1. Introduction to Signal Transduction
Cinderella Aquino
Cell Bio Review
Feb. 9, 2010

2. Signal transduction
Fig General features of chemical messengers.
Figure Molecular Biology of the Cell (© Garland Science 2008)
Signal transduction: Detection of specific signals at the cell surface and the mechanism by which such signals are transmitted into the cell’s interior, resulting in changes in cell behavior and/or gene expression

3. Different types of chemical signals can be received by cells
Ligand: substance that binds to a specific receptor, thereby initiating the particular event or series of events for which that receptor is responsible
Primary messenger: A molecule that binds to a receptor, thereby beginning the process of transmitting a signal to the cell
Second messenger: any of several substances, including cyclic AMP, calcium ions, inositol triphosphate, and diacylglycerol, that transmit signals from extracellular signaling ligands to the cell interior

4. The binding of extracellular signal molecules to either cell-surface or intracellular receptors
Most signal molecules are hydrophilic and are therefore unable to cross the target cell’s plasma membrane directly; instead, they bind to cell-surface receptors, which in turn generate signal inside the target cell
Some small signal molecules, by contrast, diffuse across the plasma membrane and bind to receptor proteins inside the target cell- either in the cytosol or in the nucleus.
Many of these small signal molecules are hydrophobic and nearly insoluble in aqueous solutions; they are therefore transported in the bloodstream and other extracellular fluids bound to carrier proteins, from which they dissociate before entering the target cell.
Fig Intracellular versus plasma membrane receptors. Plasma membrane receptors have extracellular binding domains. Intracellular receptors bind steroid hormones or other messengers that are able to diffuse through the plasma membrane. Their receptors may reside in the cytoplasm and translocate to the nucleus, reside in the nucleus bound to DNA, or reside in the nucleus bound to other proteins.
Figure 15-3a Molecular Biology of the Cell (© Garland Science 2008)
Figure 15-3b Molecular Biology of the Cell (© Garland Science 2008)

5. G-protein signaling
G-protein-coupled receptors
(also called Heptahelical receptors)
Act by indirectly regulating the activity of a separate plasma-membrane-bound target protein, which is generally either an enzyme or an ion channel
A trimeric GTP-binding protein mediates the interaction between the activated receptor and this target protein
Remember the whole G-protein being active when bound to GTP and inactive when bound to GDP?

6. Gs signaling http://www.youtube.com/watch?v=V_0EcUr_txk Ligand binds
A subunit loses GDP and bind GTP
Targets adenyl cyclase and produces cAMP
PKA has 4 regulatory subunits
cAMP phoshodiesterase degrades cAMP
cAMP binds regulatory and frees catalytic to go phosphorylate their targets.

7. Cyclic AMP is a second messenger used by one class of G proteins
Adenylyl cyclase (found associated with plasma membrane) forms cyclic AMP (cAMP) from cytosolic ATP
Gs signaling pathway activates adenylyl cyclase
Phosphodiesterase degrades cAMP
Phosphodiesterases cleave cAMP to AMP
Table Molecular Biology of the Cell (© Garland Science 2008)

8. Cyclic AMP is a second messenger used by one class of G proteins
cAMP-dependent kinase or protein kinase A (PKA)
PKA is a serine/threonine kinase
PKA has two regulatory subunits and two catalytic subunits. When cAMP binds to PKA, the regulatory subunits disassociates from catalytic subunits
The cAMP pathways are important for regulating glycogen metabolism, heart contraction, blood clotting and secretion of salts and water in the gut

9. Intracellular effects of the cAMP pathway: Control of glycogen degradation
Important
Epinephrine binds b-Adrenergic receptor
Epinephrine is a key hormone in the fight-flight response
Gs protein binds GTP, dissociates from other subunits
Activated Gs binds and activates adenylate cyclase
Adenylate cyclase produces cAMP from ATP
PKA binds cAMP and becomes activated
PKA phosphorylates and activates Phosphorylase Kinase
Phosphorylase Kinase phosphorylates phosphorylase b into the active phosphorylase a form
Phosphorylase a catalyzes the phosphorolytic cleavage of glycogen into glucose-1-phosphate
Meanwhile, the enzyme system responsible for glycogen synthesis is inactivated by cAMP pathway. PKA also phosphorylates the enzyme glycogen synthase and inactivates it
Epinephrine is the ligand. Has different effects in different organs.
This happens in the liver

10. Important

11. PKA Also Phosphorylates Transcription Factors
Cyclic AMP-inducible gene expression The free catalytic subunit of protein kinase A translocates to the nucleus and phosphorylates the transcription factor CREB (CRE-binding protein), leading to expression of cAMP-inducible genes.
Figure Molecular Biology of the Cell (© Garland Science 2008)
CREB: cAMP response element binding protein

12. Cholera toxin poisoning disrupts normal Gs protein signaling
Cholera toxin alters secretion of salts and fluid in the intestine, which is normally regulated by hormones that act through the Gs to alter intracellular cAMP levels. A portion of the cholera toxin is an enzyme that chemically modifies Gs, so that it can no longer hydrolyze GTP. Thus cAMP levels stay high and cells stimulate to secrete salts and water
The Details
Cholera A toxin is absorbed into mucosal cells, where it is processed and complexed with ARF (ADP-ribosylation factor), a small G-protein that is normally involved with vesicular transport.
Cholera A toxin is an NAD-glycohydrolase, which cleaves NAD and transfers the ADP ribose portion to other proteins.
It ADP-ribosylates the Gas subunit of heterotrimeric G-proteins, thereby inhibiting their GTPase activity.
As a consequence, they remain actively bound to adenylyl cyclase, resulting in increased production of cAMP.
The CFTR channel is activated, resulting in secretion of chloride ion and Na+ ion into the intestinal lumen.
The ion secretion is followed by loss of water, resulting in vomiting and watery diarrhea
Interferes with Gs signaling so that a subunit is stuck to GTP which keeps binding andenlyl cyclase, etc

13. Methylxanthines Methylxanthines: Two modes of action:
caffeine from coffee and tea
theophylline from tea
theobromine from chocolate
Two modes of action:
The principal mode of action of caffeine is as an antagonist of adenosine receptors
The caffeine molecule is structurally similar to adenosine
Methylxanthines are also cAMP phosphodiesterase inhibitors
Images from
Binds to adenosine receptors. This makes it an antagonist of adenosine.
cAMP stay elevated bc can’t be broken down.

14. Gi Signaling
Gi signaling can inhibit Adenylyl cyclase activity by the Ga subunit
Gi bg subunits act mainly by regulating ion channels
For example stimulation of muscarinic acetylcholine receptors by acetylcholine released by the vagus nerve, results in the bg subunits binding to K+ channels in the heart muscle plasma membrane and opening them.
Whooping cough (pertussis toxin)
The inhibitory protein Gi is inactivated by pertussis toxin, Gi can no longer inhibit adenylyl cyclases

15. Gq Signaling
Many G proteins use inositol triphosphate and diglycerol as second messengers
Phospholipase C cleaves PIP2 into inositol triphosphate and DAG
The inositol-phospholipid-calcium pathway
Ligand binds to receptor
Gq is activated
Gq then activates phospholipase Cb
PIP2 is cleaved into InsP3 and DAG
InsP3 binds to InsP3 receptor (ligand-gated calcium channel) in ER
Calcium is released into cytosol and binds calmodulin and modulates other pathways
Meanwhile DAG activates protein kinase C (PKC)
PKC then stimulates other pathways

16. Gq Signaling Pathway
Important

18. The calcium-calmodulin complex
In the cytosol, calcium binds to proteins like calmodulin.
Calmodulin is one of the most important calcium-binding proteins in the cell and can constitute as much as 1% of the total protein mass.
When calmodulin is activated after binding calcium, it targets and regulates kinases and phosphatases

19. Nitric Oxide couples G protein-linked receptor stimulation in endothelial cells to relaxation of smooth muscle cells in blood vessels
Nitric Oxide (NO) produced by NO synthase, which converts amino acid arginine to NO and citrulline
NO stimulates guanylyl cyclase to make cGMP
Has short half life so can only act locally
NO can diffuse out of the cells and affect smooth muscle.
Figure 15-12b Molecular Biology of the Cell (© Garland Science 2008)

20. Important

21. NO, erectile dysfunction and angina pectoris
Nitroglycerin for angina to release constricted coronary arteries.
Glycerol trinitrate decomposes to NO, which activates a guanylyl cyclase.
This will relax arterial smooth muscle cells
Viagra (sildenafil): an inhibitor of cGMP-specific phosphodiesterase (breakdown of cGMP)
Nitric Oxide released by the neurons in the penis results in the blood vessel dilation responsible for penile erection.
Viagra helps maintain elevated cGMP in erectile tissue, this pathway is stimulated for a longer time period following NO release.
Since cGMP can’t be broken down, cGMP levels stay high, so dilation is sustained so erection remains.

22. Prostaglandins: typically activate G protein receptor
Prostaglandins such as thromboxane A (TXA2) can stimulate Gq signaling pathways
Prostaglandins are synthesized from arachidonic acid.
Cyclooxygenases are key enzyme in the pathway
Aspirin blocks cyclooxygenases, thereby inhibiting synthesis of thromboxane A2 and other prostaglandins form arachidonic acid
Aspirin significantly reduces incidence of heart attacks by reducing blood clots

23. Key points Signal transduction is important!
3 major G protein pathways: Gs, Gi, Gq
Which is involved with cAMP production and glycogen breakdown?
Which is involved with Viagra?
Which is affected by pertussis toxin? Cholera? Caffeine?
Which involves the cleavage of PIP2? What molecules does PIP2 break down to?
Which causes release of calcium?
What causes the activation of PKA? PKC? PKG? What do those do? Huh?