• Glucocorticoids are examples of steroid hormones.  
• There receptors lie in the cytoplasm of the cell. 
• The hormone diffuses through the plasma membrane and binds the receptor. 
•  The hormone receptor complex enters into the nucleus and binds the hormone response element to upregulate (and in some cases downregulate trx)

A patient has external female phenotype, but also has testes.  Androgens are released by the testes but the tissue does not respond, giving allowing for the female phenotype.  What is the probable reason for this?

This is testicular feminization.  Androgens are examples of steroid hormones.  Because we know that the testes are releasing the androgens the most probable cause for the female phenotype is that the adrenergic receptor is defected.

A deficiency of a receptor will make the cell unable to respond to the hormone.

Describe the 3 subunits of the G protein

α= can bind GTP or GDP

β&γ = subunits are tightly bound to eachother and can act as a single unit

Note: the alpha subunit is only loosely associated with the beta and gamma subunits

Describe the action of a G protein

The G protein is associated with an unstimulated receptor and the alpha subunit is bound to GDP. Once the a water soluble hormone binds the receptor, then the G protein undergoes a conformational change.  This causes the affinity of the alpha subunit for GDP to decrease, and the GDP is replaced with a GTP.  Once GTP is bound then the alpha subunit looses its affinity for the receptor and βγ subunit.

Both products alpha-GTP and βγ can bind effector proteins in the plasma membrane.  They affect these proteins by allosteric mechanisms.

How is the G Protein signal terminated

When the alpha-GTP binds the effector protein, then the GTPase activity of the alpha subunit is stimulated.  The GTP is hydrolyzed to GDP and and the alpha-GDP complex no longer acts on the effector but now goes back to bind with the βγ subunit, forming in the inactive G protein again.

Steroid hormones are lipid soluble which means that they can diffuse through a membrane.  Their receptors lie inside the cytoplasm or the nucleus.  (thyroid hormones, calcitonin, and retinoic acid also act like steroid proteins)

Water soluble proteins cannot diffuse through the membrane, thus their receptors must be located extracellularly.

[image]cAMP is formed when a stimulus binds an extracellular receptor that is coupled to a G-protein. Alpha subunit of G protein associates with the membrane protein, adenylate cyclase which works to produce cAMP.

cAMP then diffuses through the cytoplasm and binds Phosphokinase A.  Phosphokinase A then goes into the nucleus and binds the threonine and serine residues of the CREB protein phosphorylating it.. the activated CREB protein can then bind response elements and influence transcription.

Phosphodiesterase breaks down cAMP

Coffee works to inhibit phosphodiesterase

Hormone

receptor

G protein

Adenyl Cyclase

cAMP

Protein Kinase A

nucleus

CREB

response element

transcirption

translation

new protein

For example, the cholera toxin causes diarrhea because of the high amounts of cAMP that are maintained in the cell.  This causes the cell to release lots of water and electrolytes into the intestine which results in diarrhea.  Endorphins work to inhibit adenylate cyclase.  Thus, morphine works to activate more endorphin receptors in the cell, to bring down the cAMP concentration.

Describe what part of the G protein is affected by the following:

a. Cholera toxin

b. pertussis toxin

a. cholera toxin causes the G protein to loose its GTPase activity

b. pertussis toxin causes the inhibitory G protein to be dysfunctional, which means that their is no inhibition of adenylate cyclase

Although the G protein is affected differently for both of these, they both result in the same problem = over production of cAMP = diarrhea

There could be a problem in the PTH receptor in the cell.

There could also be aproblem with the G protein coupling to adenylate cyclase, which will cause no cAMP to form.

1. cholera toxin

2. toxic thyroid nodules

TSH normally produces hormones in the thyroid gland.  

In the case of toxic thyroid nodules, there is over production of the hormones regardless of TSH, resulting in benign tumors on the thyroid gland.

In some cases this is due to a somatic mutation in the TSH receptor.

In other cases it has to do with the G protein, where it is constitutively on do to a defect in the GTPase activity.

If there is a defect in the G protein than any time a receptor activates the G protein, that G protein won't work .

Ex: pseudoparathyroidism, where the G protein is compromised due to its GTPase activity.  This results in other problems as well because of the G protein problems.

(Nor)Epinephrine can bind different receptors.

alpha1: raises IP3 and calcium levels causing contraction

alpha2: reduces cAMP levels causing contraction

beta: raises cAMP levels causing  of relaxation

Where is phospholipase C located? and how is it stimulated?

Phospholipase C is located in the membrane, 

A calcium stimulating hormone binds a receptor G protein and the alpha subunit, it then stimulates phospholipase C

Calcium elevating hormone binds cell receptor

G protein is activated

Alpha subunit then binds phospholipase C

formation of 2 molecules: IP3 and DAG (diacylglycerol)

IP3 then binds endoplasmic reticulum to have release of Ca 2+

The presence of Ca2+ then stimulates DAG to stimulate Protein kinase C (PKC) to phosphorylate Ser & Thre residues 

IP3

DAG

Diacylglycerol (DAG) stimulates the production of Protein Kinase C (PKC) in the presence of CA2+

1. Hormones acting through a G protein and IP3 system

2. Calcium ligand gates Ion channel (ex. NMDA receptor in brain)

3. Voltage gated calcium channels (i.e. an extracellular stimulus depolarizes the cell)

1. Protein Kinase C (PKC)

2. Troponin C

3. Calmodulin

1. 

Nitrovasodilator drugs metabolize into nitrous oxide (NO) which is a smoothe muscle relaxer, thus it allows for blood vessels to relax and dilate decreasing blood pressure

Ex: Viagra

inhibit cGMP-specific phosphodieseterases in the corpora cavernosa.  

This allows for cGMP to stay active in the cell longer.

A stimulus binds a receptor on the extracellular membrane.  The receptor is the extracellular portion of Guanylate cyclase.  Upon binding, GTP is converted to cGMP.

NO G-PROTEIN INVOLVED!

ANF= Atrial Natriuretic Factor

The ANF receptor is a ligand activated guanylate cyclase

note: ANF is made by the heart atrium in response to high blood pressure, high blood volume, and high salt.  ANF acts on the kidney to increase Na+ secretion.  Acts on vascular blood vessels to relax vascular smooth muscle.

Describe the difference between:

1. Protein Kinase A (PKA)

2. Protein Kinase C (PKC)

3. Protein Kinase G (PKG)

1. PKA= activated by cAMP in response to a G protein acting on adenylate cyclase

2. PKC= activated by an increase in intracellular Ca2+ and DAG.  

3. PKG= activated by cGMP

ANF is produced in the atrium

It is release in response to high: blood pressure, blood volume, and salt

ANF binds ANF receptor

ANF receptor is a ligand activated guanylate cyclase.

Upon binding to the receptor, GTP is made into cGMP

cGMP binds PKG, which then works to phosphorylate proteins

In endothelial cells: ACH, bradykinin, and histamine work to increase intracellular Ca2+to cause NO formation.

NO diffuses into smooth muscle and activates guanylate cyclase, which causes formation of cGMP.  

cGMP works to relax smooth muscle by dreasing TPR and decreasing BP.

Note: By increasing intracellular Ca+ concentration this process can be reversed and cause contraction of smooth muscle.

For an erection to occur NO diffuses in and causes relaxation of the smooth muscle via the cGMP cascade.

Viagra works to inhibit the cGMP phosphodiesterase, to allow NO to linger longer to maintain the erection.

Protein Kinase A, C, and G phosphorlyate which type of residue?

Which residue does the kinase for insulin and other growth factors phosphorlate?

PKA, PKC, and PKG all phosphorylate Ser and Thr residues.

the kinase for insulin and other growth factors phosphorylate tyrosine residues.

Autophosphorylation is a process typical of insulin and other growth factor receptors.

Upon signal from a stimulus, this causes membrane proteins to group together, become allosterically activated and phosphorylate eachother on tyrosine residues.

Growth factors:

Stimulus binds receptor (monomer in unstimulated state)

Receptor undergoes oligomerization

Receptor groups and autophosphorylation occurs on tyr residue

Any signaling protein with an SH2 domain can bind the autophosphorylated receptor

The signaling proteins are then allosterically activated or also autophosphorylated on their tyr residues.

Insulin: (tetramer in unstimulated state)

Very similar mechanism, however, autophosphorylation of the insulin receptor  does not allow for SH domain binding proteins to bind.  Instead and IRS comes in and binds and it is autophosphorylated.  SH domain docking proteins can now bind IRS.

Growth factors are able to stimulate the IP3 system to proceed with a signaling cascade.

Growth factor binds receptor

NO G-PROTEIN INVOLVED because participates in autophosphorylation, which stimulates PLC-gamma

PIP2 is formed into IP3 which will increase intracellular CA2+ levels

Growth hormone binds the receptor

transmembrane proteins gather together 

Autophosphorylation of the tyrosine residues

causes PI3Kinase to catalyze the conversion of

PIP2->PIP3 which in turn activates PKB (AKT)

Inhibition of PKB (AKT) would be a great approach to an anti-cancer drug, as it would stop the cascade.

1. Calcium stimulator binds receptor, G protein stimulates PLC-beta which forms IP3 and DAG. IP3 binds endoplasmic reticulum to release Ca2+

2. Growth factor binds receptor, autophosphorlation stimulates PLC-gamma which converts PIP2 to IP3 ...

Growth factor binds the receptor

Autophosphorylation on tyrosine residue

GRB2 (which is an SH2 domain binding protein) binds the inner receptor

SOS is then recruited by GRB2 and binds

SOS then binds RAS (G protein)

which signals for phosphorylation cascade

which starts the MAP kinase cascade

1. BARK and/or PKA (negative feedback loop)

will phosphorylate the intracellular domain of the stimulated receptor on Ser or Thr residues which will still allow binding of the stimulus to the receptor but will not allow for any intracellular modifications.

2. Receptor mediated endocytosis

ligand stimulated receptors aggregate on cell surface and are internalized in an endocytotic vesicle.  This vesicle will fuse with a lysosome where it will be degraded

1. Desensitization caused by BARK or PKA can be reversed via dephosphorylation using phosphatases

2. loss of receptors by endocytosis can only be remedied by the production of new receptors