WEEK 2 Flashcards
describe neurotransmitters
rely on diffusion
made and released from neurons
used for rapid communication over short distances
effects are short lived
describe hormones
made and released from endocrine cells
act over long distances
take a long time to act and effects are long lasting
describe drugs
exogenous substances
used to mimic (agonist), increase or block (antagonist) effects of first messengers
name the interactions that occur in receptor-ligand interactions
- hydrophobic force
- van der Waals
- hydrogen bonding
- electrostatic interactions
what does it mean when Kd is high
the dissociation constant is high when the receptor and ligand have a low affinity for each other
what does it mean when Ka is high
the association constant is high when the receptor and ligand have a high affinity for each other
Ka (or Kf) equation
[RL] / [R] *[L]
what is pharmacokinetics
the study of the time course of drug absorption, distribution, metabolism and excretion
what is pharmacodynamics
the relationship between drug concentration at the site of action and the resulting effect
what is the relationship between EC50 and drug potency?
the lower the EC50 the more potent the drug.
meaning less of the drug is required to produce a meaningful effect
what do agonists do?
mimic the effect of the endogenous ligand’s activation of its receptor to produce a response
what do partial agonists do?
partial agonists bind and activate a receptor but do not elicit a full response
what do superagonists do? what is their relative efficacy?
they show a higher efficacy than full agonists. their efficacy is at 200%
what do antagonists do?
they bind to the endogenous ligand’s receptor and block the signal molecule from producing a biological response.
what do inverse agonists do?
they bind to the endogenous receptor and exert the opposite biological response.
examples of non-chemical signals
mechanical forces, light, temperature
receptors can be on the _____ and in the _____
cell surface ; intracellular space
what are the types of cell surface receptors
ligand gated ion channels
GPCR
Catalytic receptors
- Guanylyl cyclase, RTKs, extrinsic protein tyrosine kinase, RTSKs, and Receptor tyrosine phosphatase
types of intracellular receptors
nuclear and cytoplasmic
describe ligand gated ion channels
- aka ionotropic
- binding domain for a specific ligand is at the extracellular domain of the receptor
- binding event opens the ion channel
- two states are open and closed
- regulate ion flow across the plasma
describe G-protein coupled receptors
- aka metabotropic
- diverse signals and effects
- extracellular ligand binding domain
- intracellular binding domain for a G protein
- Seven transmembrane spanning domains
- No pore
- signal transducer by GTP binding proteins known as G proteins
how do catalytic receptors signal
through intrinsic enzyme activity or closely associated enzyme activity
describe catalytic receptors
- have an extracellular binding domain
- a single membrane spanning domain
- a catalytic domain
- endogenous agonists = diverse peptides and proteins
- ligand binding triggers dimerization
Guanylyl cyclase is also known as a
natriuretic peptide receptor
describe atrial natriuretic hormone
- ANP
- secreated from atrial myocytes
- synthesized by magnocellular neurons of the brain
- receptors are dimers
- activation increases guanylyl cyclase activity
largest subclasses of catalytic receptors are____ . Describe it.
- Receptor tyrosine kinases (RTKs)
- respond to diverse ligands (EGF, insulin, VEGF, etc. )
- play important roles in cell growth, differentiation, cell survival, and metabolism
- mutations in RTKs and aberrant activation of their intracellular signaling pathways have been linked to many diseases and cancers
- important targets for therapeutic effects
what is the mechanism of RTK activation
- when ligand binds to RTK, neighboring RTK dimerize
- dimerization activates the tyrosine kinase activity in RTKs through phosphorylation (autophosphorylation or cross-phosphorylation)
- phosphorylated receptor now serves as a site for activation and assembly of intracellular signaling proteins
Activated RTKs serve as docking sites for intracellular signaling proteins that contain _____ and attract ______
SRC homology 2 (SH2)
phosphotryosine binding (PTB) ;
intracellular signaling proteins and docking proteins
describe Ras in the Ras-MAPK pathway
- small, monomeric G protein
- anchored to the inner leaflet by covalent bound to lipid
- it’s activation is regulated by Guanine exchange and hydrolysis through GEF and GAP proteins
- its activation requires adaptor proteins called Grb2 and Sos
- Ras swaps GDP for GTP
- it’s activation, activates several downstream targets
What kind of domain does Grb2 in the Ras-MAPK pathway have
an SH2 domain
Sos in Ras-MAPK pathway is what type of protein?
GEF protein
Sos binds ____
Grb2 and Ras
what targets are activated by Ras activation
Raf kinase (activated by Ras)
MEK (activated by Raf-P)
MAP kinase (activated by MEK-P) and ERK
*SEE SLIDE 25
Why is MAP Kinase important in the Ras-MAPK pathway
- phosphorylated MAP kinase is a transcription factor that upregulated proteins important for cell proliferation
Ras hyperactivation is associated with what disease
cancer
Name an example of when the Ras-MAPK pathway is used
epidermal growth factor
cell migration and adhesion
JAK-STAT is what type of catalytic receptor
extrinsic protein tyrosine kinase
what is the use of JAK-STAT in the body
-hematopoiesis (formation of blood cellular components)
- tissue repair
- inflammation
- apoptosis
describe Janus Kinase (JAK)
- non-covalently associated with cytokine receptors
- doesn’t bind to the receptor until the receptor is activated by the ligand
- JAK autophosphorylates and adds phosphates to receptor
- recruits STAT
describe Signal transducer and activator of transcription (STAT)
- STAT binds to the phosphorylated tyrosine on the receptor
- once bound they leave and take the phosphate with it to dimerize with the other STAT
- JAK-STAT process continues as long as ligand is bound to the receptor
- this is the most rapid activation of transcription factors
- loss or mutation of JAK/STAT components is related to many human diseases
intracellular receptors can only interact with ligands that can _____ the plasma membrane
cross
what type of ligands interact with intracellular receptors
- lipophilic vitamins
- steroid hormones
- nitric oxide
Inactive intracellular receptors are bound to _____
heat shock protein 90
what are the three steps of signal transduction
- receive first messenger
- signal transduction
- response
- signals that effect cytoskeleton proteins produce what kind of effect
- signals that effect gene regulatory proteins produce what kind of effect
- signals that effect metabolic regulatory proteins produce what kind of effect
- altered cell shape or movement
- altered gene expression
- altered metabolism
who discovered second messengers, specifically cAMP
Earl sutherland & ted Rall
glycogen phosphorylase is the key enzyme in breaking down
glycogen. this reaction is called glycogenolysis
where does glycogen breakdown (or glycogenolysis) happen? and what is cleaved off from the glycogen polymer
in hepatocytes ; glucose 1-phosphate is cleaved from the polymer
types of second messengers include
hydrophobic molecules- cannot leave the membrane (DAG)
Hydrophilic molecules- can diffuse throughout the cytosol (cAMP, cGMP, IP3)
Ions (calcium)
Gases- can diffuse through aqueous and lipid settings (NO, CO)
Soluble proteins (JAK/STAT)
G-proteins are found in the genomes of _______
bacteria, yeast, plants, invertebrates, and all mammals
describe GPCRs
- is a monomeric proteins.
- a multi-pass transmembrane protein (7 spanning)
- has an extracellular binding domain
- the intracellular G-protein binding domain includes C3, C4, and sometimes C2
- GPCR activation actives G-proteins
G-proteins are ______ binding proteins and act as molecular switches
heterotrimeric guanine nucleotide
describe the subunits of G-proteins
- alpha subunit contains the nucleotide binding domain and has endogenous GTPase activity. (where GTP/GDP bind)
- the alpha and gamma subunits are covalently anchored to the membrane
- the beta and gamma subunits remain bound, alpha does not, and move as a unit
describe “off” state of G-protein
- attached to the GPCR at the inner membrane
- GDP is attached to Ga
describe ligand binding to GPCR
- helps release of GDP from Ga
- binds GTP; this causes conformational change that results in dissociation of Ga-GTP and GBy
describe the timing mechanism
the intrinsic GTPase activity of the Ga subunit means that the G protein can only be active for a short period of time.
- GTP is hydrolyzed to GDP returning Ga to its inactive state which is Ga-GDP
what happens when GBy reassociates with Ga-GDP
terminates all effector interactions
describe PKA
- major target of cAMP
- PK-A is a heterotetramer: it has 2 regulatory subunits, and 2 catalytic subunits
- catalytic subunits are phosphorylated by adding phosphate groups to serine and/or threonine residues
- C subunit can also diffuse to the nucleus where it phosphorylates transcription factors.
what happens when GPCR is coupled to Gai
- Gai-GTP inhibits adenylyl cyclase
- intracellular cAMP levels fall
- no longer sufficient cAMP to bind subunits of PK-A
- regulatory and catalytic subunits reassemble, inactivating PK-A
intracellular cAMP levels can be regulated by cAMP phosphodiesterase (PDE). True or False
True. PDEs mediate termination of cAMP and cGMP
what are the four major classes of second messengers
- cyclic nucleotides
- lipid messengers that signal w/in the cell membranes
- ions that signal within and between cellular compartments
- gases and free radicals
what types of first messengers produce second messenger cAMP
Neurotransmitters (epinephrine, acetylcholine)
hormones (ACTH, CRH, Glucagon, LH)
What is the signal transducer for cAMP
GPCR-Gs, GPCR-Gi
what is the primary and secondary effector of cAMP
primary: adenylyl cyclase
secondary: PKA, cyclic nucleotide gated ion channels
Viagra (Sildenafil citrate) is an example of
an cGMP PDE inhibitor. it increases cyclic nucleotides, specifically cGMP.
- it helps treat erectile dysfunction
How is cAMP regulated
- stimulating and inhibiting adenylyl cyclase
- cAMP PDE
PKA ______ cAMP phosphodiesterase
activates
intracellular cAMP concentration levels regulate _____ activity
PKA
active phosphoprotein phosphatase 1 (PP1) vs. inactive PP1? what inactivates PP1
active PP1 dephosphorylates activated proteins
inactive PP1 does not.
PP1 is inactivated by phosphoprotein phosphatase inhibitor (I-1)
glycogen breakdown in skeletal muscle: what do the catalytic subunits of active PKA do?
once cAMP binds to PKA, activating it the catalytic subunits help:
- activate phosphorylase kinase (PK); this activates glycogen phosphorylase allowing for glycogen breakdown
- inactivates glycogen synthase; this inhibits the build up of glycogen
- inhibits PP1, by phosphorylating I-1 which binds to PP1
How does the cholera toxin protein work
- cholera passes through gastric acid barrier and colonizes the small intestine
- the A subunit of Cholera Toxin (CT) bind to the G-protein causing ADP-ribosylation of arginine residues
- Gs is locked in its active form
- Na+/H+ exchanger (NHE) is blocked, prohibiting Na from entering
- in addition to the build up of Na outside the cell, Cl-, HCO3-, and H2O remains on the outside
- cells dehydrate
- body dehydrates, diarrhea, death
how does the Pertussis toxin (PTX) work (whooping cough)
- the “A protomer” from PTX and NAD+ uncouples the Ga-inhibitory from the GPCR
- this increase adenylyl cyclase activity.
- cAMP levels rise and accumulate
- cascade continues to be stimulated b/c of the high levels of intracellular cAMP
cGMP has a ______ nucleobase, whereas cAMP has a ______ nucleobase.
guanine ; adenine
cGMP is derived from _______ triphosphate
guanosine. (GTP)
the two primary pathways for cGMP synthesis (hint: receptors)
- membrane bound guanylyl cyclase
(this is a natriuretic peptide receptor; a subdivision of catalytic receptors) - soluble guanylyl cyclase via Nitric Oxide activation
How is cGMP degraded
cGMP PDEs:
- specific cGMP PDEs, such as PDE-5A
- non-specific PDEs, such as PDE-2 and PDE-3 which can degrade cAMP and cGMP
PDE-5A is a _____
cGMP specific PDE
PDE-2 and PDE-3 can hydrolyze
cGMP and cAMP (into GMP and AMP respectively)
cGMP activates cGMP ion gated channels and which protein kinase?
Protein Kinase G
PKG (protein kinase G) is expressed in which areas
- brain
- lungs,
- vascular tissue
where are cGMP gated ion channels found in the body?
in photoreceptors which are important for phototransduction (eyes).
histamine is ligand that can activate _____
phospholipase A2
Phospholipase A2 hydrolyzes ________ to produce Arachidonic acid
membrane phospholipids
Arachidonic acid is a precursor to which pathways
cyclooxygenase pathway and lipoxygenase pathway
the cyclooxygenase pathway can be blocked by what?
Aspirin and NSAIDs
Why does aspirin have anti-inflammatory effects and anticoagulant effects
Aspirin blocks the cyclooxygenase pathway, inhibiting the production of prostaglandins which cause inflammation and vascular effects and thromboxanes which cause blood clotting and other vascular effects
the Lipoxygenase pathway produces _____ which cause ________ and _______
Leukotriennes; allergic ; inflammatory reactions
Phospholipase C-β is activated by ______
GPCRs
Phospholipase C-γ is activated by ______
tyrosine kinase
Name examples of phospholipase C-β hormonal ligands
Hormones: angiotensin, GnRH, GHRH, Oxytocin, TRH
name the secondary messengers and effectors of phospholipase C-β
secondary messengers: IP3; DAG; Ca2+
Secondary effector: Protein kinase C and calmodulin
IP3 stimulates the release of Ca2+ from ______
the SER (Smooth ER) and the Mitochondria
DAG is a derived from _____ and stimulates _____
PIP2 ; PK-C (protein kinase C)
PLC hydrolyzes PIP2 to produce DAG and IP3. True or False
True.
DAG diffuses through the cytosol. True or false
False. DAG is a hydrophobic second messenger and therefore remains in the membrane while IP3 is hydrophobic and diffuses through the cytosol.
what does the SERCA do in the Smooth ER
it pumps Ca2+ back into the SE.
- it is an anti-porter. it utilizes ATP to do this
IP3 is bi-phasic. True or False
False. DAG is bi-phasic.
Explain what the second wave of DAG comes from.
- the slow DAG wave is caused by the breakdown of Phosphatidylcholine (PC) by PLCβ and PLD
- the first wave is caused the DAG released from the hydrolysis of PIP2 by PLCβ
What are Protein Kinase C functions (HINT: S.S.I.R.R L)
Smooth muscle contraction
Secretion
Immune responses
Receptor desensitization
Regulating transcription and Cell growth
Learning and memory
name the three types of functional protein kinases C
conventional , novel, and atypical
describe the main differences between conventional, novel, and atypical PKCs
conventional: are activated by Ca2+ and DAG
novel: only activated by DAG
atypical: not activated by either Ca2+ or DAG
What is an extracellular and intracellular source of calcium?
extracellular: calcium from the ECF can be transported the ICF through calcium channels
intracellular: calcium stores in the SER and mitochondria can be released into the cytoplasm
How can calcium be removed from the ICF
- pumps: Ca-ATPase and Ca-Na exchangers can move calcium out of the cell or into the ER
How can calcium concentration be buffered in the ICF
- use of binding proteins such as parvalbumin and calbindin
What is the glucose equivalent of 1 epinephrine molecule
10^8 Glucose 1-phosphate molecules
What is an example of first messengers interacting with different receptors thus producing different responses in different cells
Acetylcholine (ACH):
1. binds ionotropic/nicotinic in skeletal muscle = muscle contraction
- binds metabotropic/GPCR in heart muscle = reduces rate and force of muscle contraction
What is the name of a molecule that is small, highly diffusible, a non-protein organic molecules or ions that have very specific target proteins they regulate by altering their targets’ activity.
second messengers
why are second messengers not rate limited? and what is their effect because of this
- they are produced from abundant precursor molecules or rich intracellular stores
- b/c they are not rate limited they can provide rapid and efficient signaling that is temporarily precise
True or False:
1. compared to proteins, second messengers are tightly and rapidly controlled. True or False.
- Second messengers are not spatially controlled
- True
- False. Second messengers are spatially controlled
Convergence vs. Divergence
Convergence - several different second messenger pathways can converge on a single target effector, producing an amplified effect.
divergence-
1. describe the different effects of a single first messenger on different cell types.
2. It can also be used to refer to diffusible intracellular second messengers that can also target different downstream effectors leading to a single first messenger having divergent effects.
what is the time scale for diffusion and opening ion channels
10 um/1-10 seconds; milliseconds
what is the time scale for Mammalian transcription?
10-100 nucleotides/sec or 10 min/gene
what is the time scale for Mammalian translation?
10 amino acids/sec or 1 min/protein (300 aa)
categorize mTOR in terms of kind of receptor
Catalytic receptor: RTSKs (receptor serine/threonine protein kinase)
what is the catalytic subunit of mTOR complex 1 & 2
mTOR
Fill in:
mTOR stands for ________________
mTOR is the major regulator of ____________
mTOR Controls ___________ processes
mTOR is a member of the __________ family
- mammalian target of rapamycin
- growth in animals
- anabolic and catabolic
- PI3K-related kinase (PIKK)
True or False. mTOR Responds to nutrients and nutrient-induced signals
True. such as insulin
What is the difference between mTOR1 & mTOR2 subunits
mTOR1 has subunits Raptor and PRAS40
mTOR2 has subunits mSin1 and Rictor
compare mTOR1 and mTOR2
mTOR1:
- mTOR complex 1 promotes cell growth & proliferation
- Growth factors (insulin/insulin like), Energy status, Oxygen, Amino acids activate mTOR complex 1
- Inhibited by rapamycin
Promotes: protein synthesis, de novo lipid synthesis, synthesis of nucleotides needed for DNA replication and ribosome genesis, a shift away from oxidative phosphorylation to glycolysis, lysosome and mitochondria biogenesis
suppresses: protein catabolism (autophagy)
mTOR2:
- mTOR complex 2 controls cell survival, proliferation and regulates cell growth
- signals primarily come through insulin/PI3K signaling
Regulates: Lipogenesis, Glucose metabolism, the actin cytoskeleton, and Apoptosis (cell death)
insensitive to rapamycin
does fasting inhibit or activate mTOR1
inhibits
At high nutrient levels mTOR1 ________ catabolism and ________ anabolism
inhibits catabolism and activates anabolism
True or false. Dysregualtion of mTOR is associated with negative physiological outcomes
True.
select the right answer.
mTOR effects are ______
A. gene-specific
B. ligand-specific
C. nutrients-specific
D. cell-specific
D. cell-specific
When is mTORC1 activity optimal?
at caloric restriction
explain chronic mTORC1inhibition and activation
