4. Hormone Signaling Pathways

Question 1

Hormones are __________ molecules made and secreted by _______ cells.

• Released into blood.
• Bind to specific receptors
• Result in: __________.

Answer 1

Hormones are MESSENGER molecules made and secreted by ENDOCRINE cells.

• Released into blood.
• Bind to specific receptors
• Result in: SIGNAL TRANSDUCTION, ALTER GENE EXPRESSION, CAUSING A. CELL-TYPE SPECIFIC RESPONSE.

Question 2

We only need a small amount of hormone to have an effect. Why?

Answer 2

Their effect is magnified by amplification!

Question 3

What are the steps in hormone signaling? (9)

Answer 3

1. Biosynthesis
2. Storage
3. Secretion
4. Transport to target tissue/cell
5. Recognize and bind to R
6. Activation of signal transduction path (on switch)
7. Amplification
8. Cellular response
9. Degradation (off switch)

Question 4

How is the signal terminated?

Answer 4

1. Removal of a signaling molecule/receptor
2. Attenuation/inactivation of signaling events.

Question 5

What are the general signaling steps?

Answer 5

1. Signaling cell makes and secretes a signaling molecule (ligand) d/t a stimulus.
2. Signaling molecule (ligand) binds to receptor on target cell.
3. Ligand + receptor activates or inhibits cellular pathway to cause a response.
4. Signal terminated.

Question 6

Describe the last type of signaling:

1. Endocrine signaling- signaling molecule is released to a cell far away via blood. Ex. EPI
2. Paracrine signaling- signaling molecule is released by one cell type and binds to neighboring cell of a different type. Ex. Testosterone
3. Autocrine signaling- signaling molecule acts on the same cell type as the secreting cell. Ex. IL-1
   4.

Answer 6

Juxtacrine signaling- signaling molecule stays attached to the secreted cell and binds to a receptor on an adjacent target cell. Ex. Heparin-binding epidermal GF.

Question 7

Do signaling molecules only participate in ONE type of signaling?

Answer 7

NO. Some signaling molecules can participate in more than 1 type of signaling.

Question 8

Hydrophilic hormones (water soluble) cannot pass the phospholipid bilayer, and thus need receptors on the cell surface. When it binds to the receptor, it does what?

Answer 8

initiates production of second messenger in the cell to cause a cell response.

Question 9

Hydrophobic hormones (such as the steroid hormones), by contrast, can pass directly into the cell and have an effect there. This receptor could either be cytosolic – forming a complex that translocates to the nucleus, or already in the nucleus. Both act like a what?

Answer 9

A transcription factor, regulating transcription.

Question 10

Are cytoplasmic receptors always active?

Answer 10

No. They exist as an inactive complex with HSP 90. When ligand binds, HSP 90 dissociates. The ligand+ receptor -> nucleus and binds to DNA sequence called the hormone response element (HRE) in the promotor.

Question 11

Examples of hydrophilic hormones include:

Answer 11

1. AA derived (melatonin, DA, NE, EPI, Histamine, 5HT) ,
2. Those from lipid metabolism (ACh)
3. Polpeptides such as insulin, glucagon, TSH

Question 12

Examples of hydrophobic hormones include:

Answer 12

1. steroid hormones (include cortisol, aldosterone and vitamin D),
2. thyroid hormones (thyroxine)
3. retinoids.

Question 13

What is the difference between hydrophilic medications and lipophilic medications?

Answer 13

• Hydrophilic medications have short half-lives that range from seconds- minutes. Examples are EPI and insulin.
• Lipophilic medications have long-half lives, from hours- days. Examples are oral contraceptives.

Question 14

Describe the activation/inactivation cycle of trimeric G-protein.

Answer 14

GCPR is a trimeric G-protein with 3 subunits (a, b, y)

Activation:

• Inactive G protein has GDP bound to alpha subunit. To become active, it exchanges GDP -> GTP via GEF (guanine nucleotide exchange factor). Once active, GTP-bound alpha subunit separates from b and y subunits.

Inactivation:

• Intrinsic GTPase activity of G-protein hydrolyzes GTP-> GDP and Pi. GAP (GTPase activating protein) accelerates this activity.

Question 15

What are the 4 types of G-proteins and what is their mechanism of action?

Answer 15

• Gs: ATP -> cAMP via adenylate cyclase (AC)
  
  • cAMP -> activates PKA
  • PKA phosphorylates protein
• Gt: light activates cGMP PDE to convert cGMP-> 5’GMP
• Gq: signal activates PLC to activate PIP2 -> DAG and IP3.
  
  • IP3 will cause the release of Ca2+ from ER/SR.
  • Ca and DAG activate PKC-> phosphorylates
  • Ca+ also forms a complex with calmodulin (ca2+-calmodulin complex) which activates proteins.
• Gi: cAMP and PKA are not activated.

Question 16

Gs examples:

Answer 16

• EPI is a non-selective agonist of all adrenergic receptors and can undergo multiple GCPR pathways. It can bind to B-adrenergic receptor
• Histamine binds to H2 receptors and causes bronchoconstriction and sx of allergic reactions

Question 17

Gi examples:

Answer 17

• EPI/NE bind to a-adrenergic receptor and cause constriction of smooth muscle.
• DA binds to D2 receptors and causes increase HR.

Question 18

Gq examples:

Answer 18

• ACh binds to M3 receptors and causes bronchoconstriction and stimulation of salivary glands

Question 19

Gt examples:

Answer 19

Light binds to Gt receptor and influences vision.

Question 20

What is the structure of a RTK?

Answer 20

• Extracellular domain that binds ligand
• Single alpha helical transmembrane domain
• Intracellular domain with TK activity.

Question 21

What are the steps of RTK activity?

Answer 21

1. Ligand binds to extracellular domain
2. Dimerization
3. Dimerized receptor phosphorylates tyrosine residues
4. Adaptor and docking proteins recognize the phosphotyrosines
5. Activates downstream signaling: RAS dependent (MAPK) and independent.
6. Both trigger phosphorylation of a specific protein in the cytoplasm/nucleus -> alter gene transcription and protein activity.
7. Signal terminated by
   
   • Degradation of ligand by extracellular proteases
   • Ligand-induced endocytosis of receptor and degradation
   • RAS inactivation
   • Dephosphorylation

Question 22

How is inactive insulin stored in the body?

Answer 22

Hexamer.

Active form is a monomer.

Question 23

How is insulin made?

Answer 23

1. Begins as preproinsulin mRNA
2. Translated into preproinsulin protein
3. Translated to ER lumen where it is cleaved by a protease to make proinsulin
4. Folded and goes to Golgi.
5. Packaged into granules
6. Cleaved by proteases to make [insulin and C peptide]
7. Released together from beta cells.

Question 24

In the blood we see a rise in glucose, this causes insulin to be secreted.

How is it secreted?

Answer 24

Insulin is released in two phases:

• a first rapid but transient phase from our readily releasable pool (5%)
• a second sustained phase from our reserve pool, where granules have to undergo mobilization before they can be released.

Question 25

How do we release insulin from the pancreatic B cell?

Answer 25

1. Glucose is going to bind to GLUT2 R on the pancreatic B cell and move in via faciltated diffusion
2. Glucose goes inside and Glu–> G6P via glucokinase.
3. G6P will undergo glycolysis, TCA cycle and oxidation–> ATP
4. ATP will close KATP channels
5. Cell depolarizes
6. Depolarization reaches the VGCa2+ channels and allows Ca2+ influx
7. Triggers exocytosis of secretory vesicle
8. insulin and C peptide are released

Question 26

What is the target of sulfonylurea drugs to treat type 2 non-insulin dependent DB?

Answer 26

KATP channels; increase insulin secretion

Question 27

Describe RAS-dependent insulin signaling.

Answer 27

1. Insulin binds to insulin receptor, a RTK, which is already dimerized.
2. Tyrosine residues are autophosphorylated
3. IRS-1 (insulin receptor substrate-1) binds to phosphorylated receptor tyrosine kinase
4. Insulin receptor phosphorylates IRS-1 on its tyrosine.
5. IRS-1 then recruits an adapter protein called GRB-2, which activates the RAS and MAP kinase pathway.
6. Multiple proteins are phosphorylated which leads gene transcription – specifically ↑ transcription of glucokinase.
7. This results in increased glucose uptake and glycogen synthesis.

Question 28

Describe RAS-independent insulin signaling:

Answer 28

1. Insulin binds to insulin receptor, a RTK, which is already dimerized.
2. Tyrosine residues are autophosphorylated
3. IRS-1 (insulin receptor substrate-1) binds to phosphorylated receptor tyrosine kinase
4. Insulin receptor phosphorylates IRS-1 on its tyrosine.
5. IRS-1 then recruits an adapter protein called PI 3-kinase (phosphoionositide-3-kinase).
6. P1 3-kinase phosphorylated phosphoinositides to make PIP2 and PIP3.
7. PIP2 and PIP3 act as second messengers and recruit PKB and activate it via phosphorylation.
8. RESULT: ALTER PROTEIN AND ENZYME ACTIVITY
9. 1. increase in GLUT4’s movement to the plasma membrane,
   2. • glycogen synthase.
10. Both result in an increase in glucose uptake and glycogen synthesis.

Question 29

How can we terminate insulin signaling?

Answer 29

1. Insulin receptor complex is internalized in target cells via endocytosis. There it is degraded by proteases or recycled back into membrane.
2. Negative feedback
3. Insulin can increase rate of degradation

Question 30

How can we test insulin resistance?

Answer 30

Measure the amount of glucose cleared from blood in response to a fixed dose of insulin. If insulin does not cause the expected response -> insulin resistance.

Question 31

Why does insulin resistance occur?

Answer 31

NO CLUE. Possible explanations:

1. Downregulation of insulin receptor
2. Defects in the insulin receptor (mutation)
3. Defects in insulin binding
4. Defects in IRS1 and IRS2
5. Phophorylation of serine/threonine kinase instead of tyrosine of IRS, which inhibits its activation and prevents activation of PI-3 kinase.
6. Serine threonine kinase is activated by cytokines, FFA, DAG, ceramind, inflammatory molcules.

Question 32

Nuclear Receptors include _________ receptors and _______ receptors that bind lipophilic hormones such as __________, __________, _______, _______ and _________. They are receptors and effectors for the signal.

Answer 32

Nuclear Receptors include nuclear receptors and orphan receptors that bind lipophilic hormones such as glucocorticoids, mineralocorticoids, estrogen, progesterone and androgens. They are receptors and effectors for the signal.

Question 33

Where are nuclear receptors located?

Answer 33

Nucleus or cytosol.

Question 34

What are NR important for?

Answer 34

DRUG TARGETS.

Question 35

What is the architecture NRs?

Answer 35

1. LGB (ligand binding domain)
2. AF1 domain (activation function 1 domain) aka transcription activating domain
3. DBD (DNA binding domain)

Question 36

What happens when a ligand binds to LBD?

Answer 36

When it binds, the LBD undergoes conformational change and recruits and binds coactivators and corepressors that regulate transcription.

Question 37

What does AF-I do?

Answer 37

modifies the conformation of the entire receptor

Question 38

What does DBD do?

Answer 38

highly conserved and binds to HRE (hormone response element) in the nucleus.

Question 39

How do nuclear receptors change before and after binding of a ligand?

Answer 39

• Before the ligand binds, the nuclear receptor is surrounded by inhibitory proteins.
• After the ligand binds, the inhibitory proteins dissociate, the activation function domain (AF 1 / “transcription activating domain”) changes confirmation, and co-activator proteins bind.
• The nuclear receptor complex then binds to the receptor binding element of the HRE and alters gene expression.

Question 40

Describe the primary and secondary responses to steroid hormones.

Answer 40

• In the primary (early) response, primary response genes are activated by the steroid hormone receptor complex, and synthesis of primary response proteins is induced.
• In the secondary (delayed) response, the primary response proteins shut off the primary response genes if they persist long enough and activate the secondary response genes to transcribe the secondary response proteins.
• Both the primary and secondary response proteins can have an effect – testosterone, for example, increases spermatogenesis in the short-term, but increases to form secondary sexual characteristics as a secondary response long-term.

Question 41

What are the two types of estrogen receptors?

Answer 41

ER-alpha and ER-beta, which are both estrogen-dependent transcription factors.

Question 42

Where are both ER-alpha and ER-beta located?

Answer 42

• ER-alpha -> female reproductive tract (uterus, vagina and ovaries) and mammary glands, hypothalamus, endothelial cells and vascular smooth muscle.
• ER-beta -> ovaries and prostate with lower expression in the lung, brain, bone and vasculatire.

Question 43

Describe the binding onto estrogen receptors.

Answer 43

• The estrogen receptor is already present in the nucleus (however, there is some evidence that ER is located in the cytosol and binding causes dimerization and translocation -> nucleus).
• When estrogen binds to the estrogen receptor, it dimerizes.
• This allows it to bind to the estrogen response element (ERE) on DNA and recruit co-activators that modify chromatin structure via histone acetyltransferase (HAT).
• Loosens up chromatin and activates transcription.
• Also, it recruits proteins that make up the GTA (general transcription apparatus), causing enhanced transcription to form mRNA.

Question 44

Describe the importance of tamoxifen.

Answer 44

Tamoxifen is a estrogen receptor ANT.

Tamoxifen, which is used to treat breast cancer, is metabolized by cytochrome P450 to 4-hydroxy-tamoxifen, which promotes histone deacetylase activity (HDAC1), which tightens chromatin and prevents interaction with GTA, thus, preventing transcription of cancerous cells.