularly important for
α subunit. When an agonist binds to the GPCRs, it induces switching on enzyme activity. The PLCγ2 isoform is ac-
a conformational change that is transmitted to the under- tivated by phosphorylation of tyrosine residues 753 and
lying G protein, causing it to dissociate. At this time the 759, mainly by the tyrosine kinase Btk (Module 2: Figure
GDP bound to the α subunit is exchanged for GTP. The ROS effects on Ca 2 + signalling).
active GTP/α subunit complex can then activate PLCβ The enzymatic activity of PLCγ is also stimulated by
and this activation process is terminated when the intrinsic the PtdIns3,4,5P 3 produced by the PtdIns 3-kinase sig-
GTPase activity of the α subunit hydrolyses GTP back nalling pathway and thus represents a major point of in-
to GDP allowing the GDP/α subunit to once again bind teraction between these two signalling pathways. This in-
to the βγ subunit to form the inactive complex. The in- teraction is particularly important during lymphocyte ac-
trinsic GTPase activity of the α subunit is rather low, but tivation, where the Tec tyrosine kinase family are activ-
is greatly enhanced by two mechanisms. There is a GAP ated by PtdIns3,4,5P 3 to phosphorylate and activate both
activity associated with the long C-terminal tail of PLCβ. PLCγ1and PLCγ2. Both isoforms bind strongly to this
Therefore PLCβ plays a direct role in terminating its own highly charged lipid through the N-terminal PH domain,
activity. In addition, there are regulators of G protein sig- and this interaction enhances enzymatic activity.
nalling (RGS) proteins. There are about 20 of these RGS The Src homology 3 (SH3) domain, which binds to
proteins, of which RGS2--RGS4 seem particularly effect- proline-rich sequences, may enable PLCγ to bind to other
ive in interacting with Gα q . The GTPase activity of Gα q components, such as the cytoskeleton and the protein dy-
is enhanced 25-fold by RGS4. There is considerable dis- namin.
crimination with regard to how individual RGS proteins
bind to specific receptor/Gα q complexes enabling them
to exert an agonist-specific inhibitory mechanism. This Phospholipase Cδ (PLCδ)
mechanism could explain the different types of Ca 2 + sig- Less is known about the activation of PLCδ. Unlike PLCβ
nals observed in hepatocytes following stimulation with and PLCγ,PLCδ appears not to be regulated by receptors
different agonists. directly. Once PLCδ1 has associated with the membrane,
it appears to be activated by an elevation in cytosolic Ca 2 + .
Modulation of PLCβ by other signalling pathways This sensitivity to changes in intracellular Ca 2 + is thought
In general, the phosphorylation of PLCβ, particularly iso- to depend on the C2 domain (Module 2: Figure PLC struc-
forms 2 and 3, by protein kinase A (PKA) results in inhib- ture and function).
ition of the enzyme. Crystallographic studies of PLCδ1 have begun to reveal
how PLC enzymes function to cleave their lipid substrates
Phospholipase Cγ (PLCγ) that are embedded in the plasma membrane.
PLCγ functions predominantly in early development and
in the signalling pathways that control cell proliferation.
PLCγ1 knockout mice die at embryonic day 9, by which Phospholipase Cε (PLCε)
time the embryos appear normal, albeit somewhat smaller. Phospholipase Cε (PLCε) differs from the other isoforms
PLCγ is characterized by having a large insert between by having two Ras-association domains (RA) at its C-
the X and Y domains (Module 2: Figure PLC structure and terminus (Module 2: Figure PLC structure and function).
function). This insert contains an additional pleckstrin ho- It binds specifically to the GTP-bound forms of Ha-Ras
mology (PH) domain, which is itself split by another insert and Rap1A. The enzyme is targeted to the membrane
containing two Src homology 2 (SH2) domains and a single through its ability to bind to these G proteins. It also has
Src homology 3 (SH3) domain. The SH2 domains play a a Cdc25 homology domain that is a guanine nucleotide
critical role in the activation of PLCγ because they provide exchange factor (GEF) motif for Rap1, which enables it to
a docking module that enables the enzyme to translocate to translocate to the perinuclear region following epidermal
the membrane to dock to phosphorylated tyrosine residues growth factor (EGF) stimulation, where it can activate the
on activated receptors or associated scaffolding proteins. mitogen-activated protein kinase (MAPK) signalling path-
This tyrosine phosphorylation occurs during activation of way. There is evidence that cyclic AMP acting through the
protein tyrosine kinase-linked receptors (PTKRs) or the exchange protein activated by cAMP (EPAC) can stimu-
non-receptor protein tyrosine kinases (e.g. Src, Syk, Btk, late Rap1 to activate PLCε (Module 2: Figure cyclic AMP
Lck, Fyn). This sequence of events is well illustrated by signalling) and this mechanism has been implicated in the
the activation of PLCγ1 during activation of the T cell re- control of autophagy (Module 11: Figure autophagy sig-
ceptor (Module 9: Figure TCR signalling)orPLCγ2fol- nalling mechanisms).

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