G-Protein Coupled Receptors (GPCRs) are plasma membrane proteins with seven α-helical transmembrane domains. They have an extracellular N-terminus and an intracellular C-terminal tail. The extracellular region binds to ligands and are a means of transmitting extracellular signals into the cell. The cytoplasmic regions interact with a multisubunit G-protein. The G-proteins bind to GDP when associated with the GPCR. The GPCR is a guanine nucleotide exchange factor (GEF) and exchanges GDP for GTP when an activating ligand binds. This acts as an off/on switch for the signalling activity of G-proteins. G-proteins coupled to GTP are active and transmit the signal while G-proteins coupled to GDP are inactive and remain receptor-associated.
GPCR are a huge family of receptors, participating in a multitude of signalling pathways from calcium sensing to endocrine activity. There are almost a thousand different types of GPCR in the human genome, and for many of them the function is not yet known. Some GPCR will be “pleiotropic” which means they can utilise more than one pathway. Their ligands are highly variable and include hormones and neurotransmitters. Loops between the helical domains act as binding sites both extracellularly for signal molecules and intracellularly for G-proteins.
The G protein itself is made up of an α, β and γ subunit. The α-subunit has the GTPase activity, and carries out signal transduction. The β and γ subunits normally inhibit the GDP-bound α-subunit and localise it to the plasma membrane. Specificity of the α-subunit is the main determinant in signalling activity and there are a number of different isoforms of this subunit including:
Gαs exchanges GDP for GTP upon ligand binding to the GPCR (e.g. glucagon binding to glucagon receptor). This allows it to dissociate into the cytoplasm and activate adenylyl cyclase. This converts ATP to cAMP, the second messenger of the Gαs pathway. The increase in cAMP activates PKA, a protein kinase which phosphorylates many different targets. cAMP also activates EPACs and regulates calcium ion channels which in turn have a broad spectrum of targets.
The Gαi pathway antagonises the Gαs through the inhibition of adenylyl cyclase. Upon ligand binding to the GPCR and Gαi exchanging GDP for GTP, Gαi dissociates and inhibits adenylyl cyclase and consequently prevents the activation of PKA.
Some GPCR are pleiotropic and will activate both Gαi and Gαs as a way of controlling signalling. Gαi also controls ion channels for example it leads to the opening of potassium and closing of calcium channels.
The Gαq pathway has two mechanisms of signalling. Gαq activates phospholipase C (PLC) which is anchored to the membrane and converts phosphatidylinositol 4,5-bisphosphate (PIP2), to diacylglycerol (DAG) and inositol 3-phosphate (IP3).
DAG is found in the membrane and leads to the activation of protein kinase C (PKC), a protein kinase with many targets. IP3 dissociates into the cytoplasm and activates a membrane receptor localised to the sarcoplasmic reticulum, IP3R. This is a calcium channel, and it's activation leads to an efflux of Ca2+ ions from the sarcoplasmic reticulum store. Rises in cytoplasmic Ca2+ can activate or inhibit numerous targets regulating processes such as gene regulating and contraction.
G12/13 α-subunits signal through Rho family GEFs. Dysfunction of this signalling mechanism leads is involved in cancer, asthma and vascular diseases. Their targets also include cytoskeletal contraction and gene regulation though a protein called ROCK.
Activation of GPCR is typically by ligand association. There are many different types of ligand for GPCRs, with novel research identifying steroid hormones (GPR30 and oestrogen), ions (Calcium Receptor on parathyroid chief cells) or fatty acids (GPR40 in glucose homeostasis tissues such as Beta islets or liver). With light sensing, retinal is permanently associated to its GPCR and converts from an inactive 11-cis to active all-trans upon light stimulation. The conformation change causes the activation of the associated rhodopsin protein, a GPCR.
Turning off the GPCR signalling pathways allows for complex fine tuning. Dissociation of the Ligand will inactivate the receptor, while the intrinsic GTPase activity of the Gα-subunit will turn off the G protein signalling. The GPCR can be activated or inactivated using other control mechanisms such as phosphorylation or phosphate removal. This is particularly important in the lusitropy of cardiac calcium transients (calcium influx during systole), where phosphorylation of the beta-adrenergic receptor reduces the duration of activity. Downstream kinases themselves will have further regulatory mechanisms, also normally involving phosphate removal.
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