PSIO 404 Exam 2 Flashcards
Very briefly summarize signal transduction: receptor-ligand binding, signal transduction by second messengers, and cellular responses as rapid or long-term.
Cell data processing happens is three steps, step 1 is Receptor-ligand binding which is the binding of a ligand, or primary messenger, that binds to a receptor at the cells surface membrane and then step 2, signal transduction by second messengers which carry on the message in a different form inside the cell. Lastly is step 3, cellular responses which can be rapid or long term or both. Rapid responses happen immediately and they help make rapid decisions through pre programed actions. Long term responses help adjust the data processing network that lead to an accumulation of changes that can lead to a change in gene expression.
Describe in general how receptors are controlled by allosteric regulation.
As ligand bind to receptors, receptors will alter the conformations.
When the correct ligand binds the conformational change will lead to the activation of signaling activities. This is allosteric regulation. With allosteric regulation, there are ligands that push the receptors to be active and there are also ligands that push the reports to be less active. An example is receptor tyrosine kinases.
Very briefly discuss three uses for post-translational modifications of receptors.
Post translational modifications of receptors is the cell control the allosteric regulation of its receptor. It can do this three ways
1. Control and fine tuning of signaling activity, which is control the the amount of signaling activity the receptors get
2. oligomerization (noise filtering, forming complexes, etc. ), which is make it easier or more difficult to form signaling complexes.
3. Communication with other proteins via interaction domain, which is creating interaction partners for interaction domains.
Discuss the structure and signaling protein functions of receptors.
They have three primary regions
1. discriminator domain with ligand binding site, which determines whether receptor should become activated or not
2. transmembrane domain which reaches from the outside of the cell to the inside of the cell and connect the discomintor and effector domain
3. effector domain which mediates the effects of an activated receptor.
The functions require energy input and require linked to switching reactions
-These are used to transform external input like endogenous stimuli (hormones) or environmental stimuli (ligh, touch) into intracellular signals
Explain the versatility of receptors, in terms of discriminator and effector domains, for input/output signals and activation/deactivation
When the right input/signal binds to the discominator domain this will lead to a conformational change that alters the shape of the effector domain to interact with downstream elements that lead to output signals.
-this interaction has to be highly specific, non covalent, and structural complementarity
Define agonist and antagonist in terms of interactions with receptors.
agonist = is the proper ligans that accomplishes the same effects that favor the active conformation of a receptor (allosteric equilibrium) so there are chemical outputs
antagonist = these favor the inactive form of the receptor or counter effect the natural effects of agonist, so that there is no chemical output
Explain how various cell types respond differently to same stimulus.
a particular signal is going to give you a particular reaction thank to the receptor it binds to.
It might give you a common reaction that you see with other ligands binding to other receptor, but a pathway initiated by one receptor versus a different receptor will still give you a difference in ultimate influence over the cell.
List and very briefly describe the three general regions of a prototypical receptor for hydrophilic ligands
- discriminator domains which contain binding sites for ligands
- transmembrane domains which physically connect the external receptors regions with the internal receptor regions
3, effector domains which are coupled with switching domains
List and very briefly describe the three most abundant membrane receptor types, including an explanation of the source of energy which powers each type.
- protein kinase coupled receptors - the most common for plants second most common for animals, and the energy proved for these switch is from ATP for the phosphorylation Switch
- G-protein coupled receptors- the most common in animal cells, and the energy provided for these switch is from GTPase hydrolysis switch
- ion channel coupled receptor - the energy is provided by membrane potential discharge from the primary active transport that is powered by ATP
In addition to the three most abundant membrane receptor types, list and very briefly describe the function of three additional receptor types, including an explanation of the source of energy which powers each additional type
- Guanylate cyclase - coupled receptor - which when activated catalyzed production of cGMP and the energy is supplied by GTP
- Protease - coupled receptors - which when activated release peptide second messengers and the energy is supplied by protein hydrolysis / ATP (very expensive but effective)
- E3 ubiquitin ligase coupled receptor - which when activated trigger the degradation of signaling proteins and the energy is supplied by ATP dependent activation of ubiquitin by ubiquitin activating enzyme E1
Discuss the usefulness of considering combinations of receptor-triggered switching events when a cell processes information
the combination of receptor - triggered switching events rather than the specific number of them, which determines successful performance of the cell’s signaling processing network
because there are 4 million receptors per cell, if the have a combination of receptor triggered switching events, these is an infinite amount of possible recognition and this allows the cell to be very specific witch what influences it.
Discuss, in general, how signaling pathways are investigated.
Signaling pathways are investigated by artificially dissecting them into linear cascades and consistently activate a mutant protein that activates 1 protein in the pathway and see what else it activates or upregulates or what it does not and look at the downstream effects.
List the methods used to determine interaction & cellular localization.
determining interaction you can use co -immunoprecipitation to develop an antibody to a particular protein and run it through a column and see what sticks to the protein to see what is capable of interacting with it.
For localization you can use immunohistochemistry which you label the particular protein with an antibody with fluorophore and look at it on the slide of the dead cells and see what part of the cell lights up.
To determine interaction:
- co-immunoprecipitation
- yeast two-hybrid system
- fluorescence resonance energy transfer (FRET)
To determine localization:
- cell fractionation
- immunohistochemistry
- GFP tagging
Discuss the twofold goal of investigations into cellular signaling.
- to understand in detail the molecular mechanism of a given cellular signaling process
- to understand in detail the biological significance of a given cellular signaling process
Identify the model organisms which are best for studying human cellular signal processing as it applies to human physiology and human disease, and explain why that is the case.
- yeast - allow us to look at basic signaling mechanisms because they are the simplest eukaryotic
- slime mold - allows us to study multicellularity, chemotaxis, aggregation of cells, and how cells move
- nematode - differentiation and development of cells to study cell proliferations and programmed cell death
- fly - short generation time and used cytogenetic and molecular genetic studies
- plant - easily bred used for genetics, not useful for modeling signal pathways in humans
- frogs - gene expression alterations through MAP kinases same for mice / rats
- human cell cultures - behave differently than intake tissue but used for cell signaling
Briefly discuss the contributions of Carl and Gerty Cori and Bernardo Houssay which were recognized by the 1947 Nobel Prize in Physiology or Medicine
because they found the discovery in glycogenolysis pathway
Describe the post-translational modification of glycogen phosphorylase-a which is responsible for its activation. Which two researchers discovered this modification
fischer and krebs found that form a was phosphorylated at a specific Ser reside in a reversible reaction by glycogen phosphorylase kinase
Describe the two crucial observations made by Earl Sutherland which enabled him to study hormone action in vitro, and briefly discuss his subsequent discovery of second messengers
- glycogen phosphorylase exists in two forms: active and in active b
- found epinephrine and glucagon worked on tissue homogenate not just intact tissue and for the first time hormones could be studies in vitro
-found that magnesium dependent enzyme was stimulated in the presences of epinephrine (the first messenger) and produced a low molecular weight product of ATP (the second messenger) which stimulates the enzymes of glycogenolysis (the was determined to be cAMP)
Describe the two questions which led researchers to the discovery of G- proteins
- how does epinephrine receptor active adenylate cyclase?
-adenylate cyclase stimulation required ATP and GTP which shows the GTP-hydrolyzing enzyme was an interconnecting molecule between receptor and adenylate cyclase - How does cAMP stimulate the phosphorylation of glycogen phosphorylase?
-cAMP-dependent protein kinase A (PKA)
Explain the signaling effects of epinephrine (adrenaline) which lead to a change in glycogen phosphorylase activity
epinephrine binds the adrenergic receptor (GPCR) which catalyses the exchange of old GTP for brand new GTP and this change leads to structural alteration of alpha subunit which enable it to interact with effects such as adenylyl cyclase and produces cyclic AMP and leads to the activation of PKA which then phoporlates phophorylase kinase adn that phosphorylates and activates glycogen phosphorylase
Summarize the structure and function of trimeric G-proteins: include subunits, switching, and regulators of switching.
subunits :
Ga = contains GTase region ,
Gb = has lipid anchor and is alway links to Gy adn Ga
switching = GTPase is Ga and it mediates G-protein signaling
Gby subunits also mediate signals
regulators =
List the major effectors of Gα subunits
- adenylate cyclase (AC), Gi,0, Golf, , Gs
- phosphatidylinositol (PI) specific phospholipase C (PLC) type ,beta, Gq,11
- cGMP-specific phosphodiesterase (PDE6) in retinal cells , Gt, Ggust,
- the GEF of the small G-protein Rho, G12,13
List the major effectors of Gβγ subunits.
- adenylate cyclases (ACs) , Gt, Gi,o
- phospholipase C (PLC) type beta, Gs
- phosphatidylinositol 3-kinase (Pi3K) type gamma (PIPs), Golf
- beta-adrenergic receptor kinase (BARK), Gq,11
Describe the 4 major subfamilies of trimeric G-proteins, noting any bias towards overall promotion of activation vs. inhibition, and noting which subfamily is the most abundant
- Gs family (includes Golf) where s = stimulatory (cholera)
- Gi,0 (includes Gt and Ggust) where i,0 = inhibitory with 0 referring to a subfamily in the brain (pertussis) (MOST ABUNDANT FAMILY)
- Gq,11 family
- G12,13 family
Discuss the regulation of trimeric G-proteins by AGSs.
Regulated by three families that alternatively activated G protiens
1. AGS family = true GEF’s which resemble GPCRs, do not need signaling molecule
2. AGS family 2 = GDP - dissociation inhibitor for Ga which block reassociation of Gby to stimulate Gby-controlled signaling, these proteins take the place og Gby adn set free Gby so only get the effects of Gby
3, AGS family 3 = form complexes with Gby and have unknown function, maybe a way to trap adn shield Gby that are set free for a window of time allowing Ga to be stimulated
Discuss the regulation of trimeric G-proteins by effector proteins
Proteins like PKA can encourage the conversion, the hydrolysis of GTP and alpha subunits which leads to a negative feedback loop
Discuss the regulation of trimeric G-proteins by RGS proteins.
In order to combat the auto- inactivation of Ga from being to slow, RGS proteins are promotion proteins which quickly hydrolyze GTP
-are heterogenous with only RGS domain in common
-have interaction domains for crosstalk
-they shorten signaling by direct stimulation of GTPase activity
Discuss the effects of cholera toxin on G-protein signaling
-is a disease caused by the disturbance of G protein signaling
-the toxin cholera gives off leads to major and deadly symptom of extreme loss of fluid across the intestinal wall
-it is an oligomeric protein that binds to the intestinal membrane that’s inserts its catalytic subunits into the target cell
- the cholera toxin injects its A subunits into our cells and the A subunit is a ADP ribosylation cataylis ARg of Gs, which turn the GTPase hydrolysis to be constitutively active which causes all of the PKA active with leads to a over distribution of CFTR which causes Cl- to to exit the intestines which leads to the exist of sodium and water follow sodium
Discuss the effects of pertussis toxin on G-protein signaling.
pertussis toxin is an ADP-ribosyltransferase with one catalytic A subunit and 5 membrane binding B subunits
-targets a Cys residue in Gai0 subunits disturbing its association with the receptor and suppressing Gio effects
-At the ER, the B subunits bind to ATP and dissociates from A
-A attaches to exosomes and heads to the membrane where ADP-ribosylation the alpha subunit of heterotrimeric G proteins, causing the G protein to be unable to be activated which now can no longer inhibit cAMP production causing cAMP levels to elevate
Consider the situation in which just 30 of the total number human receptor genes are being used by a cell (we can assume just 1 isoform for each). If each receptor recognizes a different signal, how many combinations of signals would the cell be able to identify?
30^2 (which equals 900)
2^30 (which equals 1,073,741,824)
2 x 30 (which equals 60)
an infinite number
2^30 (which equals 1,073,741,824)
Which of the following types of membrane receptors has seven transmembrane domains and is the largest receptor family in animals?
GTPase-coupled receptors
Ion channel-coupled receptors
Protease-coupled receptors
Guanylate cyclase-coupled receptors
Protein kinase-coupled receptors
GTPase-coupled receptors
Which of the following alters allosteric equilibrium in favor of the active state for receptors?
agonists
antagonists
discriminators
effectors
agonists
This organism possesses about 6000 genes, is useful for investigating basic signaling mechanisms, and is unsuited for investigating signals used in the process of tissue formation.
Saccharomyces cerevisiae
This organism forms aggregates of up to 100,000 cells, are best suited for studies of signaling in chemotaxis, and is said to be an organism standing at the border between uni- and multicellularity
Dictyostelium discoideum
This organism possesses about 19,000 genes, is a standard model for studies on cell proliferation and programmed cell death, and is composed of exactly 959 somatic cells
Caenorhabditis elegans
This organism is by far the best investigated higher invertebrate, and in regards to signal transduction exhibits striking similarities to mammalian cells.
Drosophila melanogaster
The human genome harbors approximately ____ genes encoding putative receptor proteins
1500
For E3 ubiquitin ligase-coupled receptors, what supplies the energy for the initial step(s) of data processing?
ATP
GTP
PEP
NADPH
ATP
In almost all cases, receptor tyrosine kinases (RTKs) must form a dimer after binding a ligand in order to initiate signaling. This is in actuality a way to
amplify signals.
filter out noise.
detect combinations of signaling reactions.
avoid the effects of agonists.
filter out noise.
As we learned in module 1, the amino acid which is required for GTP hydrolysis by small G-proteins (and is provided by GAPs for small G-proteins) is
Arg
Glu
Cys
Asp
Arg
Which of the following enzymes is activated by the beta-gamma subunits of G-proteins of the GS subfamily?
β-adrenergic receptor kinase
cGMP-specific phosphodiesterase (PDE6)
phospholipase C (PLC), type β
adenylate cyclase (AC)
phospholipase C (PLC), type β
The members of this G-Protein family are inhibited by pertussis toxin.
G12,13 family
Gs family
Gq,11 family
Gi,0 family
Gi,0 family
