Cell Signalling
The binding of a ligand to a receptor leads to a change in the phosphorylation of a target protein, changing its activation state
Protein Kinase
Adds a phosphate group to the hydroxyl of serine, threonine or tyrosine (amino acids with a hydroxyl), activating it
Takes the input of an ATP molecule, which is turned into ADP
Protein Phosphatase
Removes a phosphate group from a target protein, deactivating it. Does this to serine, threonine or tyrosine. Turns the phosphate back into hydroxyl group.
Takes the input of one H2O molecule, and releases one inorganic phosphate (Pi)
Signalling cascades
1)Transfer
2)Transform
3)Amplify
4)Divergence
The chemical messenger is transferred onto the receptor of the receptor cell
In order to enter the receptor cell, the primary messenger(ligand) is transformed into a secondary messenger through a conformational change in the receptor. This turns it into a signal inside the cell
A small chemical change on the cell surface can be amplified through modulations within the cell to turn into a large chemical change
A change at one receptor can have an effect on many different target proteins in the cell
The signal pathway can be modulated by other factors along the way
Two different types of signalling mechanisms
Intracellular signalling
Small hydrophobic signal molecule moves through the cell membrane and binds to a receptor within the cell (in cytoplasm or in nucleus). Usually hormones
Cell surface receptors
A large hydrophilic signal molecule binds to a receptor on the outside of the cell membrane
Receptor sets of secondary signalling cascade within the cell.
Are 1) G-protein coupled, 2) enzyme linked or 3) ion-channel linked
Different Types of Signal molecules
Endocrine Signals
Act over a long distance. Signals (hormones) are released by an endocrine cell into the circulatory system. Travel through the blood to the receptor cell.
Paracrine Signals
Act over short distances between the signalling and receptor cells. Often in development
Juxtacrine Signals
The signalling and receptor cells must be attached to each other.
Autocrine Signals
The same cell sends out signal and receives it
4 characteristics of signal-transducing systems
Specificity: A signal and receptor only bind to each other. Signal does not fit in any other receptors. Receptor can't bind any other signals
Amplification: The result of the signal binding is amplified exponentially throughout the signalling cascade. A small change at the cell membrane can lead to a large change within the cell, and potential for many different proteins to be affected
Desensitization/Adaptation: Is how building a tolerance works. When a signal binds to the receptor, it activates a feedback loop which endocytes the receptor from the cell membrane or shuts it off, preventing the signal cascade from starting
Integration: The cell has to deal with multiple signal cascades at the same time. Some signal cascades have opposite effects on a system, so the overall effect is the net sum of the two signalling cascades.
Signal Transduction
Is the ability of a cell to change its behavior or gene expression based on ligand-receptor binding
The primary messenger binds to a receptor on the target cell-either on the cellular membrane or within the cytoplasm or nucleus
The binding to the receptor changes the conformation of the receptor, releasing secondary messenger molecules within the cell
Receptor-Ligand Binding
Is very specific
Receptor contains specific amino acids with side chains that form specific non-covalent bond with its ligand
Resembles enzyme-substrate binding
Place where ligand binds to receptor is the "binding site"
The receptor specific for a ligand is called that ligand's "cognate receptor"
A receptor bound to a ligand is said to be occupied
Receptor Affinity
Receptor is measured using Kd. Kd is a measurement of the concentration of free ligand in the solution needed to have half of the receptors on the cell occupied
A receptor with high affinity will have a low Kd (will not take much ligand to have lots bind. A receptor with low affinity will have a high Kd.
In drug design
Synthetic drugs can be designed that bind to a receptor even more strongly than its specific ligand
Agonists bind tightly to a receptor and induce its signal-eg. Fentanyl (was originally used as a strong pain killer. Binds to the receptors for opiates.
Antagonists bind tightly to a receptor and do not set off a signal cascade. They prevent other signals from binding to the receptor. Eg. Naloxon has a Kd that is lower than fentanyl, so it binds to the opiate receptors and prevents fentanyl from binding
Activation of signal transduction events
When a ligand binds to its cognate receptor, it either changes the conformation of the receptor, or causes receptors to cluster.
This sets off a preprogrammed sequence of events within the cell
Signalling crosstalk is when signalling molecules from the pathway of one receptor have an effect on the pathway of another receptor.
One receptor could have an effect on multiple processes in the cell
Two receptors can have the same effect in the cell
Very small amounts of ligand are needed to cause a response in the cell
At each step in the signalling cascade, intermediates promote the production of lots of new molecules. This causes an amplification effect
Receptor down regulation
Receptors are programmed to sense changes in ligand concentration, rather than specific concentrations
When receptors have been bound to ligands to long periods of time, the cell adapts so that the receptors no longer respond as strongly to the ligand.
Receptor-mediated endocytosis removes receptors from the cell surface so that there are not as many available to interact with the ligand
Desensitization changes the receptor so that it does not bind as strongly to ligands. Common method is through phosphorilation
Basic types of signalling pathways
Ligand gated ion channel
A ligand binds to an ion channel, increasing or decreasing the flow of ions through it
G-protein coupled receptor (GPCR) (Plasma membrane receptor)
Receptor Kinase (plasma membrane receptor)
Nuclear receptor
eg. acetylcholine, glutamate, GABA
Regulation of glycogen metabolism
Adrenaline/epinephrine binds to G-protein coupled receptor (GPCR). This causes adenylate cyclase activity (effector molecule), which causes the production of cAMP. cAMP creates cascades for different types of kinases which affect different targets in the cell. This promotes the glycogen synthesis. Is part of "fight or flight response"
Binding of a ligand causes a conformational change in the receptor which activates a specific G protein (specific to that receptor)
A G protein is a guanine nucleotide binding protein
Part of an activated G protein binds to a specific target protein, changing its activity
opioid receptors are a type of G-protein coupled receptors
All GPCRs have a similar structure, but differences in amino acid sequence cause differences in activity.
GPCRs have 7 transmembrane alpha helices, connected by alternating extracellular or cytosolic loops
The extracellular portion of a GPCR has a specific conformation which allows for specific binding (unique messenger binding site)
the cytosolic portion of the GPCR has a part that interacts with the G-protein, as well as phosphorylation sites for protein kinase A and phosphorylation sites for G-protein coupled receptor kinases (GRKs). The phosphorylation sites are for regulation.
Large heterotrimeric G-proteins
Small monomeric G-proteins
Activation depends on the binding of GTP or GDP (GTP usually = active, GDP usually = inactive)
Mediate signal transduction through G-protein linked receptors
Have G alpha, G beta and G gamma subunits
When ligand binds to GPCR, the conformational change causes a G-protein to associate with it, releasing its GDP
The G alpha subunit then binds to a new GTP and detaches from the complex
Depending on the G protein, either the G alpha subunit or the G beta-gamma subunit (which are still attached to each other) initiate signal transduction. Can be through either activation or inactivation of target molecule
GAPs, GEFs and GDIs all regulate G protein function
The G alpha subunit hydrolyzes its GTP molecule, changing it back into GDP. It then rebinds to the G beta gamma subunit, reforming an inactive G protein.
eg. Novochok
Inhibits acetylcholinesterase, so that acetylcholine (Ach) can't be removed from the synapse, and potassium ion channels are permanently opened
This causes the cells to be flooded with potassium, and the cell is unable to function
Is an example of interaction of the effector with the G beta subunit rather than interaction with the G alpha subunit
GTP hydrolysis to cause the reassociation of G alpha subunits with G beta gamma subunits is enhanced by GTPase activating proteins (GAPs)
Regulation
GRKs (G protein linked receptor kinases) phosphorylate amino acids in the cytosolic domain of the GPCR (G-protein coupled receptor). Act on activated GPCRs.
cAMP phosphodiesterase (PDE) is activated
beta-arrestin binds to the phosphorylated active site of the GPCR, further desensitizing it to the ligand, and in some cases causing endocytosis.
Protein kinase A
Is an example of negative feedback during signalling
Is activated through the signalling cascade of adenylyl cyclase, which is mediated by g protein coupled receptors, but then phosphorylates amino acids on the GPCR, inhibiting it
Secondary Messengers
cyclic AMP
Is produced from adenylyl cyclase from cytosolic ATP when the adenylyl cyclase is activated by the binding of a G protein alpha subunit+GTP.
Posphodiesterase degrades cAMP into AMP
Activates PKA (protein kinase A) by binding to its regulatory subunits, which releases its pseudosubstrates.
Usually only remain active for a short period of time so that they can respond to changing conditions
After the G protein is inactivated, adenylyl cyclase stops producing cAMP. The remaining cAMP needs to be degraded by phosphodiesterase. Eg. cAMP is activating targets that allow you to contract your muscles. When you no longer want to contract your muscle, phosphodiesterase degrades the cAMP.
PKA phosphorylates serine and threonine residues on proteins using ATP as its phosphate source.
Is a secondary messenger signal responsible for many functions in the body. eg. glycogen degradation
Inositol-1,4,5-trisphosphate and diacylglycerol
Inositol-1,4,5-trisphosphate (IP3) and diacylglycerol are both second messengers
Are formed when the enzyme phospholipase C (specifically C beta) cleaves PIP2 into IP3 and diacylglycerol. C beta (the phosphlipase C) is activated by the binding of the alpha subunit + GTP from the G protein Gq (like the magazine)
PIP2 is a lipid membrane
IP3 moves through the cytosol and binds to the IP3 receptor channel, which is a ligand gated calcium channel.
This releases calcium ions into the cytoplasm
Both the calcium released by IP3 receptor channel and diacylglycerol activate various protein kinase As (PKAs), which phosphorylate serine and threonine residues on a variety of target proteins.
The concentration of calcium ions in the cell is regulated by calcium ATPases in the plasma membrane and ER, which transport calcium ions out of the cytoplasm
Calcium Induced Calcium Release (CIRC): IP3 receptor channels and ryanodine receptor channels are both sensitive to calcium concentration in the cell. A rapid increase of calcium ions in the cell will activate them, causing them to release more calcium ions into the cytoplasm from the ER or SER. The ryanodine receptor channel is directly sensitive to calcium ions, whereas the IP3 receptor channel's sensitivity is mediated by the binding of IP3 to it
Calmodulin mediates calcium ion activated processes in the cell. Binds four calcium ions, two in each hand.