CARBOHYDRATE BIOSYNTHESIS April 14, 2015 BC368 Biochemistry of the Cell II.
Signaling Chapter 12, pp. 433-460, 471-472 February 19, 2015 BC368 Biochemistry of the Cell II.
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Transcript of Signaling Chapter 12, pp. 433-460, 471-472 February 19, 2015 BC368 Biochemistry of the Cell II.
Signaling Chapter 12, pp. 433-460, 471-472
February 19, 2015
BC368Biochemistry of the Cell II
Signal transduction
Binding of acetylcholine to its receptor opens an ion channel.
Signal = acetylcholine Response= ions flow
Signal transduction
Binding of insulin to its receptor results in recruitment of GLUT4 in certain cell types.
Signal = insulin Response= increased glucose transport
Signal transduction
Binding of insulin to its receptor results in recruitment of GLUT4 in certain cell types.
Signal = insulin Response= increased glucose transport
Here’s why you shouldn’t trust everything on the web!
Signal transduction
Amplification
FeedbackRegulation
Types of signals
Intercellular signaling
Intercellular signaling can occur over long distances (endocrine) or short distances.
hormone
neurotransmitter some growth factors
some cytokines
Properties of signal transduction
Fig. 12-1
Properties of signal transduction
Fig. 12-1
Properties of signal transduction
Fig. 12-1
Properties of signal transduction
Fig. 12-1
Properties of signal transduction
Fig. 12-1
In other words, signaling proteins can interact with more than one target, forming complexes with different properties.
Types of chemical signals
Cells communicate with each other by sending out signaling molecules.
In order to respond to the signal, target cell must have a receptor.
Mechanism of signal transduction depends on the chemistry of the transmitter.
Interfering with chemical signals
Some molecules interfere with the normal signaling pathway.
Interfering with chemical signals
Some molecules interfere with the normal signaling pathway.
Agonists bind to the receptor and mimic the effects of the normal signal.
Interfering with chemical signals
Some molecules interfere with the normal signaling pathway.
Agonists bind to the receptor and mimic the effects of the normal signal.
Antagonists act as competitive inhibitors of the normal signal.
Types of signal transducers
✓
Nuclear receptors
Nonpolar signal molecules can pass through the plasma membrane.
Steroid hormones
Thyroidhormones
Nuclear receptors
Hormone-receptor complex acts in the nucleus to affect gene expression.
Nonpolar signal molecules can pass through the plasma membrane.
Receptor can be in cytosol or nucleus.
Nuclear receptors
Females who produce excess testosterone can have hyperandrogenism.
Problems with steroid signaling
Caster Semenya2009 World 800-Meter
Champion
Individuals who have a defective testosterone receptor have androgen insensitivity.
Jazz singer Eden AtwoodX,Y Genotype
Their use in sports was pioneered by East Germany, which had a systematic governmental doping program from 1965 to 1990.
Anabolic steroids are testosterone agonists used to build skeletal muscle and stimulate bone growth.
Anabolic Steroids
At the first world swimming championships, in 1973, East German women won 10 of the 14 gold medals available, setting eight world records.
Three years later at the Montreal Summer Olympics, the East German women won 11 of 13 events.
llona SlupianekShot Put World Record Holder
1980 –1984
Anabolic Steroids
Types of signal transducers
✓
✓
G-protein coupled receptors
GS-protein pathway: β-adrenergic receptor
Animation
A “GTP Switch” protein is active when GTP is bound; inactive when GDP is bound.When α subunit binds GTP, it separates from β and γ.
Activated α subunit finds AC and turns it on.
Intrinsic GTPase activity turns α subunit off- it finds β and γ.
GS-protein activation
Activation of AC
Activated AC makes cAMP.
One target of cAMP is protein kinase A (PKA).
Activation of Protein Kinase A
PKA is active only when 4 cAMP are bound, freeing the two catalytic subunits.
Active PKA has many effects, depending on cell type.
Activation of Protein Kinase A
cAMP: second messenger
Overview
G-protein summary
Inhibitory G proteins
Case Study
Peter T., a teacher aged 42, had just returned from Calcutta. Within 24 h of his arrival, he had suddenly developed diarrhea. It was not associated with pain, and as it was watery and voluminous, P.T. took plenty of tea to make up the body fluid. As his condition worsened, with fluid losses up to 1 L/h and with vomiting and muscle cramps, he called his doctor.
On examination, P.T. was apathetic, his cheeks were hollow, and his eyes sunken; no peripheral pulse was palpable, he was cyanotic and in a state of profound shock. He had tachycardia and his respiration was rapid and shallow. The diarrheal fluid was colorless and turbid, like “rice water.”
An isotonic solution containing Na+, K+, Cl-, and HCO3- was infused
intravenously at a rate of 100 mL/min until a strong pulse was restored and thereafter in quantities sufficient to maintain normal pulse, blood pressure, and skin turgor. When the patient stopped vomiting, a solution of similar composition but containing 2% glucose was given by mouth.
Microscopic examination of the diarrheal fluid established the presence of masses of typical short, comma-shaped rods of Vibrio cholerae. A course of antibiotic therapy was started.
After 24 h the fluid loss began to decline and the patient made a rapid and complete recovery. He was estimated to have lost about 20 L of fluid containing 2.8 mol NaCl.
Toxins that Target G proteins
•Vibrio cholerae produces the cholera toxin, which ADP-ribosylates GSα, inhibiting GTPase activity.
Box 12-2
Net result: cAMP is high
Toxins that Target G proteins
•Bordetella pertussis produces the pertussis toxin, which ADP-ribosylates GIα, inhibiting nucleotide exchange.
GI
GI
Pertussistoxin
GI is kept off, so AC is on and cAMP levels increase
Net result: cAMP is high
Drugs that Enhance cAMP
•Caffeine and theophylline inhibit cAMP phosphodiesterase.
cAMPphosphodiesterase
Net result: cAMP is high
Termination of Response
Removal of signal
Desensitization of receptor
Hydrolysis of GTP (promoted by GTPase activator proteins [GAPS])
Degradation of 2nd messenger
Hydrolysis of phosphates
Pathway can be terminated at any step!
Mechanisms of DesensitizationFig. 12-8
Strategy #1: Receptor Level
Strategy #2: Downstream Effects
Morphine receptor works through GI.
Body responds to morphine by increasing AC and PKA expression.
Mechanisms of Desensitization
Gq activates phospholipase C
DAG and IP3
DAGIP3
DAG and IP3
DAG and IP3
Gq acts through DAG and IP3
~Fig. 12-10
DAG/IP3 as Second Messengers
http://www.youtube.com/watch?v=2bbBrpgeheY
Ca2+ as Second Messenger
Variable Ca 2+
increases depending on the amount of IP3
“Because of the calcium ion’s diverse roles in cell function, it has plagued investigators from many biological disciplines for the last 30 years. Although its steady state concentration in the cytosol ranges from only 0.01 – 0.1 μM, the calcium ion triggers such diverse phenomena as lipid and glycogen degradation, the release of neurotransmitters, muscle contraction, and cell division.
How does calcium perform its varied cellular functions when it exists only in such minute quantities? It was first suggested in 1964 that the calcium ion itself is inactive; it must form a complex with one of a homologous class of calcium-binding proteins.”
-Julie T. MillardChemistry Honors Thesis
Amherst College, 1984
Ca2+ as Second Messenger
~Fig. 12-11 Ca2+ as Second Messenger
Calmodulin Animation
Types of signal transducers
Receptor Tyrosine kinases
Receptors for many growth factors, cytokines, and hormones.
Binding of signal initiates a kinase cascade, beginning with autophosphorylation.
Receptor Tyrosine kinases
Activated receptor is a dimer.
Monomer that dimerizes (e.g., epidermal growth factor receptor)
Starts out as dimer (e.g., insulin receptor)
Receptor Tyrosine kinases
Ligand binding causes activation of the dimer (turns on kinase activity).
Autophosphorylation results
Receptor Tyrosine Kinases #1: Growth Factor Receptor
Epidermal growth factor stimulates cell growth, proliferation, and differentiation by binding to its receptor, which is an RTK.
This pathway is an excellent example of signal amplication.
Receptor Tyrosine Kinases #1: Growth Factor Receptor
Growth Factor Signaling
Receptor Tyrosine Kinases #1: Growth Factor Receptor
Epidermal growth factor stimulates cell growth, proliferation, and differentiation by binding to its receptor.
Signal binding to the receptor leads to a kinase cascade.
http://www.youtube.com/watch?v=OvvXgzf58MQ
Receptor Tyrosine Kinase #1: Epidermal Growth Factor Receptor
Ligand binding leads to receptor dimerization, autophosphorylation, and recruitment of adapter molecules:
GRB2 Sos
Binding of adapter molecules to receptor recruits and activates Ras (GTP switch).
•Activated Ras recruits Raf.
•Binding to Ras activates Raf.
•Raf phosphorylates MEK.
•MEK phosphorylates MAP kinase (aka ERK).
•MAP kinase/ERK phosphorylates its targets.
Receptor Tyrosine Kinase #1: Epidermal Growth Factor Receptor
Receptor Tyrosine Kinase (RTK)/Ras GTPase/MAP kinase (MAPK) signaling pathway
Raf
MEK
ERKAnimation
Receptor Tyrosine Kinase #1: Epidermal Growth Factor Receptor
•Activated ERK has many substrates in the cytosol [e.g. cytoskeletal proteins, phospholipase A2, signalling proteins, and activation of transcription proteins (STATs).
•ERK can also enter the nucleus to control gene expression by phosphorylating transcription factors such as Elk-1, growth-factor- receptor-binding protein 2, and SRF, serum response factor.
Regulation of GTP-Switch Proteins
Guanine nucleotide Exchange Factor
GTPase Activating ProteinRegulator of G-protein Signaling
Ras: a GTP-Switch Protein
https://www.youtube.com/watch?v=NL3ndoSzFo4
RTK Example 2: Insulin Receptor
•Insulin receptor is already a dimer
•Insulin binding triggers conformational change and autophosphorylation of Tyr residues in the cytosolic region
•Receptor then binds and phosphorylates target proteins
RTK Example 2: Insulin ReceptorFig. 12-15
•Activated IRS-1 recruits Grb2. •Grb2 binds Sos.
•Sos binds Ras, leading to nucleotide exchange.
•Ras activates Raf-1.
•Raf-1 phosphorylates MEK.
•MEK phosphorylates ERK/ MAPK, which phosphorylates transcription factors.
•One target is IRS-1.
RTK Example 2: Insulin ReceptorFig. 12-16
•Insulin signaling has many effects. Here’s another one.
Erythropoietin (EPO) is a peptide hormone that stimulates red blood cell production in the bone marrow.
RTK Example 3: EPO Receptor
An estimated 70% of professional cyclists in Europe used EPO in the mid 1990s.
EPO receptor has no intrinsic protein kinase activity, but recruits a tyrosine kinase (JAK).
RTK Example 3: EPO Receptor
EPO receptor has no intrinsic protein kinase activity, but recruits a tyrosine kinase (JAK).JAK phosphorylates STAT, which dimerizes, goes to nucleus, and affects gene expression.
RTK Example 3: EPO Receptor
EPO receptor has no intrinsic protein kinase activity, but recruits a tyrosine kinase (JAK).JAK phosphorylates STAT, which dimerizes, goes to nucleus, and affects gene expression.
RTK Example 3: EPO Receptor
JAK also binds Grb2, initiating the MAPK cascade.
Types of signal transducers
Guanylyl Cyclases
When activated, these receptor enzymes convert GTP to the second messenger cGMP.
Guanylyl Cyclases
When activated, these receptor enzymes convert GTP to the second messenger cGMP.
Two types:Single- transmembrane pass receptorCytosolic NO receptor
Guanylyl Cyclases
The effects of cGMP are often mediated by protein kinase G (cGMP-dependent protein kinase).
cGMP is degraded by a specific phosphodiesterase.
Case Study
A 56-year-old man presents to the emergency department complaining of substernal chest pain described as "something very heavy on my chest." The pain started while carrying boxes up a flight of stairs. Originally, the pain was a 9 out of 10 but after rest is now a 4 out of 10. The pain radiates to his left shoulder and is accompanied by shortness of breath. The patient states that he's had similar but less severe pain in the recent past with exertion. He wasn't going to come to the ED ("I told her it was just indigestion”), but his wife, concerned about a heart attack, made him. He has hypertension, hypercholesterolemia, and smokes a pack of cigarettes a day.
The ED nurse gives him nitroglycerine tablets, 0.4 mg sublingual (under his tongue), every 5 minutes three times. She also gives him chewable aspirin (324 mg), oxygen by nasal cannula, nitroglycerine paste on his skin, and obtains an EKG. The EKG demonstrates changes consistent with ischemia (low oxygen). Ten minutes later when the ED physician sees him, his pain is nearly gone at a 1 out of 10. Blood work, including tests for cardiac enzymes, is obtained.
The results of the blood work are normal, but due to concerns of angina the patient is admitted to the hospital. Further blood work rules out a myocardial infarction, and a stress test reveals reversible cardiac ischemia. He is diagnosed with unstable angina and is prescribed nitroglycerine to take as needed, a beta-blocker, daily aspirin, an exercise regimen, and told to stop smoking. He is scheduled for a cardiac angiogram and sent home.
Enhancing cGMP
NO
Vasodilation
Case Study
Researchers at Pfizer realized that a phosphodiesterase (PDE) inhibitor could be a therapeutic option for treatment of angina. Screening of existing compound collections resulted in several lead compounds that acted as potent inhibitors of PDE-5, a cGMP-specific phosphodiesterase in coronary smooth muscle. Further optimization lead to a single candidate, which underwent clinical trials but proved to be ineffective for angina.
During these trials, several patients noted experiencing enhanced penile erections. Subsequently, PDE-5 was identified as the main cGMP-degrading enzyme in the corpus cavernosm. Researchers at Pfizer then reshifted their focus towards developing a drug for erectile dysfunction. Sildenafil (Viagra) was approved by the FDA in 1998. In 2013, Viagra generated $1.88 billion of revenue.